Induction of MHC class II expression in recipient tissues caused by allograft rejection

MHC class II antigens (DR) are not commonly expressed on parenchymal cells of kidney and liver except when they are allografts undergoing rejection. The objective of this study was to determine whether allograft rejection can also induce DR upregulation in parenchymal cells of autologous recipient organs. Dogs had unilateral renal autografts to facilitate kidney sampling. All kidneys were tubular cell DR-negative. After 8-14 days each dog received a tubular cell DR-negative allograft. Tubular cell DR became positive in both allograft and autograft simultaneously, its onset and intensity correlating with blast cell infiltration and rejection in the allograft. Blast cells were first detected in the autograft after allograft nephrectomy, and then disappeared as autograft tubular cell DR diminished over the next 6-8 days. This was reproduced on repeat allografting. In 2 untreated dogs hepatocytes became positive on day 4, with no hepatic blast infiltrate. Four other dogs received cyclosporine immunosuppression. Allograft and autograft tubular cell DR, and hepatocyte DR, increased in all dogs, but were delayed while on CsA until onset of rejection despite transient earlier allograft blast infiltration. Downregulation in autograft and liver occurred together after allograft nephrectomy. An interferon-like substance appeared in plasma after allografting in association with the DR changes in native kidney and liver. Renal allorejection therefore induces upregulation of parenchymal DR expression in autologous liver and kidney of the recipient. It is probably mediated by an interferon-like substance derived from cells infiltrating the allograft. The effect is modified by CsA.     

Donor pretreatment with FK506 reduces reperfusion injury and accelerates intestinal graft recovery in rats

BACKGROUND: FK506 alleviates warm ischemia-reperfusion injury, but it remains unknown if such protection is manifest after cold storage and transplantation. We studied the early outcome after transplantation of intestines from donors pretreated with FK506 compared to grafts from controls treated with saline (154 mM NaCl). METHODS: Sprague-Dawley rats received 0.3 mg/kg FK506 or saline intravenously 6 hours before graft retrieval. The small bowel was harvested, stored for 3 hours, and then transplanted heterotopically. Samples were taken after preservation and at 20 minutes, 6 hours, 12 hours, and 24 hours after reperfusion. Heat shock protein 72 (Hsp72) and iintercellular adhesion molecule (ICAM)-1 expression and nuclear factor kappaB (NF-kappaB) activation were assessed via Western blots and eelectrophoretic mobility shift assay (EMSA), respectively. Dissacharidase activity and enterocyte proliferation rate were also studied. RESULTS: Preservation injury was similar between groups, but pretreated grafts had better morphology already 20 minutes after reperfusion. Control grafts always had thinner mucosa and more PMN infiltration. Hsp72 expression was greater in pretreated grafts. ICAM-1 was absent after harvesting, preservation, and immediately after reperfusion but increased in control grafts at the later time points. Control grafts showed a biphasic NF-kappaB activation pattern, whereas NF-kappaB activation was inhibited effectively in pretreated grafts. Dissacharidase activity decreased during the first 6 hours after reperfusion but recovered within 24 hours in pretreated grafts but not in control grafts. Earlier enterocyte proliferation was observed in pretreated grafts. CONCLUSIONS: FK506 donor pretreatment reduced graft proinflammatory activation and neutrophil inflammation. Pretreated groups revealed a milder reperfusion injury and accelerated morphologic and functional recovery. The mechanisms involved appear to involve Hsp72 upregulation and NF-kappaB inhibition.     

Effects of fish oil on graft arteriosclerosis and MHC class II antigen expression in rat heterotopic cardiac allografts.

