Antigens in human renal allografts

CLINICAL

IMMUNOLOGY

AND

Antigens L.C.

19, 206-223

IMMUNOPATHOLOGY

PAUL,'

in Human

(1981)

Renal Allografts

L. A. VAN Es, AND W.M.

BALDWIN

III2

Laboratory of Immunogenetics and Transplantation, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts 02115, and Department of Nephrology. University Hospital. 2333 AA Leiden, The Netherlands

Received October 9. 1980 While the anatomic location of renal autoantigens is well established from routine immunopathological studies of biopsy material, until recently the location of alloantigens on renal structures was not clearly defined. In this paper we review the literature concerning the distribution of allo- and autoantigens within the kidney and their possible role in the rejection of renal allografts. Most, if not all, alloantigenic systems for which there is considerable evidence to imply them as clinically important histocompatibility antigens assessed on the basis of matching, presence of serum antibodies in relation with rejection, or antibodies in eluates from rejected grafts, are localized on the renal endothelium. These include the ABO blood group determinants, HLA-A,B,C, and DR, LB antigens, and endothelial-monocyte antigens. Other alloantigenic systems such as Lewis, Rhesus, MN, J/i, and HT-A have not been demonstrated on the renal endothelium and may be less important in graft rejection. Antigens on the tubular basement membrane, collecting duct epithelium, proximal tubular brush borders, arterial smooth muscle cells, and nuclei appear to be unrelated to rejection.

INTRODUCTION Antigens in Human Renal Allografts

Rejection is the main cause of early failure of renal allografts. This is defined as an immune response stimulated by and directed against genetically determined tissue antigens, which are present in the donor kidney but absent in the recipient. (1). These antigens are by definition alloantigens which serve as histocompatibility antigens. The strength of the donor immune response against histocompatibility antigens is controlled by the genetically determined capacity to respond to these antigens as well as by several acquired factors, such as anemia, uremia, infections, and drug therapy, including immunosuppressive treatment. By matching recipients for the donor histocompatibility antigens, an improved graft prognosis should be expected. Renal failure can be the result of an autoimmune disease due to loss of tolerance for autoantigens of the kidney. When these autoantigens are also present in the transplanted kidney, recurrence of the original disease in the graft may be expected. Thus, both allo- and autoimmunity may potentially contribute to graft damage. Because a renal allograft provides a potent and complex set of antigenic stimuli to the recipient, antibodies may be produced that have little or no effect on graft survival. Therefore, in order to demonstrate that a given antigen (immunogen) ’ Supported by a Public Health Service lntemational 2 Supported by the Juvenile Diabetes Foundation. 206

0090-1229/81/050206-18$01.00/O Copyright All rights

@ 1981 by Academic Press, Inc. of reproduction in any form reserved.

Research Fellowship

(1 F05 TW 2629-02).

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20-l

ANTIGENS

stimulates a response that is critical to graft survival it is necessary to show not only that an immune response is stimulated, but also that the relevant antigen is available as a target for the response and that the mediators of the immune response (cells or antibodies) localize to the antigen. In clinical transplantation the cellular and humoral responses are generally studied using cells (lymphocytes, erythrocytes) which express the antigen(s) under study (2). This approach elucidates the extent of cellular and/or humoral immunity against antigens in the graft which are intrinsically present in renal tissue and/or present on passenger blood cells. On the other hand, immunopathological studies of transplanted kidneys provide information concerning the existence of cellular and/or humoral responses against antigens in the transplanted kidney itself. A drawback of this approach is that immunospecificity of the responses can, in general, not be established. Ideally, monitoring of the recipient’s cellular and humoral immune responses against the tissue of the transplanted kidney with subsequent analysis of the specificity should provide direct information concerning the importance of the various antigenic systems of the kidney with respect to allografting. In this paper we review the literature pertaining to allo- and autoantigens expressed in kidneys and their relevance as target determinants which lead to immunologic failure of renal grafts. The antigens which will be discussed are summarized in Table 1. The techniques used to detect the antigens will be discussed in the various sections. Several alloantigenic systems such as the H-Y system (3) and P red cell system (4) will not be discussed since these antigens have not been demonstrated in human kidneys. Renal Alloantigens I. ABO blood group antigens. The A and B blood group antigens are by immunofluorescence localized on the endothelium of several organs, including the kidneys (5, 6). The observation that the endothelial cell surface was clearly outlined while both the cytoplasma and the nucleus did not bind the blood group antibodies strongly suggests that the antigens are expressed on the plasma membrane surface (5). Furthermore, the antigens are localized on the walls of the cells of the basal and contiguous layers of the transitional epithelium of the urinary tract (5). Recently it was found that at least the A antigen is also present in distal convoluted TABLE ANTIGENS

