Anti-idiotypic antibodies specific for HLA in heart and kidney allograft recipients

Anti- Idiotypic Antibodies

Immunol Res 1993;12:1-11

Elaine Reed a, Eric Ho a David J. Cohen b Will iam R a m e y d

Charles Marboe a Vivette D'Agati a Eric A. Rose c, M a r k Hardy c

Nicole Suciu-Foca a

College o f Physicians and Surgeons o f Co lumbia Univers i ty , Depa r tmen t s of

a Pathology, b Medicine and c Surgery; and d St. Luke 's-Roosevel t Hospital ,

Depar tmen t o f Surgery, New York, N.Y., USA

o ~ 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6

Key Words Anti-idiotypic antibodies Anti-HLA antibodies Ir gene control Heart transplantation Kidney transplantation

Anti-ldiotypic Antibodies Specific for HLA in Heart and Kidney Allograft Recipients

+.oo~176176176176176176176176176176176176176176176176176174176176176176176176176176176176

Abstract Chronic rejection is the major threat to both heart and renal allograft survival. We have explored the possibility that some patients with anti-donor HLA antibodies (Ab i) develop specific anti-idiotypic antibodies (Ab2) which suppress the production of Abl, and subsequently, the progression of chronic rejection. Analysis of Ab2 in sera obtained from Ab 1 producers showed that 22% of heart and 18% of kidney recipients produced Ab2. The 4- and 5-year actuarial graft survivals in Ab2 producers were 100% and 83%, respectively, compared to 57% in patients who formed Abl but not Ab2 (p < 0.004). Patients carrying the DR2 alleles, DRB 1 * 1501, * 1502 or * 1601 were at a lower risk of producing anti-donor HLA antibodies. ~176176176176176176176176176176

Chronic rejection, causing the persistent attrition of functioning allografts, represents the major threat to long-term survival of organ transplants [i-5]. Although the patho- physiologic mechanisms leading to late graft failures is ill understood, it has long been sus- pected that antibodies reacting with allograft antigens are involved [5].

Supporting this view is the consistent finding of immunoglobulin and complement deposition in affected vessel walls of the graft [5-8] and the presence of anti-don0r HLA antibodies in recipient sera [9, 10]. The production of anti-HLA antibodies implies the activation of T helper cells which recognize peptides derived from the processing and pre-

sentation of allogeneic major histocompatibility complex (MHC) molecules by host antigen-presenting cells (APCs) [1 I, 12]. Such a process of indirect 'allo-recognition' may in turn depend on the ability of host MHC class II molecules to bind allopeptides [11-15]. A corollary of this postulate is that an Ir gene effect may be evidenced in transplantation by the association of responder/nonresponder status with certain MHC class II genotypes. Evidence for positive or negative associations between the outcome of the graft and recipient MHC phenotype has already been described [16-22], yet it has been difficult to confirm mostly because of ambiguities in serologic typing of human HLA class II anti-

Dr. Nicole Suciu-Foca College of Physicians and Surgeons of Columbia University 630 West 168th Street New York, NY 10032 (USA)

�9 S. KargerAG, Basel 0257-277X/93/ 0121-000152.75/0

gens. Such Ir gene effects may be obscured, however, by the development in the host of immunoregulatory mechanisms suppressing rejection, such as anti-idiotypic anti-HLA antibodies, for example. In an attempt to ex- plore these possibilities, we have analyzed the HLA-DR genotype of heart and kidney allograft recipients, and evaluated their sera for anti-HLA (Abl) and anti-anti-HLA (Ab2) antibodies. The data presented in this paper sub- stantiate the existence of an Ir gene effect, as well as the regulatory role of Ab2 which may suppress the progression of chronic organ allograft rejection.

Materials and Methods

Patient Population Selection of patients for these studies was limited

to recipients who had been monitored for anti-HLA antibodies at least four times during the first year following transplantation and had available cryopre- served lymphocytes from the corresponding cadaver donor. A total of 111 heart and 59 kidney aUograft recipients fulfilled these criteria. All patients were recipients of primary cadaver grafts and received Cy- closporine-based immunosuppressive therapy.

