Abstracts of the Eleventh Annual Meeting of the European Society for Clinical Investigation, Rotterdam, The Netherlands, April 20–22, 1978

Abstracts of the Eleventh Annual Meeting of the European Society for Clinical Investigation, Rotterdam, The Netherlands, April 20-22, 1978

These abstracts have been arranged in order of presentation.

250. Role of the interferon in the nephritis due to lymphocytic choriomeningitis virus (LCM.V) P. Verroust, L. Morel-Maroger, Y. Riviere, J.C. Guillon, and I. Gresser. INSERM U64 Hbpital Tenon, Institut Pasteur, Paris, and IRSC, CNRS, Villejuif, France. It has been shown that the disease induced by neonatal infection with LCM.V was dependent - at least in part - from the early production of endogenous interferon (IF). The present work was designed to investigate the role of IF in the genesis of glomerulo- nephritis (ON) induced by neonatal infection with LCM.V. Patho- gen-free newborn Swiss-mice were inoculated with either sheep anti- interferon globulin (n = 75) or normal sheep gammaglobulin (control mice) (n = 70). Six hours later they were injected subcutaneously with 104 LDSO of LCM.V CIPV 76001 from the Pasteur Institute. IF was not detected in the sera of mice treated with anti-interferon globulin, whereas the titers were 1 5 8 and 1:74 in the LCM mice injected with control globulin. The serum virus titer at 3 days was higher in mice treated with anti-IF globulin (106.6 LDSO/ml) than in the sera of mice treated with control globulin (104.5 to 104.' LDSO/ml). Later on, at 65 and 125 days serum virus titers were identical in both groups. Glomerular lesions and immunoglobulin deposits were present in all control mice from day 15 and increased gradually in severity. By contrast renal lesions were not detectable at day 15 in anti-IF-treated mice, and were only present on a small number of mice until day 65 with a lower degree of severity. Later on, at days 125-195, the number of affected mice increased progressively as well as the severity of the disease. It is therefore suggested that the early production of IF may play a role in the induction of LCM.V-induced GN. Y. Rivikre, 1. Gresser, J.C. Guillon and M.G. Tovey: Inhibition by anti-interferon serum of lymphocytic choriomeningitis virus disease in suckling mice. Proc. Natl. Acad. Sci., USA, 1977,74,2135.

243. Hepatic T3 generation from T4 by rat and mouse liver homogenate: role of SH groups in fasting, hypothyroidism and neonatal life. A. Vagenakis, A. Harris, L. Hinerfeld, S. Fang, and L. Braverman, University of Massachusetts Medical School, Worcester, Massachusetts. We have previously reported that fasting and hypothyroidism (Tx) in the rat results in a decrease in in vitro hepatic T, generation from T, (H-T,. Studies were carried out to determine whether Sulfhydryl groups (SH) are important in H-T, during fasting, hypothyroidism and neonatal life. SH were measured in 25% liver homogenate. Non protein SH (NP-SH) were decreased during 2d of fasting (Fast, 1.14*0.04 Fed, 2.22*00.15mM p(O.Ool), associated with decreased H-T, (50% fed controls). Addition of 4mM Dithiothreitol (DTT) which increases SH groups restored the decreased H-T, observed in fasting. In vivo T, and T, replacement during fasting had no effect on the decreased H-T,. In the Tx rat and in the congenitally hypothyroid dwarf mouse (dw/dw), the NP-SH were also decreased (Tx 1.42i0.06 vs 2.2*0.15mM, p<O.001; dw/dw 1.3k0.03 vs 2.0+0.2mM, 6 0 . 0 1 ) and the H-T, was markedly decreased. DTT only minimally effected H-T, in these hypothyroid animals. T, replacement in the T x rat increased NP-SH by only 11% and H-T, by 800% and had no effect on NP-SH but increased H-T by 5oOOT'o in the dw/dw. H-T, was markedly decreased in the fetal(F) and Id neonatal rat (N) as compared to pregnant and normal adults. NP-SH were decreased in F and Nand increased with age. DTT partially restored this decreased H-T, in F and N rats. Conclusions. The decreased H-T, during fasting is due to decreased SH and not to enzyme deficiency. In hypothyroidism, decreased H-T, is probably due to enzyme deficiency since it is not effected by increased SH groups but is restored by T, and T, administration probably by newly synthesized deiodinating enzyme. In F and N rats, decreased H-T, is probably due to decreased SH groups as well as decreased deiodinating enzyme concentrations.

34. Chloride binding by human hemoglobin: its physiological role as allosteric effector in normal and mutant hemoglobins. E. Bursaux, C. Poyart, B. Bohn, P. Guesnon INSERM U.27,92150 SURESNES. As other small organic or inorganic anions, chloride is a well known allosteric effector of human hemoglobins. Increasing concentrations of C1- lead to decreased 0, affinity, increased cooperativity and increased alkaline Bohr effect. C1- binds to positively charged residues of the f i chains and most likely to those groups responsible for the binding of phosphorylated organic anions as DPG or IHP. Recently cristallographic studies by Arnone have revealed an important binding site for small anions located between the N and C terminals of the a chains. We have searched evidence for the physiological importance of this binding site by studying the effect of CI- on the functional properties of purified Hb Suresnes (a, 141 Arg -+ His fid a new high 0, affinity variant where the N-C terminal region of the a chains is disrupted. Comparative studies of Hb Suresnes at different CI- showed an inhibition of the effect of Chloride on the 0, affinity, cooperativity and alkaline Bohr effect relative to those observed in HbA solutions. By contrast p chains bound effectors (DPG or IHP) have normal effects on the functional properties of Hb Suresnes. Comparison will be made with the effect of chloride on HbA a des Arg (HbCPB digested) where the C terminus in the a chains was removed. These results point out the major importance of chloride binding on the a chains in the understanding of the functional properties of hemoglobin within the erythrocytes. This work was supported by I.N.S.E.R.M. (C.L. 75.5.145.5 and ATP 41.76.73).

124. Instantaneous - Biphasic insulin elevation after exposure to visual and olphactory stimuli in obese and normal individuals. M. Krotkiewski, L. SjOstrOm & L. Sullivan; Sahlgren Hosp., GOteborg, Sweden. There is ample evidence that obesity is associated with the deviation of apetite regulation expressed as a higher sensivity to external cues. Insulin has been postulated as an important and primary factor in the development of obesity. The effects of visual and olphactory stimuli on the insulin secretion (cephalic insulin stimulation - C.I.S.) therefore were compared in obese and normal persons. Cephalic insulin stimulation - CIS was determined after fasting from 8 p.m. to 11.30 a.m. the following day. At this time a delicious meal was put in front of the patients for 5 min. Insulin levels were determined every two minutes for 20 min. The plotted surface representing increased insulin secretion was taken as an index for CIS. Cephalic insulin stimulation test was determined in 30 obese, weight stable patients and in 23 normal subjects. Cephalic induced insulin elevation was almost 10 times higher in obese subjects than in the normal individuals, that is: 31,6 * 8,3 versus 3,8 f 1,4 mU x 1-1 x min-1 - (means 2 SE; p < 0,005) in the obese and normal group respectively. Two peaks of insulin elevation were observed during the first eight min. after smelling and viewing the meal. When atropine was ad- ministrated (0,Ol mg/kg body weight subcutaneously) only the second peak of insulin elevation was observed. A neural-vagus mediated mechanism seems consistent with the observed atropine effect, while a hypothalamic humoral factor is postulated to be involved in the inducement of the second peak of insulin elevation.

91. Intrarenal control of urine concentration by anglotensin 11. J.L. Imbs, M. Schmidt and J. Schwartz. Institute of Pharmacology and Experimental Medicine. Univ. Louis Pasteur. Strasbourg. France.

325

326 ABSTRACTS The intrarenal role of angiotensin I1 in the regulation of water excretion was examined in pentobarbital anaesthetized dogs. The recovery of urinary concentration was followed for 6 hours after intravenous injection of frusemide (20 mg/kg). Dogs treated with the diuretic in conjunction with the intrarenal infusion (Ipg/kg/min) of the competitive antagonists [l-succinamoyl-5-valine-8-phenylgIy- cine] - angiotensin I1 (Hoe 409) or [I-sarcosine-S-valine-8-alanine) -angiotensin I1 (saralasine), were compared with control dogs receiving frusemide alone. Water excretion in the angiotensin-antagonist-treated dogs was higher during the 6 h after the diuretic injection : at the 6th h after frusemide injection, urinary osmolarity in these dogs was 334 mOsm/kg (SD 25, n = 4, p < 0.05) as against 460 mOsm/kg (SD 88, n = 9) in the control dogs. There was no difference in renal blood flow and glomerular filtration rate between the two groups of animals. These data confirm prior observations* on the recovery of the urine concentration process after frusemide, which is delayed when renin secretion is inhibited with propanolol. We suggest that angiotensin 11 controls the vasa recta blood flow by means of its constrictive action on the efferent arteriole of the jux- ta-medullary glomeruli, and thus plays a role in the maintenance of the corticopapillary concentration gradient. *Clin. Sci. Mol. Med. 52: 171-182, 1977.

17. Age and pressure related increase in plasma noradrenaline and decreasing beta-adrenoceptor responsiveness in essential hypertension 0. Bertel and F.R. Biihler, Department of Medicine, University of Basel, Basel, Switzerland The role of the sympathetic nervous system in patients with essential hypertension was studied by measuring plasma noradrenaline (PNA) at rest and during sympathetic stimulation with equieffective submaximal ergometry. Responses in PNA were related to blood pressure, heart rate as well as to a test assessing beta-adrenoceptor sensitivity, i.e. the chronotropic dose of isoproterenol which increased heart rate by 25 beats/min (CD25). Resting PNA increased with age in normal subjects (n = 18, r = 0.66, p < 0.05). This age effect was obscured in 56 hypertensives by young patients exhibiting supranormal PNA concentrations. Dynamic studies in 24 hypertensives revealed a 4-fold rise in PNA with submaximal exercise from 308 & 55 SEM pg/ml to 1393f117 (p < 0.001): this rise was only to 1092+129 in the 7 patients below 35 years of age, to 1326rt164 in the 9 between 35 to 50 years, but to 1731f241 in the 8 above 50 years (p < 0.05). In contrast, older age was associated with a reduction in both exercise tachycardia (r = 0.73, p < 0.001) and isoproterenol sensitivity: i.e. CD25 was 1.20+0.47 pg/mz in the younger, 2.4250.91 in the middle aged and 6.73k6.90 in the older patient group. In turn, the exercise- induced response in blood pressure increased with older age. Thus, adrenergic nerve activity tends to be supranormal in younger patient or in the onset phase of essential hypertension. The relatively higher PNA concentrations found in the older patients enhance alpha-adrenoceptor mediated vasoconstriction consequent to reduced beta-adrenoceptor sensitivity and may thereby contribute to the transition from a hyperadrenergic form to a high resistance type of hypertension.

69. Change in haemudynamic effect of propanolol by salt depletion in hypertension. G.G. Geyskes, P. Boer, G.J. M. van Rooyen, J.C. Roos and E. J. Dorhout Mees. Department of Nephrology and Hypertension, University Hospital, Utrecht, The Netherlands. Studies were performed to wether salt depletion alteres the hypotensive mechanism of propanolol in patients with hypertension. Blood pressure (BP) was recorded by arteriosonde@ , cardiac output (CO) by impedance cardiography, and plasma renin activity (PRA) by a modified Haber method. In 11 patients short time salt depletion caused a decrease in CO of 22 Qo and an increase in calculated periferal resistance (TPR) of I 1 Qo. After 4 days additional propanolol treatment (320 mglday), CO increased by 1 1 Qo (p < 0.05) and TPR decreased 25 Qo (p < 0.001) reaching a level slightly below that before salt depletion. In 19 outpatients on chronic chlorthalidon treatment addition of propranolol in increasing dosages till optimal BP control was ar- chieved (80-640 mg/day) resulted in similar changes: CO increased by 15 Qo (p < 0.02) while TPR decreased by 18 '70 (p < 0.02). In both studies changes in TPR were significantly correlated with changes in PRA. In 4 patients the effect of saralasin@ was studied before and after salt depletion. There was a significant correlation between the

changes in TPR induced by saralasin and by subsequent propanolol treatment. The results suggest that lowering of the PRA by propranolol results in a decrease in the TPR. After salt depletion this part of the hypotensive action of the drug prevails over its depressive action on CO. The resulting improvement of the circulations supports the use of combined saliuretic beta-blocking therapy.

55. Effects of diffusion on blood pressure during hernodialysis in anephric conscious dogs. S. Draibe, N.K. Man, J.F. Liard and J.P. Grlinfeld. Clinique Ntphrologique, HBpital Necker, Paris, France. Previous studies suggested that hemodialysis-induced hypotension in patients on regular dialysis treatment was mainly due to decreased blood volume and/or autonomic insufficiency. The effects of solute mass transfer on blood pressure have not been systematically evaluated. We have therefore attempted to evaluate the effects of the solute changes obtained by high (strategy A) and low (strategy B) mass transfer hemodialysis on the blood pressure of 6 anephric conscious dogs during a 90 min period of a constant body-weight hemodialysis (diffusion process only). Dialysate composition was : Na: 147.0, K: 2.0, Ca: 3.5, Mg: 1,5, CI: 116 and acetate: 38 mEq/l; glucose: 2.0 g/l and osmolality: 290 mOsm/kg. Mean blood pressure (MBP) and heart rate (HR) were measured a t 15 min intervals whereas osmolality, plasma proteins, electrolytes, urea and creatinine concentrations were measured at 30 min intervals. The percent of change between initial and final values in the two strategies were:

urea creat osm prot Na K Ca HC03 C1 A -45 -42 -7.3 -6.1 -1.14 -46.3 -2.1 +27.7 +1.0 B -27 -28 -4.9 -7.4 0 -27.3 -6.0 +17.5 +3.5 MBP dropped dramatically in strategy A : from 101.3 -t 2.9 to 77.3 f 6.2 mmHg (+ SEM); and slightly in strategy B : from 102.8 f 4.1 to 96.5 rt 3.2 mmHg (A vs B, p < 0.005). HR increased markedly in both strategies from 71.0 2 2.4 to 131.0 f 8.0 and from 73.5 f 2.8 to 124.5 2 13.1 beatslmin respectively. Neither the changes in plasma volume nor the changes in plasma electrolytes or osmolality can explain the greater fall in blood pressure observed in strategy A. Our data suggest that solute transfer of other substances such as acetate or vasoactive metabolites should be taken into account to explain our results.

151. The pathomechanism of accelerated hypertensive vascular disease. Z. Nemes, R. Dietz, J.F.E. Mann, L.B. Liith and F. Gross Department of Pharmacology, Univ. of Heidelberg, Heidelberg, Germany. In order to discriminate between the vascular consequences of direct vasoconstriction and those of increased blood pressure accelerated vascular disease has been produced in rats by angiotensin-induced primary vasoconstriction and also by primary mechanical stress imposed on the renal vascular bed by releasing the Goldblatt clamp. In angiotensin infusion experiments the incidence and severity of vascular lesions did not correlate with the level of blood pressure elevation. The specific antagonist of angiotensin, saralasin/ P 113 / prevented the rise of blood pressure produced by angiotensin but did not alter the pattern of vascular lesions. Anaesthesia, however, without major reduction in the blood pressure, suppressed or mitigated the vascular disease in the mesenterial vascular bed and completely prevented the renal vascular diesease. In conscious rats the severity of vascular lesions in the declamped kidneys correlated with the calculated rise in the renal perfusion pressure. The lesions were generally destructive. No vascular lesions occurred after the removal of the Goldblatt clamp in the anaesthetized rats. When the blood pressure of hypertensive rats has been normalized by sodium nitro-prusside infusion prior to declamping severe destructive renal vascular lesions developed in the declamped kidneys both in conscious as well as in anaesthetized rats. It has been concluded that the destructive vascular lesions occur at a critical wall tension according to Laplace's law. The smooth muscle cell necrosis in non- destructive lesions is produced by the sustained excessive spasm of smooth muscle cells caused either by autoregulatory vasoconstriction in severe hypertension or by vasopressor agents like angiotensin.

137. Gap junctions of pancreatic &cells increase during glibenclamiae stimulated insulin release.

ABSTRACTS 327 affected by starvation than that of B-cells, an effect not due to the increase in ketones during starvation. The low secretory activity of B-cells may favor IRS-release. Alternatively, the comparatively high secretion of D-cells may cause low IRI-release.

plasma 3H HM 3H-i. LM

1494 10% 57% 33%

P. Meda, A. Perrelet and L. Orci. Institute of Histology and Embryology, Geneva, Switzerland. Gap junctions (g.j.) are plasma membrane differentiations allowing exchange of ions or small molecules between cells (ionic and metabolic coupling) and their presence between B-cells (and other endo- crine cells) of the islet of Langerhans suggested that coupling may be important for their secretory function. T o test this hypothesis, we assessed quantitatively the development of g.j. in resting and stimulating conditions of insulin release. Islets isolated from rats stimulated in vivo by glibenclamide were studied with freeze-fracture and compared to control, resting isolated islets. In stimulated B-cells, the total area of the plasma membrane occupied by g.j. is significantly (p < 0.05) higher than in controls (0.058 f 0.011 To and 0.023 _+ 0.008 % respectively). This is due to a moderate but not significant increase of g.j. frequency(l0.17 5 1.83 g.j./100 pm2 and 5.76 f 1.61 g.j./100 pm2 respectively) and to a significant (p < 0.005) increase of the g.j. mean size (0.0056 f 0.0002 pm2 and 0.0037 + O.OOO5 pm* respectively). The mean size augmentation results from the presence of more large g.j. (containing at least 100 particles) in stimulated than in resting B-cells (10.9 % and 3.7 % respectively). Our data suggest that gap junction development is influenced by the secretory activity of the B-cells and lend support to the hypothesis that coupling may play a role in this activity.

kidney liver 3H HM 3H-i. LM 3H HM 3H-i. LM 25237 11% 47% 41% 2232 41% 11% 48%

86. Effects of alloxan on insulin release and decrehe in B cells K permeability in response to various secretagogues. J.C. Henquin and A.E. Lambert. Unite de Diabtte et Croissance, University of Louvain, Brussels, Belgium. The mechanisms whereby Alloxan (A) damages pancreatic B cells are still largely unknown. A has been previously reported to inhibit insulin (IRI) release in vifro, to depolarize B cells and to inhibit 86Rb pumping. In the present studies, the effects of the drug on the changes of IRI release and 86Rb efflux (used as an index of B cells K permeability) in response to various agents were continuously monitored in perifused rat islets. Islets were exposed to 2mM A for 5 min before stimulation with glucose (G) or other agents. A introduction at G 3mM did not affect basal IRI release but produced a transient increase in 86Rb efflux. A completely suppressed the stimulation of IRI release by G l5mM while considerably delaying the reduction of 86Rb efflux normally produced by G 15mM. 3-0-Methyl-Glucose prevented the increase in 86Rb efflux rate produced by A and partially restored the effect of G l5mM on IRI release and 86Rb efflux. A was without effect on tolbutamide (T)- induced monophasic IRI release and immediate decrease in 8f'Rb efflux. The drug hardly affected the early secretory phase and the fall in 86Rb efflux induced by lOmM Glyceraldehyde (Gly) or Keto- isocaproic acid (KIC); the late phase of secretion was however markedly reduced after A treatment. These studies suggest that a correlation exists between the ability of alloxan to interfere with secretagogue-induced decrease in K permeability of B cells and to prevent immediate stimulation of IRI release. Furthermore, the drug more severely alters G effects on B cells than those of T, Gly or KIC.

187. Effect of fasting on somatostatin- and insulin-release by isolated rat pancreatic islets P. Schauder, C. McIntosh, R. Arnold, H. Frerichs Department of Medicine, Division of Gastroenterology and Meta- bolism. University of GOttingen. FRG Intra-islet cell-to-cell interaction for the secretion of somatostatin (IRS) and insulin(IR1) has been suggested by previous studies (FEBS Lett. 81,355,1977). We now studied the effect of 48 h fasting on islet content and release of IRS and IRI, and the effect of aceto- acetate, 3-OH-butyrate, acetone, hexanoate and capronate on these release processes. Collagenase-isolated islets were incubated (10 islets; 500 ul KRB- buffer; 45 min) or perifused (250 islets; flow rate 0.9 ml/min; 90 min). IRS and IRI were measured by specific RIA. Fasting decreased the IRS content, but not the IRI content of the islets. Basal (2 mM glucose) release of IRS and IRI was normal. Release of IRI, stimulated by 3.3 or 10 mM glucose, was diminished but IRS release was not impaired. However, islets preperifused for 45 min with 2 mM glucose released less IRS on subsequent stimulation with 25 mM glucose than islets from fed rats. Ketone bodies and FA had no effect on IRS- and IRI-release. It is suggested that no strong correlation exists between IRS-content and IRS-release. The D-cell's secretory mechanism appears less

19. Distribution and metabolism of intravenously injected (i.v.i.) insulin. P.A. Halban, R.E. Offord, G. Davies and M. Berger. Institut de Biochimie Clinique, University of Geneva, Switzerland, and Labo- ratory of Molecular Blophysics, University of Oxford, UK. A novel radioactively labelled insulin, semisynthetic 3H-insulin (3H-i.), was used to follow the pharmacokinetics of i.v.i. insulin in rats. A neutral solution of 3H-i. (ca. 0.02 U/kg b.wt.) was injected into the tail vein of fed rats; the animals were decapitated after various time intervals and 3H levels were determined in plasma and SDS-solubilized extracts of liver and kidney. To measure 3H-i.-levels. plasma samples were chromatographed on GSo Sephadex and organ extracts on G75 which enabled us to separate intact labelled insulin from its degradation products of low (LM) and of high molecular weight (HM). Based on 4-8 separate determinations each, at 9 time points from 0.5 to 60 min after i.v.i. the metabolic clearance rate was 23.60 ml/min/kg and the apparent distribution space 85 mVkg. The kidney concentrated 3H relative to plasma 10- to 20- fold, the liver did not. Extracts from both organs showed a different distribution of radioactivity when compared with plasma at any given time - as is exemplified for the time point lo min after i.v.i. of lO5cpm 'H-i. per 100 g b.wt. (3H is given in cpm/ml or cpm/g):

These data assist in the interpretation of i.v.i. insulin pharmacokinetics and clarify the role of the kidney and the liver in insulin metabolism: whereas degradation products in the kidney were predominantly of low molecular weight, high molecular weight material was found in the liver.

108. Familial proinsulinemia: a possible cause of abnormal grucose tolerance. Y. Kanazawa, M. Hayashi, M. Ikeuchi, K. Hiramatsu and K. Kosaka. The 3rd Department of Internal Medicine, Faculty of Medicine, University of Tokyo. Hongo, Bunkyo-ku, Tokyo, Japan. The first case of familial proinsulinemia, a genetic disorder of insulin biosynthesis was reported in a family of United States on 1975. The second family of proinsulinemia was found in Japan. The propo- situs, 65 year-old male, was mildly diabetic for a long time. His plasma insulin (IRI) levels were high throughout the oral glucose tolerance. No insulin binding antibody was detected in his plasma. His plasma IRI levels determined by two different antisera gave different values. His sera were fractionated with Bio-gel P-30 column and found that 90% of IRI was recovered in the fraction where proinsulin appeared. The percentage of proinsulin like material (PIL) in total IRI did not change either on fasting state or under stimulated conditions by glucose. Major part of c-peptide like immunoreactivity (CPR) was also recovered in the same fraction with PIL. These data strongly suggest that the PIL is proinsulin and/or proinsulin intermediates, and that the release mechanism of PIL is the same to that of insulin. His sister and a son were found to be proinsulinemia and had abnormal glucose tolerance, while two other sons had normal IRI pattern by fractionation and normal glucose tolerance. Small amount of PIL (150ng/day) was excreted in patient's urine, while major part of CPR in his urine was free c-peptide. Daily excretion of CPR (103pg/day) was in the range of the control (90*24pg/d.). The characterization of PIL in serum and urine and of CPR in urine is now in progress. However, considering the reported biological activity of proinsulin and decreased glucose tolerance, we may spe- culate that PIL of our patient is closer to proinsulin than to biolo- gically more active proinsulin intermediates.

24. Inblbition of HDL-promoted cholesterol elimination from arterial tissue in bypercholesterolemin. G. Bondjers. Department of Medicine I , University of GOteborg, GOteborg, Sweden. Hypercholesterolemia may lead to formation of atherosclerotic lesions and arterial cholesterol deposition. This may appear in

328 ABSTRACTS conflict with recent observations that high density lipoproteins (HDL) are closer related with arterial tissue cholesterol than very low and low density lipoproteins, which change most in hypercholesterolemia. The purpose of the present investigation was to study whether the capacity of HDL to promote cholesterol elimination changes when serum cholesterol increases. - Aortic tissue from rabbits with moderate hypercholesterolemia (300mg%) or human mesenteric arterial tissue obtained as peroperative bipsies, were incubated with HDL isolated with ultracentrifugation and the serum infranate at density 1.20. Cholesterol contents in control and incubated tissue and incubate were measured and compared. - When the rabbit aortic tissue was incubated with HDL from rabbits with low serum cholesterol (100rng%), elimination of cholesterol from the tissue was observed. However, when the tissue was incubated with HDL from rabbits with higher serum cholesterol ( 3 ~ m g % ) , cholesterol was deposited in the tissue. When the human arterial tissue was incubated with HDL from the same individuals, cholesterol elimination from the tissue was observed in individuals with low serum cholesterol, and cholesterol deposition in the tissue in individuals with high serum cholesterol. - These data suggest that both in man and in rabbits, one effect of hypercholesterolemia may be to inhibit the capacity of HDL to promote cholesterol elimination from the arterial tissue. The nature of such an inhibition is unclear, but one major effect of it may be to delay arterial repair as cholesterol elimination appears to be an important component in the early phases of this process.

98. The Tromsg heart study. Serum apolipoprotein A1 concentration in relation to future coronary heart disease. T. Ishikawa, N. Fidge, D.S. Thelle, O.H. Forde and N.E. Miller. Unit of Cardiovascular Metabolism and Nutrition Research, Baker Medical Research Institute, Melbourne, Australia and Institutes of Clinical Medicine and Community Medicine, University of Tromp( Norway. The concentrations of apolipoprotein AI, a major peptide of high density lipoprotein (HDL), have been measured by immunoelectro- phoresis in samples of serum from 12 subjects who subsequently developed a coronary event during two years of follow-up and compared with those in serum from 16 control subjects who remained free of coronary heart disease (CHD). Serum apolipoprotein A1 concentrations in the cases (36.9 f 2.6 pnol/l; 105 f 7 mg/dl; mean f SEM) were significantly lower (p = 0.01) than in the controls (49.5 f 3.4 pmol/l; 140 f 10 mg/dl). Values for HDL cholesterol concentration measured previously in the same serum samples(l), averaged 0.67 * 0.05 mmol/l for the cases and 0.95 f 0.05 mmol/l for the controls (p < 0.001). In several of the cases, however, the reduction of apolipoprotein appeared to be pro- portionately much less than that of HDL cholesterol. These prospective data support other evidence that low HDL cholesterol concentration associated with increased susceptibility to CHD commonly reflects a low HDL particle number, but that changes in HDL composition or in the relative proportion of different HDL subfractions may be more prominant in some individuals. (I) Miller N. E., Forde 0. H., Thelle D. S. and Mjos 0. D. The Tromso Heart Sfudy. High densify lipoprotein and coronary heart disease: a prospective case-confrolstudy. Lancet i, 965-968, 1977.

