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British Journal of Haematology, 1983, 54, 485-488 Factor VIII inhibitor and raised platelet IgG levels associated with methyldopa therapy S. DEVEREUX, D. M. FISHER, B. L. T. ROTER AND U. M. HEGDE Department of Haematology, Ealing Hospital, London Received 22 October 1982; acceptedforpublication 28 January 1983 SUMMARY. An inhibitor to the plasma coagulation factor VIII and abnormally high levels of platelet associated IgG (PAIgG) were found in a patient whilst on methyldopa. Both these parameters felI on initial withdrawal, but on rechallenge with the drug only the PAIgG rose to the high presenting levels. No inhibitory activity to factor VIIIc could be detected in a concentrated eIuate prepared from the patient’s platelets. These results may imply that two distinct antibodies were provoked by the administration of methyldopa, but could also be explained by the action of anti VIIIc alone on the patient’s platelets. A positive direct antiglobulin test (DAGT)due to IgG autoantibody on the red-cell surface is a common abnormality in patients on the anti-hypertensive agent methyldopa, although significant haemolysis causing anaemia is rare, occurring in less than 1% of these (Petz & Garratty, 1980). In contrast to observations on the red cell, there have been no reports of abnormal amounts of platelet associated antibody in non-thrombocytopenicpatients on this drug. We report a patient whose platelet count was consistently normal and in whom markedly elevated levels of platelet associated IgG as well as an inhibitor to the plasma coagulation factor VIII were observed: both seemed to be related to the administration of methyldopa. CASE REPORT A 77-year-old white Caucasian woman presented with a 2-month history of easy bruising. Apart from hypertension and mild peripheral vascular disease she was in good health. Drug therapy at this time was comprised of naftidro-furyl, labetolol and methyldopa 2 50 mg t.d.s. which she had taken for 3 years. Physical examination was normal apart from several large bruises on her extremities. Preliminary tests showed the presence of an acquired plasma coagulation inhibitor as well as elevated levels of platelet-associated IgG. Because of its known propensity to provoke autoimmune disorders, methyldopa therapy was stopped and serial measurements of the abnormal parameters were performed. Correspondence: Dr U. M. Hegde, Department of Haematology, Ealing Hospital, Uxbridge Road, Southall, London UB1 2HW. 48 5

Factor VIII inhibitor and raised platelet IgG levels associated with methyldopa therapy

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Page 1: Factor VIII inhibitor and raised platelet IgG levels associated with methyldopa therapy

British Journal of Haematology, 1983, 54, 485-488

Factor VIII inhibitor and raised platelet IgG levels associated with methyldopa therapy

S. DEVEREUX, D. M. FISHER, B. L. T. ROTER AND U. M. HEGDE Department of Haematology, Ealing Hospital, London

Received 22 October 1982; accepted forpublication 28 January 1983

SUMMARY. An inhibitor to the plasma coagulation factor VIII and abnormally high levels of platelet associated IgG (PAIgG) were found in a patient whilst on methyldopa. Both these parameters felI on initial withdrawal, but on rechallenge with the drug only the PAIgG rose to the high presenting levels. No inhibitory activity to factor VIIIc could be detected in a concentrated eIuate prepared from the patient’s platelets. These results may imply that two distinct antibodies were provoked by the administration of methyldopa, but could also be explained by the action of anti VIIIc alone on the patient’s platelets.

A positive direct antiglobulin test (DAGT) due to IgG autoantibody on the red-cell surface is a common abnormality in patients on the anti-hypertensive agent methyldopa, although significant haemolysis causing anaemia is rare, occurring in less than 1% of these (Petz & Garratty, 1980). In contrast to observations on the red cell, there have been no reports of abnormal amounts of platelet associated antibody in non-thrombocytopenic patients on this drug. We report a patient whose platelet count was consistently normal and in whom markedly elevated levels of platelet associated IgG as well as an inhibitor to the plasma coagulation factor VIII were observed: both seemed to be related to the administration of methyldopa.