Statistical Analysis Actuarial graft survival rates were computed by the

Kaplan-Meier estimate, and the log rank test was used for statistical comparison of survival curves (BMDP I-L, BMDP Statistical Software). The Z-' test was used to determine significant associations between DRBI alleles and anti-HLA antibody production (SAS/STAT software). The duration of transplant survival was computed from the date of transplantation to the date of recipient death or retransplantation in the case of cardiac allograft recipients and to the date of return to dialysis for renal graft recipients. Since unsuccessful immunosuppression may result in rejection, malignan- cies or infections, the assumption was made that immunological problems were the major underlying cause of transplant failure.

The hypothesis that anti-HLA antibody producers (i.e. responders) differ with respect to their HLA-DR genotype from patients who do not form antibodies (i.e. nonresponders) was tested by constructing 2 X 2 contingency tables for each of the 37 DRB1 genes

encountered in the patient population, p values were corrected to control for type I error introduced by the performance of multiple comparisons: the raw p value was multiplied by the number of comparisons performed, which is 37 for DRB1 analysis [23].

HLA-A, B, C and DR Typing Recipients and donors were typed for HLA-A,B,C

and DR antigens by conventional serology, as previously described [24]. The HLA-DRB1 genotype of 161 recipients was determined by polymerase chain reaction amplification of the first domain of the DRB 1 * gene and hybridization with sequence specific oligonucleotide probes, as previously described [25, 26]. Molecular typing was performed for the following DRBI* alleles: 0101, 0102, 0103, 1501, 1502, 1601, 1602, 0301,0302, 0401,0402, 0403, 0404, 0405, 0406, 0407, 0408, 0409, 0410, 0411, 1101, 1102, 1103, 1104, 1201, 1202, 1301, 1302, 1303, 1304, 1305, 1401, 1402, 1403, 1404, 1405, 07, 0801, 0802, 0803, 0804, 09 and 1001.

Evaluation of Anti-HL4 Antibodies Sera obtained at the time of transplantation and at

3-month intervals following transplantation were depleted of soluble HLA class I and class II antigens [9, 10, 27] and then screened for anti-HLA-A,B,C. DR and DQ antibodies on an HIA reference pancl of 70 cells [24]. Assignment of anti-HLA antibody spccific- ity was based on the tail analysis program (UCLA Soft- ware) indicating a combined correlation coefficient >__0.7 between serum reactivity and the presence ot" certain HLA antigens on target lymphocytes. All sera were tested for reactivity with donor T and B cells and for autoreactivity with recipient lymphocytes. To ex- clude the possibility that lymphocytotoxicity of the patients' sera was caused by antilympbocytic antibodies used for treatment, such as monoclonal antibody (mAb) OKT3 or ATG, sera were tested by immu- nocytofluorometry for mouse and horse antibodies using FITC-conjugated goat-anti-mouse Ig (Tago, Bur- lingame, Calif., USA) and FITC-conjugated goat-anti- horse lg (Chemicon, Temecula, Calif.). For this, 5 X 105 lymphocytes were incubated with 50 pl of patient serum for 30 min at 4 ~ After 30 min of incubation, the cells were washed 3 times in PBS containing 2.5% fetal calf serum and 0.05% sodium azide. Cells were then stained with 50 ~tl of the appropriate secondary antibody. After 30 min of incubation at 4 ~ cells were washed and analyzed on a FACSTAR cytofluoro- graph (Becton-Dickinson, Mountain View, Calif.). Sera containing xenogeneic antibodies were excluded from the analysis.

Reed/Ho/Cohen/Ramey/Marboe/ Anti-ldiotypic Antibodies in D'Agati/Rose/Hardy/Suciu-Foca Transplantation