13. Effect of physical training on plasma lipids and lipoproteins. D. Ballantyne, V.M. Hawthorne, D. Henry, T. Semple & F.C. Ballantyne. Cardiac Dept., Victoria Infirmary, Glasgow, Dept. of Community Medicine, University of Glasgow and University Dept. of Pathological Biochemistry. The Royal Infirmary, Glasgow. The purpose of the present study was to determine the effect of six months physical training on plasma lipid and lipoprotein concentrations. From a population survey of 511 men and women aged 44-45, subjects who were apparently free of vascular disease (negative history and normal ECG), did not smoke and were within 10% of ideal body weight were asked to take part in the study. Twenty men and 16 women volunteered. Subjects were randomly allocated to three different types of exercise of similar intensity. Prior to training fasting plasma cholesterol and triglyceride concentrations were measured together with the cholesterol concentration of very low density lipoprotein (VLDL), low density lipoprotein (LDL) and high density lipoprotein (HDL). These estima- tions were repeated six months later. The sexes were analysed sepa- rately but in view of the small numbers, the three exercising groups were combined. After six months training the males showed a significant fall in the fasting plasma triglyceride and significant rise in

HDL cholesterol concentration with no change in LDL. In the females there were significant falls in fasting plasma triglyceride and LDL cholesterol concentrations with no change in HDL. However, initial HDL levels were significantly higher in females. In terms of atherosclerotic risk the effects of training on fasting plasma triglyceride and on LDL and HDL cholesterol concentrations could be beneficial.

88. High density lipoprotein cholesterol levels in relation to cardiovascular risk factors in a dutch city. L.W. Hessel, C.M. van Gent and A.C. Arntzenius (Intr. by P. Brakman). Gaubius Institute, Health Research Organization TNO, Leiden, The Netherlands. In order to ascertain the status of high density lipoprotein cholesterol (HDL chol) levels as a cardiovascular risk factor, HDL chol levels were measured in a random sample of 1026 40-41 year old citizens of Leiden. This was part of a multi-city screening program, the ’CB Heart Project’. Samples were collected from February till August 1976. HDL chol was determined by a standardized Huang procedure after precipitation of the other lipoproteins by heparin and manganese chloride. Mean levels were < o.ool Mean levels calculated per 2 week collection periods showed a systematic variation with a minimum in March (39.2 m u d l for men and 43.1 mg/dl for women) and a maximum around the beginning of July (47.2 mg/dl for men and 54.8 mg/dl for women). HDL chol levels in subgroups of the total sample were:

men (n = 532) 42.5 f 10.7 mg/dl women (n=494) 48.0 f 11.0 mg/dl

men

non smokers ogr;f. smokers 44.1?10 41.5?11 P<O.Ol

women on oral contraceptives 46.3 ? 10 42.6 ? 10 P < 0.02

women not on oral contraceptives 5 1 . O f 10 48.1 5 13 P<0.02

Differences in levels between users and non-users of oral contraceptives were significant at the P < 0.001 level. In users of oral contraceptives there was a weak negative correlation of HDL chol with serum cholesterol levels (r = -0.16 P < 0.05).

238. Effect of estradiol on plasma lipoproteins in postmenopausal women. M. Tikkanen, E. Vartiainen and E.A. Nikkiltl. Third Department of Medicine, University of Helsinki, Finland. The cholesterol and triglyceride contents of plasma very-low- density-lipoprotein (VLDL), low-density-lipoprotein (LDL), and high-density-lipoprotein (HDL) were studied in sixteen estrogen- deficient postmenopausal women before and after three month’s treatment with estradiol valerate (Progynova) 2 mg daily. The mean LDL-cholesterol level decreased 25% (p < 0.01) accounting for the less marked fall of 10% (p < 0.02) of total serum cholesterol. The VLDL-cholesterol level increased 124% (p < 0.05) and HDL- cholesterol was raised 9%, the latter change being statistically insi- gnificant. However, in relation to LDL-cholesterol the HDL-cholesterol increased significantly, as indicated by an increase of the HDLILDL (u,/m-lipoprotein cholesterol ratio from 0.26 to 0.37 (p < 0.005). Earlier studies have shown that women in the age group below forty have less cholesterol in the LDL (/$lipoprotein fraction and more in the HDL ( a l ) fraction than older women. Thus, the present results suggest that estradiol treatment changes the postmenopausal serum lipoprotein pattern toward that found in young women. This is of interest since increased levels of circulating LDL-cholesterol and decreased levels of HDL-cholesterol have been associated with an increased risk of developing coronary heart disease.

110. Immunological blocking of endogenous secretin and exocrine pancreatic secretion in dog. L. Kayasseh, K. Gyr, J. Girard, F. Meyer, W.W. Rittmann and G.A. Stalder. Department of Gastroenterology, Kantonsspital Basel, Switzerland. There is continuing controversy whether secretin is physiologically released by a meal. In this context it was of interest to block the endogenous secretin activity in response to acid and to meal by in-

jection of secretin antibody. In a mongrel dog with chronic gastric and duodenal fistula plasma secretin or antibody levels were determined by RIA in response to intraduodenal HC1 and a testmeal. Simultaneously pancreatic secretion was measured by direct cannu- lation of the pancreatic duct. These experiments were performed before and after i.v. injection of 2 ml secretin antibody (capable of binding more than 2 pg synthetic secretin) until the secretin antibody became undetectable in the serum. After injection pancreatic response to intraduodenal acid was significantly reduced by 61 4 % for volume, 62% for bicarbonate output and 63% for protein output. All parameters returned to pretreatment levels when the secretin antibody disappeared. In contrast, pancreatic secretion following testmeal did not change. It is concluded that although i.v. injection of secretin antibody is capable of blocking the biological activity of endogenous secretin, the antibody did not change the pancreatic secretory response to testmeal.

30. Potent inhibition of intestinal sucrase. y-amylase, and maltase by a new a-glucosidase-inhibitor (M-GI). W. F. Caspary. Division of Gastroenterology and Metabolism, Dept. of Medicine, University of GOttingen, Germany. A delay or inhibition of carbohydrate absorption can be achieved by inhibiting active hexose transport (i.e. biguanides) or by affecting final digestion by brush border disaccharidases (i.e. TRIS). A recently detected complex oligo-saccharide *(M.W. 645) was found to have a potent inhibitory effect on intestinal a-glucosidases. The inhibitory effect of this a-GI on disaccharide and peptide hydrolase activity was characterized with total homogenates and isolated brush borders from rat small intestine. Lactase, trehalase, peptide hydrolase activity (substrates: Gly-Phe, Phe-Gly, Leu-Gly, Gly-Leu, Leu-Pro) were not affected even by concentration of the a-GI > 6 pg/ml(lO-5 M). Y-amylase > sucrase > maltase > isomal- tase were markedly inhibited by a-GI at concentrations of SxlO-aM. Kinetic analysis revealed a competitive reversible type of inhibition versus sucrose hydrolysis with a Ki of 0.5 .I 10-6 M. Since Km of sucrose hydrolysis is 3x10-2 M, Ki of a-GI versus sucrose hydrolysis was 0.5 .( 106 M, the inhibitor has a 60000 fold higher affinity to the enzyme (sucrase) than the natural substrate (sucrose) and lies in the same order of magnitude as the Ki of phlorizin on glucose active transport. This new a-GI may prove to be of beneficial therapeutic value to delay intestinal carbohydrate absorption. *Diabetologia 13, 1977, 426

136. Hepatic cholesterol synthesis in cholelithiasis: role of HMGCoA reductase in response to, and resistance to, medical treatment. P.N. Maton and R.H. Dowling, Gastroenterology Unit, Guy’s Hosp. Med. Sch. London, England. Modification of hepatic cholesterol synthesis, as evidence by changes in HMGCoAR, plays a central role in cholesterol gallstone formation and dissolution: HMGCoA activity, which correlates with biliary cholesterol secretiodl), is increased in patients forming gallstones@) and is inhibited during chenodeoxycholic acid (CDCA) treatment(,). Little is known about the effects of another gallstone dissolving agent, ursodeoxycholic acid (UDCA), on hepatic cholesterogenesis nor why occasional obese and non-obese patients, when given adequate doses of CDCA, fail to achieve unsaturated bile. Methods: We therefore measured HMGCoAR in operative liver biopsies, from 6 patients w’th untreated cholelithiasis, 4 after 1-24 months treatment with 12.1-, 17.0 mg CDCA Kg-lday-1, 5 after 1-6 months of 5 mg UDCA Kg-iday-1 and 2 <(resistant)) to CDCA - 1 non-obese with supersaturated (SS) bile despite 19mg CDCA Kg-lday-l and 72% CDCA in biliary bile acids and 1 obese (IBW 152%) with SS bile despite 22mg CDCA Kg-1day-l and 96% CDCA in bile. Results: In untreated cholelithiasis, HMGCoAR (pmoles mg microsomal prot-1 min-1) was 32.9 rt SEM 3:3; following CDCA therapy mean values fell by 53% to 15.5 + 1.5 and after UDCA in 1/3rd the CDCA dose, to 19.4 + 0.7. Resistance to CDCA was associated with a moderately high enzyme activity (47.1) in 1 patient whilst off treatment, and seemed inadequately suppressed (31.5) in the second during CDCA therapy. Conclusions: These results (i) confirm that CDCA induced unsaturated bile is associated with lowered HMGCoAR (ii) show that UDCA also inhibits HMGCoAR (iii) suggest that resistance to medical treatment may be due to high and unsuppressed HMGCoAR. References: (1) Keyet al, Gastroenterology 1977, 72: I182 (abstract) (2) Nicolau et al, J. Lipied Res. 1974, IS: 94 (3) Coyneet al, J. Lab.Clin.Med. 1976,87: 281.

ABSTRACTS 329 103. Trophic effect of portal blood in maintenance cultures of rat hepatocytes. U. Junge, W. Creutzfeldt. Department of Medicine, Division of Gastroenterology and Metabolism, University of GOttingen, FRG. Portal blood contains humoral factors which prevent liver atrophy. Its trophic effect could also be demonstrated in maintenance cultures of adult rat hepatocytes: DNA-synthesis as measured by the specific 3H-thymidine uptake in cultures incubated for 7 days with 10% pooled portal serum was higher than in control cultures with peripheral serum from the same rat (20578 compared to 14405 cpm/mg DNA). This difference was not observed when portal blood was treated with charcoal or when it was obtained from rats which were fasted for 2 days. Because insulin ( 1 pg/ml) caused a twofold increase of DNA- synthesis in serum-free cultures its role as portal hepatotrophic factor was further evaluated. Portal serum from streptozotocin treated diabetic rats stimulated DNA-synthesis less than portal control serum whereas after tolbutamide injection (25 mg/kg) it had a much better effect. These data suggest that portal blood has a hepatotrophic effect in vitro which seems to be at least partially caused by insulin.

203. Stimulation of human colonic adenylate cyclase by prostaglandin E 2 and vasoactive intestinal polypeptide. B. Simon, P. Czygan and H. Kather, Medizinische UniversitSltskli- nik Heidelberg; Abteilung fur Gastroenterologie, Heidelberg, West- Germany. A messenger function of CAMP in the stimulation of colonic secretion by bile acids and hydroxy fatty acids has been previously established. The role of the adenylate cyclase-CAMP-system in hormonal effects has not yet been investigated. Biopsy specimens of human colonic mucosa were obtained endos- copically (n = 8). The tissue fragments were gently homogenized and assayed for adenylate cyclase activity by the method of SALO- MON et al. (AnaLBiochem. 54: 541, 174). The protein content of the samples was determined according to the LOWRY-method (J.Biol.Chem.: 193: 265, 1951). Basal enzyme activity in homogenates of human colonic mucosa averaged 350 pmol cAMP/mg prot./l5 min. Both vasoactive intestinal polypeptide (VIP) (10 uM) as well as prostaglandin E 2 (0.3 mM) are capable of activating the human enzyme system by 220% and 3 m % , respectively. The /3-adrenergic agonists isoproterenol and epinephrine as well as glucagon had no stimulatory effects. The antisecretory compound lactamimide (RMA 12330 A) inhibited adenylate cyclase activity stimulated by VIP, prostaglandin E 2, lS(S)lS-prostaglandin E 2, NaF as well as GMP(PNP). When PG E 2 and VIP at maximally stirnulatory concentrations were added together the activating effect of both hormones on the colonic adenylate cyclase were additive. Our results suggest that VIP and prostaglandins which are important local tissue hormones have an as yet unrecognized CAMP- mediated physiological role in colonic function.

256. Sequential deiodination of thyroxine in rat liver homogenate and subcellular fractions. T.J. Visser, D. Fekkes, E. van Overmeeren, R. Docter and G. Hennemann. Department of Internal Medicine 111, Medical Faculty, Erasmus University, Rotterdam, The Netherlands. The present study was designed to reveal the presence of thyroxine (TQ) deiodinating enzymes in rat liver homogenate, their subcellular location, their mechanism of action and factors as pH and possible cofactors which may be of importance in the physiological regulation of enzyme activity. The products of the several reactions were measured by specific radioimmunoassays. During incubation with homogenate at 37°C production of 3,3’, 5-triiodothyronine (T,) from T, was highest at pH 6.0-6.5 and was markedly stimulated by dithie threitol (DTT) and effectively inhibited by 6-propyl-2-thiouracil (PTU). In the sequence T, - 3,3’, 5’-triiodothyronine (rT,) -. 3.3’-diiodothyronine (3,3’-T2) the former reaction was found to be the rate limiting step. From this finding it was deduced that conver- sion of T, into rT, was most effective at pH 8.0. DTT stimulated this reaction to a large extent and PTU was an effective inhibitor. Monodeiodination of the tyrosyl ring of T, was found to be the slowest reaction, to be optimal at pH 8.0 and to be less affected by DTT and PTU. 5’-Deiodination of rT, was extremely rapid with a pH optimum at 6.5. Due to the high reaction rate under the conditions used the effect of both DTT and PTU could not be studied

330 ABSTRACTS appropriately. Subcellular fractionation and comparison with profiles of marker enzymes revealed that the several enzymes were all located in the microsomal fraction of the homogenate. It was found that glutathione was a effective as DTT in supporting the several reactions. The results indicate that glutathione is the endogenous cofactor of these reactions and suggest that starvation-induced alterations in the peripheral deiodination of T, may be due to changes in intracellular glutathione concentrations.

33. The control of serum triiodothyronine by food intake. A.G. Burger, C. Wimpfheimer, M. Berger and E. Danforth. Department of Medicine and Institut de Biochimie Clinique, Uni- versity of Geneva, Switzerland. It has been well established that serum T, levels are influenced by caloric intake. To define what particular food component is important in T, induction, adult rats (300 to 350 gm) were fasted for 5 days, after which they were refed for 3 hours with either carbohydrate (CHO), protein, or fat (20KCal). After CHO, serum T, concentrations rose in 3 hours from 39 f 6 to 68 5 9 ng/dl (normal 66 ? 3 ng/dl) and, from thereon, decreased over 24 hours to fasting levels. The effect was reproduced by protein feeding whereas fat was without effect. The free T, fraction was .25 rt .021% in the fasted animals, and .27+ .008% after 3 hours of refeeding. The CHO and protein effect was dependent on the length of the fasting period. CHO refeeding increased serum T, by 39% after a 2 day fast, and by 114% after a 4 day fast. However, after 6 days, no T, induction occurred. Betahydroxybutyrate (BOH) fell from the high fasting concentration to normal after CHO and protein, while fat administration increased BOH. There was an inverse relationship between BOH and serum T,. To determine whether the observed T, increase was due to a de novo production or simply to a compartmental shift, ~ 3 ~ l - T 3 was injected into 20 fasted rats, and after 4 hours, 10 animals were fed with CHO. Serum I3Il-T3 was identical in both groups, thus excluding a compartmental shift. Conclusions: The decreased serum T, levels seen during fasting are rapidly reversed by CHO and protein refeeding. This T, increase results from a new production, rather than a compartmental change.

142. Produclion of some fealures of colloid goiters by chronic minimal- dose goitrogen feeding. E. Miloni, H. Biirgi, L. Siebenhuener and H. Studer. Medizinische Universitatsklinik, Inselspital, Berne, Switzerland. Naturally occurring goiters often contain large amounts of poorly iodinated thyroglobulin (Tg). Histologically they are rich in colloid. Goiters produced experimentally by an iodine deficient diet or goitrogens also contain poorly iodinated Tg, but in contrast to natural goiters they are depleted of colloid and Tg, i.e. hyperplastic. So far, there is no experimental procedure to produce genuine colloid goiters. We tested the hypothesis that minimal doses of goitrogens could lower the iodine content of Tg, but would not be sufficient to cause hyperplasia with colloid depletion, thus resulting in colloid goiters. Rats on an adequate iodine intake were fed minimal doses of methimazole (MMI, 0.0001 and 0.0002 070 in drinking water). Thyroid weight (3.5 mg per 100 g body weight) and thyroidal Tg- content (3.2 mg per 100 mg gland) remained constant for 6 weeks with both doses of MMI. By contrast the thyroidal radioiodine uptake dropped from 1.91% to 1.54% and the iodine content of Tg decreased from 0.91% to 0.57% (p < 0.001). Histologically, controls and MMI-treated thyroids had a similar colloid content. We conclude that borderline doses of goitrogens can lower iodination of Tg without causing hyperplasia and colloid depletion. Long-term exposure to very low doses of naturally occurring goitrogens should be considered as a possible cause of colloid goiter.

114. Short feed-back control of thyroid function W. Kielczynski and J. Nauman. Department of Biochemistry, Medical Centre of Postgraduate Education, Warsaw, Poland. We previously shown, that triiodothyronine T,, reverse triiodothyronine rT, and thyroxine T, specifically inhibit TSH binding to thyroid plasma membranes. To clarify further thyroid hormones- TSH receptor interrelationship, the nature of T, and T4 binding to thyroid membranes was studied and kinetic parameters of reaction estimated. Membranes were prepared from human thyroids as described by Neville and Kahn. T, and T, were labelled with 125-1 to

specific activity above 1500 mCi/mg by Chloramine T method. The reaction was carried out in medium containing T, or T, and thyroid membranes in 25 mM Tris-acetate buffer, pH 6.0 and saturation technique was employed to measure Ka, Kd and Bmax values. Both T, and T, were specifically bound to membranes. The Ka for T, was 2.26~10~M-l . Kd-4.4xlO-'OM and Bmax 4.13 xlOlOM. The same values for T, were 3 . 5 ~ l O ~ M - ~ , 2 .86~10-~M and 1 .O2xlO-9M respectively. TSH, LH and FSH specifically inhibited thyroid hormones binding however the effect of two later glycopro- teins was 1000 times weaker. The results suggest direct interaction of T, and T, with TSH receptor Present study support the existence of short feed-back control of thyroid function on TSH receptor level.

154. The effect of three different enzyme inducing agents and their influence on thyroid hormone metabolism in man. E.E. Ohnhaus und H. Biirgi. Department of Medicine, University of Berne, Switzerland. Following chronic phenobarbitone administration an increased deiodination of thyroxin was found in rats, a finding connected with increased liver microsomal activity. Therefore, we were inte- rested, whether these findings also apply to man. 3 groups of healthy volunteers were investigated and three different treatment known to induce liver microsomal enzyme activity in man were given: antipyrine (1200 mg), phenobarbitone (100 mg) and rifampicin (1200 mg) daily for 14 days. Before and after each treatment the following parameters of enzyme induction were measured: total body clearance of antipyrine, y-glutamyltranspeptidase, d-glucaric acid, &/?-OH-cortisol. In addition, thyroxin, T3-resin uptake, T,-RIA and reversed T3 were estimated. All parameters of enzyme induction and the thyroid hormones measured were comparable in all groups before drug treatment. After discontinuation of the three inducing agents different results were obtained. After antipyrine and phenobarbitone treatment an identical increase by 50% in the total body clearance of antipyrine was seen, while with rifampicin an increase of about 124% occurred. The other parameters of microsomal enzyme activity showed similar significant changes. The different thyroid hormones did not change following phenobarbitone and antipyrine administration, but after rifampicin a significant decrease in thyroxin and rT, and an increase in T, levels occurred. Therefore, rifampicin produces a shift of the peripheral thyroid hormone metabolism towards the activating pathway. The connection of this new finding to enzyme induction is not yet clear.

129. Metbotrexate (MTX) recovered from urine of high dose patients characterization and clinical reuse. A. Leyva and H.M. Pinedo, Dept. Internal Med., Univ. Hosp., Utrecht, The Netherlands, F. Spreafico, Mario Negri Inst. Phar- macol.Res., Milan, Italy. Chemotherapy of neoplastic disease with 4-6 hr high dose (50-300 mg/kg) infusion of MTX with leucovorin rescue is proving to be promising. However, the cost for such doses presents an economic problem for many European oncology centers. Since most of the MTX dose is excreted largely unmetabolized in the urine within 24 hrs after administration, the feasibility of its recovery for clinical re-use was investigated. More than half of the initial dose of MTX was isolated by acid precipitation from the urine of patients having received 10-18 g MTX. Pyrogens were removed by calcium phosphate treatment and ultrafiltration. The urine-derived drug (MTX-U5 was compared to the administered commercial drug 4MTX-C) by DEAE cellulose chromatography. MTX-U was 90-95 To MTX based on absorbance at 260 nm which was comparable to 94 To for MTX-C. MTX-U and MTX-C contained similar impurities, most with spectral properties characteristic of pteridine compounds and demonstrating inhibition of dihydrofolate reductase (DHFR). Impurities detected in MTX-U but not in MTX-C INCLUDED 7-hydroxy MTX and uric acid MTX-U was found to inhibit DHFR in vitro and to be therapeutically effective in tumor- bearing mice to a degree equivalent to MTX-C. The treatment of two patients with 1 or 7,5 g doses of MTX-U resulted in favourable antitumor responses without adverse effects. This study shows that MTX can be easily recovered following high dose therapy and made suitable for subsequent clinical re-use.

33 1 47. Melanoma protein in urine, melanoma antibody in blood in malignant melanoma and benign pigmentary skin disease. P.W.M. Copeman and K.B. Cooke, Westminster Hospital, London, SWI A specific sialo-protein, immunologically related to the cytoplasm of altered naevus cells, has been isolated from the urine of patients with malignant melanoma. This protein reacts against (i) a specific rabbit antiserum raised against fresh human malignant

melanoma cells, and (ii) the naturally occurring humoral cytoplasmic antibodies of

patients with either malignant melanoma or benign halo naevus. The protein occurs in the urine of patients with malignant melanoma: in 50To of those with isolated primary tumours; and in 83% with widespread metastatic disease. Also it is found in patients with actively developing halo naevi or with very rapidly extending vitiligo, but in no other pigmentary skin disease examined. In halo naevus the urine melanoma protein and cytoplasmic antibody can occur together and this combination is found also in long- surviving melanoma patients. We postulate that a state of symbiosis may exist between patient and tumour. Physico-chemical studies indicate that the specific protein is of cellular origin, but its occurrence in patients with isolated halo naevi would argue against it being a measure of cell death.

223. Cis-diammine-dichloro-platinum (DDP), vinblastine (VBL) and bleomycin (BLM) combination Chemotherapy in pre-treated testicular non-seminoma. G. Stoter, A. Struyvenberg, C.P.J. Vendrik and H.M. Pinedo, Oncology Unit, Dept. Internal Med., University Hospital, Utrecht, The Netherlands. The new cytostatic agent DDP has proven to be active in testicular non-seminoma. The best results have been achieved with an induction regimen with DDP, VBL and BLM and maintenance treatment with VBL and BCG (Einhorn, Ann.Int.Med., 1977). We initiated a similar study in both radio- and/or chemo-pretreated and non-pretreated patients. Chemotherapy consisted of DDP 20 mg/m2 i.v. d. 1-5, VBL 0.2 mg/kg i.v. d. 1.2 and BLM 30 mg i.v. d. 2, then weekly x 12. Courses of DDP and VBL were repeated q 3 wks (3-4 courses). No BCG was given. The purpose of the study was to compare remission-induction rate and toxicity in both patient groups and to evaluate if remission duration is comparable to that obtained with BCG. We have treated 9 pretreated (4 radiw, 3 cheme and 2 radio + chemotherapy) and 3 non-pre-treated patients with non-seminomas, stages 2 or 3.8 patients achieved complete remission, 1 relapsed after 6 months. Remission durations of the others range from 2+ - 18+ months. 3 patients are in partial remission but still on induction regimen. 1 patient in near-complete remission died of sepsis. We observed severe toxicity in the pretreated group: 4/9 mild and 1/9 severe renal failure, 2/9 paralytic ileus, 8/9 severe leukopenia (WBC < 1000/mm3 longer than 1 wk), 8/9 severe trombopenia (< 30.000/mm3), 3/9 sepsis requiring leukocyte and trombocyte transfusions. 1/3 non-pre-treated patient had severe leukopenia during 3 d. No other signs of severe toxicity have been observed in these 3 patients. In conclusion, induction chemotherapy consisting of DDP, VBL and BLM in testicular non-seminomas seems effective in both patient groups. Severe toxicity was only observed in the pretreated groups. Up to now omission of BCG has not influenced remission durations as compared to those published.

29. Failure of vitamine E (E) lo prevent adriamycine (A) -induced cardiotoxicity in the rabbit model. J.G.S. Breed, A.N.E. Zimmerman and H.M. Pinedo. Oncology Unit, Dept.Int.Medicine, Univ.Hospital. Utrecht, The Netherlands. A has been hampering chemotherapy because of its cardiotoxicity above a cumulative dose of 550 mg/m2. Recently Meyers (Science 197:165,1977) reported the prevention of A cardiotoxicity by E in CDF,mice. Phase I studies have been proposed. We studied the hearts of 29 NZW rabbits after 3 months in vivo exposure to A,A + E AND SALINE4S) by analysis of 1) contractility of the isolated perfused heart (Kruta curves) and 2) light microscopy(LM) and electronmicroscopy(EM). 23 rabbits were treated with A i.v. (1.0 mg/kg twice a week, cumulative dose 25-30 mg/kg) of which 10 were given in addition E (300 mg/kg, 5 times a week). Six control rabbits received only S. Animal food contained usual amounts of E (80 mg/kg and Se (0.45 ppm). Kruta curves revealed no significant improvement in contractility in the A + E group as compared to the A

ABSTRACTS

'7 group. LM and EM of the hearts of the A group showed characteristic A pathology,

of the severity of, the lesions was seen in the

Only slight reduction

I A + E group. These 3 1 results indicate that 3

A cardiomyopathy is

Thus, it is not war- g 90-

ranted to initiate phase I studies to in- LT I

protection of E against = m -

likely species specific. - ... A + E n . K

5 n- 6

Sl IHUlAl lNG INlERVAl IN H SEC

500 LW LOO 350 300 2% 100

vestigate the protective effect of E in human. ,-,-,-,-!-,-I

"

131. Studies on biosynthesis of albumin and hepatic proteins in cancer patients. K. Lundholm, I. Karlberg, T. Schersten Department of Surgery, Sahlgrenska sjukhuset, Gdteborg, Sweden. The mechanisms for hypoalbuminemia in cancer patients are still unknown. The aim of the present investigation was to study biosynthesis rate of albumin and other hepatic proteins in cancer patients. Liver tissue slices were incubated at optimal in vitro conditions at + 37'C. Albumin was isolated by monospecific rabbit antiserum. Irnmunoprecipitated albumin in hepatic tissue consisted of proalbu- min and ((serum albumin)). Albumin isolated from the incubation medium consisted only of ccserum albumin)). Cancer patients (n = 23) showed statistically significantly increased incorporation rate of leucine into hepatic proteins. The protein synthesis rate was correlated to the concentration of hepatic RNA, which was increased in cancer disease. The incorporation rate into hepatic albumin was of the same order of magnitude in cancer and controls. Medium albumin on the other hand had significantly increased specific activity in cancer patients. This was concomitant with decreased extrusion of unlabelled albumin into the incubation medium. The relationships (regression coefficients) between hepatic protein synthesis rate and albumin synthesis rate in cancer patients and controls differed significantly. The results showed that cancer disease was associated with increased hepatic RNA content probably preferentially used for biosynthesis of other hepatic proteins than albumin. The translational capacity for albumin synthesis was not altered in cancer patients. This means that previously suggested impaired albumin synthesis in cancer disease can not be ascribed to depressed cellular function. The present results were most closely compatible with increased turnover of albumin.