The effect of fish oil on accelerated graft coronary arteriosclerosis was assessed in Lewis to Brown-Norway rat heterotopic cardiac allografts. Twelve Brown-Norway rats were supplemented with 2 ml/kg/day of fish oil (68.3 mg eicosopentaenoic acid and 47.5 mg decosahexaenoic acid per milliliter). Eleven additional animals, receiving an isocaloric amount of safflower oil, served as control. All diets began 1 week before operation. Immunosuppression was obtained with low-dose cyclosporine (2 mg/kg/d). When killed (100 days), there were no significant differences in percentage weight gain, graft function, or histologic rejection score. Although lipid profiles were comparable, total cholesterol:high-density lipoprotein ratio was marginally higher in animals treated with fish oil (p = 0.069). Mean percentage luminal occlusion (before and after correcting for differences in size between coronary vessels analyzed) and average intimal thickness were similar between animals treated with fish oil and safflower oil as assessed by computer-assisted digitized, morphometric planimetry. In all allografts, donor interstitial dendritic cells were repopulated with recipient dendritic cells. The major histocompatibility complex class II cell density in the fish oil group did not differ significantly from rats supplemented with safflower oil (1.48 +/- 0.68 vs 1.48 +/- 0.65 cells per mm2, p = 0.995). In conclusion, fish oil did not exert any beneficial effect over safflower oil in terms of graft coronary arteriosclerosis, histologic rejection, or plasma lipids.(ABSTRACT TRUNCATED AT 250 WORDS)     

The study of peripheral blood mononuclear cell MHC I and MHC II gene mRNA expression in acute graft rejection

BACKGROUND: Early diagnosis of acute graft rejection is important in the clinic. To explore a reliable diagnostic marker, we selected skin-grafted rabbits as an animal model to study peripheral blood mononuclear cell (PBMC) major histocompatibility complex 1 (MHC I) and MHC II gene mRNA in acute graft rejection (AGR). METHODS: Fifteen New Zealand white rabbits were randomly divided into three groups to observe skin graft rejection: three rabbits were in the autograft control group; six rabbits in a cyclosporine (CsA) treated allografted group; and the other six rabbits in untreated allografted group. The CsA-treated allografted group was given CsA (5 mg/kg) daily intramuscularly. PBMC samples were obtained every 2 days to detect by real-time polymerase chain reaction, PBMC MHC I and MHC II gene mRNA. RESULTS: MHC I and MHC II gene mRNA levels did not show any obvious change in the autografted controls. MHC I gene mRNA levels showed a slow increase in the CsA-treated allografted group, but no obvious change in the untreated allografted group. MHC II gene mRNA reached the highest level at 2 to 3 days before graft rejection appeared macroscopically in the CsA-treated allografted group and untreated allografted group, then decreasing to a low level. CONCLUSION: Compared with MHC I gene mRNA expression, PBMC MHC II gene mRNA expression may be considered to be an earlier marker for AGR.     

The influence of MHC class II antigen blockade by perfusion with a monoclonal antibody on rat renal graft survival

To decrease immunogenicity of the rat kidney, grafts were perfused with an anti-MHC class II monoclonal antibody (mAb). How effectively this procedure blocked class II-positive cells, which were mainly dendritic in appearance, was checked by immunostaining renal sections after perfusion and comparing them with in vitro stained sections. Optimum conditions were applied for graft pretreatment before transplantation. This procedure prolonged graft survival, though not satisfactorily from the biological point of view (9.6±0.8 versus 7.7±0.5 days in the control group; P<0.02). The dendritic cells were not killed but blocked. Several hours after transplantation, the mAb dissociated from these class II-positive cells. It was also shown that donor cells migrate into the recipient's spleen early after transplantation. The number of these cells was smaller when the transplanted organ was perfused with the mAb. Further studies are suggested to deplete the graft of donor dendritic cells more adequately. They should also combine graft perfusion with anticlass II mAb and recipient immunosuppression at reduced doses.     

The influence of MHC class II antigen blockade by perfusion with a monoclonal antibody on rat renal graft survival

Abstract. To decrease immunogenicity of the rat kidney, grafts were perfused with an anti-MHC class II monoclonal antibody (mAb). How effectively this procedure blocked class II-positive cells, which were mainly dendritic in appearance, was checked by immunostaining renal sections after perfusion and comparing them with in vitro stained sections. Optimum conditions were applied for graft pretreatment before transplantation. This procedure prolonged graft survival, though not satisfactorily from the biological point of view (9.6 ± 0.8 versus 7.7 ± 0.5 days in the control group; P < 0.02). The dendritic cells were not killed but blocked. Several hours after transplantation, the mAb dissociated from these class II-positive cells. It was also shown that donor cells migrate into the recipient's spleen early after transplantation. The number of these cells was smaller when the transplanted organ was perfused with the mAb. Further studies are suggested to deplete the graft of donor dendritic cells more adequately. They should also combine graft perfusion with anti-class II mAb and recipient immunosuppression at reduced doses.     