Determination of Anti-Idiotypic Antibodies The procedure used for ascertaining the presence of

anti-idiotypic antibodies specific for HLA in patient sera has been previously described [9, 10, 24]. Briefly, the following steps were followed: (1)all sera were depleted of soluble HLA class I and class II antigens by incubation with magnetic beads coupled with mAbs to common epitopes of HLA class I (mAb W6/32) and class II (Dynabeads HLA Cell Prep II) (Dynal Inc., Great Neck, N.Y., USA). The completeness of soluble antigen depletion was monitored as previously described [9, 10]; sera were considered free of antigen if they no longer inhibited murine anti-HLA mAbs. (2) Following HLA-antigen depletion sera were cross- matched with donor lymphocytes by complement- mediated lymphocytotoxicity and by immunofluorescence cytofluorometry to identify complement-fixing and non-complement-fixing antibodies [24]. (3)Sera containing anti-donor antibodies were titered in serial dilutions using donor T and B lymphocytes as target cells. Sera with antibody titers greater than 1:2 were selected as Abl reagents; where more than one serum with the same specificity was available, the specimen with the highest titer was chosen. (4) Sera showing no reactivity with donor lymphocytes either by cytotoxic- ity or immunofluorescence testing were assayed for Ab2-blocking activity. For this. serial dilutions of Abl in McCoy's medium were plated at 1 gl/weU in 60-well Terasaki plates. One microliter of test serum (putative Ab2) was added to each well. Following a 60-min incubation at 24 ~ C, target lymphocytes were plated. Com- plement was added after l h, followed by ethidium bromide. For each patient, the serum obtained at the time of transplantation was used as a baseline-internal negative control in blocking assays. The test serum was considered inhibitory only when it inhibited Abl above the baseline value. (5)The specificity of Abl blocking was ascertained using reference human anti- HLA antisera of the same and of a different HLA specificity as controls [28, 29].

Results

Actuarial Survival of Heart and Renal Allografts in Patients with and without Anti-Donor HLA Antibodies The deve lopment of ant i -HLA antibodies

coincides with the onset o f a complex chain of immunologic events, presumably involving the provision o f T cell help for B cells and rec-

ognition by the helper T cell receptor of allopeptides bound and presented by B cells [ 12]. To determine the impact o f an t i -HLA antibodies on allografts, we analyzed the actuarial graft survival at 5 years in recipients with and without ant i -donor HLA antibodies.

Patients were considered ant i -HLA antibody producers if at least 1 of their sera reacted with donor and panel lymphocytes. In heart allograft recipients without ant i -HLA antibodies (n = 48) the actuarial survival at 5 years was 86%. The rate of t ransplant failure was 9, 2 and 3% after 1, 2 and 3-5 years, respectively. In recipients o f heart allografts who developed ant i -HLA ant ibodies (n = 63) during the first year following transplantation, the long-term survival rate was significantly lower (p = 0.05), decreasing f rom 84% after 1 year to 56% after 5 years. There was a steady and progressive rate of at tr i t ion rang- ing f rom 3 to 12% per year (fig. la). Similar results were obtained f rom the analysis o f allograft survival in kidney recipients with (n = 40) or without (n = 19) ant i -donor HLA antibodies. The actuarial graft survival at 5 years was 100% for patients who formed no ant i -donor antibodies. Patients developing such antibodies following t ransplantat ion showed a survival rate of only 66% after 5 years, a result which is significantly different (p = 0.01) f rom that seen in the patients who did not produce such antibodies. After the first year, when 18% of the grafts were lost, the annual rate of at tr i t ion ranged f rom 8 to 10% (fig. l b). The combined actuarial survival at 5 years in heart and renal recipients was 92% for non-ant ibody producers (n = 67) and 62% in ant ibody producers (n = 103). The difference between the groups is highly significant (p = 0.003) (fig. lc).

Fig. 1. a Actuarial survival of heart allografts in patients with (n =

63) and w i t h o u t (n = 48) a n t i - d o n o r

H L A ant ibod ies , p = 0.05. b Ac-

tuar ia l su rv i va l o f renal a]]ografts

in pat ients w i t h (n = 40) and w i th -

ou t (n = 19) Abl . p = 0.01. c Com- bined actuarial survival of heart and renal allografts in patients with (n = 103) and without (n = 67) Abl . p = 0.003.