71. Effect of Ro 11 1781, calcium antagonist, on paroxysmal supraventricular tachycardia. R. Gmeiner, C.K.Ng. C.Schwenninger and M. Gstdttner Depart- ment of Internal Medicine, University of Innsbruck, Austria. The effect of Ro 1 1 1781 (N-(3,4-Dimethoxyphenthyl)-2-(3,4-dime- thoxyphenyl)-II-methyl-m-dithiane-2-propyIamine 1,1,3,3-tetraoxide hydrochloride), a new calciumantagonist, was studied by His bundle electrography and atrial and ventricular stimulation in 8 patients with paroxysmal supraventricular tachycardia (PSVT). Analysis of A,-A,, HI-H2 curves revealed smooth conduction curves in 3 patients with WPW syndrome and 2 Patients with concealed bypasses conducting ocly in ventriculoatrial direction. In 3 patients discontinuous (dual pathway) curves suggested AV nodal reentry. Before Ro 11 1781 tachycardia zones (TZ) were demonstrated in all patients. An induced tachycardia was terminated by 2 mg/kg Ro 11 1781 i.v. Block occurred in the antegrade pathway in 7 cases after prolongation of the A-H interval. After Ro 11 1781 the tachycardia could not be initiated in 3 patients with AV nodal reentry. The effective refractory period of the fast pathway was shortened, the slow pathway curve was shifted rightward and upward in 2 and abolished in 1 patient. In the smooth pathway cases the TZ was reduced in 2, unchanged in 1 and increased in 1 patient. After Ro 1 1 1781 the ability to sustain the tachycardia was lost in 2 patients. Beneficial effects of Ro 11 1781 included the interruption of the tachycardia, the abolition or decrease of the TZ, and loss of the ability to sustain the tachycardia and the reduction of rate, the widening of the TZ may be harmful.

332 ABSTRACTS 262. Uptake of 42K by cardiac muscle from normal and potassium depleted rabbits. J.P.T. Ward and I.R. Cameron. Department of Medicine, St. Thomas’s Hospital Medical School, London. During dietary K + depletion cardiac muscle maintains the level of intracellular [K+] ([K+ i]) at control values even though skeletal muscle may have lost up 25% of [ K + i ] . Following 1 week of K + repletion cardiac muscle [K+ i] exceeds control values, returning to this level only after 3 weeks repletion (Cameron and Hall, J. Phy- siol. 1975, 251, 70p). The mechanism whereby cardiac muscle [ K + J is protected against K + loss and the cause for the repletion ’overshoot’ remains unknown. The uptake of 4*K by cardiac muscle has been studied using the arterially perfused interventricular septum from normal and K + depleted rabbits. The septum was perfused with a specially prepared Ringer solution until 42K uptake reached a steady state. The change in the uptake following alteration of the [K+] in the perfusate was studied. During perfusion, tem-

perature, tension and the differential of tension were monitored. Intracellular electrolytes and the ECS of ventricular septal muscle were measured from control and depleted rabbits. In the septum from depleted rabbits [K+i] was 334.4 k 6.1 mmol kg~l FFDS (+ SEM) and 343.7 2 5.7 mmol kg-l FFDS in controls (p 3 0.25). Skeletal muscle (quadriceps) [K+ i] in the depleted rabbits was 75% of that in the controls. In the isolated septal preparation from both control and depleted rabbits 42K uptake changed with variation of [K + ] in the perfusate. The relationship between the change in uptake and perfusate [ K ] differed ‘in the depleted and control animals; that in the depleted animals being shifted so that the curve is set at a new low level of external [K+] (shift to the left). The mechanism whereby this shift occurs which allows normal K + uptake at low levels of plasma [K+] is as yet unknown.

213. Effect of aldosterone antagonists on the potassium transport of the heart muscle. J. Somogyi, H. Mattern, F.H. Schmidt, B. Szab6 and F. Willig. Inst. Biochem.I., Semmelweis Univ.Med.Schoo1, Budapest, Inst. Physiol.Univ. Med.School, Debrecen, Hungary, Res.Lab. of Boehringer-Mannheim and Speyererhof Hospital Heidelberg, federal Republic Germany. Aldosterone antagonists have a favorable influence on the arrhytmic state accompaning the intoxication caused by the cardiac glycosides. Since the tolerance for cardiac glycosides is closely related with the K + level and the receptor for these glycosides is probably identical with the Na+ + K+-ATPase, we investigated the influence of different aldosterone antagonists on the K f transport of heart muscle as well as on the Na+ + K+-ATPase. Isolated hearts of guinea pig were perfused by the Langendorf’s procedure with Ty- rode’s solution, under standardised conditions. The electrolyte balance was calculated from the difference in the K f content of the effluent solution in comparison with the initial value. On addition to the perfusion fluid Spirolactone or Canrenone, the myocardium took up K + . 147 pM Canrenone could completely abolish the loss of K + caused by 2 pg/ml ouabain, and it significantly reduced the K release from myocardium due to 5 pg/ml/3-methyl-digoxin and increased the survival time about 50 per cent when 10 pg/mlfi-methyl-digoxin were added with 147 pM Canrenone. On the other hand, Canrenone failed to influence the Na+ + K+-ATPase activity, it had no essential action on the ouabain induced inhibition of enzyme and did not influence the extent of 3H-ouabain binding to the enzyme. I t may be assumed that the influence of aldosterone antagonists upon the intoxication by cardiac glycosides is based on the increased K + concentration of the heart muscle cells and not on their action on the digitalis receptor.

269. Does imaging with thallium-201 improve the value of stress tests for diagnosis of coronary artery disease? A. Withagen, M.L. Simoons, W. Bakker, P. Kooy, and H. Reiber. Thoraxcenter and Department of Nuclear Medicine, Erasmus Uni- versity, Rotterdam, The Netherlands. The relative values of exercise electrocardiography (XECG), corn- puter assisted XECG and myocardial perfusion imaging (MPI) with TL-201 were compared in 40 male patients with a normal ECG at rest. All patients underwent selective coronary arteriography, 30 had 70% or greater coronary artery obstructions (CAD). Exercise was performed on a bicycle ergometer with stepwise workload increments of 10 Watts per minute until complaints or near exhaus- tion. The sensitivities for prediction of CAD were: XECG 18/30,

computer assisted XECG 21/30, MPI 19/30. The specificities for prediction of absence of CAD were 7/10,7/10 and 6/10 respectively. Two patients had angina pectoris during the exercise test with ST depression and new MPI defects, in spite of normal coronary arteries. This suggests that these patients had myocardial disease or small vessel disease in spite of the normal coronary angiogram. MPI appeared to be neither more sensitive nor more specific than XECG for prediction of CAD in patients with a normal ECG at rest.

240. Sinus arrythmia and respiratory modulation of the carotid baroreceptor cardiac reflex in man. A. Trzebski, M. Raczkowska and L. Kubin (intr. by S. Majcherczyk). Department of Physiology, Medical Aca- demy, Warsaw 00 658, Poland. In 38 men 21-24 years old carotid sinus baroreceptors were stimulated with neck suction of -30 up to -60 mmHg as described by Eckberg et al.ECG R-R intervals were measured, integrated and

gain of the reflex ~ ~ ~ a ~ ~ ~ ~ ~ ~ l was calculated. Respiratory move- ments and end-tidal CO, concentrations were continously recorded. During baroreceptor stimulation over 5 respiratory cycles amplitude of the sinus arrythmia was increased. Baroreceptor reflex was reduced if short stimuli of 2 seconds were applied during inspiration and the inhibitory effect was positively related to the rate of inspiration. I f the stimuli were applied during expiration a facilitation of the reflex was observed. On the peak of the prolonged supranormal inspiration spontaneous bradycardia appeared. At the end of supranormal expiration a shortening of R-R intervals and reduction of the gain of the reflex was observed. During apnea rhythmical changes of R-R intervals and the gain of the baroreflex were recorded. These apneic periodicities were of similar or lower frequencies than the preceding respiratory rate and the sinus arrythmia of the heart rate. It is concluded that central respiratory activity modifies sinus node responses to carotid baroreceptor stimulation.

56. Lactate and myocardial metabolism. Angela J. Drake, D. Papadoyannis and M.I.M. Noble. Midhurst Medical Research Institute, Midhurst, Sussex, England. Lactate is metabolised entirely through aerobic pathways and one might therefore expect that a switch to lactate consumption would increase myocardial oxygen consumption for a given amount of work. In order to test this idea, lactate was infused up to an arterial concentration of 9 mmol.l-l. This produced an initial depression of myocardial contraction followed by elevation of the contractile performance above control, i.e. increased cardiac output, work, and rate of rise of left ventricular pressure. This was accompanied by a small increase in myocardial oxygen consumption (from a mean value of 0.137 ml.min-i.g-l to 0.164) and a large increase in lactate consumption (0.664 mmol.1-I .g-I to 1.834). However, when norma- lised for external work performed or pressure generated (tension time index x heart rate), there was no change in myocardial oxygen consumption. Thus, a switch to lactate did not produce metabolic inefficiency. In 16 closed chest anaesthetised dogs the myocardial oxygen consumption of lactate exceeded that of glucose and free fatty acids (mean values:- lactate 4.9%, glucose 32.5%, FFA 20.6%). Myocardial consumption of lactate was consistent whereas glucose of FFA consumption was often zero (coefficients of variation:- lactate 0.49, glucose 0.77, FFA 1.09%). In the presence of lactate the heart did not switch to glucose when the latter was infused up to an arterial concentration of 26 mmol.1-l (normal range 4.0 - 6.0 m m ~ l . l - ~ ) . Lactate is thus the major myocardial substrate for these normal dog hearts.

245. Fractional clearances of dextran and dextran sulfate in a new model of proteinuria in the dog. Y. Vanrenterghem, M. Lammens and P.P. Lambert. Queen Elisabeth Medical Foundation, Brussels, Belgium. In a variety of glomerulopathies, glomerular polyanion content is diminished. This reduction of fixed negative charges in the capillary wall could explain the enhanced passage of anionic macromolecules and plasma proteins observed in rats after injection of nephrotoxic serum. For the present study, unilateral proteinuria was induced in the dog by injecting a 4 Mol urea solution in a 1 : 1 mixture of plasma and

water into the left renal artery during a 5 to 6 min interruption of renal blood flow. 30 min later, mean protein excretion in the left kidney increased from 0.36 mg/min to I5 mg/min. GFR and CPAH decreased respectively to 43% and 36% of the control values. Frac- tional clearances of D and DS with a mean Einstein Stokes radius of 37 A" (4 37) are shown in the table.

D l e f t n = 7 D r i g h t DSleftn=8DSright Control 0.161 0.163 0.073 0.078 Proteinuric 0.193 0.159 0.171 0.078 Conclusions : in the normal kidney, ultrafiltration of polyanionic macromolecules is more retarded than for neutral macromolecules. In the proteinuric kidney, a significant increase of 4 37 is observed for charged molecules only. In this model also, proteinuria results at least partly from a reduction of the electrostatic repulsion of polyanions. The observed morphological alterations fit with this conclusion.

75. Vasopressinomimetic action of psychotropic drugs. A. Grosso and R.C. de Sousa. Departments of Physiology and Me- dicine, School of Medicine, Geneva, Switzerland. It has been recently reported that patients taking psychotropic drugs can develop a syndrome of water intoxication. Two types of general mechanisms may be envisaged to explain this condition : a central action leading to an inappropriate release of vasopressin and a peripheral action due to a vasopressinomimetic effect of these drugs on water transport. In an attempt to examine this last hypothesis, water flow measurements were performed with automatic, volume- tric techniques in two models in vitro - the urinary bladder and the skin of toads Bufo marinus. Several psychoactive agents - harmaline and other harmala alkaloids, amitriptyline and chlorpromazine - at concentrations ranging from 0.1 to 1 mM induced a significant (p < 0.02) hydrosmotic effect in toad skin, competitive with that of vasopressin. In contrast, harmaline and amitriptyline, in toad bladder, elicited only minor changes in basal water flow but blocked its stimulation by vasopressin (p < 0.001). The finding of these opposite effects on water permeability in two vasopressin-sensitive epithelia of the same animal, emphasizes the pitfalls of generalizing from results obtained with a single model. Further work is necessary however to elucidate the cellular mechanism(s) underlying this striking discrepancy.

126. Clinical evaluation of hyperuricemk patients. F. Lang, E. Griss, K. Lang, H. Oberleithner, R. Greger, P. Deetjen, Institute for Physiology, Innsbruck, Austria and Klinisches Sana- torium Dr. Birkle, uberlingen, Germany. A test program was designed to evaluate urate production and renal elimination in normal subjects (n = 13). patients with moderate hyperuricemia (n= 14) and with marked hyperuricemia ( n = 17, including 9 patients with clinical gout). All patients were kept on an identical purine intake (1 mmol/die) and the groups were similar in age, sex and renal function. Urate (CuA) and Creatinine (CCJ clearances were monitored during 3 1/2 days. At the beginning of the third day probenecid (500 mg) or benzbromarone (lo0 mg) were administered. The decline of plasma concentration and the increase of urate excretion following the uricosuric agents allowed an estimate of the rapid miscible urate pool of the patients. The results are depicted in the following table (mean f SEM): group plasma urate C,,/C,-, urate

urate excretion pool (mmol/l) (mmol/die) (%) mmol

Normal 0 ,28r0,02 2 ,6f0 ,2 10,5f1,1 6.6*0,7 ModerateH. 0,35+0,02 2,81t0,2 7,0&0,5 6.8+0,7 MarkedH. 0,45*0,03 2,9+0,2 5,4+0,5 10,0+0,9 (H. = Hyperuricemia) Urate clearance was increased some fourfold after administration of the uricosuric agents. A strong correlation (r = 0.90) was apparent between urate clearance before and after application of the uricosuric agents indicating that the agents did not interfere with the mechanism responsible for the differences of urate clearance among different patients. Furthermore reduced urate clearances appeared to be the major cause underlying the variations in plasma urate concentration.

ABSTRACTS 333 246. Acid-base and metabolic consequences of the gastro-intestinal chelation of cations. C. van Ypersele de Strihou and J.P. Dieu, Renal Unit, Department of Medicine, Cliniques Universitaires St-Luc, 1200 Brussels, Belgium. <<Endogenous acid production)) (EAP) is partly determined by the gastrointestinal handling of electrolytes. The present study was undertaken to assess the possibility of altering EAP by modifying the digestive absorption of cations. Balance studies were performed in 30 dogs maintained on a constant electrolyte free diet with different electrolyte solutions. After a control period a cationic exchange resin, Kayexalate (Ka), (30 g/d), is added for 8 to 18 days (Per. I). Ka is interrupted and observations are continued for 4-10 days (Per. 11). In dogs (N = 6) given a mineral supplement (CaCO,, P O 3 sodium Ka produced metabolic alkalosis (PHCO, + 12 meg/l), a reduced net acid excretion (- 46 meq/d) and increased urine phosphate excretion (+ 8 mmol/d). These changes were independent of potassium balance : they disappeared after withdrawal of Ka despite the persistence of potassium deficit. The same changes were induced in dogs given NaCl (N = 3) and in adrenalectomized dogs (N = 9) given a constant dosage of DOCA. They are thus independent of Na intake or changes in adrenal secretion. However, they are markedly reduced (plasma HCO, and urinary phosphate no change, net acid - 16 meq/d) if the animals are given a calcium free diet (N = 6). These data suggest that calcium chelation by Ka significantly aug- ments digestive HCO, and PO, absorption and thus EAP. Conversely, provision of calcium Ka induces the opposite changes : metabolic acidosis (PHCO, - 6 meqll), increased net acid excretion ( + 7 meq/d), decreased urine PO, (- 4 mmol/d). The development of kayexalate alkalosis and acidosis demonstrates that alteration of digestive handling of cations and especially calcium may profoundly alter EAP.

235. Is arginine vasopressin (AVP) involved in the pathogenesis of idiopathic oedema of women? M. Thibonnier and P. Corvol. Inserm U 36, Paris - France. Idiopathic oedema (10) of women are associated with an impairment of water excretion, especially in upright position, suggesting the AVP'responsability in this disorder. Urinary AVP, plasma renin and plasma aldosterone were measured by radioimmunoassays in 10 normal women and in 10 1.0. before and during water loading (15 ml/kg) in supine and upright positions under a 120 mEq Na+ diet. Glomerular filtration rate (GFR), osmolar clearance (Cosm), free water clearance were determined throughout the study. In 1.0. urinary AVP excretion significantly increased in upright position (p 0.01). This increase was higher than that observed in normals (p 0.05). Whereas a correlation could be established between plasma osmolality or protides and urinary AVP in normals, such a relation could not be found in 1.0. Finally this study showed a transient but significant decrease of G.F.R., a significant decrease of Cosm and a stimulation of renin aldosterone system in 1.0. standing up. In summary, 1) AVP is increased and abnormally regulated in 1.0. 2) Other factors are involved in water retention : disturbances of renal function and stimulation of renin aldosterone system.

70. Effect of inhibition of prostaglandins(PG) on renin(PRA) and aldosterone(UA) response to salt deprivation. Ph. Glasson and M.B. Vallotton. Div. Endocr. Dept. Medecine. Geneva, Switzerland. Previously we demonstrated that pretreatment with indomethacin (I) prevents all response of PRA and UA to diuretic-induced Na + depletion. To investigate further the relation between renal PG, PRA and UA, we studied normal volunteers during 10 days on low salt diet (10-15 mmol/d) receiving I(150 mg/d). In group A (6 males, 5 females) I was given from day 6 to 10, while in group B (4 males, 2 females) I was given from day 1. Results: in both groups Na+ balance was achieved at day 5 and I suppressed urinary P G from control values (PGE2 385 i 59 and PGF2 D 1013 116) to 109 F 22 and 285 f 40 ng/d respectively (RIA with antiserum from Dr. F. Dray, INSERM FRA8, Paris). In group A PRA elevated to 5.5 f 0.6 ON DAY 6 decreased to 3.1 f 0.4 ng/rnl/hr (p < 0.01). UA decreased from 64.5 + 10 to 50.5 f 7.5 ng/d, (p < 0.5) concommitantly with a rise of plasma K + from 3.7 to 4.2 Inmol/l. In group B two patterns of response were observed: in 3 subjects increments of PRA was 5 f 1 ng/ml/hr and of UA 33 k 8 ng/d whereas in 3 others both PRA and UA were unchanged despite similar plasma and urinary electrolytes. Conclusion: in subjects with prior stimulation of

334 ABSTRACTS the RAAS by volume depletion I fails to completely suppress PRA and UA (the latter possibly through a rise of plasma K + ) . Simul- taneous suppression of PG and stimulation of the RAAS result in responses with and without PRA and UA increase. Therefore the time-sequence of PG inhibition and PRA stimulation appears important to evaluate the relation, which can be abolished, between both systems.

123. Studies on the metabolism of the lipoprotein Lp(a) in man. F. Krempler, G. Kostner, K. Bolzano and F. Sandhofer. First Department of Medicine, Landeskrankenanstalten, Salzburg, and Institute of Medical Biochemistry, University of Graz, Graz (Austria) The lipoprotein Lp(a) can be found in almost all human sera. It has been postulated tt,at the appearance of LNa) in high serum concentrations might be a risk factor for atherosclerotic vascular disease. Virtually nothing is known about the metabolism of Lp(a). The aim of our study was to investigate the behaviour of Lp(a) labeled by reductive alkylation. In four subjects labeled Lp(a) was injected intravenously. A linear decline of the specific activity of Lp(a) in the serum was found when plotted semilogarithmically against time. Half-lives of Lp(a) in the serum were 35,38,53,58 h. The ctsoluble)) and the ((insoluble)) apoproteins showed the same half-life as the whole Lp(a) molecule. At different times after the injection of LHa) only small amounts of radioactivity could be found in other density fractions. After in vitro incubation of labeled Lp(a) with whole serum for 24 h, more than 99 Olp of the radioactivity of the incubated sample were found in Lp(a). The chemical composition of Lp(a) is similar to that of low density lipoprotein (LDL). Our data indicate that Lp(a) is metabolized slightly faster than LDL as measured by others. N o exchange of Lp(a) apoproteins with other lipoproteins takes place in the plasma.

143. Regulation of 3-hydroxy-3 methylglutaryl coenzyme a reductase in mononuclear cells by dietary cholesterol in man. P. Mistry, R.A. Bacchus and B. Lewis. Department of Chemical Pathology and Metabolic Disorders, St. Thomas’ Hospital, London, England. Seventeen normal subjects were fed a cholesterol supplement of 1.5 gm/day for 14 days, as egg-yolk. The serum cholesterol increase varied widely (-0.075 to 0.121 mmol/L/day); levels of all lipoprotein classes increased, particularly LDL,. HMG CoA reductase activity was measured in freshly isolated blood mononuclear cells and in cells derepressed for 72 hours in RPMI 1640 medium containing 10% lipoprotein-deficient serum (LPDS). Low levels of reductase activity could be detected in fresh cells (RJ and this was elevated 3.8-26 fold by incubation in LPDS medium (R7,. R, was reduced by cholesterol feeding (1.55 f 0.19 to 0.92 i 0.12 pmol/ min/mg protein, mean 2 SEM, p < 0.001). The degree of inhibition ranged from 0-73%; this was unrelated to the cholesterolaemic response. R,, was similar in cells obtained before and after the cholesterol feeding (12.8 f 1.5 and 11.9k 1.1). Suppression of reductase in circulating mononuclear cells was reflected in R7,/R, ratio, which increased from 9.3 k 1.6 to 15 f 2.7 (p < 0.05). The data suggest that cholesterol synthesis in mononuclear cells, like that in hepatocytes, is subject to regulation by dietary cholesterol. Mononuclear cells may provide a useful model for studying dietary factors influencing cholesterol metabolism.

35. Regulation of cholesterogenesis in rat hepatocytes. S. Calandra, P. Tarugi, N. Battistini, Department of General Pathology, University of Modena, Modena, Italy. Recent studies indicate that hepatic cholesterogenesis may be under the control of serum lipoproteins as the inravenous infusion of high amounts of chylomicrons, VLDL and LDL reduces hepatic cholesterogenesis. Since the complexity of the in vivo studies pre- cludes the understanding of the mechanism of regulation of cholesterol synthesis we investigated this problem by utilizing adult rat hepatocytes. Rat hepatocytes (RH) were isolated by perfusing the liver with collagenase and were incubated in Eagle’s medium sup- plemented with labelled acetate or mevalonate. Human, rat and rabbit serum lipoproteins were added to the medium at concentrations ranging from 10 to 200 pg/ml. RH were exposed to serum lipoproteins for 5-24 hours. In the short term incubations we found that human VLDL and LDL as well as rat and rabbit VLDL did not

reduce cholesterogenesis. Rat LDL instead (15 pg/ml) reduced cholesterogenesis by 50%. When RH were incubated for 24 hours a partial reduction of cholesterogenesis was also produced by human LDL. An almost complete suppression of cholesterogenesis was produced by VLDL isolated from cholesterol fed rabbits, which chemically resemble chylomicron-remnants. These observations indicate that both LDL and cholesterol rich VLDL may be the physiological modulators of cholesterogenesis in rat liver.

141. Is secretion of lecithin. cholesterol acyltransferase by the liver linked to that of very-low-density lipoproteins? J. P. Miller, Department of Medicine, University Hospital of South Manchester, Manchester, U.K. Lecithin: cholesterol acyltransferase (LCAT) and very-low-density lipoproteins (VLDL) are both secreted by the liver and a relationship between LCAT activity and plasma triglyceride concentration has been demonstrated frequently. In an attempt to determine if secretion of LCAT by the liver is linked to that of triglyceride - rich lipoproteins the effect of feeding orotic acid, which blocks hepatic secretion of such lipoproteins and leads to fat accumulation in the liver, has been investigated in Sprague-Dawley rats. Rats were fed a synthetic ((purine free)) diet for periods of 11-17 days, half of the animals receiving orotic acid (2% w/w) in addition. Orotic acid feeding was associated with significant reductions in serum lipid concentrations, due predominantly to reduction in the concentrations of lipoproteins of d < 1.063, and with a highly significant increase in liver weight (expressed as a percentage of body weight). These changes could be prevented by the addition of 0.25% adenine sulphate to the diet as shown by others. PIasma LCAT activity was 69.6 f 4.1 (mean f SD) n mol. ml.-Ih:l in control rats and 71.7 f 7.2 in orotic acid fed animals. It is concluded that, under the conditions of these experiments, either secretion of LCAT by the liver is not linked to that of triglyceride - rich lipoproteins or that such linkage occurs at a stage before that at which orotic acid interferes with lipoprotein secretion.

200. On the extravascular pool of low density lipoprotein (LDL) in the liver of rats and humans. G. Sigurdsson, D. Shames and R.J. Havel. (1ntr.by Th.V.Gud- mundsson) Department of Med. and Nuclear Med. Univ. of Cali- fornia, San Francisco. Radioiodinated rat LDL was injected intravenously into intact rats and the extravascular pool of LDL in the liver quantified by deter- mining the radioactivity remaning in the extravascular tissues of liver and in plasma at the time of equilibrium between intra-and extravascular radioactivity in the liver (equilibrium time method). The size of the extravascular pool of LDL in the liver thus determined was about 6% of the total intravascular pool. Total extravascular pool of LDL in 8 intact rats, obtained by multicompartmental analysis of the disappearance curve of I 3 I I-LDL from blood plasma was 43% + 15 (S.D.) of the intravascular pool size. Hence the extravascular pool in the liver is about 14% of the total extravascular pool of LDL in the rat. The size of the extravascular pool of LDL in the liver of humans was determined by comparing external scanning over liver to that over head or over heart after an injection of 131 I-LDL intravenously. No significant increase in liver/heart radioact. ratio was observed and only a small increase in liverlheart radioact. ratio did occur. These studies in humans therefore support the studies in rats suggesting that only a small fraction of the total extravascular pool of LDL is in the liver.

176. On the ((rapid beta lipoprotein)) variant. S. Rbssner, L. Holmquist and K. Carlson. King Gustaf V Research Institute, Karolinska Hospital, Stockholm, Sweden. Analysis of serum lipoproteins in grossly obese subjects revealed that about 50% of these patients exhibited a beta lipoprotein. which moved more rapidly than control lipoproteins on agarose electrophoresis, ((rapid beta, r /h . In randomly selected healthy subjects the frequency of r/3 was found to be about 8%. After bypass surgery and subsequent lowering of the LDL cholesterol concentrations in the obese patients, the lipoprotein variant was observed in 80% of these patients. The possible causes for the increased /3 mobility were studied in detail, and the following factors that could

possibly affect the LDL mobility were analysed: Total LDL concentration did not differ between rp and non-rp samples. No significant differences were found with regard to the concentrations of soluble apoproteins and sialic acid. Detailed analysis of the LDL fraction revealed that in rp the LDL, concentration was higher. In some but not all r/3 samples, an increased plasma FFA concentration was found. Drugs did not systematically affect the rp appearance. Dilution of the LDL fraction increased the /l mobility. This was the most likely reason for the increased rp frequency in obese patients after bypass operation but did not explain the high incidence in obesity before any intervention. The rp variant was observed in samples of patients with Type 11 and 111. The rp variant thus may result from several, independent changes of the LDL composition. The cause of the high incidence in obesity is not known as yet.