CMV infection, class II antigen expression, and human kidney allograft rejection

After successful transplantation, the major histocompatibility complex (MHC) antigens of kidney parenchymal cells are lost and no longer detectable in the graft, presumably due to administration of glucocorticosteroids. During rejection, the MHC antigens reappear in the graft parenchymal cells. The upregulation is possibly due to gamma-interferon released in situ by the allograft activated T (blast) cells. In this communication we demonstrate that cytomegalovirus (CMV) infection is invariably associated with an upregulation of the antigen display in the graft. In 12 of 14 (86%) cases with proved CMV disease, the display of class II antigens was associated with a cytological and/or clinical episode of rejection. In 223 transplant recipients without proved CMV disease transplanted during the same period, the frequency of late rejections was 17% (P less than 0.001). The results suggest that the display of class II antigens on the graft, mediated presumably by gamma-interferon as a consequence of CMV infection, is the reason for graft rejection in context of CMV disease.     

Influence of donor MHC class I antigen expression on graft survival after rat parathyroid allotransplantation

BACKGROUND AND AIMS: We investigated the influence of donor MHC antigen expression on graft survival after parathyroid transplantation in three different strain combinations. METHODS: MHC class I and II expression on parathyroid tissue of Lewis (LEW), Dark Agouti (DA), and Wistar-Furth (WF) rats was first analysed semiquantitatively by immunohistochemistry. Additionally, five groups were transplanted: (1) LEW to LEW, (2) DA to DA, (3) LEW to DA, (4) WF to LEW, and (5) DA to LEW. METHODS: MHC class I expression was strong in DA, moderate in WF, and weak in LEW rats; MHC class II expression was negative in all three strains. In the interstitium of all investigated tissue specimens, the proportion of MHC class II-expressing cells was low. RESULTS: After syngeneic transplantation, graft survival could be documented over the whole observation period. A mean graft survival of 20 (+/-2) days was observed following transplantation from LEW to DA, grafts in the group WF to LEW were rejected after 13 (+/-1) days, and graft function lasted 8 (+/-2) days in the group DA to LEW. The number of intragraft leukocytes expressing MHC class II molecules was equal in all groups, whereas increased levels of MHC class I on rat parathyroid tissue before transplantation resulted in a more rapid rejection. CONCLUSION: These results demonstrate that immunogenicity of rat parathyroid tissue seems to be determined by the amount of MHC class I expressed on donor parenchymal cells.     

Suppression of kidney allograft rejection across full MHC barriers by recipient-specific antibodies to class II MHC antigens.

The aim of these studies was to see if recipient-specific antibodies to class II MHC antigens might be effective in suppressing kidney graft rejection in rats. For these experiments, the polymorphic BMAC-4 mouse IgG1 monoclonal antibody to RT1-D class II MHC antigens was raised. This antibody reacts with the DA, LEW, PVG, and SHR strains, but not the BN or WAG strains, and is therefore recipient-specific in the WAG to PVG combination. Initial in vivo titrations demonstrated that 1 ml doses of the BMAC-4 and also of the MRC OX6 (monomorphic mouse IgG1 anti-RT1-B class II) antibody resulted in the maintenance of free antibody levels in blood for greater than 24 hr. Treatment of PVG recipients of WAG kidney allografts with the BMAC-4 antibody, but not the MRC OX6 antibody, resulted in greatly prolonged graft survival. To examine possible mechanisms, several experiments were performed. After intravenous injection, the antibody was found to have ready access to the connective tissues of nonlymphoid organs, to the red and white pulp of the spleen, and to the medulla of lymph nodes. However, there was poor early access to the cortex and paracortex of lymph nodes. Both MRC OX6 and BMAC-4 could completely suppress PVG anti-WAG and WAG anti-PVG mixed lymphocyte culture reactions. Both antibodies were also equally effective for opsonisation of class II-positive cells from the blood circulation. However, only the recipient-specific, anti-RT1-D BMAC-4 antibody suppressed graft rejection. Thus, while the BMAC-4 antibody is likely to have had a variety of different effects on RT1-D positive recipient cells, the locus specificity of the immunosuppression is consistent with an important component of those effects being the blocking of presentation of WAG donor alloantigens by PVG RT1-D class II antigens on PVG antigen-presenting cells.     