% Survival

% Survival

% Survival

100

80

60

40

20-

0 0

Abl -

Abl+

I i i i i

1 2 3 4 5

100 '

80'

6 0 -

40=

20 -

0

s : = Abl-

rt Abl+

i i i 1 i

I 2 3 4 5

100 ~ " ~ - " - - - - - - - - ~ ~ a [] Abl -

80

60 ~ -': Abl+

40

20

0 0

i / i i i

1 2 3 4 5

Years Post-Transplantation

Relationship between Anti-Idiotypic A n tibodies Specific for HLA and A llograft Survival Although the actuarial graft survival at 5

years was significantly lower in patients developing anti-donor HLA antibodies than in those who did not there was still heterogeneity

in the antibody-producing group, as some of these patients remained in quiescence after 5 years. We explored the hypothesis that anti- idiotypic antibodies may suppress the production ofanti-HLA antibodies slowing down the progression of chronic rejection. Of the 63 heart allograft recipients producing anti-HLA

Reed/Ho/Cohen/Ramey/Marboe/ Anti-ldiotypic Antibodies in D'Agati/Rose/Hardy/Suciu-Foca Transplantation

Tab le 1. Patterns of Abl and Ab2 production in heart and renal allograft recipients

Patterns of Recipients with Abl and Ab2 anti-HLA antibodies

production heart kidney (n = 63) (n = 40)

Persistent Abl 10(16%) 20(50%)

Cyclic Abl 53(84%) 20(50%) with Ab2 14(22%) 7(18%) withoutAb2 39(62%) 13(33%)

Fig. 2. a Actuarial survival of heart allografts in patients with (n = 14) and without (n = 49) anti-idiotypic antibodies, p = 0.01. bAc- tuarial survival of renal allografts in patients with (n = 7) and without (n = 33) Ab2. p = 0.08. c Combined actuarial survival of heart and renal allografts in patients with (n = 21) and without (n = 82) Ab2. p = 0.0O4.

lOO

8o,

% Survival 60 -

4 0 -

2 0 -

o

lOO

80

% Survival 60

40

20

o

lOO

8o

% Survival 60

4 0 -

2 0 -

o

| i i i

3 4 5

Abl+Ab2+

Abl+Ab2-

=3

Abl+Ab2+

~ Abl+Ab2-

i ! i

i i i i

2 3 4, 5 Years Post.Transplantation

Abl+Ab2§

Abl+Ab2-

antibodies 10 (l 6%) had persisting antibodies in all sera, while 53 (84%) showed cyclic fluctuations of Ab 1. In heart recipients with cyclic fluctuations o fAbl only some of the sera con- tained anti-donor antibodies while other sub- sequent sera were negative. Such variations were associated with the presence of Ab2 in

14 patients (22%), while 39 patients (62%) showed no Ab2 in their sera (table t). The actuarial graft survival was 100% at 4 years and 80% at 5 years in Abl producers who developed Ab2 (fig. 2a). In contrast, heart allograft recipients who showed persistent levels of Abl or cyclic variations that were not

Fig. 3. Comparison of actuarial survival of heart and renal allografts in patients with cyclic fluctuations of Abl associated with Ab2 (n , n = 21); cyclic fluctuations of Abl not associated with Ab2 (El, n = 52); or persistent levels of Abl (#, n = 30). p = 0.01.

% Surv iva l

100-~

8 0 -

6 0 -

4 0 -

2 0 -

0 j l | l I

1 2 3 4 5

Years Post-Transplantation

accompanied by the production of Ab2 had an actuarial survival of 57% at 4 and 5 years. The difference between the groups is statisti- cally significant (p = 0.01) and indicates that the development of anti-idiotypic antibodies improves the prognosis of patients producing anti-HLA antibodies (fig. 2a).

Of the 40 kidney allograft recipients who developed anti-HLA antibodies following transplantation, 20 showed persistent and 20 showed cyclic variations of Abl in sequential sera. Cyclic fluctuations of Abl were associated with the presence of Ab2 in 7 patients (18%), while the remaining 13 (33%) never showed Ab2 (table 1).

The 5-year actuarial survival was 100% in patients with Ab2, compared to 58% in Abl producers without Ab2 (fig. 2b).

The combined analysis of heart and renal allograft recipients producing Ab 1 shows that the actuarial graft survival was 100% at 4 years and 83% at 5 years in patients developing Ab2, compared to 57% at 4 and 5 years in patients without Ab2 (p = 0.004) (fig. 2c). Graft survival at 5 years was lowest in patients with persistent Abl (47%) and highest (83%) in patients with Ab2 (fig. 3). These data

indicate that the evaluation ofanti-HLA antibodies and of specific anti-idiotypic antibodies is of prognostic importance for allograft recipients.