111. Effect of therapeutic doses of somatostatin (SST) on splanchnic blood flow in man. U. Keller, A. Perruchoud, L. Kayasseh and N. Gyr. Department of Medicine, Kantonsspital, Basel, Switzerland. A preliminary report from our clinic showed that somatostatin (250 pg/hr) is effective in controlling bleeding in patients with peptic ulcer haemorrhage (Z.f.Gastroenterol., Sept.77, Abstr.83). The present studies were performed to evaluate if the dose of SST used has a significant effect on splanchnic blood flow, and if a higher dose (500 pglhr) is more effective. Furthermore, the dose-dependent suppression of gastrin, insulin glucagon and splanchnic glucose production was studied. 10 human subjects (age 27-68 yrs) without hepatic or metabolic disease underwent arterial-hepatic-venous catheterisation. Hepatic blood flow was measured using indo- cyanine green infusion. During 60 min of SST infusion (250 pg i.v. followed by 250 pg/hr) mean estimated splanchnic blood flow decreased from 1.05 f 0.14 to 0.80 f 0.14 L/min (24% decrease, p < 0.01). Blood pressure and pulse rate were unaffected. This dose of SST also suppressed gastrin, insulin and glucagon levels and splanchnic glucose production. The higher dose of SST (500 pg/hr) lowered splanchnic blood flow and suppressed gastrin, insulin and glucagon to a similar degree as the lower dose. The studies demonstrate that SST at the dose of 250 pglhr exerts a significant effect on splanchnic blood flow which is not more pronounced when the dose is doubled. The results suggest that SST in the dose used is capable of significantly lowering splanchnic blood flow in addition to the known suppression of gastric acid output aud gastrin release. These combined effects probably contribute to the therapeutic efficacy of SST in peptic ulcer bleeding.

85. Plasma levels of human pancreatic polypeptide (HPP) under conditions of normal food ingestion and fasting. J.A. Hedo, M.L. Villanuevaand J . Marco. Clinica Puerta de Hierro, Universidad Aut6noma de Madrid, Madrid, Spain. In this work we have examined the daily fluctuations of circulating HPP in normal individuals subjected to a conventional meal schedule as well as during prolonged food deprivation. Volunteers were admitted to our metabolic unit. One group (n=7) received customary meals (breakfast at 9:00 a.m.; lunch at 1:OO p.m.; dinner at 8:OO p.m.). In a second group (n=6) fasting was maintained for 84 hours. HPP was estimated by radioimmunoassay using standard and aatiserum donated by Dr. R.E. Chance. The mean basal (after an overnight fast) H P P concentration ranged from 61 f 15 (SEM) to 73 f 30 pg/ml. Ingestion of each meal was followed by a sustained HPP elevation (peaks: 158 f 31 pg/ml at 11:30 a m . , 551 f 131 pg/ml at 3:00 p m . , 640 2 153 pg/ml at 8:30 p.m.; p < 0.001). Between meals, HPP levels did not return to basal values. In the fasted group, plasma HPP rose significantly 24 hours after the last meal (201 f 67 pg/ml, p < 0.03) and persisted elevated during the remainder of the experimental period, oscillating between 152 f 48 and 218 k 66 pglml. Blood sugar decreased progressively from 83 f 1 to 50 f 2 mg/dl (p< 0.OOOl). Pancreatic polypeptide is secreted after food ingestion and it is thought to possess gastrointestinal functions. Our present data show that plasma HPP remains above basal values throughout the day while on an habitual meal schedule which is compatible with a tonic influence of this hormone on some digestive process. Howe- ver, this concept is challenged by the intriguing rise of H P P observed after prolonged fasting. Such a rise, independently of its significance, might be a consequence of the decline of plasma glucose, as a fall of blood sugar provokes HPP release.

ABSTRACTS 335 146. Elevation of high molecular weight glucagon immunoreactivity (IRG) in plasma of patients with idiopathic hemochromatosis (IH). W.A. Muller, M. Berger, H.J. Ciippers, P. Berchtold, G. Stroh- meyer, H.R. Hofstetter and J.-J. Gonvers. Universities of Geneva, Lausanne, Switzerland and Dllsseldorf, FRG. Diabetes mellitus associated with IH has been thought to be secondary to an iron induced destruction of pancreatic betacells. To test whether the syndrome, IH-diabetes, includes a perturbation of pancreatic alpha-cell function, IRG was measured in 11 patients with IH and diabetes (4 treated with insulin, 7 with abnormal glucose tolerance, GT) and compared with that of 4 patients with IH and normal GT, and with 6 normal subjects. At 30 min of an arginine infusion (460 mg/kg/40 min), all groups with IH exhibited higher IRG levels (antiserum 30-K) than normals (IH: nl GT 496 f 61, abnl GT 593 f 142, insulin treated 451 5 53 pg/ml; normals: 327 f 32; p < 0.05) suggesting hyperglucagonemia in IH regardless of GT. When, however, plasma was treated with charcoal extracting thereby glucagon and similar weight polypepti- des, the remaining IRG (of high mol weight) was elevated in all groups with IH (IH: nl GT 149 f 15; abnlGT 207 f 32, insulin treated 194 f 44; normals: 99 k 11 pg/ml; p < 0.05. The calculated levels of true glucagon (total IRG - high mol weight IRG) were similar in all groups. The data suggest that a) IH associated with diabetes mellitus does not include a disturbed pancreatic alpha-cell function; b) high mol weight IRG is elevated in plasma of patients with IH confirming that (thigh plasma IRG (30-K))) does not necessarily represent high plasma glucagon.

16. Association between functional and morphological findings in insulinomas. M. Berger, P. Berchtold, A.v. Sttilpnagel, H. J. Ctippers, F. A. Gries, D. Griineklee, H.J. Mitzkat, W.A. Miiller, H. Miintefering, W. Wiegelmann, and H. Zimmermann. Universities of Dussel- dorf; Hannover, FRG, and Geneva, Switzerland. To investigate possible relationships between morphological and functional characteristics in insulinomas we have assessed the suppressibility of serum insulin (IRI) by somatostatin (SS, i.v. injection of 250 pg followed by an infusion of 250 pg/h) and diazoxide (D, 600 mg/h i.v.) in 13 patients. In 2 patients with islet cell carci- nomas, SS had no apparent effect on IRI, in the 11 patients with islet cell adenomas the responses were variable. IRI-suppression by the two agents, SS and D, correlated significantly (r = 0.77, p < 0.005). - For further morphological analysis, electron-microscopy has been performed and nine tumors were classified according to the aspect of the beta-cell granules as described by Creutzfeldt et a1 (Diabetologia 9:217, 1973), class I being closest to normal. In patients bearing insulinomas of class I (n=2) and I1 (n=3) , a substantial fall of IRI had been induced both by SS (mean maximal suppression from basal in I: -85% and in I1 -75 f 5 % ) and by D (in I: -79% and in 11: -83 +. 4%). In class 111, however, (n=4, including 1 carcinoma) the abnormal granules were associated with a significant reduction in suppressibility of IRI both by SS (-34 f 3%) and D (-39 f 8%). - In contrast to the variations in IRI suppression plasma glucagon reduction by SS was similar in all patients (- 47 f 3%). These results suggest an association between functional and morphological characteristics of insulinomas, i.e. a decrease of suppres- sibilty of insulin secretion with increasing de-differentiation of the pancreatic beta-cell.

249. Behaviour of carbohydrate tolerance and insulin secretion after manifestation of maturity onset diabetes (controls over 5 years). H.-J. Verlohren, D. Lohmann and W. Heilmann (Intr. by H.-J. Hahn), Stadtkrankenhaus Leipzig, GDR. Immediately after clinical manifestation of maturity onset diabetes long-time investigations of the glucose tolerance and insulin secretion were carried through. In the probands no insulin secretion could be stimulated by administration of glucose. But administration of tolbutamide and glucagon resulted in a clear insulin secretion. Altogether 52 probands were observed for up to three years and 20 patients for up to 5 years. The patients were grouped in depen- dence on the treatment indicated (either purely dietetic treatment or additional treatment with glibenclamide). For judging the above parameters a combined stimulation test was applied (100 g glucose per os, subsequently 1 g tolbutamide intravenously and finally 1 mg glucagon intravenously). After an improvement at the starting period

336 ABSTRACTS the carbohydrate tolerance grew worse in both treatment groups. The insulin secretion, on the other hand, did not show qualitative changes nor was there a quantitative reduction of maximum stimulability. The stimulability of the b-cell is obviously dependent on the type of diabetes concerned and not on its duration.

253. Urinary albumin excretion during conventional treatment and during subcutaneous insulin infusion in diabetic subjects. G.C. Viberti, J.C. Pickap, R.J. Jarrett and H. Keen. Unit for Metabolic Medicine, Guy's Hospital, London, England. Urinary albumin excretion may be elevated in diabetics prior to the development of clinical proteinuria (i.e. Albustix positive). The increased leakage of proteins could induce the development of structural changes which eventually lead to clinical diabetic nephro- pathy. We have studied the 24 h urinary albumin and /I2-microglobulin excretion in 8 clinically non-proteinuric insulin-dependent diabetics (IDD) during conventional treatment and during subcutaneous insulin infusion and in 10 normal controls (C). Meta- bolic control was assessed in the diabetics by three indices: mean blood glucose, M-value and 24 h urinary glucose excretion. During subcutaneous infusion of insulin metabolic control improved in all the patients as judged by the improvement of at least two of the three indices. Urinary albumin excretion rate was higher in IDD during conventional treatment than in C. Subcutaneous insulin infusion significantly reduced urinary albumin excretion in IDD and in most this occurred after only one day of infusion. Urinary albumin excretion rate was completely normalized in 3 patients. The excretion rate of 02- microglobulin in IDD during conventional treatment was not different from that of C and was not uniformly changed by the subcutaneous infusion of insulin. These findings indicate that during conventional treatment the IDD had an elevated transglomerular leakage of albumin which could be reduced or normalized by a short period of good metabolic control.

121. Characteristics of adenylate cyclase of cardiac and lung tissue. W. Krawietz, M. Weinsteiger, E. Erdmann. Medizinische Klinik I der Universitat Miinchen, Germany. Binding studies with labeled /%blocking agents have been successful in analysing specific /l-adrenergic binding sites coupled to adenylate cyclase (AC). AC was measured and experiments fullfilling the criteria of specific pharmacon-receptor-binding were performed with 3H-alprenolol in cardiac and lung tissue (guinea-pig). AC of cardiac (lung) tissue shows a basal activity of 594 f 30 (145 f 9) picomoles cAMP/mg prot.xlOmin, which could be stimulated by isoprenaline, Gpp(NH)p, NaF and maximally by isoprenaline l&M/Gpp(NH)p 6pM to 3740 f 176 (679 ? 50) picomoles cAMP/prot.xlOmin. The stimulation of AC of cardiac tissue by Gpp(NH)p/isoprenaline induces a threefold increase, contrary to that of lung tissue with only an additive effect on the increase by Gpp(NH)p and isoprenaline. Addition of propanolol to tissue was ineffective to inhibit the maximally stimulated AC. These data suggest that in the case of Gpp(NH)p/isoprenaline the hormone or drug may be required only transiently to initiate an interaction of the AC with the nucleotide. MgZ+ concentration-dependently stimulated AC of cardiac and lung tissue distinctly. A maximal effect was measured in lung tissue at 5 mM MgCI,, increasing concentrations of Mg2+ inhibit AC of lung tissue but stimulate that of cardiac. These data suggest the existence of different types of receptor- coupled AC, and may explain the measured distinct effects and binding affinities of some /3-receptor-blocking drugs in heart and lung.

133. Propanoiol withdrawal. T.J.B. Maling and C.T. Dollery. Department of Clinical Pharma- cology, Royal Postgraduate Medical School, London. There is a temporal relationship between sudden withdrawal of chronic propanolol therapy and the development of acute coronary artery syndromes in some patients with ischaemic heart disease. The mechanism is not clear but one possibility is that there may be a period of rebound sympathetic hyperactivity following propanolol withdrawal. Five hypertensive patients without ischaemic heart disease were investigated to elucidate this possibility. Blood pressure, heart rate, plasma propanolol concentration, plasma noradrenaline, plasma renin activity and urinary 24 hour catecholamine excretion

were measured before and for 4 days after propanolol withdrawal. Supine systolic blood pressure rose from mean f SEM 131.2 f 3.2 mm Hg to 153.0 f 3.3 mm Hg, p < 0.001, and on standing from 116.1 f 5.7 mm Hg to 144.5 f 5.6 mm Hg, p < 0.001. Supine heart rate rose from 59.3 f 1.4 beats/min to 77.8 f 1.9 beats/min, p < 0.001 and on standing from 67.4 f 1.8 to 103.2 k 4.3 beatdmin, p < 0.001. There was a 47% reduction in plasma noradrenaline from 0.55 % 0.06 ng/ml prior to withdrawal, to 0.29 f 0.05 ng/ml 3 days after withdrawal, p < 0.02. Urinary catecholamines tended to fall after withdrawal. There was no significant change in plasma renin activity from chronic treatment levels. There was no evidence of rebound sympathetic hyperactivity after propanolol withdrawal.

130. Cardiac interaction between parathyroid hormone (PTH) and beta- adrenoceptor agents in chronic renal failure (CRF). F. Lhoste, T. Drueke, S. Larno, A. Ulmann and J.P. Tillement. (Intr. by F. Lemaire). Department of Pharmacology, Henri Mondor Hospital, Creteil, France. In CRF inconsistant and unreliable action of beta-adrenoceptor agents have been noted. Therefore the effect of /3+ and /3- adrenergic agents were investigated during CRF in the presence and in the absence of severe HPT. In 7 patients with CRF and severe HPT, chronotropic action of isoproterenol has been studied before and after parathyroidectomy (PTX). Tachycardia induced by isoproterenol was increased after PTX. (p < 0.02). Cardiodepressive effect (CDE) of d.1. propanolol was studied in vitro on isolated guinea pig atria (IGPA). Plasma ultra-filtrates from uraemic patients with HPT reduced CDE of d.1. propanolol as compared with ultra- filtrates from uraemic patients after PTX. Pure synthetic PTH decreased the effects both of f i + and /% adrenergic drugs. Similar PTH effects were obtained with calcium influx antagonists; D. propanolol and verapamil : PTH also reduced o+ adrenergic drug effect on rat seminal vesicles. The results show that PTH in vivo during CRF and in vitro interferes with adrenoceptor agents. An interaction between PTH and adrenoceptor agents on transmembranous calcium transport is suggested.

173. Acute and chronic beta adrenoeeptor blockade: influence on haemw dynamic effects and plasma noradrenaline changes after sodium nitroprusside infusion in conscious rabbits. J.L. Reid, B. Fraser and C. Hamilton Department of Clinical Pharmacology, Royal Postgraduate Medical School, London. The mechanism of hypotensive action of beta adrenoceptor antagonists remains disputed. Neurogenic mechanisms have been proposed both on the central nervous system (Reid et at, J.P.E.T. 188, 294, 1973) and at a peripheral presynaptic site (Langer, B.J.Pharm. 60, 481, 1977). Such actions should result in impaired sympathetic function with reduction in noradrenaline release. We have investigated the influence of short term (2 hours) and long term (3 weeks) treatment with propanolol on mean arterial pressure (MAP), heart rate (HR) and plasma noradrenaline (NA) in conscious rabbits (n = 6) before and after intravenous (IV) sodium nitroprusside (SNP) 40 pg/kg/rnin. SNP reduced MAP from 81.4 f 3.9 to 57.3 f 3.4 mm Hg and increased HR from 223 f 8 to 343 f 6 beats/min and plasma NA from 0.24 f 0.06 to 0.63 f 0. I8 &I. IV propranolol (2 mg/kg/hr) for 2 hours lowered MAP to 75.4 f 4.4 mm Hg (p < 0.05) and increased plasma NA to 0.56 f 0.17 pgA (p < 0.05). SNP still lowered MAP (54.3 f 4.8 mm Hg) and increased plasma Na (1.20 f 0.26 pg/l) although HR rise was significantly reduced (p < 0.01). Propanolol2 mg/kg IP twice daily for 3 weeks did not significantly alter resting plasma NA or MAP. The changes in MAP, HR and NA after SNP infusion were not different to untreated controls. Sino-aortic denervation (SAD) increased MAP, HR and plasma NA. The fall in MAP after SNP was increased (p < 0.01) but HR and plasma NA did not rise. IV propanolol did not alter plasma NA in SAD rabbits. These results d o not reveal effects of beta blockers on baroreflex mediated increases in plasma noradrenaline.

145. Long-term monitoring of propanolol blood levels in essential hypertension. Their relevance to clinical effect. P.L. Morselli, G. Bianchetti, J.L. Elghozi., P. Meyer*. Synthe- labo L.E.R.S., 75013 Paris, F:ance. 'Physiologic & Pharmacologie, INSERM U.7, Hapital Necker, 75015 Paris, France.

Although a number of studies have established a relationship between peak plasma levels of pblockers and bradycardia or hypotensive response, there appears to be little data related to steady state plasma levels and clinical effect. We have monitored plasma blood levels at regular intervals, for 4 to 6 months, in ten newly treated patients with essential hypertension. Plasma propanolol was measured with the use of gas chromatography. No other drugs were associated, and doses ranged from 40 to 160 mg b i d . Blood samples were collected at 8-9 hours after drug intake. A clear dissociation between effect on heart rate and on arterial blood pressure was observed. In 6 cases reduction of arterial blood pressure was observed only when steady state blood levels reached values of 20-40 ng/rnl. At levels inferior to I5 ng/ml, hypotensive response was practically absent but a decrease in cardiac frequency was always present. Blood levels higher than 50 ng/ml were not associated with an increase of hypotensive response but on the contrary could be related either to severe side-effects or paradoxical responses. In one case a complete lack of effect was associated with an extremely low bioavailability. In two other cases where no hypotensive response was observed despite blood levels of 40-50 ng/ml and a diuretic treatment was also unsuccessful. The data indicate that optimal blood levels range from I5 to 40 ng/ml at steady state. That is to say a relationship exists between plasma steady state blood levels and hypotensive response. Routine monitoring of plasma blood levels can therefore help in the management of hypertensive patients treated with /%blocking agents.

54. Effects of somatostatin on lipolysis of isolated human fat cells. H.H. Ditschuneit, S. Raptis, R. Fussganger, H. Ditschuneit and

J. Rosenthal. Div. of Nutr. and Metab., Dept. of Med., University of Ulm, FRG. Recently we observed in vivo that in man Somatostatin (Growth Hormone Inhibiting Factor (GIF) ) enhances the orciprenaline induced elevation of plasma FFA. This effect could be due to a GIF- induced inhibition of insulin secretion or to a relative glucagon increase caused by orciprenaline or to a direct lipolytic action of GIF. In order to elucidate the action on lipolysis we studied the effect of GIF on isolated human fat cells. GIF increased catecholamine-induced lipolysis at concentrations of 2-50 pg/ml. The antilipolytic action of insulin was not affected, indicating that GIF has no insulin antagonistic effect. Binding studies showed that GIF has an effect on the binding of H3-isoproterenol to the fat cell membrane. The apperent number of binding sites as well as their affinity for H3-isoproterenol was increased. This observation could be the cause of the enhancement of the orciprenaline induced elevation of plasma FFA by GIF.

162. Effecls of gold salts, aspirin and betamethasone on the in vim chemw taxis of human neutrophils. A. Pecoud, A. Leimgruber and P.C. Frei. Division of Immunology and Allergy, Department of Medicine, CHUV, Lausanne, Swit- zerland. In order to examine the mechanisms of the action of various antiinflammatory drugs, we tested the effect of gold salts, aspirin and betamethasone on the chemotaxis of neutrophils. Using a modified Boyden chamber, the drugs were added to either the upper chamber (uc) which contained 106 neutrophils from healthy human volunteers, or to the lower chamber (lc) which contained homologous plasma activated by aggregated gamma-globulin. Both chambers were separated by a filter (Millipore, 3 p) and chemotaxis was assessed by counting the total number of neutrophils reaching the Ic. The addition of gold salts (300 ug% of gold) to the uc reduced chemotaxis by 27 + 3% (mean * sem; n = 16; p < 0,001) and to the l c by 1 1 + 5% (n = 9; p < 0,05). This inhibitory effect was significantly reduced by washing neutrophils after incubation with the drug and was found to be dose-dependant. At concentrations varying from 1.5 to 150 mg%, aspirin was not found to have any effect when added either to the uc or the Ic. Betamethasone (1 pg%) decreased neutrophils chemotaxis by 21 + 3% (n = 9; p < 0,001) when added to the uc, but had no effect when added to the Ic. These data tend to indicate that the antiinflammatory drugs tested might act through different mechanisms and that neutrophils chemotaxis reduction might be an important mechanism for the action of gold salts.

a. DNA excretion by resting or stimulated human blood lymphocytes P. Anker, M. Stroun, and P.A. Maurice, Division of Onco- Hematology, Hapital cantonal, Geneva, Switzerland. Unstimulated human blood lymphocytes have been shown in this

ABSTRACTS 337 laboratory to excrete in vitro a nucleoprotein complex containing DNA and RNA, both nucleic acids being released according to a homeostatic mechanism. We now report on the effect of mitogens or specific antigens on the release mechanism of DNA by the same cells. Lymphocyte suspensions (lo6 cells/ml) were cultured for 3 days in presence of PHA or for 5 days in presence of PPD and labeled with [3"] TdR at the end of this culture period. After medium renewal the culture was extended for 2 additional days in absence of mitogen or antigen and [3"] TdR, and the radioactivity of he DNA was determined daily in the acid-precipitable material obtained from the cells and from their supernatant. In parallel cultures the amount of DNA excreted daily in the medium was determined after its extraction and purification from the released material. Peak cellular radioactivity was observed at day 3 after PHA stimulation and at day 5 after PPD stimulation. During the following two days a sharp decrease in cellular acid-precipitable radioactivity was observed whereas extracellular counts increased to values representing 77 to 95% of the radioactivity originally present in the cells. The amount of DNA released at the same time by the lymphocytes was of 50 to 100pg/day/100 ml of culture. These results suggest that mitogens or antigens suppress the homeostatic mechanism controlling DNA release in unstimulated lymphocytes.

51. Modulation of the kinetics of human non specific cell mediated cytoloxicity (CMC) by corticosteroids. E. Dupont (introduced by J.-L. Vanherweghem), Department of Nephrology, UniversitC Libre de Bruxelles, Brussels, Belgium. Kinetic studies have provided useful informaions on the mechanisms of lysis induced by CMC. Introduction of corticosteroids (CS) in vitro (methylprednisolone 10-3 M/ml) after interaction between effectors and target cells is characterized by a corticosensitive period during which cytolysis can still be inhibited. Antibody-dependent cell-mediated cyto-toxicity (ADCC obtained whith 51Cr labelled lymphocytes sensitized with rabbit ALS) and mitogen induced cytotoxicity (MICC with PHA or con A and 51Cr labelled CRBC) were compared. Lysis by ADCC is almost immediate and the corticosensitive period does not exceed 10 min while MICC is slower (lag period of lysis as well as corticosensitive period exceeding one hour). These results suggest that beside their metabolic action, CS affect the cellular bridging processes leading to lysis either at the specific receptors level or by a membrane protecting effect.

65. Exploration of suppressor T cell function in the human. P. Galanaud, J.F. Delfraissy, P. Segond and J. Dormont. INSERM U 131, Hapita1 Antoine Bbclkre, Clamart, France. As shown in several experimental models, T suppressor cells play an important role in the regulation of the antibody response. Study of the suppressor function of T cells in the human has been impaired by difficulties in obtaining a specific in vitro antibody response from human Peripheral Blood Lymphocytes (PBL). We have recently described a method allowing the induction of a primary anti- hapten IgM antibody response in human PBL cultures. The addition of Concanavalin-A (Con-A) at the initiation of the cultures induces a suppression of the B lymphocyte response: 0.5 pg/ml of Con-A led to a suppression of more than 50% (mean suppression 73 f 3%) in all of the 20 normal individuals studied. This suppression can be obtained when Con-A activated T cells (cultured for 48 hours in the presence of Con-A) are added to fresh cells, and does not require histocompatibility between siAppressor and responding cells. Although the suppressive concentrations of Con-A are below the optimal concentration for the induction of T cell proliferation (20 pg/ml), cell proliferation is required for the generation of suppressor cells. Thus, the suppressive effect of Con-A activated T cells on the in vitro antibody response can be proposed as a method to evaluate the functional state of T suppressor cells. Such an evaluation should be important in several immunopathologic situations.

211. Serum complement abnormalities and androgen therapy of hereditary angioedema. A.T. Sobel, M. Moisy, C. Blanc. D. Belghiti, Y. Gabay, L. Intrator and G. Lagrue. Nephrology and lnserm 139, HBpital Henri Mondor, Creteil, France. Hereditary angioedema (HANE) is a rare, life-threatening disease due to the deficiency of C1 inhibitor (cl Inh). Androgen therapy has been recently shown to be effective for prophylaxis of HANE attacks. Since lifelong androgen therapy may be hazardous, this study was designed to define the minimal does required for

338 ABSTRACTS effectiveness. Nine patients from 5 different families were treated during cumulative 41 months by Danazol (an impeded derivative of ethinyltestosterone) and/or methandrostenolone. They received various doses of both drugs ranging from 112 tablet to 3 tablets/ day. Serum levels of CI Inh, CH50, C4, C3, C2 and Clq were regularly determined by standard immunochemical or hemolytical methods. Before treatment, the patients had an average of 2.7 attackdmonth. Their complement profile was typical, characterized by low CI Inh (mean 5mg/100 ml), low C4 (3mg/lD0ml), low C2 /49 % of normal) and normal vallles of CH50, C3 and Clq. One tablet of either Danazol (200 mg) or methandrostenolone (5Omg) was the minimal requirement to prevent any attack in all patients. When 3 tablets day of either drug were given, complement abnormalities were rapidly reversed : CI Inh rose to 14mg/100ml, C4 to 36mg/lOOml and C2 to 100 %. It was noticed that simultaneous CH5O levels were significantly higher than normal (60u/ml) and that C3 fell to 80mg/100ml. When 1 tablet/day was given the biological effect was barely detectable : CI Inh : 8mg/ml, C4 : 6mg/ml, normal values for CH50, C3 and Clq. Thus, this study confirmed the known clinical and biological efficacy of 3 tablets/day of two different androgen derivatives. The data show that an excellent clinical result can be obtained with much lower doses, even if biological abnormalities are not simultaneously reversed. They seemed important with respect to the serious dose- dependent risks of androgens.