Red cell antigens ABO Lewis Rhesus MN Iii Pr Gd

LOCATED

Leukocyte antigens HLA-AB (C) HLA-DR HLA-LB/MB Endothelial monocyte

I

IN HUMAN

KIDNEYS

Renal (auto) antigens GBM TBM RTE

Miscellaneous

antigens

HT-A Henle’s loops antigens Epithelial antigens Nuclear antigens Smooth muscle antigens

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tubules. Considerable variations in intensity of the staining of these structures are found in the same kidney. In individuals who are secretors of blood group substances, the antigens are, in addition to the above-mentioned sites, also present in a water-soluble form in and on the collecting ducts (5, 6). The early observations in clinical transplantation that major ABO incompatibility between donor and recipient usually results in early vascular rejection (7, 8) suggest that these antigens are important histocompatibility antigens. In one such case we demonstrated after an AB kidney was transplanted into an A individual that the anti-B antibodies decreased in the circulation. The graft was rejected shortly after transplantation and large amounts of IgM antibodies were deposited on the graft endothelium. When eluted, these antibodies agglutinated B erythrocytes (9). These data provide sufficient evidence to conclude that antibodies to ABO blood group antigens can cause renal allograft rejection. 2. Lewis antigens. Several recent retrospective studies have suggested that Lewis blood group determinants may constitute a histocompatibility system in clinical renal transplantation (10, 11). Graft survival in Lewis-negative recipients, who have on the basis of the population distribution of Lewis-positive individuals about a 90% chance of receiving a Lewis-positive kidney, is lo- 15% worse than in Lewis-positive recipients (11). Oriol ef al. (12) provided data that Lewis antigens are synthesized in the kidney. Using an indirect immunofluorescence technique, the antigens were localized in the cytoplasma of epithelial cells of distal convoluted and collecting tubules but not in endothelial cells. The mechanism(s) accounting for the apparent increase in rejection rate in Lewis-incompatible combinations has not yet been elucidated. 3. Rhesus (Rh) antigens. Boorman and Dodd concluded on the basis of hemagglutination -inhibition experiments with kidney homogenates that the Rh antigens are present in kidneys (13). The degree of inhibition of the anti-Rh by kidney homogenates was found to be comparable with the inhibition of anti-A and anti-B blood group antisera, which may suggest that the density of Rh antigens in kidneys equals that of the A and B antigens (13). In interpreting these data, however, it should be borne in mind that tissue homogenates contain variable amounts of antigens derived from circulating blood cells, including erythrocytes. In view of the persisting interest in Rh antigens as histocompatibility antigens in renal allografting, confirmation of the presence of Rh antigens in kidneys is certainly warranted. The significance of Rh antigens in transplantation was suggested by the observations that graft survival was poorer in Rh-negative recipients compared to Rhpositive recipients (14, 15); other groups, however, found no differences in graft survival (4, 16). Huestis and Zukoski (17) reported that the production of Rh antibodies following the transplantation of Rh-positive kidneys into Rh-negative recipients was associated with a greater than 6-year graft survival in two of three patients. The third allograft was rejected in 2 weeks, but a second Rh-positive kidney survived more than a year. Thus, the production of Rh antibodies is not associated with allograft rejection. 4. MN erythrocyte antigens. MN antigens, which are products of a diailelic gene at one locus, were found in a low concentration in human kidney homoge-