The specificity and titer of the anti-idiotypic antibodies found in heart and renal allograft recipients is summarized in table 2. The anti-idiotypic antibodies found in heart allograft recipients blocked specifically an anti- donor HLA class I antibody in 5 cases, a class II antibody in 3 cases, and both a class I and a class II antibody in 6 cases.

In renal allograft recipients, Ab2 was specific for an Abl to donor class I in 2 cases, class II in 2 cases and both a class I and a class II antigen in 3 cases. Hence, HLA class I and class II alloantigens expressed by the graft elicit the production of anti-HEA antibodies which can in turn stimulate the development of Ab2.

Although idiotypes are individual anti- genic specificities, among antibodies developed by one individual in response to a particular foreign antigen, certain functionally related antibodies carry cross-reactive idiotypes (Idx) which are shared among different individuals and often display regulatory func-

Reed/Ho/Cohen/Ramey/Marboe/ AntMdiotypic Antibodies in D'Agati/Rose/Hardy/Suciu-Foca Transplantation

Table 2. Study of anti-idiotypic autoantibodies to donor HLA atloantigens in heart and renal allograft recipients

Patient Autologous Ab 1 to donor HLA Allogeneic Ab 1 to Allogeneic Ab I to No. donor HLA control HLA specificity titer in serum a

baseline test speci- titer in serum a speci- titer in serum a

ficity baseline test ficity baseline test

Heart allograft 1 B7, DR4 1:2 _b B7 1:8 1:2 DR2 1:64 1:64

DR4 1:16 1:16 2 DR7 1:8 1:2 DR7 1:8 1:2 DR2 1:64 1:64 3 DR5 1:16 1:4 DR5 1:16 1:2 DR2 1:64 1:63 4 B7, A29 1:4 _b B7 1:4 _b DR5 1:16 1:16

A29 1:64 1:64 5 DR3 1:32 1:2 DR3 1:64 1:8 B8 1:32 1:32 6 B40 1:4 _b B40 1:8 1:8 DR7 1:8 1:8 7 B8 1:8 1:2 B8 1:32 1:32 DR3 1:64 1:64 8 B 5 1:4 _b NT NT 9 B 18 1:4 1:2 NT NT

10 A28, B5, DR6 1:4 _b NT DR7 1:8 1:8 11 A33, B35, DR3, DR5 1:4 _b NT DR7 1:8 l:8 12 A24, BI5, B35, DR6 1:8 1:4 NT DR7 1:8 1:8 13 A3, A23, BI2, DR7 1:8 1:2 NT DR2 1:64 1:64 14 A3, B7, B35, DRI, DR2 1:4 1:2 NT DR5 1:16 1:16

Renal alh)gr~t/'t 1 A2, DR7 1:4 _b A2 1:4 _b DR2 1:64 1:64

DR7 1:8 1:8 2 B35, DR7 1:8 _b B35 1:4 _b DR2 1:64 1:64

DR7 1:8 1:8 3 A28, B7 1:16 _b A28 1:8 1:8 DR5 1:16 1:16

B7 1:8 1:8 4 A23 1:8 1:2 A9 1:8 1:8 DR2 1:64 1:64 5 DR4 1:8 1:4 DR4 1:16 1:16 DR2 1:64 1:64 6 DQwl 1:16 1:2 DQwl 1:16 1:16 DR7 1:8 1:8 7 A30, BI7, B42, DR6 1:8 _b NT DR2 1:64 1:64

NT = Not tested due to insufficient amount of test serum. a Indicates the highest dilution of ant ibody with lymphocytotoxic activity. b Indicates the absence oflymphocytotoxic activity.

tion [30]. To determine whether Ab2 found in transplant patients recognize such Idx, we tested the ability of sera containing Ab2 to block anti-HLA antibodies (Abl) of the same and of unrela ted specificity derived from other individuals [28, 29].