Provocation of skin graft rejection across murine class II differences by non--bone-marrow-derived cells

We have evaluated the relative contribution of bone-marrow-derived cells to skin allograft immunogenicity in mice differing only at class II major histocompatibility genes by using bone marrow radiation chimeras as donors. The mouse strains used were C57BL/6Kh (B6) and B6.C-H-2bm12 (bm12), which differ only at at A beta gene of the I region of the mouse H-2 complex. Our results demonstrated that skin from (B6----bm12) chimeras was accepted by bm12 recipients and rejected by B6 mice in a manner indistinguishable from that of normal bm12 skin. Likewise, naive bm12 mice rejected (bm12----B6) chimeric skin and normal B6 skin equally well, and B6 animals accepted both types of skin grafts. Our data argues that the donor cell-type leading to graft rejection across limited I region differences is not of bone marrow origin, and that these cells must--at least under certain circumstances--express class II antigens.     

Needle biopsy evaluation of class II major histocompatibility complex antigen expression for the differential diagnosis of cyclosporine nephrotoxicity from kidney graft rejection

Twenty needle biopsies from 14 patients were taken at times of renal dysfunction, and frozen sections were stained for class I and class II major histocompatibility complex (MHC) antigen expression using the immunoperoxidase technique and monomorphic mouse monoclonal antibodies. Eight of the 9 biopsies taken during periods of dysfunction attributed to cyclosporine toxicity had normal levels of class II expression. In contrast, 9 of the 10 biopsies taken during episodes of rejection had easily recognized increases in class II expression. In the one case where no definite clinical diagnosis was possible, no class II induction was present. Class I levels were less definitive but tended to be markedly raised in the cases of rejection, and only mildly raised in the cases of nephrotoxicity. Biopsy results can be available within 1 1/2-2 hr. The test is therefore likely to be of value in the correct diagnosis of the cause of renal dysfunction and thereby improve the management of cyclosporine-treated renal transplant patients.     

Effect of cyclosporine on MHC class II antigen expression on arterial and venous endothelium in vitro

MHC class II antigens play a crucial role in immunological responses. The expression of MHC class II antigens on monocytes and endothelial cells is reported to be variable and able to be induced by gamma-interferon. In this study we report on MHC class II antigen expression in vitro by arterial and venous canine endothelial cells, as detected with FACS analysis and indirect immunofluorescence with a monoclonal antibody against canine MHC class II antigens. It appears that cultured endothelial cells do not express MHC class II antigens. Their expression could be induced during a three-day incubation period in lymphokine-containing supernatant produced in mixed leukocyte culture (MLC). Cyclosporine (CsA) added to allogeneically stimulated or unstimulated canine lymphocytes in MLC inhibited the induction of expression by the MLC supernatant. The addition of CsA to MLC supernatant did not have an inhibitory effect. It is concluded that CsA inhibits the production of an MHC class-II-antigen-inducing lymphokine produced by lymphocytes in mixed cultures; allogeneic stimulation is not necessary for production of the lymphokine. It is postulated that a possible mode of action of CsA in prolongation of allograft survival is based on prevention of the induction of MHC class II antigen expression by endothelial cells.     

Donor class I and class II major histocompatibility complex antigen expression following liver allografting in rejecting and nonrejecting rat strain combinations