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nates using a hemagglutination-inhibition technique (13). Van Hooff (4) concluded that the MN antigens are not important transplantation antigens, since the l-year graft survival for MN heterozygous recipients, who would always receive MN-compatible grafts did not differ from that for M- or N-homozygous recipients who would have a 75% chance of receiving a MN-incompatible kidney. No data have been reported concerning the development of immunity against MN antigens in renal allograft recipients. 5. Erythrocyte antigens reacting with cold agglutinins. Cold agglutinins, which are usually autoantibodies, may bind in vitro to sections of human kidneys at low temperatures (18). Depending upon the specificity they bind to the epithelium of Henle’s loops and the distal tubules (Ui antigens) (18), to the endothelium of glomerular and peritubular vessels (Pr antigens) (18), or to the glomerular, peritubular and interstitial vessels as well as the brush borders of the tubular epithelium (Gd antigens) (18). Graft failure in the presence of autoagglutinins has been reported and seems to be the result of autohemagglutination within the graft immediately after release of the vascular clamps when the graft temperature is still below body temperature (19, 20). The available data do not indicate that these antibodies induce damage through binding to the antigens intrinsic to the graft itself. 6. Human leukocyte antigens (LHA). Since the original observations of Dausset (21) and Van Rood (22), the work of many investigators led to the recognition of the HLA system as a genetic region on a part of the short arm of the sixth chromosome consisting of several separate loci with a series of multiple alleles at each of these loci (reviewed in Ref. (23)). The serologically defined HLA antigens (the “Classical” HLA antigens) are coded for by a highly polymorphic system of three closely linked loci: HLA-A, B, and C. These antigens have a widespread tissue distribution. In an attempt to devise a direct in vitro measure of histocompatibility Bach and Hirschhorn cultured lymphocytes from different individuals together and related the degree of blast transformation in this mixed lymphocyte culture to histocompatibility (24). Genetic analysis of the mixed lymphocyte reactions (MLR) established that the “MLR-antigens” are coded for by a separate HLA locus, called HLA-D (25-27). After the definition of MLC antigens, it was attempted to define these antigens serologically (28). Anti-B-cell antibodies have been identified which are directed against antigens controlled by a locus or loci which are either identical or very closely linked to HLA-D. These antigens are designated HLA-DR antigens (29, 30). Recently several public B-cell alloantigenic systems have been discerned which are closely associated with HLA-DR. At present the molecular relations among these systems defined primarily by patterns of serologic testing, are not clear (31, 33). In addition to HLA-D associated B-cell alloantigens (HLA-DR), there is evidence for the existence of B-cell alloantigens linked to the HLA-A locus (34) as well as B-cell alloantigens which segregate independently of HLA (35). HLA-ABC Antigens Several approaches have been used to solve the question as to whether the classical HLA antigens are expressed in kidneys. Early absorption experiments

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by Van Rood et al. (36) indicated their presence in kidney homogenates. Absorptions with isolated disrupted glomeruli demonstrated the antigens in the cellular fractions of the glomeruli but not in the basement membrane (37). Quantitative absorption experiments using kidney homogenates and a monoclonal antibody against a nonpolymorphic xenodeterminant of HLA-ABC demonstrated that the kidney carries approximately 15% of the amount of HLA-ABC found in spleen (38). Hayry et al. applied the staphylococcus rosette assay on single-cell suspensions prepared by enzymatic digestion of kidneys. Using a xenoantiserum against HLA-ABC common determinants positive reactions were observed with presumed passenger lymphocytes, endothelial cells, and tubular cells, whereas with alloantisera positive reactions were obtained with only passenger cells and endothelial cells (39). Immunofluorescence studies of kidney sections with monoclonal antibodies against nonpolymorphic xenodeterminants show staining of the glomerular and arterial endothelium; the peritubular capillary endothelium stains only moderately (38, 40). (Fig. la). However, monoclonal antibodies against HLA-A and B allospecificities or typing alloantisera stain renal vascular endothelium only weakly or undetectably (40).

FIG. 1. Photomicrograph of kidney sections incubated with (a) a mouse monoclonal antibody to common antigenic determinants of HLA-AB molecules, (b) an alloantiserum to HLA-DR-1, (c) an alloantiserum to HLA-LB-13, and (d) a postrejection alloserum containing endothelial-monocyte antibodies. The sections were stained with fluorescein-labeled antisera. The anti-HLA-AB xenoantibody gives intense staining of the glomeruli and weak staining of the peritubular capillary endothelium (a). Alloantisera to HLA-DR stain ptc endothelium weakly (b). whereas alloantisera to HLA-LB give a moderate staining (c). Endothelial-monocyte antisera give an intense staining of the ptc endothehum (d).