Five heart (cases 1-5) and two renal (cases 1, 2) allograft recipients had Idx since the anti-idiotypic antibodies which they developed inhibited anti-HLA antibodies of the same specificity from other HLA reference sera. None of the Ab2 inhibited anti-HLA

Table 3. Association between the HLA-DR2 alleles DRBI*1501, "1502, "1601 and anti-HLA antibody production

Responder Anti-HLA antibody

positive negative

DR2 positive 35 t 6 DR2 negative 103 7 Tetal ! 38* 23*

X 2 = 17.798. *p = 0.00006; Bonferroni's adjusted p = 0.002.

antibodies specific for an antigen other than that recognized by the patient's own anti- donor Abl.

the population negative for that allele) and responsiveness (i.e. Abl) versus nonresponsiveness to donor HLA antigens. After correc- tion for the number of comparisons made, a significant association was found both in heart and in kidney allograft recipients between the HLA-DR2 genes DRBI*1501, *1502 and *1601 and nonresponsiveness to the donor (p = 0.002) (table 3). The DRBI* 1602 allele was not carried by any of the heart recipients and showed no significant association with the potential to produce antibodies in kidney allograft recipients. There was no difference between DR2-positive and -negative recipients with respect to the mean number of mismatched donor HLA antigens and with respect to allograft survival.

Relationship between Recipient HLA-DRB1 Genotype and Ability to Produce Anti-Donor HLA Antibodies The immune response to alloantigens is

likely to be controlled by Ir genes because T helper cell recognition of peptides derived from the processing of allogeneic MHC molecules is restricted by the MHC molecule used by the host for allopeptide presentation [I 1- 15]. We explored the possibility that responsiveness versus nonresponsiveness, as re- flected in the production of anti-donor HLA antibodies, may be influenced by the recipients' HLA-DRB1 genotype. The size of the population used in this analysis was increased to 161 heart and renal recipients who have been monitored for anti-HLA antibodies. In- dividuals showing anti-HLA antibodies in at least 1 of the 4 sera obtained during the first year after transplantation were considered responders, while patients forming no antibodies were considered nonresponders. Two- by-two contingency tables were constructed to determine the ~2 for association between a particular HLA allele (compared to the rest of

Discussion

A large proportion of organ allografts fail several month or years after transplantation because of chronic rejection, a progressive and irreversible host immunological attack on the graft. The common denominator of chronic rejection is the development of fibro- obliterative arterial lesions accompanied by immunoglobulin and complement deposition [5-8]. In both heart and kidney transplant recipients a correlation between the development of chronic rejection and anti-HLA antibodies has been observed in previous studies [9, 10] as well as in the present one. The mechanism by which anti-HLA antibodies may damage the graft is still unknown. It is possible that such antibodies attach to and damage vascular endothelial cells whose level of MHC class I and class II expression might have been upregulated by cytokines such as TNF and Y- interferon [31]. Exposure of endothelium to anti-MHC antibodies may result in endothelial cell dysfunction: cellular mechanisms supporting the function of the anticoagulant pro-

Reed/Ho/CohenlRamey/Marboe/ Anti-ldiotypic Antibodies in D'Agati/Rose/Hardy/Suciu-Foca Transplantation

tein and fibrinolytic pathways may be sup- pressed, while expression of the procoagulant cofactor, tissue factor may be enhanced, pro- moting deposition of fibrin on the vascular surface and within vascular lesions [32-37]. The production of lymphokines enhancing the level of allogeneic MHC expression on endothelial cells and the production of anti- HLA antibodies may be mediated by T helper cells. It was also shown that increased host immune responsiveness causes intracellular release of arachidonic acid, a precursor of thromboxane A2 which can enhance T-cell cytotoxic activity [5].