109. Occurrence of IgA-deposits in clinically normal skin, liver and glomerular mesangium of patients with alcoholic hepatic disease. L, Kater, A.C. Jbbsis, E.H. Baart de la FaiUsKuyper, S.S. Wagenaar, R. Grijm and A.J.M. Vogten. Div. Immunopathology, dep. of Medicine and Pathology, University Hospitals Utrecht and Amsterdam, The Netherlands. We investigated the extent to which the immunofluorescent phenomenon of homogeneous deposition of IgA in the liver sinusoids (so called continuous pattern) was specific for alcoholic liver disease. In 66 of 320 liver biopsies a continuous IgA pattern was observed. Alcoholism was mentioned in 50 of these 66 patients (76%). The biopsies of the remaining 254 cases showed no or scanty detectable IgA (discontinuous pattern). In the latter group only 8 patients (30/0) gave a history of alcoholism. A direct correlation between continuous IgA pattern in the liver sinusoids and alcohol is therefore concluded (p < 0.001). An additional finding is the concomitant occurrences of IgA in the perisinusoidal linings of the liver, in the walls of superficial cutaneous capillaries, in capillaries of the gut and in the glomerular mesangium of patients with alcoholic liver disease. There is a striking similarity between these observations and those found in skin, gut and kidney of patients with either anaphylactoid purpura or recurrent, often benign, hematuria. The experiences strengthen the assumption that either IgA aggregates or antigen complexed with IgA may circulate in the bloodstream of these patients.

191. The influence of changes in blood volume on the haemodynamic effects of positive end expiratory pressure (PEEP). J.J. Schreuder, J.M. Bogaard, J.R.C. Jansen and A. Versprille. Dept. of Pulmonary Diseases, Erasmus University, Rotterdam, Holland. In our study the decrease of cardiac output (CO) with PEEP was investigated at three different levels of bloodvolume in 8 young healthy curarized pigs (7-10 kg) during artificial ventilation under pentobarbital anaesthesia. The levels of bloodvolume were normovolaemia, hypervolaemia (+ I5 ml dextran per kg body- weight) and hypovolaemia (- I5 ml kg-'). PEEP was rised continuously with .7 cmH20 min-1 up to 15 cmH20 in all three volaemic states. The absolute decrease of CO with PEEP is significantly larger at a higher bloodvolume. The relative decrease shows a positive trend with bloodvolume. In the normovolaemic state the relation of CO v. PEEP shows a three phasic decrease with a steeper slope between approximately 2 to 9 cmH20 (phase 2) than below 2 (phase 1) and above 9 cmH20 (phase 3). Aortic pressure (Pao) decreased only significantly in phase 2; heart rate (HR) did not change. Measurements of the transmural pressure of the pulmonary artery (Ppa,t) cancelled the hypothesis of a higher afterload of the right ventricule as an explanation for phase 2: In the hypervolaemic state phase 2 is shifted to a higher level of PEEP. Only in this phase an obvious decrease of Pao and HR was observed. During hypovolaemia CO v. PEEP shows a gradual

decrease over the whole range of PEEP, without a change of P a o but with a continuous rise of HR. In the hypovolaemic state the autonomic control of bloodpressure seems to be more active than in circumstances with higher bloodvolumes. This effective control might probably also explain the tendency to a relatively smaller decrease of CO with PEEP in the hypovolaemic state.

125. Trial of systematic neutralization of shunt with positive-end- respiratory-pressure (PEEP) in acute respiratory distress syndrome (ARDS). J . Labrousse, A. Tenaillon, J. Chastre, J.L. Bourgain and J. Lissac (Intr. by A. Versprille). Department of intensive care, Boucicaut Hospital, Paris, France. The aim of this study was to obtain a Pa 022400 mm Hg (Fi 02 = 1) by maximum alveolar recruitment using PEEP in ARDS. In 12 artificially ventilated patients the level of PEEP needed to achieve this goal ranged from 10 to 35 cm H20, in most cases 20 cm H20. When the cardiac output (Swan - Ganz catheter, thermodilution) decreased on PEEP it was systematically supported by vascular replenishment and/or dopamine. The Pa 02 raised from 99.3 f 50 mm Hg ( x * SD) on IPPB to 430 f 41 mmHg on PEEP (p < .001); the shunt simultaneously decreased from 40.7 f 11.8% to 15.3 f 3.7% ( p < .001). Although the wanted leved of Pa 02 was reached in all patients, Pa C 0 2 and/ or total lung compliance were impaired in some cases at a high level of PEEP. No pneumothorax occured in the present series. In 5 patients PEEP was progressively decreased until weaning from the respirator. In the other cases it was impossible to stop PEEP and patients died. These findings point out that in ARDS: a) PEEP appears to be able to neutralize the pulmonary shunt, but it needs sometimes a high level; b) cardiac output has to be simultaneously supported; c) respiratory mechanics are not a reliable test of the best level of PEEP.

152. Modifications of left ventricular end diastolic pressure and pulmonary capillary pressure by changes in positive end expiratory pressure. A. Net, G . Vazquez, R. Martinez, G. Grange, A. Artigas, L. Marruecos, E. Quintana, S. Benito (intr. by A. Versprille). Unidad de Cuidados Intensivos Generales del Hospital de la Santa Cruz y San Pablo. Barcelona. Espana. To observe the modifications on the Mean Pulmonary Capillary Pressure (PCP)/Left Ventricular End Diastolic Pressure (LVEDP) gradient introduced by increase of the mean intrathoracic and alveolar pressures, 21 intubated and ventilated mongreal dogs had their pulmorary artery and left ventricule catheterized, their PEEP, increased by 3 cm H20 increments every ten minutes up to 24 cm H20. They all had an esophageal balloon placed to allowe recording of esophageal pressure. A maximal difference of PCP/LVEDP < 3 mmHg was accepted as normal. Dogs were divided into 4 groups: Control (4). norm0 (S), hyper (5) and hypovolemic (4). Hypervolemia was obtained with Ringer infusion loading, and hypovolemia was produced with i.v. furosemide. In normovolemic dogs PCP/LVEDP gradient with PEEP 18 cm H20 was 4.38 * 1.09 mmHg (p <0.05). In hypervolemic dogs the gradient was 3.06 f 0.81 (p < 0.01) with PEEP of 15 cm. H20. In the hypovolemic group no statiscally significant differences were observed. These results suggest that with high values of PEEP the PCP does not reflect the concomitant changes of LVEDP, but probably is influenced by a change in pressure originated in the alveoli or pulmonary veins.

206. Cardiorespiratory responses to peep challenge. A. Sladen, M. Klain and A.J. Sweatman. SICU, Montefiore Hospital, Pittsburgh, PA, USA. Cardiorespiratory responses to PEEP challenge were observed using computerized physiologic profiles. 253 profiles were performed with incremental PEEP from 0 to 15 cm H20 in a heterogeneous group of 44 adult patients mechanically ventilated in the Surgical ICU (I6 survivors4, 28 non-survivors--NS). S: 114 studies in 16 patients. Increasing PEEP 0 to I5 cm H,O resulted in the following mean changes. Effective compliance (CEf) 29.1 * 2.8 to 36.2 f 3.0 ml/cm H20. Cardiac index (CI) 2.94 * 0.24 to 2.40 f 0.41 L/min/M2, left ventricular stroke work (LVSW) 38 2 2.3 to 28 * 3.6 gm/M/M2, total peripheral resistance (TPR) 2500 f 274 to 2600 * 354 dyne.sec.cm-5M2, oxygen consumption (Q03 114 f 7.9 to 99 f 1 1 ml/min/M2, intrapulmonary shunt 31 f 3.2 to

ABSTRACTS 339 23 f 2.9'70, arterial oxygen tension (pa03 (FPZ = 1 .O) 201 f 27 to 285 f 43 torr. NS: 139 studies in 28 patients. C,, 27.6 f 2.6 to 31.8k 1.9. CI 3.03 20.28 to4.45 f0.38, LVSW 28 f 3.4 to47 k 4.8,

* 3.0 to43 f 2.3, paO,122 -c I5 to 175 k 19. There was no significant difference between the changes in C,, in the S and NS groups when subject to PEEP challenge. In the NS group CI, LVSW, and Qs/QT increased with PEEP while the reverse was seen in the S group. The hypothesis that the diverse responses in CI to PEEP challenge in the two groups was directly related to differences in C,, prove to be unfounded. PaO, increased in both groups but the mechanisms were entirely different in each group. In the S group Qs/QT decreased whereas in the NS group Qs/QT increased and the rise in paO, resulted from an increase in C1. Caution is advised that with PEEP challenge an improvement in pa0, may be due to an increase in CI rather than a decrease in Qs/QT and that an increase in CI, LVSW, QOz and Qs/QT coupled with a decrease in TPR in response to PEEP challenge predict an unfavorable outcome.

TPR2036f398 tO1150f 119,Q,z93f 11.8 tO100+6.2,Q,/Q,37

67, Positive end-expiratory pressure (PEEP) effects on the magnitude of pulmonary shunt (PS). F. Garrigosa, V. Diez Balda, J.R. Oncins, J.F. San Juan and J.M. Cavanilles (Intr. by A. Versprille). Servicio de Medicina In- tensiva. Hospital Principes de Espana. Barcelona. Espana. To assess the beneficial effects of PEEP on pulmonary alterations causing PS this ventilatory modality was used during short periods of time in order to avoid the influence of other therapeutical interventions. 35 patients that needed mechanical ventilation where included in the study. Analytical determinations were done in spontaneous breathing, on IPPB and with PEEP of 5 and 10 cmH20: Every study was realized after breathing 100% oxygen over a 10 minutes period of time. The four steps followed in each study were separated by rest periods of at least 15 minutes. The FiO, in each was accurately checked. PS was calulated with the Bergree's equation. The standard alveolar gas equation was used. HbOz saturation and Hb were measured with a properly calibrated co- oximeter. PO,, PCO, and pH were obtained with standard electrodes. Following the change from spontaneous breathing to OPPB, PS decreased in more than 5% in 33.3% of the patients, did not change in 44.4010 and increased in 22.2%. Following the change from IPPB to PEEP of 5 cmH20, PS decreased in 20 To of the patients, increased in 6.7% and did not change in the remaining 73.3qo. Following the pass from PEEP 5 to PEEP 10 the amount of PS decreased in 19.2% of the cases, increased in another 15.4% and remained invariable in the remaining 65.4%. The differences on the magnitude of PS with IPPB and different degrees of PEEP were not stadistically significant (P < 0.1). PEEP by itself might have a detrimental effect on patient's oxygenation when used during short periods of time. The beneficial effects of PEEP on patient's oxygenation when used during long periods of time may be directly related to it or to other therapeutical interventions. We can not deny the possible beneficial effects in preventing and improving atelectasis.

1. Intermittent mandatory ventilation (IMV). Preservation of normal patterns of thoraco-abdominal coordination, and avoidance of muscle fatigue during weaning from artificial ventilation. J.B. Andersen, J.P. Rasmussen, P. Howardy, and T. Kann (Intr. by A. Versprille). Department of anaesthesia, Herlev Hospital, Herlev, Denmark. Thoraco-abdominal coordination was studied by means of loop traces of anterior posterior diameter changes of chest and abdomen (Konno and Mead's method) during weaning of patients from artificial ventilation, conventionally or by an IMV system. Simulta- neous studies were made of the phenomenon of muscle fatigue as reflected by decrease in the ratio of high and low frequency of EMG signals recorded from the 6th and 7th intercostal spaces (Lindstr4ms method, modofied by Macklem). Fourteen patients have been studied, six with acute exacerbation of chronic obstruc- tive pulmonary disease, and eight with acute respiratory failure with no history of respiratory disease. During conventional weaning (progressively increasing periods of spontaneous respiration) abnormal loop changes of two types were seen in 10-20 minutes - wide clockwise loops (chest out, abdomen in), and loops resembling a disordered isovolume line (reflecting gross incoordination; shift of air from thorax to abdomen and virtually no lung ventilation). Nine patients displzyed more or less abnormal patterns. Signs Of muscle fatigue were observated in ten patients. During IMV weaning with the respirator set to give one half of the predetermi- nated respiratory frequency (equivalent to the patient being out of the respirator for 30 minutes every hour), no abnormal loops were seen, and muscle fatigue was not observated. I t would seem that

thorac*abdominal incoordination and muscle fatigue are implicated when conventional weaning fails, and, further that IMV is a very promising prophylactic alternative.

64. Increased plasma adrenaline concentrations in benign essential hypertension. R. Franco-Morselli, J.L. Elghozi, E. Joly, S. Di Giulio', P. Meyer. Physiologie & Pharmacologie, INSERM U7, Hapita1 Necker, 75015 Paris, France. *Service de Therapeutique Nephrologique, Hbpital Necker, 75015 Paris, France. Plasma adrenaline, noradrenaline, and dopamine concentrations and plasma renin activity were measured in the supine position and after standing for 10 minutes in 14 patients with sustained benign essential hypertension and in five patients with labile hypertension. Results were compared with values obtained in 11 normotensive control subjects. In controls plasma noradrenaline concentrations increased with age, while plasma adrenaline values tended to decrease with age. No significant difference in mean plasma noradrenaline was found between hypertension and control subjects, but plasma noradrenaline seemed slightly increased in a proportion of hypertensive patients aged less than 50. Plasma adrenaline was considerably raised in both supine and standing positions in eight patients with sustained hypertension and in two with labile hypertension. Dopamine concentrations and plasma renin activity were similar in all groups studied. The finding of significantly raised plasma adrenaline concentrations in a large proportion of hypertensive patients supports the hypothesis that the activity of the sympathetic nervous system is increased in essential hypertension. Measurement of plasma adrenaline seems to be a more sensitive index of this activity than that of plasma noradrenaline.

119. Serum and arterial tissue cholesterol in severely obese patients. J.G: Kral, G. Bondjers and A. Gustafson. Depts of Surgery and Medicine, Univ. of GOteborg, Sahlgrenska sjukhuset, GOteborg, Sweden. Serum lipoprotein levels in 19 normolipoproteinemic severely obese (105-210 kg) patients, 14 women and 5 men aged 19-43 yrs, were determined before end-side jejuno-ileostomy (J-I) and at 6 month intervals following the operation. In 11 of the women and 2 of the men cholesterol content of mesenteric arterial tissue obtained during the operation was related to the cholesterol content of different serum lipoprotein fractions. Serum HDL cholesterol levels were significantly lower than in normal weight controls. There was a statistically significant negative relationship between serum HDL cholesterol and that in arterial tissue (rr = -0.80). Already 2 weeks after J-I there was a staistically significant (p < 0.01) decrease in serum HDL cholesterol from 1.2 to 1.0 mmol/l persisting throughout observation periods of two years (n = 7). Even though there was a significant decrease in total serum cholesterol from 5.5 to 3.3 mmol/l, the operation might have an unfavorable effect on arterial tissue cholesterol.

23. Baroreceptor control of atrioventricular conduction in man. 0. Bonazzi, A. Ferrari, L. Pozzoni and 0. Mancia, lstituto Ricerche Cardiovascolari, Patologia Medica I and CNR, UniversitA di Milano, Milano, Italy. Although there is plenty of evidence that in man baroreceptors expert a marked influence on sino-atrial node, little is known of their influence on atrio-ventricular node. In 8 subjects with normal cardiac conduction we altered baroreceptor activity by i.v. injection of phenylephrine and trinitroglycerine and recorded simultaneously intraarterial blood pressure, ECG and His bundle electrogram. We related changes in mean arterial pressure (& 25 mmHg, MAP) to resulting changes in R-R, A-H, H-V intervals (msec). At sinus rhythm increase and decrease in MAP caused linear lengthening and shortening of R-R interval (mean regression ccefficients 12.7 f 2.3 and 9.1 f 1.5 respectively) but no change in A-H interval. However when in the same subjects heart rate was kept constant by pacing, increase and decrease in MAP caused linear lengthelling and shortening of A-H interval (regression ccefficients l . S f 0.3 and 1.1 k 0.2). In both cases there were no changes in H-V interval. Thus baroreceptors have a marked influence on supra-though not on infra-hissian conduction. This influence is unmasked when pacing prevents baroreceptor influence on si noatrial node.

62. Quantitation of cardiac glycoside receptors in human heart and erythrocyte. E. Erdmann, W. Krawietz and P. Presek. Medizinische Klinik I der Universitat Miinchen, Germany.

340 ABSTRACTS Cardiac glycoside (CG) binding to their membrane-bound specific receptors preceeds their pharmacological effects. Previous in vitro experiments (animal) on electrically stimulated ventricular strips have shown that 50% of maximal positive inotropic effect of 3H-ouabain is obtained if 50 % of total cardiac glycoside receptors have bound a drug molecule. Thus, the quantitation of CG- receptors and their affinity for glycosides is of importance. Using radioactively labeled ouabain (digoxin) the total number of receptors in human right and left ventricular tissue was determined as - 1014/g wet weight or as - 1000/pm2 cell surface at optimal equilibrium binding conditions. Half maximal 3H-ouabain (3H- digoxin) binding occurred at 3nM (2nM) in the absence and at 9nM (50nM) in the presence of serum. The amount of CG-receptors measured postmortem in left ventricle is greatly reduced in infarcted areas, the receptor affinity remains unchanged, however. In order to investigate changes in patients in disease, a method was developed to measure CG-receptors quantitatively on their erythrocytes. Experimental results so far show that the number of CG-Receptors is increased in chronic hypokalaemia (513 f 110, n= 14, in acute hypokalaemia 256 f 54, n = 2 in control 259 f 51, n= 41). After treatment the normal nomber of CG-receptors/single erythrocyte was found again within 75 - 130 days. It is concluded that some forms of hyper - or hyposensitivity to cardiac glycosides may origi- nate from an altered number of cardiac glycoside receptors.

182. Metabolic and Electrophysiological Effects of Glucose in Acute Myocardial lschaemia in Dogs. D.C. Russell, H.J. Smith, M.F. Oliver. Departments of Cardiology and Medicine, Royal Infirmary, Edinburgh. Coronary occlusion produces rapid alterations in cardiac metabolism, blood flow and electrophysiological properties of the myocardium that rnay lead to ventricular fibrillation (VF). The effect of glucose infusion during successive 5 minute occlusions of the anterior descending coronary artery has therefore been examined in dogs by combined measurements of metabolic gradients (arterial - local coronary venous differences of glucose, potassium, lactate and free fatty acids), regional myocardial blood flow and intracellular and extracellular potential change. Data was obtained during successive occlusions following an initial sham occlusion. Glucose infusion (arterial levels 12.5 f 1.2mM) transiently reduced epicardial action potential (AP) shortening and endocardial- epicardial conduction time (CT) after high coronary occlusion (severe ischaemia) but had no effect on incidence of VF. By contrast following lower coronary occlusion (moderate ischaemia) AP shortened from 178 f 3 msec to 154 f IS msec compared with a shortening from 175 f 2 msec to 129 f 8 ms during control occlusions (p < 0.05). CT prolongation was also less during glucose infusion. These changes were associated with increased arterial- local coronary venous differences of glucose. Regional myocardial blood flow and other metabolic gradients were unchanged. It is suggested that increased availability of glucose is protective against early ventricular arrhythmias in moderate but not severe myocardial ischaemia.

48. Evidence for HLA linked genetic susceptility in type I Diabetes. A.G. Cudworth (Intr. by C.J. Dickinson). Medical’ Unit, St. Bartholomew’s Hospital Medical College, London. Four hundred and forty patients with classical ’juvenile’ type insulin dependent diabetes (age of onset 1 - 79 years) have been HLA typed. There is a significant positive association between this type of diabetes and particular HLA phenotypes (BS, B18, BlS, B40; and a significant negative association with other HLA B alleles (BS, B7). An additive relative risk occurs in subjects who possess two ’high risk’ alleles, which has an important effect on the prevalence of the disease in sibships and possibly on the concordance rate in identical twins. There is suggestive evidence that particular HLA combinations are associated with increased Coxsackie neutralising antibody titres in 173 newly diagnosed diabetics and also with the persistence of islet cell antibodies. Forty families with a pair of affected siblings have been HLA genotyped. There is a significant departure from the expected random zygotic assortment of HLA haplotypes in the affected siblings. These data are consistent with the presence of at least two genes in the HLA chiumosomal region which determine susceptibility to this disease (Type 1 diabetes).

18. Clinical evaluation of high serum levels of the aminoterminal type 111 procollagen peptide in Fibrotic liver disease. E. Hahn, L. Vargas, H. Rohde., R. Timpl*, Ch. Bode and G.A. Martini. Medical University Hospital, Marburg/Lahn and

Max-Planck-Institute for Biochemistry*, Martinsried, Fed. Rep. Germany. A highly sensitive radioimmunoassay had been developed which allowed the detection of the aminoterminal type I11 procollagen peptide (N-111 PP) in sera of patients with liver disease (Rohde. Hahn, Timpl: Fresenius Zschr. Analyt. Chemie, in press). The precise relationship between the fibrotic process and serum levels of the procollagen peptide remained to be established and will be presented here. N-111 P P values in serum are expressed as percent inhibition of rabbit antibodies reacting with iodine-125 labeled N-I11 P P of calf origin. In healthy persons (n = 8) the range was 0 - 10% inhibition. In all sera from patients with chronic active hepatitis (n = 5 ) N-I11 P P activity was markedly elevated (20 - 51%) with high values in those patients without histologic evidence of fully developed fibrosis. In patients with liver cirrhosis (n = 13) there was a marked variation of N-I11 PP activity. Highest values were found for patients with active cirrhosis (16 - 517’0). Normal levels could be detected in 4 patients with inactive cirrhosis (3 -12%). These results suggest a close correlation between clinical and histologic activity of the fibrotic process in the liver and the serum level of N-111 PP.

181. Response of inactive renin to inhibition of prostaglandin synthesis. K.W. Rumpf, B. Schkhterle, S. Schmidt, K. Becker and F. Scheler (Intr. by P. Schauder). Department of Medicine, University Hospital, GOttingen, Germany. Plasma renin activity has been shown to decrease after inhibition of prostaglandin synthesis (Rumpf et al., Prostaglandins 10, 641, 1975). However, the response of inactive renin -- a higher molecular weight form of renin, which is activated by acidification -- is not yet clear. We therefore investigated the response to inhibition of prostaglandin synthesis by indomethacin in 20 normal volunteers using a modification of the method introduced by Derkx et al. (Lancet 11, 496, 1976). Furthermore, the response to stimuli (furosemide, orthostasis) was tested. -- We found a high percentage of inactive renin in plasma averaging about 80% of total renin. Inactive renin was not changed by inhibition of prostaglandin synthesis (28.5 f 2.9 ng/ml/h before and 26.2 f 3 ng/ml/h after inhibition of prostaglandin synthesis; p < 0.6). Stimulation of renin by orthostasis led to a significant increase in inactive renin (33.5 f 3.6 ng/ml/h; p < 0.01) as well as in active renin. This increase in inactive renin was diminished after inhibition of prostaglandin synthesis (29.4 ng/ml/h; p < O.l) , whereas active renin still increased significantly. Furosemide (20 mg i.v.) did not induce a further increase in inactive renin concentration either before (33.9 ng/ml/h; P < 0.9) or after (29.4 ng/ml/h; p < 0.9) inhibition of prostaglandin synthesis. Active renin, however, increased significantly. --We conclude, that inactive renin remains unchanged after inhibition of prostaglandin synthesis. There is furthermore no evidence from our experiments, that increases in active renin are accompanied by parallel decreases in inactive renin, which has been claimed by other authors. The physiological significance of inactive renin still remains to be clarified.

185. The effect of indomethacin on PRA and aldosterone in essential hypertensive patients. A. Salvetti, R. Pedrinelli, L. Poli, P. Sassano, F. Arzilli. 1st Medical Clinic, University of Pisa, Italy. The effect of indomethacin (IND) (200 mglday) on supine (s.PRA) and upright (u.PRA) PRA, urinary aldosterone (u.Ald), 17 OH and plasmacortisol (F) was studied in 23 essential hypertensive pts and compared (11 pts) to Oxprenolol (OXP) (100 mg/day) by 2 x 2 factorial trial. IND decreased s.PRA (1.20 2.23 vs. 66 f .11; p < ,015 u.PRA (3.27 f .61 vs 1.81 f .23; p < .01) and u.Ald (14.55 f 1.64 vs 11.43 f 1.42; p < .025): its effect was maintained when IND was given for 3 days (8 pts) and was related to basal values (s.PRA vs s. A PRA:r= .87; p < .001; u.PRA vs u. A PRA:r= .94; p < ,001; u. Ald vs u.A Ald:r = 3 3 ; p < .01). In low PRA pts (N=9) PRA (1.12f .24 vs 1.31 ? .28) and u.Ald (12.56 f 2.58 vs 12.19 f 2.69) were unchanged, while they were decreased in normal-high PRA GROUP 4N= 14) (u.PRA:4.65 ? .SO vs 2.12 % .31; p < ,005; u. Ald:15.84 f 2.31 vs 10.94 ? 1.65; p < .025). Aldo sterone changes were related to those of PRA ( r = .53; p < .01) like u.Ald/u.PRA ratio before and after IND (r= .82; p < .001) were. 17 OH (8.23 f .86vs7 .34f .82N.S. )andF(12.93f .01 vs 13.13f .96;N.S.)were unchanged. Both IND and OXP decreased PRA but no interaction was found between the two drugs (IND + OXP: .62 f .46; F = .45; pN.S.). IND decreased creatinine clear. (1 18.7 f 6.3 vs 100 ? 4.99; p < .01) diuresis (1 142.4 f 78.2 vs 998.7 f 70.05; p < .02), urinary Na + (64.6 f 5.66 vs 40.1 f 5.2; p < .001) but did not modify B.P. (S.B.P. 164 f 4.6 vs 144.2 f 48.92; D.B.P. 92 f 3.13 vs 92.8 f 2.55).

ABSTRACTS 34 1

insulin in subsequent short-term incubations. At high cortisol concentrations basal glucose metabolism was reduced. The long-term effects of insulin to enhance basal lipolysis and acute lipolytic effect of noradrenaline were also prevented by the presence of cortisol. The acute antilipolytic effect of insulin in short-term incubations was also inhibited by cortisol. Since all these effects of insulin were reduced by cortisol the possibility of a reduction in the number of insulin receptors caused by cortisol was investigated. Cortisol reduced the number of insulin receptors about 40%. However, since rather high insulin concentrations were used this finding can not alone explain all effects of cortisol. One effect of insulin, however, was potentiated by high physiologic cortisol concentrations, viz., lipoprotein lipase (LPL) activity was increased several-fold in abdominal fat cells in the presence of both cortisol and insulin. This finding may explain the redistribution of the adipose tissue in Cushings disease. However, the ratio between cortisol and insulin seems of importance for this effect.

Conclusions: IND may decrease PRA by acting at sites different from OXP. U.Ald changes are explained by PRA ones. The PRA unresponsivness to IND in low PRA pts may suggest renal prostaglandin deficiency in these pts.

93. Prostaglandins in renovascular and renal hypertension. A. Hornych, F. Fontaliran, M. Safar, J . BariCty, P. Milliez. Hypertension Laboratory, U-28 INSERM. Hapital Broussais, Paris, France. This study was designed to examine more closely the role of prostaglandins A,, B, , E,, F, , alfa and plasma renin activity (PRA), measured by radioimmunoassay, in the pathophysiology of renovascular hypertension with unilateral renal arterial stenosis and renal hypertension with unilateral renal atrophy. Prostaglandin (PG) A, is significantly increased in both renal veins and vena caval blood of patients with renal arterial stenosis whose PRA is increased, but not in patients with renal atrophy, whose PRA is, in general, in normal range. There is a positive significant correlation between PRA and PGA, and PGB,, but not with PGE, OR PGF alfa. PGE, is significantly increased in both groups of patients, with the highest concentration in the blood from affected kidney. PGF alfa is increased in renal venous blood from pathological kidney, especially in patients with renal atrophy. Prostaglandin levels are, in general increased in renal venous blood from pathological kidney, especially in patients with renal atrophy. Prostaglan- din levels are, in general increased, only in few patients are decreased or undetectable. The increase of PGA, and PGE, representes a secondary antihyper- tensive, diuretic and natriuretic mechanism. The increase of PGF alfa from pathological kidney may contribute to the increase of systemic blood pressure. All prostaglandins are involved in volume- blood pressure regulation. The simultaneous measurement of plasma renin activity and different prostaglandins may improve the diagnostic of renovascular and renal hypertension.