CASE REPORT A 77-year-old white Caucasian woman presented with a 2-month history of easy bruising. Apart from hypertension and mild peripheral vascular disease she was in good health. Drug therapy at this time was comprised of naftidro-furyl, labetolol and methyldopa 2 50 mg t.d.s. which she had taken for 3 years. Physical examination was normal apart from several large bruises on her extremities. Preliminary tests showed the presence of an acquired plasma coagulation inhibitor as well as elevated levels of platelet-associated IgG. Because of its known propensity to provoke autoimmune disorders, methyldopa therapy was stopped and serial measurements of the abnormal parameters were performed. Correspondence: Dr U. M. Hegde, Department of Haematology, Ealing Hospital, Uxbridge Road, Southall, London UB1 2HW.

48 5

Page 2: Factor VIII inhibitor and raised platelet IgG levels associated with methyldopa therapy

486 S. Desereux ef a1

METHODS

Platelet-associated IgG was determined by enzyme-linked immunoassay (Hegde et al, 1981). Concentrated extracts of platelet IgG were prepared as follows. Platelet concentrates were obtained after high-speed centrifugation of platelet-rich plasma from patient and normal controls. resuspended in phosphate-buffered saline, pH 7.4, and then submitted to freeze-thawing, sonication and ultracentrifugation procedures to elute the PAIgG in the supernate (Hymes t?f al. 1979). This was then dialysed overnight against Owren's buffer and further concentrated five times using polyacrylamide gel. All other tests were performed by standard methods.

RES tJLTS

Investigations were as follows: Hb 13.2 g/dl, WBC 6.8 x 10y/l, platelets 320 x lo9/[, normal differential and red cell morphology. The direct antiglobulin test was negative. No antibodies to red cells or to tissue antigens were detected in the serum and the serum immunoglobulin levels were normal. The prothrombin time, thrombin time and plasma fibrinogen levels were normal. The partial thromboplastin time with kaolin (PTTK) was prolonged at 7 3 s (control 3 5 s) and this failed to correct with the addition of equal quantities of normal plasma. A one-stage factor VIIIc assay showed only 2 3% activity, with normal factor V, IX and X levels. Prothrombin times using serial dilutions of thromboplastin did not differ significantly from control values. These results were interpreted as showing the presence of an acquired

Methyldopa]

- 50 E - 40

'C 30

20 2 10

vl c

2

v, - W M

IMethyldop 1

. m l

'March 'Apr i l 'May 'June 'July 'August ISept. '82 I

Fig 1. Changes in PAIgG. PTTK and factor 1'111 inhibitor levels on withdrawal and rechallenge with methyldopa.

Page 3: Factor VIII inhibitor and raised platelet IgG levels associated with methyldopa therapy

Methyldopa, Factor VZZZ and Platelet IgG 48 7

inhibitor to factor VIII which was initially measured at 43 Bethesda units/ml. In addition, the platelet-associated IgG was quantitated and found to be markedly elevated at greater than 150 ng/lOh platelets ( n 3.0-17.5).

Methyldopa therapy was stopped and serial measurements of PTTK, PAIgG and the inhibitor to factor VIIIc showed significant falls in these levels over a period of time. Rechallenge with methyldopa failed to reinduce the VIIIc inhibitor but reproduced the very high levels of PAIgG which fell subsequently on withdrawal again (Fig 1). Two months after finally stopping methyldopa, when the inhibitor could no longer be demonstrated, factor VIIIc activity had risen to 67%. The platelet count remained normal throughout the period of study.

Further tests were performed to determine whether the abnormal platelet associated IgG and the factor VIII inhibitor were identical. Aliquots of normal human plasma were incubated for 1 h at 37OC with equal quantities of the concentrated platelet IgG extracts prepared as described above. Factor VIII levels were determined before and after incubation; however, no inhibitory activity could be demonstrated in extracts from either the patient’s or control platelets. Furthermore, the incubation of normal pooled platelets in our patient’s plasma (tested when anti VIIIc was demonstrable in high titre) failed to provoke any rise in platelets associated IgG above their normal preincubation levels.

DISCUSSION

Therapy with methyldopa is known to be associated with autoantibody production although clinically manifest autoimmune disease such as haemolytic anaemia or systemic lupus erythematosus is unusual (Petz & Garratty, 1980; Dupont & Six, 1982). Reports of methyldopa-induced platelet antibody have been confined to thrombocytopenic patients, in one case associated with a positive DAGT (Manohitharaja et al, 1979) and in another in whom drug dependent antibody was demonstrated in the patient’s serum (Marcus & Stevenson, 19 75). In our patient, the abnormally elevated PAIgG without concomitant thrombocytopenia clearly did not require presence of a drug for its demonstration, and it is tempting to postulate that the abnormal PAIgG represented true autoantibody directed against platelets, analogous to the positive DAGT seen in approximately 20% of patients on methyldopa.