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The role of the HLA-AB antigens as transplantation antigens was suggested by the excellent survival of kidneys transplanted between HLA-identical siblings (41, 42). However, the survival of kidneys transplanted from unrelated donors which are well matched for the HLA-A and -B antigens (i.e., having three or four antigens in common with the recipient) is only lo-30% better than the results obtained with poorly matched kidneys (43), depending upon factors such as presensitization (44) and pretransplant blood transfusions (45, 46). Matching for HLA-B antigens seems to give a better graft prognosis than matching for HLA-A antigens (44, 46), whereas matching for HLA-C appears to be unimportant (47). Furthermore, even in the face of a total HLA mismatch, 30% of renal allografts survive more than 5 years. Since the HLA system displays the phenomenon of nonrandom association of alleles of linked loci, (linkage disequilibrium) it is possible that the improved results are not necessarily due to identity of the HLA-AB antigens themselves but due to compatibility for other antigens. Support for this view comes from studies in which complete family typings for both cadaver donor and recipient combinations allowed genetic analysis of haplotype inheritance (48). In such cases where there is matching for one A locus and one B locus antigen, and they are on the same chromosome, cadaveric graft survival is better than in cases where the A and B compatibilities were inherited as different haplotypes. The role of humoral immunity against HLA-ABC antigens in renal allograft rejection has been suggested by the observation that donor-specific pretransplant lymphocytotoxic antibodies are associated with hyperacute rejection in 80% of the cases (49). No conclusive evidence, however, has been presented that the antibodies were directed against HLA-ABC antigens. Morris et al. (50) showed that donor specific anti-HLA antibodies may be found after graft nephrectomy. No lymphocytotoxic antibodies, however, were found in patients whose grafts were removed in the period shortly after transplantation, when most rejections occur. In recent years in which post-transplant immunologic monitoring of recipients found widespread application, it became evident that donor-specific lymphocytotoxic antibodies may be found in association with rejection (2). Relatively few studies have addressed the question of whether HLA-ABC antibodies are bound to rejected grafts. Lucas et al. (51) demonstrated anti-HLAABC antibodies in 10 of 11 eluates from rejected grafts. In contrast, other groups found anti-HLA-ABC antibodies in only lo-30% of eluates (52-55). Thus, humoral immunity against the classical HLA antigens can be a factor in renal allograft rejection. In vitro cell-mediated cytotoxicity may be directed against HLA-ABC antigens themselves or antigens linked very closely to them (56, 57). Whether HLA-ABC antigens are the target of cell-mediated damage of renal allografts is not known. However, cells isolated from rejected grafts are cytotoxic in vitro for target cells that have donor-specific HLA antigens (58, 59). Moreover, the cytotoxicity of these cells exceed that of circulating cytotoxic cells, which suggests that an accumulation of donor-antigen specific cells occurs in the graft (59). Bendixen et al. (60), however, failed to demonstrate that the specificity of the peripheral cellular immune response is directed against the HLA-AB antigens themselves.

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HLA-D and HLA-DR Antigens The HLA-D antigens have not yet been demonstrated

in human kidneys, although suspensions of rat or pig kidney cells stimulate allogeneic lymphocytes in mixed-kidney cell-lymphocyte reactions (58-60). The importance of cellular defined antigens in human transplantation is supported by reports that graft survival is better for donor-recipient combinations with a negative or weak response in mixed-lymphocyte reactions than for those combinations with marked reactions (64-67). Some centers have, however, not been able to confirm these findings (68-70). In the past few years several investigators have demonstrated the expression of HLA-DR or, in analogy to the mouse nomenclature, Ia antigens in human kidneys. After the initial demonstration of Ia antigens in detergent-solubilized cell membranes derived from kidneys, Koyama et al. established that the isolated antigens are indistinguishable from Ia antigens derived from cultured B-lymphoid cells as far as molecular as well as xeno- and alloantigenic properties are concerned (71, 72). In a radioimmunoassay these authors showed that kidneys have amounts of Ia antigens comparable to spleen and lymph node. Similar results were obtained by Williams et al. (38), who demonstrated in quantitative absorption experiments with a monoclonal antibody against a nonpolymorphic xeno-Ia determinant that kidneys contain on average about 90% as much Ia as spleen. Cytological localization of the antigens using the staphylococcus rosette assay and cell suspensions prepared by enzyme treatment of kidneys showed that the xenoantigenic determinants of Ia antigens are located on the surface of endothelial cells and a population of passenger lymphocytes (39). Criteria necessary for the identification of endothelial cells in these cell smears were not reported. It is noteworthy that the allospecific determinants of the Ia antigens could not be demonstrated using this technique (39). Furthermore, it was noted that considerable differences were observed in the amount of antibodies bound by the endothelial cells of various kidneys. Several laboratories have localized the Ia antigens using an indirect immunofluorescence technique. Koyama et al. (72) used a xenoantiserum and observed prominent staining in the mesangioendothelial cells of the glomeruli and in the peritubular areas, apparently of the capillary endothelium. Williams et al. (38) failed to localize the antigens in sections of normal perfused kidneys using a monoclonal antibody against a nonpolymorphic Ia determinant. We recently reported that monoclonal antibodies against nonpolymorphic determinants of Ia glycoproteins stain the glomerular and arterial endothelium strongly but peritubular capillary endothelium only moderately (40). In a panel analysis of typing alloantisera to antigens of the DR locus, weak staining was observed (Fig. lb) with a high frequency of false-negative reactions. In contrast, 11 of 18 alloantisera directed against LB antigens (HLA-DR linked B-cell antigens identical or very similar to MB antigens) stained the peritubular endothelium moderately strongly (40) (Fig. lc). There is an apparent discrepancy between the detection of xeno- and alloantigenic determinants of both HLA-ABC and HLA-DR antigens. This may be due