183. Modulation of prostaglandin secretion in cultured rat renomedullary interstitial cells. F. RusseMarie, M. Paing & D. Duval. INSERM U.90 & INSERM U.7, Hbpital Necker, 75015 Paris, France. Rat renomedullary interstitial cells were isolated and grown in tissue culture. Using a highly sensitive radioimmunoassay we demonstrated that these cells synthesise prostaglandin E, and F, at rates (20 ng PGE and 2 ng P G F 2M/mg protein/hour in monolayer tissue culture) 20-fold higher than fibroblast cultures from the same origin. We therefore used this model to study in vitro the effect of various drugs on prostaglandin production. At a concentration of 10’M glucosteroids produce a marked inhibition of PGE, production which parallels their anti-inflamma- tory effects, whereas sex steroids were uneffective at the same concentrations. Using submaximal concentrations of dexamethasone and indomethacine (a potent non-steroidal inhibitor of PG synthesis), we demonstrated that the inhibitory effects of the two drugs were additive. Isoproterenol, a /J’- stimulating agent, appears also to decrease prostaglandin production. Conversely, angiotensin I1 at 2 x 10-9M induces a 50% stimulation of PGE, production. The parallelism of hormonal concentrations which modulate PO secretion in vitro with their circulating levels indicates that the in vitro system described here may be useful in the understanding of renal hemodynamics.

209. Cortisol antogonizes the effects of insulin on human adipose tissue metabolism. U. Smith and M. Cigolini. Department of Medicine 11, Sahlgren’s Hospital, Gothenburg, Sweden. Cortisol administration to man leads to insulin resistance. In the present study we investigated the metabolic effects of cortisol on human fat cells in vitro. Specimens were incubated wxith cortisol (30 x 10-8 M) and/or insulin (lo00 pU/ml) for different periods of time. After 2 hrs incubation no metabolic effect of cortisol could be shown. However, when the incubation period was prolonged using the tissue culture technique clear metabolic effects were noted. The long-term effect of insulin to increase ((basal)) rate of glucose metabolism was overcome by cortisol. Furthermore, explants cultured with cortisol alone did not respond to the acute effect of

205. The influence of cortisol (C) and insulin (I) on glucose metabolism and noradrenaline (NA) induced lipolysis (NAIL) in long-term perifused human adipose tissue (HAT). L. Sjbstrbm and P. Hallgren. Clinical Metabolic Laboratory, Department of Medicine I, Sahlgren’s Hospital, Gothenburg, Sweden. The purpose of the present study was to examine some short- and long-term effects of C and I on adipose tissue metabolism during strictly controlled conditions excluding hormonal influences from the donor of tissue. Adipose tissue was incubated in a continuous flow of Parker’s medium 199 and lipolytic rate was monitored continuously for 10 days under the following conditions, a) no additions, b) I (150 mU/I), c) C (500 nmoles/l) d) I + C. On the 1st and 10th days NAIL ( 5 . 10-5 moles NA/I) was measured in one serie of experiments and glucose incorporation into IG (GIITG) in another. On the 1st day NAIL:s were b) < a) = c). On the 10th day NAIL was zero in a) and close to zero in c) while b) resulted in a detectable NAIL. In d) NAIL was completely restored. On the 1st day a), b), c) and d) had the expected effects on GIITG. On the 10th day GI1TG:s were a) = 0 < b) < c) < d). In spite of the long-term promoting effect of C on glucose metabolism, the expected inhibiting effect of C on GIITG was possible to demonstrate in short-term experiments with HAT treated according to d) for 10 days. It is concluded that perifusion of HAT for 10 days results in a washout of factors which are essential for NAIL and GII’IG. This is not due to decreased viability. By adding factors one by one or in combinations, the prerequisites for NAIL and GIITG can be studied. The short-term effect of I was antilipolytic but the long-term effect was lipolytic. The presence of I was necessary for the permissive effect of C. C had a promoting effect on GlITG which was possible to separate from its generally accepted inhibiting effect.

251. Plasma lipoprotein lipase and hepatic lipase activity in patients on haemodialysis. L. Verschoor, H. Jansen. A.J. Zonneveld and J.C. Birkenhlger. Department of Medicine 111, University Hospital c<Dijkzigtn, Rotterdam, The Netherlands. Elevated triglyceride levels in patients on chronic intermittent haemodialysis have been explained mainly by a removal defect. Overall post-heparin lipase activity has been found low. We studied the effect of long-term heparin administration in hepatic lipase and lipoprotein lipase activity in order to see whether one or both activities contribute to the overall low activity. Fifteen patients on haemodialysis treatment and a control group were studied with longterm heparin stimulation (Brunzell et al., Metabolism 24, 1123, 1975). Basal triglyceride levels were significantly elevated in the patients. In the control group there was a sharp rise in both lipase activities, which was maintained for hepatic lipase up to four hours in contrast to a continuing decrease in lipoprotein lipase activity. Although all values for both lipase activities appeared to be lower in the patient group, only the initial lipoprotein lipase activity (i.e. 30’ value) was significantly decreased (54 ? 8 pM FFA/min/L* vs. 98 & 21 pM FFA/min/L, p < 0.05). No correlation existed between basal triglyceride concentration and lipase activity, nor between one of the parameters and basal insulin levels. Conclusion: the overall lipase activity in patients on haemodialysis

342 ABSTRACTS is decreased, the decrease in lipoprotein lipase is more pronounced than that of hepatic lipase.

mean f SEM.

106. Substrate extraction by the human myocardium before and after /%blockade. S.M. Juul, R.H. Swanton, B.S. Jenkins, C. Lowy and M.M. Webb- Peploe. Departments of Medicine and Cardiology, St. Thomas' Hospital, London, U.K. The metabolic effects of p-blockade on the human myocardium have not been extensively investigated. We have measured the myocardial extraction of lactate and free fatty acids (FFA) before and after acute administration of a /%blocker in a group of patients undergoing diagnostic cardiac catheterisation. Arterial and venous blood samples were obtained from the left ventricle and coronary sinus respectively at different heart rates (pacing). After the administration of /3-blocker, the procedure was repeated. Of the 30 patients studied, 14 received propranolol and 16 acebutolol. Ten patients were found to have normal coronary arteries. The mean lactate extraction ratio, [a-v/a] (LER), was lower at basal heart rate in ischemic patients compared to ctnormalw and this difference was greatest a t maximum heart rate (p < 0.02). Propranolol increased LER only at maximum heart rate in ctnormals)) ( p < 0.05) but at all heart rates in ischemics. Acebutolol had little effect on LER in ctnormals)) and increased this value in ischemics at maximum heart rate only (p < 0.05). Unexpectedly, the extraction of FFA was not linearly related to the arterial concentration, [a], pre /3-blockade. Following /%blockade, a correlation between [a-v] and [a] did exist, particularly in the acebutolol- treated group. Propranolol appears to ttnormalisen LER in the ischemic patients by increasing extraction and/or decreasing production of lactate. The results for FFA suggest and inhibition of endogenous myocardial lipolysis after p-blockade, more evident with acebutolol. We have concluded that the two drugs may have different effects on myocardial metabolism.

165. Effect of metoclopramide on urinary xylose test. O.S. Popovic, Z. Samcovic, D. Marinkovic, M.M. Andrejevic V. Vukeevic and A.T. Paljm. Gastroenterolosko odeljenje, Klinieka bolnica, Beograd, Jugoslavija. Very little is known about the relations between intestinal motility (transit) and absorption. Metoclopramide is known to enhance gastrie emptying and duodeno-jejunal transit. Five grammes d- xylose test was performed twice, the second time 10 mg Metoclo- pramide was given in before the test. The results are expressed as the percentage of the given dose excreted in urine in two and five hours. Patients with atrophic jejunal mucosa (7 coeliac disease, 2 alpha chain disease) were tested two to five times. The results were classified according to the appearance of jejunal mucosa when the tests were performed. When subtotal villous atrophy was found (15 tests) both2 h(5.I k 2.5 - 1.92 1.2) and 5 h (14.5 k 0 . 2 - 8.1 k4.0) xylose excretion diminished significantly (td. p < 0.01 and p < 0.005) when Metoclopramide was administered. When partial villous atrophy was present ( 1 1 tests) only 2 h excretion (12.9 f 5.1 - 9.9 f 3.0) diminished significantly (td, p < 0.05) after Metoclopramide, while 5 h excretion did not (31.8 i- 5 .5 - 29.4 * 6.4). When jejunal mucosa became normal on gluten free diet (6 tests) both 2 and 5 h xyloseexcretion(22.4k2.1 - 2 2 . 0 2 1.4,38.2k2.3-40.1 +2.5)did net change significantly after Metoclopramide; as well as in 9 patients with malabsorption but normal jejunal mucosa (21.0 * 8.2 - 25.0 k 5.2; 35.0 k 11.2 - 36.4 k 6.7) and in 9 controls (21.5 f 5.1 - 24.0 f 4.9; 38.0 k 7.4 - 41.8 f 10.0). The influence of Metoclo- pramide on renal function, and therefore on xylose excretion, was excluded by simultaneous endogeneous creatinine clearance determination. Metoclopramide, which probably acts through speeding up the gastrointestinal transit, diminishes xylose absorption only when jejunal mucosa is atrophic. Therefore, the contact time may be a rate h i t t i n g step for xylose absorption only when jejunal mucosa is atrophic.

138. Is bile a amicellar sink))? S. Menage and S. Erlinger. Uniti de Recherches de Physiopathologie Hepatique (INSERM), Hapital Beaujon, Clichy, France. It has been suggested recently that incorporation of endogenous or

exogenous compounds into mixed micelles could account for their concentrative transport into bile (1) and explain the reported increase in the biliary transport maximum (Tmax) of sulfobromophthalein (BSP) by bile salts. To test this hypothesis, the influence on BSP Tmax in the hamster of taurocholate (TC) and taurochenodeoxy- cholate (TCDC), two micelle-forming physiological bile salts, has been compared to that of taurodehydrocholate (TDH), a bile salt which, in vitro, does not form micelles.'In a first sene of expen- ments, the effect of the three compounds on biliary lipid secretion has been examined. It has been observed that TC and TCDC increased the secretion of cholesterol (recpectively + 49.8% and 109.7%; P < 0.05) and of phospholipids (+ 40.0% and + 52.6%; P < 0.05), while TDH decreased the secretion of cholesterol (- 43.1%; P < 0.05) and of phospholipids (- 30.9%; P < 0.05). These results suggest that, in vivo, TDH (or its metabolites) does not form mixed micelles. In a second series of experiments, under the same experimental conditions, the influence of the three compounds on BSP Tmax has been studied; a similar increase of Tmax has been observed with TC (+ 63.2%; P < 0.05), TCDC (+ 51.8%; P < 0.05) and TDH (+ 51.2%; P < O.OS).In conclusion, although TDH does not form mixed micelles in vivo, it increases the Tmax of BSP as well as TC and TCDC; these observations suggest that mixed micelle formation is not an important determinant of maximal BSP secretion into bile. ( I ) Scharschmidt BF, Schmid R: The ttmicellar sink)). Gastroente- rology, 1977, 75: 1182 (abstract).

214. Determination of gastric mucosal blood in man by 1%. aminopyrine: pitfalls and limitations. A. Sonnenberg, K. Weber, W. Mattle, I. Benes and A.L. Blum. Triemli Hospital, CH-Zurich. The clearance of aminopyrine (APC) of the acidified stomach is thought to reflect mucosal blood flow. This assumption was tested in man. Six healthy volunteers received a bolus injection of 0.133 pCi/kg (2.17pg/kg) AP. For the next 330 minutes, AP was infused at a rate of 0.0083pCi/kg/hr. For the last 90 min. pentagastrin (PG, 0.667 pg/kg/hr) was also given. The stomach was perfused with O.ISn HC1 (1.25 ml/min) containing 2% polyethylene glycol. Acid secretion was 0.027 mmol/min 2 0.010 SD before PG. It rose to 0.533 f 0.179 45 min. after starting PO and fell to 0.502 * 0.151 thereafter. APC exponentially (r =0.97) dropped from 52 * 25 ml/ min. to 14 f 5 within the first 90 min. and remained at this level for 150 min. P G caused an immediate but transient increase to 54 f 33 ml/min. While acid secretion continued to rise APC fell to 20 2 7. In six additional volunteers, PO was infused for 180 min. During the last 90 min., isoproterenol ( IP 2pg/kg/hr) was also given. IP inhibited acid secretion by 63 2 14% but did not reduce APC. Conclusions: The initial drop and the PG induced high but transient rise of APC do not reflect changes in blood flow. A model which explains these phenomena includes initial A P loading of the mucosa and washout by high volume flow. These transients may obscure the changes due to increased flow during PG and IP infusion.

174. Treatment of hepatic coma in cirrhotics by infusion of a special amino acid solution. H.J. Reiter and J.Ch. Bode. Medizinische Universitiltsklinik Marburg, West Deutschland. Application of an amino acid solution containing a large amount of branched chian amino acids to patients with chronic liver disease and hepatic encephalopathy has shown a significant improvement in the clinical and neurological symptoms. The aim of the present study was to determine if the parenteral or oral application could be an alternative to the current standard therapy with neomycine and lactulose. Six patients in coma grade II/III and 111 were treated with the amino solution parenterally; in addition four patients suffering from hepatic encephalopathy with an oral amino acid mixture. Besides the clinical findings plasma amino acids and NH, were determined. EEG was carried out every other day. Results: a distinct improvement within 12-18 hours could be observed in all but one patients treated parenterally, a similar improvement in three patients treated orally could be seen. Parallel to the return to consciousness an increase in the EEG frequency was noted, an elevation in the levels of the branched chain amino acis (valin, leucin, isoleucin) and a slight decrease in the cyclic amino acids (tyrosine, phenylalanine, tryptophan) and methionine was seen. The results document that application of a special amino acid solution/mixture can bring about a rapid improvement in hepatic coma/encephalopathy of patients with cirrhosis of the liver.

198. Radioimmunoassay for human type I1 (cartilage) collagen in syoovial fluids of patients with rheumatoid arthritis and osteoarthritis. B. Seewbster, E. Moshudis, G. Loben*, F. Neurathg and E. Hahn (Intr. by Ch. Bode). Medical University Hospital, Orthopaedic Hospitalg and Behringwerke', MarburdLahn, Fed. Rep. Germany. Human type I1 collagen in native conformation was labeled with iodine-125. Very high activity could be introduced into the molecule by applying the Bolton-Hunter reagent (0.3-0.5 molecules of iodine- 125 per molecule of collagen). 90-95% of labeled antigen (1.5-3 ng) could be bound by purified antibodies raised in goats against human type I1 collagen. 50% inhibition could be achieved with 50-100 ng of type I1 collagen. No inhibition was observed with type I or 111 collagens. The assay allows the detection of type I1 collagen antigen in a concentration as low as 100 ng/ml. In joint fluids of patients with rheumatoid arthritis and osteoarthritis a distinct inhibitory activity could be found. The highest inhibitory activity found was equivalent to 300 ng of type I1 collagen antigen per ml. These results establish a new method that could be applied to measure cartilage destruction in chronic joint diseases.

216. HLA-B27 homozygosity and ankylosing spondylitis: genotypic and phenotypic studies of three families. D.G. Spencer, H.M. Dick, P. J. Rooney and W.C. Dick. Centre for Rheumatic Diseases. Glasgow, Scotland, U.K. Homozygosity for the histocompatibility antigen HLA-B27 has been reported to lead to more severe ankylosing spondylitis (A.S.). Three families have been studied, consisting of eleven homozygotes, thirteen heterozygotes and six persons lacking HLA-B27. Aii were examined clinically and radiologically for AS. , and eight were found to be affected, three homozygotes, four heterzygotes and one person without the antigen. Only three members belonging to two of these kindred are as yet of insufficient age to exclude presence of disease. From these studies it is evident that homozygosity for HLA-B27 is not universally associated with AS. , nor does it seem that homozygosity necessarily leads to earlier or more severe disease. A S . in one of these families was not associated with any particular haplotype which strongly suggests that development of disease is not solely due to an inherited closely linked immune response gene.

2. A solid phase radioimmunoassay for anti-soluble deoxyribonucleo- protein (sNP). C. Adam, P. Verroust, F. Pontillon and P. Robitaille. INSERM U.64, Hapital Tenon, Paris, France. Anti-sNP antibodies (Ab) occur frequently in systemic lupus ery- thematosus (SLE). A primary binding radioimmunoassay method which requires labelling of the sNP with 1251 and density gradient ultracentrifugation has already been described. Avoiding labelling and denaturation of the sNP, a highly sensitive and discriminatory new method has been set up which enables studies of the classes of anti-sNP. Polystirene tubes were coated with crude sNP, incubated with the test sera and the fixed Ab detected by a double layered technique using a rabbit anti-human-gamma-globulin antiserum and a I125 labelled sheep anti-rabbit gammaglobulins antiserum. The binding of 33 qormal human sera was below 24 ng of sheep anti-rabbit gammaglobulins / 30 ul of sera. Among 50 SLE, 22 sera showed binding over 24 ng with 1 up to 11 times more than the normal. Inhibition of the binding occurred when positive SLE sera were preincubated with sNP, native or denatured deoxyribonucleic acid (NDDNA, SSDNA), but not with ribonucleic acid. Good correlation was found with anti-sNP or anti-DNA Ab detected by the Farr assay. Kinetics studies of the action of deoxyribonuclease I, and S1 nuclease were performed on the sNP-coated tubes. This method has also been applied to the detection of antibodies to SSDNA, Sm and ribonucleoprotein. This method provides the most sensitive and reproducible assay to distinguish the various nuclear antigens involved in connective tissue disorders.

199. Impaired in vitro antibody response in patients with rheumatoid arthritis. P. Segond, J.F. Delfraissy, P. Galanaud and P. Massias. INSERM

343 U.131 and service de Rhumatologie, Hapital Antoine B&Kre, Clamart, France. The ability of rheumatoid arthritis (RA) patients to mount an antibody response has been studied in vivo and led to conflicting results. In order to better characterize the immunological status of these patients, we examined their antibody response toward the TNP hapten using the in vitro method we recently described. Pa- tients satisfied the ARA criteria for definite RA, and had never received immunosuppressive drugs nor corticosteroids during the month before testing. Antiinflammatory drugs were stopped for at least 72 hours before blood collection. The mean response was 66 ? 21 plaque forming cells (PFC) per 106 cells in 1 1 RA patients as compared to 180 & 38 PFC in matched normal controls (p < 0.01) and to 185 ? 42 PFC in I5 control patients (p < 0.05). Thus peripheral blood lymphocytes from RA patients have a decreased ability to develop a specific IgM antibody response in vitro. The cellular basis and mechanisms of this impairment in humoral immunity are under study.

ABSTRACTS

113. The contribution of bronchial hyperreactivity, allergen-specific IgE and histamine release in allergy. K.F. Kerrebijn, H.J. Neijens and H.J. Degenhart. Department of Pediatric Respiratory Disease, Sophia Children Hospital, University of Pediatric Respiratory Disease, Sophia Children Hospital, Uni- versity of Rotterdam, the Netherlands. In 20 children with a positive history of cat allergy (sneezing and/ or wheezing within a short period after contact with a cat), the relationship was studied between the result of bronchial provocation with cat dander on the one hand and bronchial hyperreactivity (histamine threshold), specific IgE intradermal skin titer and histaminerelease from leucocytes with the same batch of allergen on the other. Methods have been described earlier (Kerrebijn et al., Arch. Dis. Childh. 1976, 51, 252-258). The children were studied in a symptom-free period and were admitted in hospital because of the study. Provocation was only found to be positive with an increased bronchial hyperreactivity (histamine threshold < 8 mg/ml; normal value 0 32 mg/ml). There is a statistically significant correlation between a positive provocation and the titer of specific IgE (P < 0.01), the skin titer (0.01 < P < 0.05) and the histaminerelease (0.01 < P < 0.05). By means of a multiple correlation analysis with nonlinear terms the contribution of each of the parameters in the achievement of a positive bronchial provocation has been calculated. The bronchial hyperreactivity is the far most important; with an increased hyperreactivity the quantities of specific IgE and released histamine seem to be of secundary importance. The chance on a positive provocation has been calculated if values of histamine threshold allergen specific IgE and skin titer are known.

170. Postural changes in human pulmonary circulation. B. Raffestin, P. Fournier, J. Mensch-Dechene, and B. Ranson-Bitker (Intr. by A. Lockhart). Department of Physiology, Faculte de Me- decine Kremlin-BicEtre, Paris, France. Whereas postural changes in central blood volume are well docu- mented there are no data with respect to postural changes in pulmonary blood volume (PBV). Therefore, pulmonary vascular pressures, blood flow, and PBV were measured in the supine and sitting posture in 8 patients suffering from small unilateral lung carcinoma. Supine cardiac output (CO), pulmonary wedge and arterial pressure, pulmonary vascular resistance, and PBV were normal. Also normal were postural changes in CO and heart rate. The difference between mean pulmonary arterial acd mean pulmonary arterial and mean pulmonary wedge pressure did not change significantly with sitting up. Pulmonary vascular resistance rose from 92 ? 25 CGS units supine to 122 & 49 sitting (p = 0.05). PBV fell from 517 k 122 ml supine to 360 ? 43 ml sitting (p < 0.01). The reduction in PBV with sitting-up is best explained by closure of alveolar vessels in the upper lung, and by the concomitant cessation of flow in corresponding extraalveolar vessels. Circumstantial evidence suggests the latter vessels remain open under the large expanding stresses which prevail in the upper part of the lung.

68. Perfusion changes modify intra-pulmonary shunting (QS/QT) in patients with adults respiratory distress syndrome (ARDS). H. Gastine, B. Regnier, B. Teisseire and F. Lemaire. Henri Mondor. Hospital ICU, Creteil, France.

344 ABSTRACTS

QS/QT calculation is largely used to assess severity of respiratory failure in ARDS. This study has been undertaken in order to evaluate the variations of Q Y Q T determined by acute pulmonary hemodynamic changes. In 28 cases of ARDS (24 first hours) (92 measurements), QS/QT variation (FiO, = 1) have been correlated with simultaneous changes of cardiac index (CI), pulmonary arterial (PAP) and wedged pressures (PWP), and pulmonary vascular resistance (PVR), induced by the treatment of respiratory and/or circulatory failure (dehydration, dopamine, isoproterenol or neosynephrine infusion, veno-arterial bypass with membrane lung oxygenation). 1) QS/QT was linearly correlated with CI (r = 0.81), p < 0.001) and with I/PVR ( r = 0.74, p < 0.001), which are prominent factors of recruitment. 2) QWQT was correlated with PAP only when PVR was low, that is when PAP and CI varied concomitantly. 3) The relationship with PWP was more complex: QS/QT was correlated with PWP only when it varied in the same way than CI. Conversely, neesynephrine, a pure u agent (n=5) . which increased PWP from 7 to 17 torr and decreased CI from 3.0 to 2.5 I/min/m2, decreased QS/QT from 22 to 16%. These data indicate that QS/QT variations have to be correlated with pulmonary hemodynamics in unstable hemo- dynamical situations. Acute shunting variations may indicate large changes of vascular recruitment in unhomogenous lung, rather than improvement or extension of pulmonary damage.

36. Immunological study of human lungs in non hemodynamic pulmonary edema (NHPE) during severe sepsis (SS). J. Carlet, J.F. Bernaudin, C. Sabatier, A. Sobel, F. Lange, C. George and F. Lemaire. Henri Mondor Hospital ICU, Creteil, France. This study was performed to precise the immunological invol- vement in the physiopathology of NHPE occuring during severe sepsis (SS). Lung biopsies of 15 patients who died with massive NHPE were studied in routine histology, electron microscopy (EM), and immunofluorescence (IF). Patients with normal lungs or hemodynamic pulmonary edema composed two control groups. Biopsies, performed within minutes after death through a bedside thoracotomy, with maintained artificial ventilation were fixed or deep frozen at -80°C. Before death, search for circulating immune complexes (CIC) was done with two methods, Polyethylene glycol precipitation test (PEG) and Clq* deviation test. Results: Intra- alveolar fixation was observed in lungs with NHPE with the following antisera: Anti-Fibrinogen (80% of the patients), Anti- Albumin (30%). Anti-IgG (70%), Anti-lgA (40070). Anti-IgM (lo%), AntiLC’3 (60%), Anti-a-Antitrypsin (30%). Fluorescence was locali- zed on the hyalin membranes identified by routine histology. Morphological aspects evocative of CIC deposition were not abser- ved either in IF or in EM. We conclude that in NHPE occurring during SS intra-alveolar protein content indicates an increased capillary permeability mechanism. However the fixation with Anti- IgG and Anti-C’3 is more frequent than with Anti-Albumin, suggesting a specificity in protein fixation. The results do not support a role for CIC deposition in the physiopathology of NHPE.

30. Prolonged blood transit time of leukaemic B lymphocytes in patients with non-hodgkin’s lymphomas. K. Rremer, A. Engeset’ and S. Frblandz. Div. of Haemat., Dept. of Med., Univ. of Essen, Germany; ‘Haemat. Lab., Norwegian Radium Hospital, Oslo; 2lnst. of Immun. and Rheumat., Univ. of Oslo, Norway. To study the intravascular fate of human blood lymphocytes (ly) autotransfusion studies with Wcytidine labelled blood ly have been performed in 6 groups of patients (pats): in untreated or irradiated pats with Hodgkin’s disease (HD), in similarly irradiated pats with carcinoma of the testis (CT), in untreated pats with chronic lymphocytic leukaemia (CLL) or with other leukaemic non-Hodgkin’s lymphomas (NHL) and in haematologically normal controls. Both groups of irradiated pats were characterized by an absolute increase of blood B ly, whereas the 2 groups with NHL showed leukaemic blood levels of neoplastic B ly. In all pats studied the initial blood ly labelling indices (LI), observed immediately after completion of rapid retransfusion of labelled autologous blood ly, decreased considerably within the following hour then levelling off into a plateau of stable LI. The halving times of these Ll decreasing rates of 139 min in the CLL pats and of 146 min in the NHL pats resp. were significantly prolonged compared with the mean values of 22 to 30 min in the remaining 4 groups of aleukaemic pats. Using these data the calculated blood transit

times revealed analogous differences: 201 and 21 1 min for the CLL and NHL pats resp. and 32 to 43 min for the other 4 groups of pats. Since these 4 groups of pats with normal blood ly counts exhibited similar short mean ly blood transit times irrespective of their different B and T blood ly concentrations, it is concluded that the observed prolonged blood transit times of the leukaemic B ly must be attributed to their neoplastic nature, possibly to an altered surface membrane.