Current views on methyldopa induced autoimmunity propose a non-specific defect of immune regulation mediated by inhibition of T suppressor lymphocyte function (Kirtland et al, 1980), which allows multiple autoantibody production. Thus the anti VIIIc found in our patient’s plasma could have been a distinct entity, since we could neither detect inhibitory activity to factor VIII in a concentrated platelet extract nor could we provoke an increase in PAIgG of normal platelets after incubation in our patient’s plasma when it contained anti VIIIc in high titre. Furthermore, the falls in PAIgG and factor VIII inhibitor levels did not parallel one another after withdrawal of methyldopa and indeed on rechallenge only a rise and fall in PAIgG was demonstrated. It is also interesting that the VIIIc inhibitor provoked both in vivo and in vitro plasma coagulation abnormalities whilst the high PAIgG levels did not cause thrombocytopenia. However, the association of factor VIII inhibitors and abnormal

Page 4: Factor VIII inhibitor and raised platelet IgG levels associated with methyldopa therapy

488 S. Devereux et nl amounts of platelet surface IgG has previously been described (Ali &I Blajchman, 1972) and these authors suggested that the inhibitor found in their patients’ plasma was responsible for the elevated IgG levels on the platelet membrane. i.e. that only one autoantibody, that against VIIIc, was involved. If so, the elevated PAIgG in our patient could be accounted for by the presence of anti VIIIc-VIIIcAg complexes on our patient’s platelets. Also, if only such complexes were present in the concentrated platelet extract, they would not be expected to exert any inhibitory effect on normal plasma coagulation. The inability of the inhibitor to VIIIc to bind to normal platelet could either be because of our failure to reproduce the in vivo conditions of this particular antgen-antibody reaction or because washing procedures removed sufficient VIIIc from donor platelets to make it impossible to measure precisely any elevation of PAIgG.

In summary. the administration of methyldopa in our patient provoked an inhibitor to the plasma coagulation factor VIIlc and also abnormally high levels of platelet associated IgG. Although it is theoretically feasible that these were two distinct autoantibodies, our results could equally be interpreted on the assumption that only one autoantibody, that against factor VIIIc, was produced.

A C K N O W L EDG M E N T S

We thank Mrs Linda Byrne for secretarial assistance. This work was supported by the Localized Organized Research Scheme, North West Thames, RHA.

REFERENCES

Ai.1, M.A.M.&BLA~cHMAN.M.A. (1972)Studiesof the antibody on the platelet surface in patients with factor VIII inhibitors. British Joitrtirrl of Hrrematology, 22, 365-368.

DUPONT. A. & SIX. K. (1980) Lupus like syndrome induced by methyldopa. British A4rdical]ourrinl. 285. 693-694.

E ~ E G ~ E . U.M.. Bows. A.. POWELL. D.K. & JOYSER.

M.V. (1981) Detection of platelet bound and serum antibodies in autoimmune thrombocy- topenia by enzyme linked assay. Vox Sanguiriis. 41. 306-312.

HY.MIES. K . . SHULMAS. S. & KARPATKIS. S.A. (1979) Solid phase radio immunoassay for bound anti platelet antibody. ]oiirrial o/ Laboratory orid C/iriirn/ Medkirir. 94, 6 39-648.

KIRTLAND, H.M.. MOHLER, D.N. & HOROWITZ. P.A. ( 1980) Methyldopa inhibition of suppressor lymphocyte function. New England journal of Medicine. 302, 825-832.

ASOH OH IT HA RAJA. S.M.. JENKINS, W.J., ROBERTS, P.D. & CLARKE, R.C. (1979) Methyldopa and associated thrombocytopenia. British Medical Joitrtinl. i , 494.

MARCIS. G.J. & STEVENSON, M. (1975) Alpha methyldopa induced thrombocytopenia. Ameri- m i Joirrrial of Clinical Pathology, 64, 11 3-1 15.

PETZ. L.D. & GARRATTY. G. (1980) Acquired lm- tniine Huemolytic Anaemias, pp. 185-289. Chur- chill Livingstone, Edinburgh.