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to differences in concentrations of xeno- and allodeterminants, differences in affinity between xeno- and alloimmune sera, or to cross-reactions of xenoimmune sera with other antigens than HLA which are expressed in high concentrations on renal endothelium. Xenoimmune sera to highly purified mouse H-2 glycoprotein have been proposed to cross-react with other structurally similar antigens that are linked to the MHC such as TL and other, yet undefined, antigens (73). In spite of technical difficulties in typing for these recently defined antigens, early clinical data have shown improved graft survival results for kidneys well matched for the HLA-DR antigens (74-76). In a recent retrospective study of patients who received one HLA haplotype graft from a family member, Duquesnoy et al. (77) showed that matching for the HLA-DR linked MB system appears to be more important than matching for HLA-A, B, or DR, a finding which awaits confirmation. The clinical significance of pretransplant antibodies against donor B-cell antigens is a subject of controversy. Some centers have reported an improved graft prognosis (78) in the presence of these antibodies, whereas others have found accelerated graft loss (79, 80) or no influence at all (81, 82). Pooled data from prospective and retrospective studies in several centers, however, indicate a low failure rate for cadaveric grafts with a positive B-cell but negative T-cell crossmatch (83). Soulillou (84) and others (54) have shown that the early development of donorspecific B-cell antibodies after transplantation correlates with graft failure. In addition, anti-donor B-cell antibodies have been detected in eluates from rejected grafts (54, 55). Ting and Morris (75), however, failed to confirm the association between anti-B-cell serum antibodies after transplantation and graft outcome. Thus, the clinical significance of anti-B-cell antibodies, both before or after transplantation, is not yet clear. This may be due to several causes, such as testing of unabsorbed sera, the presence of several alloantigenic systems on B lymphocytes, or contamination of the B-cell preparations with monocytes. The presence of B-cell antibodies in eluates from rejected grafts strongly suggest, however, that at least some B-cell alloantibodies played a role in the rejection process. 7. Endothelial-monocyte antigens. When we studied serial serum samples of transplant recipients in an indirect immunofluorescence technique using a pretransplant biopsy of their graft as a substrate, ailoantibodies were found which gave an intense staining of the endothelium of the peritubular capillaries and venules (85) (Fig. Id). Unlike the antisera to HLA determinants, these antibodies stain glomeruli less intensely than the peritubular capillaries, and stain arteries very weakly or not at all (40, 86, 87). These antigens are localized on the plasma membrane of the endothelial cells (87). Absorption experiments demonstrated that the endothelial antigens are also present on monocytes but not on platelets and lymphocytes, and thus are not HLA-A, B, C, or DR antigens (88, 89). This was contirmed in a panel analysis of endothehal-monocyte antibodies eluted from rejected grafts (89). This antigenic system probably is the same as that described by Moraes and Stastny on umbilical cord vein endothelium and monocytes (90-92). Preliminary data from Stastny (93) suggest that most endothelialmonocyte antigens segregate with HLA antigens, whereas others do not. Al-

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though there is increasing evidence that endothelial-monocyte antigens are independent from the HLA-A, B, C, DR antigens, much work remains to be done concerning the tissue distribution, the extent of polymorphism, the chemical composition, and the genetic inheritance. The relation of the endothelial-monocyte antigens with monocyte -granulocyte antigens (94) as well as with monocyteassociated antigens (95) remains to be clarified. The role of antibodies against the endothelial-monocyte antigens in the process of graft rejection is suggested by the observation that pretransplant donor-specific antibodies in the absence of lymphocytotoxic antibodies is associated with accelerated acute graft rejection (89) and the appearance of circulating antibodies in close association with irreversible vascular rejection in about 50% of the patients (85). Moraes and Stastny found that 95% of patients who had rejected a renal allograft have antibodies against a panel of umbilical vein endothelial cells; about half of these patients also reacted with a lymphocyte panel (96). It is possible that in this group positive reactions with umbilical endothelial cells were attributable to anti-HLA antibodies, since HLA antigens are expressed on umbilical endothelium (97). The patients without lymphocytoxic antibodies (53%) might have had antibodies against the endothelial-monocyte antigens. Rohr et al. found that 36% of patients with graft loss because of rejection had antibodies reactive with a panel of isolated umbilical vein endothelial cells (98). Three sera were absorbed with platelets; two of these sera were no longer cytotoxic to umbilical cord vein lymphocytes, and one serum remained cytotoxic; the antiendothelial cell activity was, however, retained in all sera. Cerilli et al. found that four out of seven patients who had pretransplant antibodies against donor intercostal arteries rejected their kidney shortly after transplantation, compared with a graft loss of 17% for those patients who did not have these antibodies (99). Furthermore, 88% of the patients who had rejected their grafts had antiendothelial antibodies detected with a panel of umbilical vein endothelial cells and/or sections of arteries; in 25% of the cases lymphocytotoxic antibodies were detected. The relationship of this antigenic system (99, 100) with the endothelial-monocyte antigens is still unclear. The demonstration that endothelial-monocyte antibodies can be eluted from rejected grafts (55, 85, 88, 89) indicates that these antibodies participated in the process of graft rejection. Preliminary data indicate that the endothelial-monocyte antibodies are formed especially in recipients which are HLA-DRw6 positive (lOl), which may indicate that the formation of these antibodies is under Ir gene control. No data with respect to matching for endothelial-monocyte antigens are yet available. 8. Heterophile transplantation antigens (HT-A). A heterophile antigen system, designated HT-A, has been demonstrated on red blood cells of outbred Wistar rats, in Enterobacteriaceae and in human tissues, including kidneys (102, 103). The antigen was demonstrated in human tissues in a hemagglutination-inhibition assay with rat erythrocytes after absorption of the test sera with ethanol extracts of the tissues (103). In rat kidneys, the antigen was localized by fluorescence in the stromal tissues and basement membranes of proximal and distal tubules and colletting ducts (104), whereas the localization in human kidneys is unknown (102, 103). Anti-HT-A antibodies, however, do not bind to human endothelial cells from