Orthotopic liver allografts in the nonrejecting DA-to-PVG strain combination and in the DA-to-LEW strain combination were studied at various times after transplantation for donor class I and class II MHC expression using immunohistological techniques and quantitative analyses. DA-to-DA isografts were also studied. In the isografts, weak class I induction on hepatocytes and biliary epithelium was noted from day 5, and this persisted to day 15, the last time point examined. In DA-to-PVG allografts, class I induction also appeared on hepatocytes and biliary epithelium from day 5, but was more intense than in the isografts. Nevertheless, the induction was patchy within most grafts, and in some grafts was not prominent. Quantitative absorption analyses demonstrated that the maximum increase in donor class I expression was only 3-fold over the normal liver. In the strong DA-to-LEW combination, class I induction on hepatocytes seemed to appear earlier, beginning at day 3, and was more uniform and intense than in the DA-to-PVG model from day 5. In the isografts, there was no induction of class II antigens on hepatocytes or biliary epithelium at any stage, but from days 5 to 15 there was a marked increase in the number of isolated, class II-positive cells in the hepatic lobule, probably representing class II induction in the Kupffer cells of the isografts. In DA-to-PVG allografts, biliary epithelium became class II-positive from day 5, and this persisted to day 30, the last time point examined. Weak but definite class II induction was seen on some hepatocytes from day 5 through day 30. However, the majority of hepatocytes remained class II-negative. By day 30, there was virtually no donor class II staining the sinusoids, but isolated class II-positive cells of recipient type were seen, the pattern suggesting a replacement of the graft Kupffer cells by recipient Kupffer cells at this stage. By quantitative absorption analysis, donor class II expression in the grafts increased approximately 5-fold. In DA-to-LEW allografts, class II induction was not noticeably different from that seen in the DA-to-PVG model, except that induction of class II antigens on the Kupffer cells possibly appeared earlier in this strain combination.     

Multiple patterns of MHC class II antigen expression on cellular constituents of rat heart grafts. Lack of correlation with graft survival, but strong correlation with vasculitis

The patterns of induced major histocompatibility antigen expression on indigenous cellular elements of heterotopic rat cardiac grafts were determined by immunohistologic methods in a variety of donor-recipient combinations. Heart grafts were studied in combined full-MHC- and non-MHC-disparate combinations, isolated intra-MHC-disparate combinations, and non-MHC-disparate combinations. The pattern of class II expression on cellular constituents of the grafts was highly variable and critically dependent upon the nature of the specific unidirectional donor-recipient combination. No uniform pattern of class II expression emerged that was clearly predictive of rapidity of rejection or of protracted survival. However, vasculitis was confined to grafts in combinations in which induced class II expression on graft large vessel endothelium was present. Sites of vasculitis were never encountered in the absence of induced class II expression on overlying endothelium. Vasculitis and associated induced class II expression on large vessel endothelium were present in rapidly rejecting grafts and in grafts with indefinite survival. In the latter, vasculitis was shown to progress to a late phase of occlusive intimal thickening. Induced class I expression on graft cardiac myofibers was present in all the genetically disparate donor-recipient combinations examined in this study, irrespective of the length of graft survival. This investigation has shown that no uniform stereotyped pattern of MHC antigen expression on cellular constituents correlates with the length of graft survival. However, induced class II expression on graft large vessel endothelium is closely associated with vasculitis, which can directly progress to occlusive intimal thickening in grafts with prolonged survival.     

Altered distribution of class I and class II MHC antigens during acute pancreas allograft rejection in the rat

The expression of MHC class I and class II antigens was investigated in a model of acute pancreas allograft rejection in the rat. Pancreaticoduodenal and duct-ligated DA(RT1a)-to-LEW(RT1(1] and LEW(RT1(1]-to-LEW.1U(RT1u) pancreas grafts were compared with normal organs and with LEW(RT1(1] isografts at daily intervals from day 1 to day 10 after transplantation. The results show profound changes of MHC antigen distribution in allografts during the process of rejection. Exocrine acinar cells, being class-I-antigen-negative in the normal pancreas, strongly express these antigens during rejection. Class II antigens, normally not found in pancreatic endothelia or parenchymal cells, appear in duct epithelia, acinar cells, and endothelia of big vessels. Endocrine islet cells and smooth muscle cells stay Ia-negative throughout the rejection process. Focal class I reactivity is also observed in acinar cells of pancreaticoduodenal isografts; but class II antigens are neither seen in parenchymal cells nor in endothelia of any isograft. Thus, in the rat pancreas allograft model, the induction of class II antigens is an early phenomenon characteristic of an ongoing immune response, and it provides a valuable new diagnostic criterion. Antibodies reactive exclusively with donor-haplotype antigens demonstrate an increase in donor-derived class I and class II antigen-positive interstitial cells in addition to parenchymal antigenic changes. A possible effect of the antigenic alteration described on the course of the rejection process is discussed.