52. Neutrophil kinetics in pernicious anaemia. K.A. Deubelbeiss and P. Roth. Division of Haematology, Insels- pital, Berne, Switzerland. Erythrokinetics are fairly well understood in pernicious anaemia. Much less is known about neutrophil kinetics. In this study marrow neutrophil cellularity was calculated from a ferrokinetic estimate of marrow normoblasts and from the neutrophil-erythroid ratio determined from marrow sections. In seven untreated patients the mean values (.109/kg * 1 SEM) for mitotic (MMN) and postmitotic (PMMN) marrow neutrophils were 5.25 * 0.42 and 12.04 * 1.73. Using 3H-thymidine the transit time through the PMMN pool was found to be 6.34 I 0.12 days. From the number of PMMN and the transit time, marrow neutrophil production rate was calculated to be 1.49 & 0.26 .lo9 neutrophils/kg/d. MMN and PMMN were increased 2.5 and 2.1 fold, marrow neutrophil production rate l . S fold from the normal values previously determined (1). In recent studies of neutrophil release from the marrow following intravenous injection of hydrocortisone we have shown a linear relationship between PMMN and maximal neutrophil increment in blood over a wide range of marrow cellularity. In these seven patients however, significantly fewer neutrophils were mobilised despite increased PMMNs. Our studies not only suggest ineffective neutropoesis, but they also point to an abnormal mobilisation of PMMNs, not previously recognized in this disorder. (I) Dancey J. T., Deubelbeiss K.A. , Harker L .A . and Finch C.A.: Neulrophil Kinetics in Man. J . Clin. Invesl. S8, 705.715, 1976.

160. In vivo studies utilizing human leucocytes selectively labelled by in vitro phagocytosis of 99mTc sulfur colloid. R. Paridaens, J. Friihling, D. Gangji and E. Schell-Frederick. Institut de Recherche Interdisciplinaire and Institut Bordet, ULB, Brussels, Belgium. The purpose of the present study was to prepare functionally intact neutrophils, labelled in a selective and stable manner, for use in hemodynamic studies and for localization of sites of infection. 200 ml whole blood was incubated with 16-20 mCi 99mTc sulfur colloid for 40 minutes at 37°C. Labelled neutrophils were isolated by plasmagel sedimentation, low speed differential centrifugation and two normal saline washes with removal of non-cell bound radioactivity and contaminating labelled platelets. All procedures were performed under sterile conditions. In the final preparation greater than 75% of the recovered radioactivity (ca. 500 FCi) was present in the neutrophils. In vitro functional studies demonstrated I ) normal random migration and chemotaxis; 2) normal phagocytosis of paraffin oil droplets; 3) normal metabolic changes associated with phagocytosis. Preliminary results of dynamic scanning and static pictures at 5 and 24 hours in 3 patients without infection after reinfusion of autologous labelled cells have shown 1) no release of the label into the circulation as measured by absence of labelling of the thyroid and no appearance of label in the urine; 2) initial accumulation of labelled leucocytes in the lungs with subsequent passage into the liver, spleen and bone marrow; 3) no accumulation of labelled cells in primary or secondary tumors. Further investigations are in progress to determine the clinical usefulness and accuracy of this technique for localizing sites of injection.

12. Hemolytic implications of alcoholism in liver disease. K.M. Goebel, F.D. Goebel, G.V. Manteuffel and J. Schneider. Department of Medicine, Philipps University, Marburg, Germany. To evaluate the effects of ethanol in vivo as a hematologic and oncologic toxin and to delineate further the pathogenesis of red cell (RBC) metabolic disorder in hemolytic syndromes of chronic alcoholism, 20 male patients were studied on acute (groupe A) and remittent (group B) phase of Zieve’s syndrome. Chronic alcoholics with hyperlipoproteinemia, jaundice and liver disease but without

hemolysis served as control (group C). RBC were separated into old (dense) and young (buoyant)-cells-by-means of density-layer centrifugation by Stractan 11 and Renografin gradients. In group A the older RBC disclosed a pyruvate kinase enzyme instability, influenced by a protein bound and hence not dialyzable hemolytic factor. Dialyzed patient’s RBC also reveiled changes of membrane lipid composition, as indicated by increased cholesterol and decreased PUFA levels (group A). Alcohol induced serum and membrane- linked vitamin E deficiency with reduced PUFA provoked increased intracellular oxidation with a decrease of GSH and ATP levels. The hence impaired RBC membrane stability was demonstrable by experimental H,O, hemolyis in group A, as compared to group B and C. Thus, it is conceivable that a linkage of extra-intracellular factors form a chain reaction: Causing target cell formaion, vit. E/PUFA deficiencies, increased oxidative metabolic disorder, and finally the susceptibility to hemolysis of alcoholic patients.

164. The mechanical response to acute acidosis of myocardium from rats acclimatised to 5% CO,. P.A. Poole-Wilson, C.H. Fry, N.T. Bateman and I.R. Cameron. Department of Medicine, St. Thomas’Hospital Medical School, London and Cardiothoracic Institute, London. When isolated myocardium is exposed acutely to an increased PCO, the intracellular pH (pHi) falls and developed tension is rapidly reduced. Rats acclimatised to 5 % CO, 20% 0, for 28 days have a normal myocardial pHi, intracellular K + and Na+ and have increased intracellular buffering on acute exposure to increased PCO,: If the fall in pHi causes the observed fall in developed tension during acute exposure to increased PCO,, developed tension should fall less in myocardium from acclimatised animals. We have compared the fall in developed tension induced by raising PCO, in myocardial muscle from control animals, and animals acclimatised to 5 % CO,, 20% 0, for 28 days. Isolated papillary muscles were maintained at 37°C and stimulated at 60 beatdmin. Maximal developed tension was 5.7 & 0.9 kN/Kg muscle weight (n = 8 : mean f SEM) in control and 2.9 f 0.5 kN/Kg (n= 11) in acclimatised rats (p < 0.02). The mechanical response to raised PCO, of the bathing fluid was expressed as the slope of the regression line for the plot of developed tension against fluid pH. The slope was 105 2 3 kN/Kg/pH unit for control and 82 f 9 kN/Kg/pH unit for acclimatised rats (p < 0.02). There was a significant correlation between the reduction of the maximum developed tension and the reduction of the mechanical response to acute acidosis in acclimatized animals (r=0.82, p < 0.01). In control muscles lowering the Ca+ + of the bathing fluid did not alter the response to CO,. The altered response of acclimatised myocardium to CO, is best explained by increased intracellular buffering.

25. Endothelial injury and growth of atherosclerotic lesions in rabbits with moderate hypercholesterolemia. G. Bondjers, S. BjOrkerud, R. Brattsand, A. Bylock, G.K. Hansson and H.A. Hansson. Department of Medicine I, University of GBteborg, GOteborg, Sweden. Atherosclerotic lesions first appear at sites with endothelial injury when rabbits are fed cholesterol. However, later in the process, the lesions grow into areas which are covered by intact endothelium in unmanipulated rabbits. In the present investigation, we have studied the relationship between endothelial injury and plaque growth in cholesterolfed rabbits. The rabbits were fed 0.1-0.3% cholesterol in the diet for 6 months. The cholesterol supplement to the diet was modified, if necessary, every second week to maintain stable and individually different serum cholesterol levels. Endothelial injury was assessed by three different methods: ( I ) a dye exclusion test, (2) direct immunofluorescence for immunoglobulin G, and (3) scanning electron microscopy. The lesions were covered with intact endothelium but bordered with a zone of injured endothelium. This finding was consistent with all three methods. In the endothelium. This finding was consistent with all three methods. In the zone with injured endothelium, surface defects with platelet adhesion was observed. These data suggest that dietarily induced experimental atherosclerosis grows by the formation of a zone of injured endothelium in the periphery of the lesions. Platelet adhesion in areas with injured endothelium may lead to release of growth stimulating factors and subsequently to smooth muscle cell proliferation as it daes in experimental atherosclerosis induced by mechanical injury. The present data provide further support for the hypothesis that atherogenesis could be regarded as a response to injury.

ABSTRACTS 345 228. Angina in patients with normal coronary arteries: a metabolic disorder? W. Stubbs, C. Brearley, N. Conway, G. Sharratt and G. Albcrti. Departments of Medicine, St. Bartholomew’s Hospital, London and Southampton General Hospital, Southampton, England. The cause of angina in patients with normal coronary arteries is uncertain. We have, therefore, examined the myocardial uptake and production of certain metabolites in two groups of patients with severe angina; one with normal coronary arteries (NCA) and the other with coronary artery disease (CAD). Cardiac catheters were placed in the aortic root and coronary sinus. Paired blood samples were taken at rest, during and following incremental atrial pacing and analysed for blood metabolites pH, pCO, and &. A coronary angiogram was performed and patients allocated to NCA (n = 5 ) and CAD ( n = 6) groups. The mean f SEM per cent extraction at maximal pacing for glucose was -9 f 6% in the NCA and 1 f 2% in the CAD group (P < 0.05) and for lactate 26 2 6% and -25 f 21% respectively (P < 0.05). The coronary sinus [lactate]/[pyruvate] on maximal pacing was 9.4 f 0.3 in the NCA and 17.9 k 6.5 in the CAD group ( P < 0.02). Thus significant differences in the handling of certain metabolites by the heart have been demonstrated between these two groups of pa ti en t s .

41. Suppression of adrenergie response by glucose, insulin, K + /CIK/in experimental and clinical myocardial infarction/MI/. L. Ceremuzyhski, K. Herbaczyhska-Cedro, M. Dluzniewski, T. Zaleska, L. Falecka. Research-Teaching Centre, Medical School Research Centre of Experimental and Clinical Medicine, Polish Academy of Sciences, Warsaw, Poland. Clinical value of GIK was reported by many authors but precise mechanism of GIK action has not been definitely established. It is recognized that increased secretion of catecholamines is one of the most harmful consequences of MI. In acute coronary occlusion in open-chest dogs blood adrenaline/AD/ level, measured continuously by bioassay, increased in 12 out of 15 dogs. Glucose/G/ 0 .6/kg and Insulin/I/ lu/kg infusion, and to some extent G alone, suppressed enhanced AD release. GI effect was confirmed spectrofluorime trically in 5 dogs. Subsequently, GIK/G5O.O 115~1, K+40 mE/infusion was given for 6h to 18 unselected patients with recent MI and AD content in urine was measured spectrofluorime trically for 3 consecutive days. In 7 patients GIK was administered during the first day after onset of symptoms and AD content in urine was lower than on the following day/mean 34.14 and 48.84pg/24h respectively, p < .05/. In 11 patients GIK was infused on the 2nd day and AD content was lower as compared with the first/mean 20.22 and 55.25pg/24h respectively, p <.005/and the 3rd day/mean 41.33vg/24h, p<.O25/. These results point to existence of the mechanism for inhibition of enhanced AD secretion by GI. This might be one of the reasons for beneficial effect of polarizing solution in acute MI.

253. Influence of thoracic epidural anesthesia on the severity of acute myocardial ischemia in open chest dogs. H. Vik-Mo, S. Ottesen and H. Renck. Department of Physiology and Department of Anesthesiology, University of Tromsg, Norway. Thoracic epidural anesthesia (TEA) is widely used for surgical operations and relief of postoperative pain. WE studied the effect of TEA on the severity of acute myocardial ischemic injury in 8 thoracotomized dogs. Ischemic iiijury was measured as the sum of ST-segment elevations (1 ST) in epicardial ECG recordings from 10-15 sites 15 min after occlusion of a branch of the left anterior descending coronary artery. TEA reduced mean arterial blood pressure (AP) by 26%, heart rate (HR) by 20%. left ventricular dP/dt by 37%. and myocardial oxygen consumption by 38%. Although arterial concentrations of free fatty acids, glucose and lactate were unchanged, their myocardial uptake was reduced in proportion to the reduction in mechanical activity. In 7 of 8 dogs a substantial reduction of the severity of the acute myocardial ischemic injury was effected by TEA. In all 8 dogs, Z ST was reduced from 34.0 f 3.4 to 23.3 2 2.8 mV (mean k SEM), p < 0.01. After correction of AP and HR to values before TEA with phenylephrine intravenously and actrial pacing, the favourable effect of TEA on myocardial ischemic injury was abolished. It is concluded that TEA effected a reduction in the severity of myocardial ischemia in open chest dogs, mainly through reduction of myocardial mechanical activity with a consequent reduction of myocardial metabolism.

346 ABSTRACTS 217. Dopamine-beta-hydroxylase (DBH) activity in renal failure. U. Spohr, E. Ritz and R. Schrack. Dept. Int. Med., Krehl-Klinik, Heidelberg, Germany. In terminal renal failure, there is conflicting evidence with regard to the state of activity of the peripheral sympathetic nerve system. While plasma noradrenaline (NA) levels were elevated (Brecht et al., Proc. EDTA, 1975), plasma DBH levels were reduced (Spohr et al., Klin. Wschr. 1977). It was the purpose of the present investigation to study whether plasma DBH levels were correlated with arterial blood pressure. Patients and methods: DBH measurement with a modified micro- morphometric assay after Nagatsu (Clin. Chem. 18,980, 1972). 30 patients with preterminal renal failure under outpatient conditions without dietary restrictions and without anti-hypertensive medica- tion. Age and sex matched controls. Results and discussion: Plasma DBH was significantly (p 0.05) lower in uremic (30.5 f 30. 1.U.). There was inverse correlation of borderline significance between resting blood pressure in untreated uremic patients and plasma DBH (r=0,31). DBH, in contrast to noradrenaline (NA), is not subject to reuptake in the sympathetic synapsis and plasma DBH might therefore more accurately reflect sympathetic nerve activity than daes NA. The above data are consistent with the notion that the activity of the peripheral sympathetic system is appropriately lowered when arterial pressure rises in renal failure by non-sympathetic mechanisms.

144. Disturbed lipid metabolism in patients with nephrosis. R. Mordasini, 0. Oetliker, J . Lutschg and H. Greten. Lipidlabor, lnstitut fur klinische Eiweissforschung and Kinderklinik der Univer- sitlt Bern, Switzerland, and Medizinische Universitatsklinik Heidelberg, Germany. We have recently investigated the pathogenesis of secondary hyperlipoproteinemia in patients with chronic uremia (CU), under hemodialysis (HD) and after renal transplantation (RT). I t was possible to demonstrate a deficiency of hepatic triglyceride lipase (H-TGL) leading to triglyceride-rich low density lipoproteins (LDL) in CU and HD. Lipoprotein lipase (LPL) was normal in these patients. We now report the analysis of plasma lipid and lipoprotein composition in patients with nephrosis. In addition, specific measurement of LPL and H-TGL was performed using an immunochemical method. 24 patients with nephrosis (14 children and 10 adults) entered the study prior to drug treatment. These patients had ( I ) increased LDL -and HDL- cholesterol values, (2) elevated VLDL-triglycerides, and (3) decreased LPL activity with normal H- TGL enzyme activity. In follow-up studies these lipid abnormalities were reversible parallelling recovery from nephrosis. The pathogenesis of secondary hyperlipoproteinemia in these patients will be discussed with regard to the particular underlying renal disorder which may result in different lipid abnormalities.

261. Defective fatty acid incorporation into adipose tissue (FIAT) in uremic subjects with bypertriglyceridaernia (HTG). G. Walldius and H.E. Norbeck, Kind Gustaf V Reseach Institute, Karolinska Hospital, Stockholm, Sweden. HTG is common in uremic subjects. A low removal of serum triglycerides (TG) may be caused by a low lipoprotein lipase activity and/or a low fatty acid incorporation (FIAT). HTG may also be caused by an increased adipose tissue fatty acid mobilizing lipolysis. In the present study 24 patients (9 on dialysis) with uremia and 20 healthy controls were investigated. Eight and 14 patients in the two groups had HTG. Subcutaneous needle biopsy was taken and serum lipids were measured in the fasting state. FIAT was measured as the incorporation of labelled palmitate into extracted glycerides. Lipo- lysis was measured as glycerol release to the incubation medium. NormoTG (NTG) uremics had higher FIAT activity ( x = 345% .g:"h-l than NTG controls ( x = 147), p < 0.02. Uremic patients with HTG had higher FIAT than HTG controls (218 and 82 respectively, p < 0.02). HTG uremics had lower FIAT than NTG uremics, p < 0.05. Glycerol release was much increased in uremic subjects compared to controls ( x = 987 and 415 nmo1.g-1.h-1, p < 0.005). There was no difference in glycerol release between NTG and HTG uremic subjects. The results obtained suggest that uremic patients have an increased rate of turn-over of fatty acids in adipose tissue compared to controls. The lower FIAT activity in HTG than in NTG uremics probably contribute to their HTG.

89. The role of PTH on lipid metabolism in experimental chronic renal failure C.C. Heuck, M. Liersch, K. Stegmeier, E. Ritz, 0. Mehls. lnstitut fur Herzinfarktforschung, Heidelberg, GFR. Various hormones such as STH, glucagon and PTH have been found to be considerably elevated in uremia. Their role on lipid metabolism is only partially identified. An experimental model was developed to investigate the effect of PTH in chronic renal insufficiency. Male rats were sham op. (C), parathyroidectomized (PTX), 9/10 nephrectomized (NX) or parathyroidectomized and nephrectomized (PTXNX). Weight matched groups were kept under dietary control for 3 weeks. In 6d intervals lipids, urea, glucose were determined from serum samples. HMG-CoA reductase activity was measured in hepatic homogenates. All serum lipids were highest in NX and lower in PTXNX but still above C and PTX. No change of hepatic HMG-CoA reductase activity was observed. Whether the alterations result from a primary action of PTH or from Ca + +will be discussed.

260. Linoleic acid content in serum lipids and adipose tissue and serum lipoprotein concentrations in patients with uremia. G. Walldius and H.E. Norbeck. King Gustaf V Research Institute, Karolinska Hospital, Stockholm, Sweden. The linoleic acid (18:2) content of serum lipids separated by TLC and of subcutaneous adipose tissue (needle biopsy) were measured by GLC in 39 patients (pts) with varying degree of uremia. Serum !ipoproteins were also measured. Nineteen pts were without dietary restrictions, whereas 20 were on a protein restricted diet, 11 pts also underwent regular haemodialysis. Largest differences compared to 12 healthy normolipidaemic controls (C) were found in the cholesterol ester fraction, 18:2 was 52% ( x ) in all uremics compared to 55.6% for C, p < .01. Pts on diet only had 51.2%. p < .025. Lowest values were found in dialysis pts, x = 50.3, p < .01. In the group of pts not treated with dialysis (n = 28) those with a low 18:2 content (< 52.5%) (n = 14) had a lower HDL-cholesterol ( x = 1.14 mmol/l) than those with a high (< 52.5%) 18:2 content ( x = 1.46), p < .02. 18:2 in serum triglycerides and phospholipids and in adipose tissue were not different from C except for a lower than normal 18:2 content in the fat of dialysis pts, p < .05. In general the uremic pts were hypertriglyceridaemic with highest values in dialysis pts. The lower than normal cholesterol linoleate may be caused by a selective metabolism of this acid in circulating lipids related to the level of serum triglycerides and/or the HDL cholesterol concentrations or to an imbalanced LCAT activity. Besides the time of duration of their disease and the uremia itself also the effects of a protein restricted diet (with an inadequate 18:2 content in the fat?) may contribute to these findings.

74. '23lodine-lsbekd chdylglyeyulistnmine: a oew agent for kpatdMiary scanning and studies of bile acid transport. E.M. Grandjean, G. Karlaganis, U. Noelpp, H . Roesler, G. Paumgartner. Departments of Clinical Pharmacology and of Nuclear Medicine, University of Berne, Berne, Switzerland. Bile acids (BA) are removed from the blood and excreted into the bile by the liver with higher specificity and efficiency than most other cholephils. A BA labeled with a y-emitter might, therefore, offer advantages over conventional hepatic scanning agents for imaging of liver and biliary system and for studies of BA transport. Consequently, we have labeled a synthetic conjugate of cholic acid, cholylglycylhistamine (CGH), with 1231, an isotope with short half- life (13.2 h), favorable gamma energy (159 keV) and no beta emission. T o compare the hepatic handling of this BA derivative with a physiologic BA, 450-3000 pCi IZ31-CGH was injected together with 5 pC i i4C-cholyltaurine (CT) i.v. in anesthetized dogs with (a) intact gallbladder or (b) cholecystectomy and Thomas duodenal cannula. The initial plasma disappearance rates of 123I-CGH and I4C-CT ranged between 0.40-0.42 min-I and were not significantly different. Both BA appeared in bile in high concentration with a maximum 15-17 min after injection, 50 to 70% of the dose being excreted into the bile within 30 min. Urinary excretion of '231-activity was minimal (< 10%). Excellent scintigrams of liver and gallbladder were obtained with IZ3I-CGH. Gall-bladder emptying induced by i.v. Ceruletid (0.05 pg/kg) could be quantified by recording the radioactivity time curve over a gallbladder region of interest. These findings indicate that '23I-CGH is handled by the liver like an endogenous BA and might be useful for hepatobiliary scanning and studies of BA transport.

224. Cytosolic bile acid binding components from rat ileum and jejunum. R.C. Strange, B.J. Chapman, B.A. Williamson and I.W. Percy- Robb. Department of Clinical Chemistry, University of Edinburgh, Edinburgh. An important part of the enterohepatic circulation of bile acids is their active transport across the terminal ileum into the portal vein. We now describe a soluble binding protein present mainly in ileum which may act as a bile acid carrier. Mixtures of ileal and jejunal cytosol and lithocholic acid were chromatographed and several binding components found although one (moLwt. approx. 40 OOO) appeared mainly in ileum. The binding of glycocholic acid to partially purified preparations of the component from ileum and jejunum showed that the ileum contained more binding sites (179pn~oVg supernatant protein) than the jejunum (47pmoVg supernatant protein) although the similar values for the dissociation constant of the ileal component (2.5pmol/l) and of the jejunal component (1.6pmol/l) suggests that the same component is present in different concentrations in both preparations. In contrast to hepatic bile acid-binding proteins, the intestinal binding componnt of mol.wt. approx. 40 OOO did not possess glutathione S-transferase activity.

263. Controlled study on effect of interferon on hepatitis B virus and liver function in HBsAG chronic active hepatitis. W. Weimar, R.A. Heijtink, S.W. Schalm, M.V. Blankenstein, H. Schellinkens, M. Masurel, V.G. Edy, H. Billiau, P de Somer. Erasmus University, Rotterdam. The Netherlands and University of Leuven. Leuven, Belgium. Parenteral administration of human leucocyte interferon has been claimed to stop viral replication in HBsAG positive chronic active hepatitis (CAH). Before starting a study on long-term effects of interferon, we performed dose response studies with human fibroblast interferon under controlled conditions. Entrycriteria for ten untreated CAH patients comprised HBsAg positivity > 6 mo, SCOT > 2x upper limit of normal, HBeAg positivity, measumble HBc associated DNA polymerase activity and liverbiopsy compatible with chronic viral hepatitis. In the first study, two of four patients received interferon i.m.twice weekly during one month with increasing doses of 2-8.106U. In the second study, 3.106U interferon were administered daily for 14 days to three of six patients; no mock preparation was given to controls. HBsAg titre, HBeAg, DNA polymerase activity, transaminase, leukocytes and thrombocytes were determined several times before, during and after the treatment periods in all patients. Results: No consistent effect was observed on viral parameters in both studies. While DNA polymerase levels were relatively stable in the majority of patients, fluctuations were observed in some; one daily interferon treated patient and one control showed a marked persistent decline of DNA polymerase activity. In all three, daily treated patients SCOT levels declined significantly ( x preRx:84 1.U.; x , postRx:49 I.U.). This effect was not observed in control patients. Conclusions: We have not been able to confirm effectiveness of interferon in HBsAg pos CAH. Leukocyte interferon may be more effective than fibroblast interferon. The consistent decline of SCOT levels after daily i.m. interferon merits further investigation.

39. Effect of chenodeoxy- (CDCA) and ursodeoxycholic acid (UDCA) on intestinal transport of water, electrolytes, and oxalate. W.F. Caspary and K. Meyne. Division of Gastroenterology and Metabolism, Department of Medicine, University of GBttingen, Germany. The most common side-effect of treatment with CDCA is a watery diarrhea resulting from CDCA-induced colonic secretion. The 7-8- isomer of CDCA, ursodeoxycholic acid (UDCA) has been shown to lower the lithogenic index and to dissolve cholesterol gallstones. The effect of equimolar concentrations of CDCA and UDCA (2.5,5 mM) on colonic absorption of water, Na, oxalate, and on transmural potential difference was studied in rat colon by an open perfusion technique with PEG 4OOO as marker. Results:

ABSTRACTS 347

Condition H,O' Na" Oxaiate**' p D (mV)

Control + 1.0t0.2 i 1 5 5 ~ 2 3 +0.4+0.1 -21k0.2 CDCA (2.5 mM) -1.4t0.1 -144-eI7 + 1.2f0.1 -11f0.3 CDCA (5.0mM) -2.6f0.1 -321+19 +2.2?0.4 - 6f0.2 UDCA (2.5 mM) O.OfO.l + 28218 +0.4k0.3 -16k0.5 UDCA (5.OmM) -1.OfO.l - 76236 +0.8kO.I -1220.4

= ml, ** = mEq., *** = pmole/l5 min. x g dry weight; given are means f SEM; n = 10; + = absorption, - = secretion. CDCA induced colonic secretion of water and Na at equimolar concentrations more potently than UDCA. Colonic absorption of oxalate was more effectively increased by CDCA than by UDCA, thus supporting the clinical observation that treatment with UDCA will less frequently induce diarrhea than CDCA.

271. Hepatic albumin metabolism in chronic renal failure. S.H. Yap, R.K. Strair, D.A. Shafritz (Intr. by J.H.M. van Tongeren). Liver Research Center, Albert Einstein College of Med. Bronx, NY 10461 and Div. of Gastroent. St. Radboud Ziekenhuis, Nijmegen, the Netherlands. Previously, we have shown that in rats with chronic renal failure (CRF) albumin synthesis by membrane-bound polyribosomes is decreased by 50%. This finding is associated with a increase in the intracellular level of albumin primarily in the cytosol fraction (J. Clin. Invest. 59:869, 1977). Recently, we found that this extra albumin is in the microsomal fraction (CRF 322 + 5pg/g liver, control 146 2 8). Using (3H) DNA complementary to albumin mRNA, we have now measured albumin mRNA sequences directly by molecular hybridization and have found that albumin mRNA content in h e r membrane-bound polyribosomes of uremic rats is increased (59% per g of liver), rather than decreased. The albumin mRNA content in free polyribosomes is essentially unchanged. Accumulation of albumin mRNA in membrane-bound polyribosomes, associated with a decrease in albumin synthesis, suggests a block in protein synthesis at the level of translation. These findings indicate that uremia causes an alteration in liver membrane properties. The decreased rate of albumin synthesis in patients with chrc- nic renal failure has usually been ascribed to the deficient amino acid supply to the liver from anorexia or dietary protein restriction. Improvement of albumin metabolism in patients with CRF after hemodialysis is probably related to direct improvement in protein synthesis upon correction of uremia and is not a secondary phenomenon.