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umbilical cord veins (98). The allospecilicity in man is presumably determined by a single locus comprised of one dominant and one recessive allele (105). McDonald ef al. (106) concluded that HT-A compatibility is a major determinant of renal allograft success, since l-year graft survival for HT-A-compatible grafts was 84 compared with 44% for HT-A-incompatible grafts. The role of humoral immunity against HT-A antigens was suggested by the observation that five out of eight transplantations performed in the presence of anti-HT-A antibodies ended in rejection, starting within 1 week of transplantation (107). The rejection was almost always associated with a significant rise in the anti-HT-A antibody titer (107). When Parker (108) reviewed the role of antibodies in allograft rejection, he stated that patients who produce heterophile antibodies are responders and therefore likely to attempt to reject their allograft. He concluded that heterophile antibodies do not seem to play any direct role in rejection since they have not been identified in eluates from allografts. For completeness of this review, a summary will be given of renal antigens which are known to play a role in autoimmune diseases of the kidney. The relevance of these antigens in clinical renal transplantation is discussed. Rend Autoantigens 9. Glomerular basement membrane (GBM) antigens. Autoreactive antibodies (109) and lymphocytes (110) against antigenic determinants of the GBM can be found in patients with glomerulonephritis. The role of anti-GBM antibodies in the induction of anti-GBM nephritis has been well established (109), whereas the role of cell-mediated immunity in the pathogenesis of the disease is still not clear. Lemer et al. (109), Dixon et al. (11 l), and Wilson and Dixon (112), have pointed out the risk of recurrence of nephritis in the graft when circulating anti-GBM antibodies against graft GBM exist at the time of transplantation. On the other hand, Porter (108) reported that examples of Goodpasture’s syndrome and proliferative glomerulonephritis with circulating anti-GBM antibodies have been recorded in which the renal transplant escaped unscanthed despite initial fixation of circulating antibodies to the glomerular capillary basement membranes of the graft. Occasionally anti-GBM antibodies develop after transplantation and cause proliferative glomerulonephritis in recipients whose original disease was probably not associated with anti-GBM antibodies (108). The antibodies can be eluted from the graft (108). It is not known whether this is due to loss of tolerance for auto-GBM antigens or whether this is attributable to possible allospecificity of GBM determinants. 10. Tubular basement membrane (TBM) antigens. Circulating anti-TBM antibodies together with linear deposits of IgG along the TBM and tubulointerstitiai changes have been reported in human renal allografts (113 - 116). Such antibodies might be expected to contribute to graft damage, since there is evidence derived from experimental models as well as human disease that anti-TBM antibodies may cause tubulointerstitial lesions (117). Recently we found that anti-TBM antibodies against the graft TBM can apparently de novo occur without tubuloinstitial