Recent data from our laboratory and oth- ers [11-15] have revealed the contribution to alloreactivity of T helper cells which recognize peptides derived from the processing of allogeneic MHC molecules by syngeneic APCs. In the setting of transplantation, it is conceivable that soluble HLA antigens are shed from the injured graft after acute rejection episodes. Such antigens may bind to HgA-specific B cells, which may present the processed allopeptides to T helper cells with corresponding T-cell receptors. Because allopeptide recognition is MHC-restricted, the af- finity of allopeptide binding to host MHC may ultimately dictate the ease and likelihood of alloimmunization. Such a hpyothesis is consistent with our present observation that individuals carrying certain DR2 alleles are at a lower risk of producing anti-donor HLA antibodies. HLA-DR2 may impose higher stringency conditions to the binding of allopeptides which may be less likely to compete and displace self-peptides bound to this molecule [13]. However, because of the relatively small size of the population studied, the association of HLA-DR2 alleles (DRBI*1501, * 1502, * 1601) with low responsiveness must be substantiated in further investigations.

Our studies on anti-idiotypic antibodies in recipients of heart and renal allografts con-

firmed previous investigations [9, 10, 24, 38- 42], showing that anti-HLA antibody producers who developed Ab2 have a significantly better prognosis than the corresponding Abl-positive/Ab2-negative group. There was no correlation between the HLA-DRB1 genotype and Ab2 production, suggesting that, if real, the Ir gene effect on Ab 1 production is not masked by anti-idiotypic autoimmunity.

The finding that the presence of anti-idiotypic antibodies improves significantly the prognosis of patients producing anti-HLA antibodies suggests that Ab2 suppresses the production of Abl, which in turn delays progression of chronic rejection. The immunoregulatory role of Ab2 in allograft recipients is consistent with previous studies on the development of Ab2 to anti-HLA antibodies during pregnancy [28, 43-45] and following blood transfusions [40, 46]. Patients with a history of anti-donor Abl, who exhibit Ab2 in their sera at the time of transplantation, were shown to tolerate the graft in spite of previous alloimmunization [24, 39-41]. Thus, anti- idiotypic autoimmunity may play a crucial role in determining the outcome of organ allografts.

Of particular interest is the finding that in both heart and kidney allograft recipients the specificity of Abl, and of the corresponding Ab2, is directed not only against MHC class I but also against MHC class II antigens. This observation reinforces the view that the production of anti-HLA antibodies is secondary to inflammatory events associated with acute rejection episodes, during which lymphokines upregulating the expression of MHC class II on endothelial cells are produced. The in- volvement of anti-HLA class II antibodies in chronic rejection also explains the significantly longer actuarial survival of heart and kidney allografts in recipients of HLA-DR- matched grafts, since such patients are un-

likely to develop anti-HLA-class-II immune responses [47, 48].

In conclusion, the data obtained in these studies show that the monitoring of anti-HLA antibodies and of anti-idiotypic antibodies following transplantation is of importance for assessing the immune status of transplant recipients and the prognosis of the graft. Heart

and kidney allograft recipients producing anti-donor HLA antibodies have relatively low long-term allograft survival rates. Hetero- geneity among the patients, with respect to Ir- gene-controUed alloreactivity and to the development of anti-idiotypic immunity, may exist, however, and should be tested in further studies.

~ 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 J . o ~ 1 7 6 ~ 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 ~ 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 ~ 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 o ~ 1 7 6 1 7 6 = . ~ 1 7 6 1 7 6 1 7 6 1 7 6 o o , ~ 1 7 6 1 7 6 1 7 6 o ~ ~ 1 7 6 1 7 6 ~ 1 7 6 1 7 6 ~ 1 7 6 ~ 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 ~ . ~ o ~ 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 1 7 6 ~ ~ J ~ 1 7 6 1 7 6 1 7 6 1 7 6

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39 Braun WE, Klingman L. Stewart RW, Ratliff N. Tubbs R. Zachary AA, Teresi GA, Rincon G, Protiva D: Two major serologic events in a successful cardiac transplant recipient-circumvention of hyperacute rejection despite a positive donor T lymphocyte crossmatch and late ap- pearance of probable anti-idiotypic antibody. Transplantation 1988;46: 153-155.

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43 Suciu-Foca N, Reed E, Rohowsky- Kochan C, Kung P, King DW: Anti- idiotypic antibodies to anti-HLA receptors induced by pregnancy. Proc Natl Acad Sci USA 1983;80:830- 834.

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48 Opelz G: Effect of HLA matching in heart transplantation. Transplant Proc 1989;21:794-796.

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Anti-idiotypic antibodies specific for HLA in heart and kidney allograft recipients