117. Causal relationship between ketonuria and glucagon-induced natriuresis in fasting subjects. J. Kolanowski, J.C. Henguin& J . Crabbk. Univ. Louvain, Medical School, Brussels, Belgium. The present study was designed to assess whether the enhancement of fasting natriuresis by large amounts of glucagon (G) (Eur. J. Clin. Invest. 7 : 167, 1977) results from a concomitant stimulation of ketogenesis and ketonuria. To this end, 6 obese women submitted to an 8-day fast received G ( 1 mg i.v. over 6h) on day 4. The daily intake of sodium (Na ; 43 mMol/d) and potassium (K ; 42 mMol/d) was kept constant throughout. During the first 3 days of fast, Na and K balances were negative, with a progressive rise in plasma levels and urinary excretion of 3-hydroxybutyrate (3-OHB) while ammo- nium (NHJ excretion was not yet increased. G infusion on day 4 raised plasma G from 223 to 2,890 pg/ml, induced an increase in blood glucose levels from 3.2 to 4.9 mMol/l and in plasma insulin (I) from 24 to 47 pU/ml while plasma 3-OHB decreased from 2.4 to 1.8 mMol/l. Contrasting with this reduction in ketonaemia, G infusion led to a marked increase in urinary 3-OHB and Na, whereas the excretion of K, NH,, chloride (CI) and creatinine (Cr) remained unchanged. The mean values for urinary excretions (in mMo1/24 h) were:

348 ABSTRACTS

3-OHB Na K CI NH4 Cr Day 3 30 68 57 71 33 11.0

(G) Day 4

141 144 59 76 30 1 1 . 1 Day 5 78 47 73 61 42 10.0

Thus, in the conditions selected, the natriuretic effect of 0 probably results from increased ketonuria. The latter reflects a direct renal effect of G on ketone body handling since ketonaemia declined, despite a 9-fold decrease in plasma I/G ratio.

196. Sensitivity to insulin of asymptomatic diabetics (A.D.) in vivo and insulin responsiveness of their adipose tissue in vitro. B. Schulz, S. Knospe, I. Rjasanowski and P. Heinke (Intro by H.J. Hahn). Central Institute for Diabetes, Karlsburg, GDR. The purpose of the present study was to assess the role of the glucose-utilizing tissues in early diabetes. Sixteen subjects who had previously been characterized by carbohydrate tolerance and insulin secretion patterns using a 2-hours glucose infusion test (12 mg/kg/min) received a standard diet for 2 days. After that the decrease of blood glucose, free fatty acids, and glycerol levels was measured during a continuous insulin infusion in which 20 mU/min and 40 mU/min were given at two 30 min intervals. The day after subcutaneous adipose tissue was removed by needle-biopsy from abdominal wall. The rate of incorporation of ( 1 -- 14) glucose into CO, and triglycerides (TG) by adipose tissue fragments was estimated in the presence of insulin (62.5 pU/ml). During insulin infusion blood glucose levels decreased by about 24 f 4% in normal subjects (n = 12) and 10 f 6% in A. D. (n = 4), respectively. CO, and TG production by adipose tissue increased up to 135 ? 8 % and 149 rt 8% in normals, but only to 117 f 4% and 110 f 4% in A. D. No differences in the metabolic clearance rate of insulin occured. The results suggest that the reduced sensitivity of substrate-utilizing tissues to insulin might be involved in the development of glucose intolerance in A. D.

266. Effects of a new disacharidase inhibitor (Bay g 5421) on adipose tissue development in the growing rat. T. William-Olsson and L. SjOstrOm. Medical Department I, Sahlgren's Hospital, Gothenburg, Sweden. Since treatment results of adult hyperplastic obesity are very poor preventive measures during childhood are of great importance. However, this prevention is also difficult to handle and new appro- aches are needed. In the present study the effect of a disacharidase inhibitor on adipose tissue development was studied. 50 mg of inhibitor was mixed with 100 g food during the production of pellets containing 24% protein, 5% fat and 48% carbohydrate. Body weight (BW), TG content of epididymal pads (TG-EP) and perirenal fat (TG-PF) as well as fat cell weight (FCW) and number (FCN) of these depots were followed between 21 days and 6 months of age (6 month not yet available). Groups containing 8 male Sprague-Dawley rats were bred under following conditions: a) Ad lib control food; b) Pairfed control food; c) Pairfed inhibitor food; d) As; c) but killed at the same body weight as b); e) Ad lib inhibitor food. Pair-feeding per se had no effects. BW in c) was only 82% and 77% of that in b) a t 40 and 90 days of age, resp. Corresponding figures for othen variables were: TG-EP 50 and 39%; FCW-EP 66 and 50%; FCN-EP 73 and 76%; TO-PF 36 and 24%; FCW-PF 31 and 5 1 %; FCN-PF 99 and 3 1 70. In d) similar results concerning adipose tissue cellularity were obtained. In e) MW was 89% and 89% of that in a). Corresponding figures for other variables were: TG-EP 56 and 63%; FCW-EP 48 and 57%; FCN-EP 116 and 97%; TG-PF 34 and 38%; FCW-PF 39 and 55%; FCN-PF 88 and 68%. Body potassium were similar in all groups at 90 days of age. Insulin was reduced in c) - e). It is concluded that the inhibitor had profound effects on adipose tissue development not only during pair-feeding but also during ad lib feeding. Thus, the drug might possess preventive properties also when administered to growing children.

112. Skeletal muscle metabolism in zucker-fatty-rats: demonstration of generalized insulin resistance. F.W. Kemmer, M. Berger, L. Herberg, T. Koschinsky and F.A. Gries. Diabetes Research Institute, Diisseldorf, West-Germany. The Zucker-fatty-rat (fafa) presents with obesity, hyperlipidaemia. hyperinsulinism and glucose intolerance, a syndrome frequently observed in patients with maturity-onset diabetes mellitus. The aim of this study was to assess whether the apparent insulin resistance in Zucker-fatty-rats is associated with insulin-insensitivity of the skeletal muscle mass. Using the technique of the isolated perfused hind- quarter we studied the glucose metabolism of skeletal muscle in fafa, their lean controls (FaFa) and Osborne-Mendel rats (OM) in absence of insulin (bs) or when 0 .5 , lo and 50 mU insulidml were added to the perfusate. Glucose uptake of skeletal muscle (ex-pressed as pnol/3o g muscle x min, means ? SEM) was (significant differences: vs bs. + vs fafa):

0 Insulin mU/ml 0.5 10 50

fafa 0.03+0.46 0.40f0.61 1.75f0.63 2.81+0.19* Fafa 0.2250.49 2.53f0.45* 5.29?0.65* + 6.37 k0.42 + OM 0.52f0.51 2.18f0.55* + 6.07k0.95. + 7.65+0.74*+

In addition, lactate oxidation was unaffected by insulin in fafa, whereas it was increased in FaFa to 190% (0.5 mU/ml) and in OM to 190% (10 mU/ml). Alanine release was decreased by all insulin concentrations in OM, but remained unchanged in fafa. In the presence of 50 mU insulin/ml, the incorporation of glucose into muscular lipids was reduced by 90% in fafa when compared with FaFa. These results demonstrate a marked generalized insulin resistance of skeletal muscle in Zucker-fatty-rats with respect to glucose uptake, lactate oxidation, alanine release and lipogenesis.

127. Effect of insulin on amino acid transport in isolated rat hepatocytes. A. Le Cam and P. Freychet. Institut National de la Sante et de la Recherche MCdicale (INSERM U.145), Faculte de Medecine (Pasteur), Nice, France. Amino acid transport in the liver is an important step in the regulation of both the protein balance and gluconeogenic process. We have investigated the effect of insulin on amino acid transport, and have compared insulin binding and insulin effect, in freshly prepared suspensions of adult rat hepatocytes. Transport was measured with the non-metabolizable analogs a - a r n i n ~ l [ l - ~ ~ C ] isobutyric acid (AIB) and I-aminocyclopentane [ 1-14Cl carboxylic acid (cycloleucine); binding was determined with I251-insulin; all experiments were performed at 37". Insulin increased the net accumulation of AIB by stimulating the influx. The onset of insulin effect was delayed by 30 to 60 min, to reach a maximum by about 120 min. Insulin increased the Vmax of AIB transport without affecting the apparent Km. Cycloheximide (100pmol/l) and actinomycin D (0.8pmol/l) largely prevented this effect. Insulin specifically stimulated the active, sodium-dependent transport of AIB (System A) without altering the sodium-independent transport of cycloleucine (System L). Dose-responses of insulin effect ranged between 0.1 and 100nmol/l, with half-maximal stimulation occuring at 2 to 3nmol/l, a concentration which also occupies about 50% of the specific binding sites at steady state. Insulin opposed the glyco- genolytic effect of glucagon, but the two hormones exerted partly additive stimulations on AIB transport. These results indicate that insulin stimulates the active component of amino acid transport into the hepatocyte. Half-maximal effect occurs at insulin concentrations within the physiological levels in hepatic portal blood in vivo. This effect is dependent on new protein (and possibly RNA) synthesis, and is partly additive to that of glucagon, a rather unique phenomenon for two hormones which usually exert opposite actions on liver metabolism.

207. Defects in testicular steroidogenesis in Klinefelter's syndrome. A.G.H. Smals, P. W.C. Kloppenborg and Th. J . Benraad. Dpt. of Medicine, Div. of Endocrinology, University of Nijmegen, Nijmegen, The Netherlands. Testicular testosterone (T) secretion and reserve are diminished in most patients with Klinefelter's syndrome (XXY) but data on the secretion of its precursor 17 (I hydroxy-progesterone (17 OHP) are scarce and controversial.

Plasma 17 OHP and T levels were measured in 8 eugonadal men and 9 hypergonadotropic XXY patients before and after 3 days of human chorionic gonadotropin administration (HCG, 1500 iU daily). Results: The mean basal plasma 17 OHP and T levels (85 f 46 and 302 f 145 ng/100 ml) in the XXY patients differend significantly from the values (136 k 39 and 605 k I80 ng/100 ml) in the controls (P < 0.05 - < 0.01). In the XXY patients a positive correlation could be demonstrated between basal 17 OHP and T levels (r = 0.87, p < 0.01), indicating that the synthesis of both steroids was similarly affected. HCG administration elicited a lower T response in the XXY patients than in the control subjects (247 vs 707 ng/100 ml) but 17 OHP increases were comparable in both groups (142 vs 121 ng/ 100 ml). Therefore the mean 17 OHPIT ratio in the XXY patients almost doubled after HCG, whereas this ratio remained unchanged in the control subjects. Conclusion: In Klinefelter's syndrome besides an early defect affecting 17 OHP and T synthesis, evidence for a partial lyase deficiency can be demonstrated.

149. Thermoregulatory responses to cold and heat exposures in patients with anorexia nervosa. K. Nazar, L. BanaS and 2. Sarol. Department of Applied Physiology, Medical Research Centre, Polish Academy of Sciences, Department of Endocrinology, Medical School of the Postgraduate Education and Military Institute of Aviation Medicine, Warsaw, Poland. Patients with anorexia nervosa are known to be very sensitive to cold but their thermal and metabolic responses to cold or heat have not been studied. In the present study, the rectal (Tr) and skin (Ts) temperatures as well as metabolic rate (MR) were measured in 20 female patients with anorexia nervosa and 9 healthy females during the 30 min exposure to cold (10°C) and heat (30°C). Before the exposures Tr and mean Ts in the patients were lower than those in the controls but there was no difference in MR (W/min/mZ). In the patients Tr decreased during cold exposure (-0.5"C) and increased during heat exposure (+ O.Z0C), whereas in the control subjects it remained unchanged. MR increased in cold in the healthy subjects but not in the patients. During the both exposures the changes in Ts and in the skin blood flow (evidenced from the skin conductance calculations) were greater in the patients than in the controls. During heat exposure a marked decrease in arterial blood pressure was found in the patients. It was concluded that both the cold and heat tolerance are impaired in patients with anorexia nervosa. The impairment of the cold tolerance may be attributed to the inability to increase the heat production.

51 Effects of transmeddian transportation on plasma cortisol circadian and ultradian variations. by D. DCsir, V.S Fang, C. Jadot, E. Van Cauter, E. Martino, S. Refetoff and G. Copinschi. Universities of Brussels, Brussels, Belgium and Chicago, Chicago, U.S.A. The duration of hormonal adaptation lags after transmeridian flights has been rarely investigated. Previous studies, based on unfre- quent determinations of plasma cortisol and urinary steroids, showed large variations in the adaptation lags of the adrenal cortex, depending on the individuals and on the direction of the time shift. In order to better assess the phenomenon, 5 healthy male volunteers, aged 21-29, were submitted to blood sampling at 15-min intervals during seven 24h spans over a total investigation period of 10 weeks, including basal study in Brussels (B), westward 7-h time shift by jet towards Chicago (C), studies 36 hours, 11 and 21 days after arrival in C, eastward shift back to B and studies 48 hours, 11 and 21 days after return in B. Standardized sleep-wakefulness schedules (sleep from 2300h to 0700h) and meal time schedules (0800h. 1230h and 1900h) were imposed according to local time (LT). Usual EEG, EOG and EMG procedures allowed to assess nocturnal sleep. Daytime naps were prevented. The cortisol circadian rhythm was still almost fitted to the B schedule, 36 hours after the westward flight, while it was partially adapted again to this B schedule 48 hours after the eastward shift. Adaptation to LT was achieved 11 days after both shifts. Mean 24h plasma cortisol levels were increased immediately after each flight in all subjects but one, then progressively decreased towards basal values. Amplitude of circadian variations, number and duration of secretory episodes were not modified throughout the investigation. Adaptation lag of cortisol circadian variations could be partially responsible for subjective discomfort of people exhibiting the jet lag syndrome.

ABSTRACTS 349 105. Calcium and phosphate metabolism in chronic renal failure (CRF). A longitudinal study. J.R. Juttmann,,J.C. W. Hagenouw-Taal, L.D.F. Lameyer, J . Ruis and J.C. Birkenhgger. Departments of Internal Medicine and Nuclear Medicine, University Hospital ctDijkzigtn, Rotterdam, The Netherlands. In 37 patients with CRF (not on regular haemodialysis) with a creatinine clearance range of 4-49mVmin significant correlations were found between Glomerular Filtration Rate (GFR) on one and serum phosphate (SeP) (p < 0.001), immunoreactive parathyroid hormone (iPTH) (p < 0.005) and intestinal fractional 47Ca absorption (Fa47Ca) (p < 0.025) on the other hand. SeP and Fa47Ca were also correlated (p < 0.025). 13 patients were followed for 2 - 5 months after institution of regula haemodialysis. The mean predialysis Fa47Ca was with 24.3 f 6.2 (SD) lower than in 10 normal subjects, 40.1 f 11.1% dose (p < 0.001). On dialysis the mean Fa47Ca rose to 36.3 k 5.3 ( p < 0.001), accompanied by a fall of SeP (p < 0.001) and iPTH (p < 0.005). In 8 patients Bone Mineral Content (BMC, photon absorptiometry) decreased to 98.0 k 3.3% over a predialysis period of 6 months, while after 6 and 10 months ofdialysisitfellto95.7i 2.8and93.1% & ofthevalueatthestartof dialysis, respectively. In 3 of these 8 patients, in whom transplantation was carried out, post-transplantation values (after 1-3 months) rose by 3.4 and 5%. In 5 non-dialyzed parients with a mean decrease of GFR from 14.6 f 2.1 to 9.6 t 1.7 mVmin over 14.0 f 5.2 months a mean BMC of 94.8 & 4.4% of the original value was found. In 9 non-dialyzed patients with a GFR of 20-50mVmin BMC did not decrease over 12.3 k 3.6 months. Conclusion: Regular haemodialysis treatment improved Fa4'Ca, but not BMC. In 3 patients BMC rose shortly after renal transplantation. In non-dialyzed patients with a GFR below 2Oml/min BMC invariably decreased.

22. Parathyroid-independent change in the renal handling of phosphate in hyperthyroid rats. J . Bommer, J:P. Bonjour, E. Ritz and H. Fleisch. Dept. Intern. Med., Heidelberg, Germany, and Dept. of Pathophysiology, University, Berne, Switzerland. Hyperthyroid patients have a higher plasma concentration and tubular reabsorption of inorganic phosphate (Pi) than normal subjects. These alterations have been attributed to reduced parathyroid activity which may result from the moderate rise in plasma calcium observed in hyperthyroid patients. In the present study we have investigated whether large doses of L-thyroxine (T4) can alter the renal handling of Pi of parathyroidectomized (PTX) rats. 8 days after PTX, hyperthyroidism (HT) was induced by injecting T, 50pg/100 g b.w. i.p. daily for 10 days. During this period HT rats were pair-fed with euthyroid-PTX counterparts. After this period of treatment the tubular capacity to reabsorb Pi was assessed by clearance study. The results show that in PTX rats HT leads to an increase in the capacity to reabsorb Pi. This effect can explain the hyperphosphatemia we have observed in these HT-PTX rats. I t was not associated with a change in urinary CAMP excretion per ml of glomerular filtrate or in the renal handling of Ca. It was maintained under marked extracellular volume expansion. In conclusion experimental hyperthyroidism can lead to a parathyroid-independent increase in the tubular capacity to re-absorb Pi. This increase does not appear to be related to alteration in extracellular volume or change in the renal adenylate cyclase system.

50. Influence of nerve impulse activity on intra-axonal transport of acetylcholine and related enzymes in rat motor neurons. A.G. Dahllof, A. Dahlstrom and P . -0 . Heiwall (Intr. by T. Schersten). Institute of Neurobiol., and Institute of Rehabi- litation Medicine, University of Goteborg, Goteborg, Sweden. In rat sciatic nerve both acetylcholine (ACh) and ACh-esterase (AChE) are transported intraaxonally at a rapid rate (100- 120 mm/d. and 200mm/d., respectively) while choline-acetyl- transferase (CAT) which is soluble, is transported at an apparent slow rate (2-3 mm/d.) The effect of impulse activity upon the intra-axonal transport of these three substances has been studied. A decreased impulse activity was obtained by sectioning the thoracic spinal cord 18 h., 6 d., or 20 d., prior to sacrifice. Increa- sed neuronal activity was obtained by exercising the rats in a

350 ABSTRACTS commercial rodent treadmill for a period of 14 d. The amounts of substances which had accumulated in the nerve segments proximal to a nerve crush during 17. h . were used to calculate the intra-axonal transport. ACh decreased markedly with time after the spinal cord transection. At 20 d. a 80 7' reduction was observed. In contrast, AChE-levels initially increased but with time the level decreased again. CAT decreased somewhat in relation to time after the spinal cord section. Physical exercise appeared to increase ACh with approximately 21 %. The AChE levels appeared unchanged. CAT- levels were not significantly altered after physical training. Maybe there was a decreased activity in the t w o segments just above the crush. Thus input to motor perikarya from supraspinal centres may regulate export from the cell body of motor neurons into their axons. Similar changes may even occur to other substances transported intraaxonally e.g. hypotnetically (ctrophic substancesn which may influence the skeletal muscle fibers in the innervated extremity.

161. Heterogeneity of the capillary flow in the brain. O.B. Paulson and M. M . Hertz. Department of Neurology, Rigshospitalet, Copenhagen, Denmark. The capillary flow in the brain might either be homogenious or inhomogenious. With homogenious flow we shall here understand that the intravascular transit time is similar in all cerebral capillaries which thould be the case if all capillaries had the same length and flow rate. lnhomogenity would be observed if the capillary transit times varied, i.e. i f either the capillry length and/or flow rate varied. In order to investigate this we have studied the venous outflow configuration curve of various tracers using the double indicator single injection technique. Studies were performed in patients in connection with diagnostic angiograms. Two ml of a solution containing a test substance, either 14C-glucose or I4C-propanolol and several intravascular reference substances, W - , zaNa + , Il'lnln-DTPA, was injected into the internal carotid artery. Simultaneously blood was sampled from the internal jugular vein for the determination of the tracer dilution curves. Corrections were made for intralaminar diffusion and for erythrocyte transport. The extraction of the test substance compared to the reference wbstance is defined as :

where CrJt) and C,,,,(t) are the relative concentrations (as compared to the injected solution) in the venous outflow of the reference and test substance respectively. The extractions of the two test substances characteristically increased with time until back- diffusion occured. This configuration must be explained by inhomogenity. The first part of the tracer dilution curve with the low extraction will then correspond to the capillaries with short transit times and the latter part to capillaries with longer transit times.

E(t) = [C,dt) - CtmJt)l / cdtct,

107. Experimental myasthenia in rats. A.M. Kamihska, K. Mrozsek and J . Rafabwska. Intr. by S. Majcherczyk. Department of Neurology, Medical School, Warsaw, Poland. The disorder of neuro-muscular transmission is recognized as a cause of myasthenia gravis and mysthenic syndrome. but the cause of block remains unknown. One of the hypotheses explaining the etiology of myasthenia qavis is the immunological theory based upon clinical, serological md histologjcal findings. We demonstrated that immunization of rats wi th homologous thymic tissue i n the way described earlier by Goldstein and Whittingham /1966/ causes insome animals the disturbances of neuro-muscular transmission characterictic for human myasthenia gravis. Electrostimulation of the medial nerve at frequencies of SWsec and 3/sec performed two weeks after immuniiation demonstrated evidence of neuro-muscular block in 40 '70 of immunized rats while in the group of healthy animals and in the group of rats thymectomized before immunization no evidence of block was found. Miniature potentials of neuro- muscular end-plate recorded with microelectrodes in the inter- transversary muscle of the tail demonstrated decreased amplitude and unchanged frequency in relation to control group. Morpholo- gical changes in the thymus and in the muscle seemed to be non qxcific. No correlation of the presence and intensity of changes with the block was found.

83. Toxicological mechanism of pesticides. F. Hatfaludi, W . Surewich, J. Somogyi, B. Karvaly. 1st Institute of Biochemistry, Semmelweis University Medical School, Budapest and Institute of Biophysics, Biological Research Centre, Hungarian Academy of Science, Szeged, Hungary. In former studies we found the inhibition of the different membrane localised enzyme systems (sodium pump, mitochondria1 electrontransport and substrate transport system etc.) by chlorophenoxyherbicides to be in the same span. Kinetic analyzes of the inhibitions exhibit an uncompetitive inhibitive nature. We supposed a possible model, in which the chlorophenoxyherbicides develop their own inhibitory effect by the alteration on the membrane lipid structure. By this model the negative temperature effect of DDT could be interpreted. The direct effect of the chlorophenoxy group and DDT on the wucture of lipid bilayers have been investigated in model systems, using ESR spectroscopic method. It was found, that all the above compounds affected the fluidity of the hydrocarbon region of bilayered lipids, depending upon pH and concentration. At physiological pH trichlorophenoxy ethanol and DDT decreased considerably the membrane fluidity. These results suggest in the case of poisoning a possible treatment by a membrane fluidising agent.

159. Delayed cellular immunity to measles antigen. V. Parameswaran, M. Robinson and H. Staunton. Depart- ment of Neurology, Richmond (St. Laurences) Hospital, Dublin, Ireland. The therapeutic use of transfer factor in multiple sclerosis has been based on a n alleged depressed cellular immune response in this condition, particularly to measles antigen. We attempted to confirm this finding, using lymphocyte transforma- tion and direct leucocyte migration inhibition as indices. This was controlled against non-specific PHA stimulation in the former and PPD and brucella antigen in the latter. No consistent blastogenic effect of measles antigen was observed in normal subjects with a history of measles (Speke's antigen). No consistent leucocyte migration inhibiting effect was observed in similar subjects (Speke's and Flow measles antigen). It was concluded that, under these conditions it would not be possible to reliably demonstrate a reduced cellular immune capacity to measles antigen in multiple sclerosis.

169. The role of efferent innervation of the carotid body in the regulation of chemoreceptor responsiveness to bleeding. A. Przybyszewski and S. Majcherczyk. Department of Physiology, Warsaw Medical Academy, Warsaw, Poland. A rise in carotid body chemoreceptor activity in response to haemorrhage was observed by several investigators. All of these results were obtained in preparations when chemoreceptor fibers were dissected from sectioned sinus nerve with sympathetic supply frequently disrupted. Recent studies have presented evidence that chemoreceptor activity is influenced both by sympathetic innervation and by efferent activity running down the sinus nerve. In the present work we investgated the effect of lowering arterial pressure by bleeding on chemoreceptor discharge in preparations with intact innervation and following selective disruption of the nervous supply. The experiments were done on cats in chloralose- urethane anaesthesia paralysed and artificially ventilated. Relevant lingual artery was cannulated for measuring carotid sinus pressure. Activity of single chemoreceptor fibers peeled away from the main nerve trunk was examined for the effect of bleeding while the innervation was intact and following section of sympathetic and efferent supply. Majority of fibers responded to bleeding with an increase in activity. This response was found to be potentiated by exclusion of efferent innervation. Suprisingly large proportion of fibers responded to bleeding with a decrease in activity. Disrup- tion of sympathetic supply markedly reduced the response in these fibers. Subsequent section of the sinus nerve had however opposite effect - the inhibifion of chemoreceptor activity to bleeding was markedly enhanced. One fiber which initially responded to bleeding with a reduction of activity, markedly changed the direction of response following section of sinus nerve.

ABSTRACTS 35 1 addition. A key requirement of the passive models for CCMS in the inner medulla is thus fulfilled. 163. High density lipoprotein cholesterol (HDGC) in relatives of coronary patients, relationship to very low density lipoprotein trlglycerldes

D. Pometta, H. Micheli, C. Jornot, J.R. Scherrer. Division de Diabetologie and Division d’lnformatique. Dkpartement de MCde- cine, Geneva, Switzerland. The role of HDL-C and its predictive value in the further development of coronary disease has been shown in coronary patients (Gordon Am. J. Med. 62: 707, 1977). The purpose of the study was to investigate HDL-C and its relationship to VLDL-Tg in men age 20 and over, first degree relatives of coronary patients (relatives) (n = 103) and in healthy controls (n = 370). C and Tg were determined after two steps preparative ultracentrifugation at density 1.006 and 1.063. The student test, the. Mann Whitney U test and comparisons of regression lines were used for the statistical analysis. Relatives of coronary patients have lower HDL-C than the controls (p < 0.001). The regression lines for HDL-C and Log VLDL-Tg are different for the relatives (-0.166 x + 0.43) and the controls (-0.191 x +0.49). It appears that close relatives of coronary patients have lower HDL-C than controls when VLDL-Tg are lower than 3.15 mmol/l. When VLDL-Tg are elevated (>3. I5 mmol/l) both relatives and controls have low HDL-C. Therefore it is suggested that low HDL-C in relatives is partly due to a mechanism independant of the level of VLDL-Tg.

(VLDL-Tg) ,

1. Function of the descending llmb of Henle’s loop in the inner medulla. M. Abramow and E. Cogan. Laboratory of Experimental Medicine (Brugmann Hospital), Free University of Brussels, Brussels, Belgium. The operation of the countercurrent multiplication system in the inner medulla (CCMS) remains controversial. Critical to the validity of passive models are the permeability characteristics of the thin descending limb of Henle’s loop (TDL). We the refore perfused TDL isolated from rabbit inner medulla. With symmetrical artificial solutions in bath and lumen, and perfusing at 2.5 .vf min-1, no transepithelial PD and no net fluid or Na movement 0ccured.h segments bathed in hypertonic medium with excess NaCl and urea, complete osmotic equilibration occured when perfusing at 10 nl min-I. In the collected fluid, the rise in concentration of the volume marker was the same as that of osmolality and of Na and C1. This could occur only if the tubules were sufficiently impermeable to NaCl and urea. As a result, a concentration gradient for NaCl of about 70 mEq/L was generated between the collected fluid and the bath. The transverse electrical resistance of tubules was very high, 700 9 cm*, revealing that TDL is one of the ((tightest)) segments of the nephron. That the Na conductance must be very low was confirmed directly by measuring isotopic Na permeability which was only 1,67 Y lo6 cm sec.1. Urea permeability was also low: 3.35 Y 106cm sec-1. We conclude that osmotic equilibration in the inner medullary TDL occurs through water abstraction with virtually no solute

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Abstracts of the Eleventh Annual Meeting of the European Society for Clinical Investigation, Rotterdam, The Netherlands, April 20–22, 1978