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changes in the graft (118). Similar observations have been made in a rat renal allograft model (119). Both in man and in the rat there are indications that the TBM may exhibit some degree of allospecificity (118, 119). 11. Renal tubular epithelial antigen. After repeated grafting of allogeneic kidneys, Klassen and Milgrom (120, 121) demonstrated in a rat model circulating antibodies against the brush border of proximal tubular epithelial cells together with granular deposits of immunoglobulins along the glomerular capillaries. This resembles the Heymann nephritis, which is a model of membranous glomerulonephritis produced by immunization of rats with an autoantigen (RTE) present in the brush border region of proximal renal tubules (122, 123). Such antibodies may either bind to circulating RTE and deposit in the glomeruli (123) or they may bind locally to RTE antigen already trapped in the split pores of the GBM (124, 125). Since RTE antigen together with immunoglobuhns and C3 have been found in the glomeruli of an occasional patient with glomerulonephritis (126-130), it has been suggested that similar mechanisms may cause glomerulonephritis in man, although most patients with membranous glomerulonephritis do not have RTE antigens deposited in their glomeruli (131). In only a few human renal allograft recipients have antibodies against the brush border of the proximal tubules of the graft been described following transplantation (1, 132, 133). The available data do not allow any conclusions concerning the occurrence of glomerulonephritis in the graft. In only one patient we demonstrated that glomerulonephritis did not occur in the presence of anti-RTE antibodies (133). In addition to the antigens present on the above-mentioned renal structures, autoantibodies against the plasma membrane of the renal collecting ducts, the renal pelvis and the ureter have been found in the sera of renal allograft recipients (134). These antibodies also bind to epithelial cells of various other organs. There is no evidence that these antibodies are harmful to the graft, possibly because the antigen is relatively unaccessible to the circulating antibodies (134). Antibodies which react with intracellular antigens of Henle’s loops have been found in sera from patients with various disorders of presumably immunological pathogenesis (135, 136). It seems unlikely that these antibodies are harmful since they cannot bind to their corresponding antigens in vivo. Histologic studies of human renal allografts have suggested that smooth muscle cells of the vasculature may be the primary target for the rejection process (137). Smooth muscle antibodies have, however, to our knowledge not been described in sera of transplant recipients in association with rejection nor in eluates of rejected kidneys. Finally, the antinuclear antibodies in sera of patients with or without systemic lupus erythematosus can be demonstrated using sections of normal human kidneys (138). There is, however, no evidence that these antibodies induce graft damage mediated through direct binding to intracellular nuclear antigens of the graft. Because of the myriad of antigens which can participate in the formation of circulating immune complexes which may induce graft damage, these antigens were not included within the scope of this review.

LOCALIZATION

2 ora 2 ora k or a -C or a a a

+ to ++ a a

Arteries

IN HUMAN

TABLE

P t ora 5 or a k + a a

P

++ a a

++ a a P P 2 ora f or a + ++ a a

Peritub. caoillaries

Endothelium

BY IMMUNOFLUORESCENCE

Glom. capillaries

KIDNEYS

2

k ora t ora + ++ a a

++ a a

Veins

OF ANTZENS

PROVEN

OR POSSIBLE

a a a a a a a a a P P*

Glomerular

P

PG

a a a a a a a a a

Tubular

Basement membranes

WITH

a a a a a a a a a a a

Glomerular

Epithelium

ALLOSPECIFICITY”

Pb PC P” a P’ a a a a a a

Tubular

’ The following notation is used: a = absence of a given antigen on a specific structure and p = the presence of the antigen. No symbol indicates that it is unclear from the cited reference(s) whether or not the antigen is present on the corresponding renal structures. In those instances in which differences in intensity of the staining of different structures were observed, the following notation is used: (++) intense staining, (+) moderate staining, (2) weak staining. ’ Present in distal convoluted tubules and in addition in secretors in the collecting tubules. C present in distal convoluted and collecting tubules. ” In the cytoplasma and on the surface of the epithelium of Her-de’s loops and distal tubules. ” Present in the brush border of proximal tubules. r Intensity relates to allospecific determinants. ’ Some antigenic determinants are shared by GBM and TBM, others are restricted to GBM or TBM.

ABO (5,6) Lewis (12) Ui (18) Pr (18) Gd (18) HLA-AB’ (39,40) HLA-DR’ (40) HLA-MB/LB (40) Endothelial monocyte (40.86) GBM (109) TBM (117,118)

ANATOMIC

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BALDWIN

COMMENTS

From the data presented in this article, it is clear that a multitude of antibodies and possibly also cytotoxic cells directed against antigens present in kidneys can be found in human renal allograft recipients. Table 2 summarizes the alloantigens of which the anatomic location in kidneys has been more or less clearly defined. Further application of monoclonal antibodies will be of value in the further development of the emerging “antigenic anatomy” of renal allografts. An assessment of the clinical importance of the various polymorphic alloantigenic systems is summarized in Table 3. This assessment was made on the basis of data obtained by matching the recipients for the various alloantigens under discussion, the presence of serum antibodies in association with rejection, and the presence of antibodies in eluates from rejected grafts. As shown, most, if not all systems for which there is considerable evidence to imply them as clinically important histocompatibility antigens (ABO, HLA-A. B, C, DR, endothelial monocyte) are expressed on the endothelium of the renal vasculature. Further support for the dominant role of alloantigens on the endothelium comes from histological studies of rejecting renal allografts (137, 139, 140) in which there is evidence that the immune response localizes in and around blood vessels. In view of the concept that antibodies can cause graft damage, provided that TABLE IMPORTANCE ALLOGRAFTING REJECTION

3

OF VARIOUS POLYMORPHIC ALLOANTIGENIC DETERMINED BY MATCHING, ASSOCIA.~ION AND

PRESENCE

OF ANTIBODIES

IN ELUATES

ABO

Beneficial

Lewis” Rhesus” MN” HLA-AB

Beneficial ( 10.11) Controversial (4,144 No Importance (4) Beneficial (43)

HLA-DR

Beneficial

(74,761

Unknown Beneficial

(106)

Endothelial HT-A”

monocyte

(7.8)

FROM

REJECTED

Serum antibodies

Matching

reviews unknown.

WITH

GRAFTS”

Eluted antibodies

Naturally occurring with increase in titers (8) Unknown 16) No correlation (171 Unknown Present (501, often polyspecific (21 Pretrdnsplant:” not harmful”” Post-transplant:’ controversial (54,75,84) Present (85.88) Naturally occurring with increase in titers (107)

ri References are made to either original observations, D Compatibility determined by indirect means. r Defined as B-cell alloantibodies, serologic specificity

RENAL.

SYSTEMS IN CADAVERIC OF SERUM ANTIBODIES

or workshop

Present

(9)

Unknown Unknown Unknown Present (51-55)

Present”

(54.55)

Present Absent

(55.85.88) (108)

reports.

RENAL

TRANSPLANTATION

ANTIGENS

219

they bind to their corresponding antigens in the graft and activate immunemediator mechanisms, the overall good survival of transplants in the presence of donor-specific B-cell alloantibodies has to be explained. As mentioned previously, the serologic specificity of the B-cell antibodies is generally poorly defined or unknown and not all B-cell alloantigens are necessarily expressed on the renal endothelium. In the rat renal allograft model, for example, antibodies against immunoglobulin allotypes can be cytotoxic for B cells. These antibodies do not bind to the renal endothelium and the graft prognosis is not influenced (141). In clinical transplantation it is, however, very likely that in a number of cases the B-cell alloantibody is directed against HLA-DR determinants. Why do these grafts escape from an early antibody-mediated vasculitis and possibly enjoy even a state of enhanced survival? The observations that in some kidneys DR antigens can be demonstrated with typing alloantisera but not in other kidneys of the same HLA-DR type, may suggest quantitative differences in the expression of these antigens in different individuals (40). The same holds true for the HLA-A and B antigens (39, 40). Quantitative and qualitative data concerning the in viva assessable alloantigenic determinants are obviously warranted to explain the abovementioned phenomenon. The excellent graft survival in recipients of an HLA-identical kidney from a family member or of an HLA-DR-identical cadaver kidney needs a final comment. Some of the genes in the HLA-complex code for cell-surface molecules (antigens) and some code for functional characteristics such as immune responsiveness. The fate of the graft may thus be influenced either by incompatibility for the antigens themselves or by the possibility that immune responsiveness against other transplantation antigens (non-HLA antigens?) is influenced by incompatibility at the HLA-DR locus. Thus, an immune response against several transplantation antigens requires not only a mismatch for these antigens but also a mismatch for immune response genes of the HLA-D related gene(s). Continuing research in the area of renal transplantation antigens is essential, since optimal management of the transplant recipient and improved graft prognosis is only possible when the targets of the rejection process have been fully identified. REFERENCES 1. Milgrom, F., Klassen, J., and Fuji, H., J. Exp. Med. 134S, 193, 1971. 2. Stiller, C. R., Dossetor, J. B., Sinclair, N. R. StC., and Rapaport, F. T., “Immunologic Monitoring of the Transplant Patient.” Grune & Stratton, New York, 1978. 3. Goulmy, E., Bradley, B. A., Lansbergen, Q., and van Rood, J. J.. Transplantution 25,315, 1978. 4. Van Hooff, J. P.. Thesis, University of Leiden, 1976. 5. Szulman, A. E., J. Exp. Med. 111, 785, 1960. 6. BariCty, J., Oriol. R., Hinglais, N., Zanetti, M., Bretton. R., Dalix, A.-M., and Mandet, C.. Kidney In?. 17. 820, 1980. 7. Gleason, R. E., and Murray, J. E., Transplantution 5, 343, 1967. 8. Wilbrandt. R., Tung, K. S. H., Deodhar, S. D., Nakamoto, S., and Kolff, W. J., Amer. J. C/in. Puthol. 51, 15, 1969. 9. Paul, L. C., van ES, L. A., Brute1 de la Rivikre, G., Eemissa, G., and de Graeff, J., Transphtutinn 26, 268, 1978. IO. Griol, R., Cartron, J., Yvart, J., Bedrossian, J., Duboust, A., Baritty. J.. Gluckman, J. C., and Gagnadoux, M. F.. Lancer 1, 574. 1978.

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VAN ES, AND BALDWIN

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