Increased expression of MHC class II antigens in rejecting canine lung allografts

Expression of major histocompatibility complex class II antigens was investigated in the normal lungs and in lung allografts of mongrel dogs after single-lung transplantation. Cryostat sections were stained with an indirect immunoperoxidase technique that used B1F6 and 7.5.10.1 as anti-MHC class II monoclonal antibodies. In the normal lungs and native lungs of the recipient dogs after single-lung transplantation, only some cells of lymphoid tissue and macrophages/dendritic cells were MHC class II-positive. During acute rejection, increased infiltration with MHC class II-positive cells in perivascular, peribronchial, and interstitial areas and intraalveolar spaces was found in lung allografts. In addition, expression of MHC class II antigens was induced on the bronchial epithelium and vascular endothelium. Induced expression of MHC class II antigens on the bronchial epithelium and vascular endothelium in rejecting lung allografts was found as early as two days after single-lung transplantation. The intensity of MHC class II antigen expression on bronchial epithelium and vascular endothelium in graft lungs increased with the progression of rejection response and directly correlated with the bronchoalveolar lavage fluid (BALF) levels of biochemical markers, as tumor necrosis factor alpha, gamma-interferon (IFN-gamma), interleukin 2 (IL-2) and soluble interleukin 2 receptor (SIL-2R). Abnormal expression of MHC class II antigens on bronchial epithelium and vascular endothelium and abnormal elevation of BALF levels of the cytokines in lung allografts could be prevented by cyclosporine (CsA) treatment. Our results suggested that MHC class II antigen expression could be induced on the bronchial epithelium and vascular endothelium of canine lung allografts during acute rejection. This abnormal expression of MHC class II antigens on bronchial epithelium and vascular endothelium of graft lungs may serve as a specific index for diagnosis of lung allograft rejection when infection as an inducing factor can be excluded. Furthermore, bronchial epithelium and vascular endothelium of lung allografts have become MHC class II-positive, and are likely to be the targets for low-grade rejection, resulting in the development of bronchiolitis obliterans and occlusive vascular disease in lung allografts.     

The role of class I and class II MHC antigens in the rejection of vascularized heart allografts in mice

We have examined the role of entire major histocompatibility complex (MHC) disparity, individual class II or class I alloantigens in the rejection of vascularized heart allografts. Our results demonstrate that entire MHC, as well as both class II and class I disparities, may induce acute heart graft rejection or severe and irreversible heart muscle destruction. However, in 1 of 2 combinations differing at class II and 1 of 5 differing at class I, hearts have shown a good function greater than 100 days postgrafting. Furthermore, each donor-recipient combination has demonstrated a unique pattern of heart allograft function as well as a degree of heart muscle damage. In conclusion, these data suggest that the rejection process depends upon multiple factors such as the immune-response-gene-regulated immunoresponsiveness of the recipient as well as the expression of alloantigens on heart grafts during the induction and effector phases of the immune response.     

Indefinite survival of MHC class I-deficient murine pancreatic islet allografts.

To examine the immune response to class I-deficient allogeneic tissue, we used beta 2-microglobulin-deficient mice as graft donors. These mice lack cell surface class I major histocompatibility complex antigen expression. Pancreatic islet allografts from class I-deficient donors survived indefinitely in a majority of fully allogeneic BALB/c recipients. In contrast, host recognition of graft class I antigen was unnecessary for prompt destruction of skin allografts of for autoimmune damage of transplanted pancreatic islet grafts in nonobese diabetic mice. These studies provide evidence that intentional genetic elimination of immunologically relevant donor antigens may prove an effective strategy for preventing allograft rejection.     

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)     

Chronic antagonism of Mig inhibits cellular infiltration and promotes survival of class II MHC disparate skin allografts

BACKGROUND: The goal of the current study was to test the ability of monokine induced by IFN-gamma (Mig)-specific antibodies to inhibit long-term T cell infiltration into class II major histocompatibility complex (MHC) disparate skin allografts and to test cellular and molecular changes in the graft during the rejection observed following cessation of treatment. METHODS: C57BL/6 recipients of B6.H-2bm12 skin grafts were treated with normal rabbit serum (NRS) or rabbit Mig antiserum (Mig AS) every other day from day 7 until day 21 posttransplant and then weekly thereafter. Allografts were retrieved during the course of treatment and following cessation. Tissue sections were prepared and stained to compare infiltration by macrophages and CD4+ and CD8+ T cells and to assess collagen deposition in the grafts. RNA was prepared and tested by ribonuclease protection assay for intragraft levels of Mig, monocyte chemotactic protein-1 (MCP-1) and regulated on activation normal T-cell expressed and secreted (RANTES). RESULTS: T cell and macrophage infiltration into allografts was inhibited and graft survival maintained as long as Mig-specific antibodies were given. Following cessation of treatment, T cells and macrophages infiltrated the allografts. In contrast to the histology of acute rejection observed in allografts from NRS-treated recipients, the resulting rejection of the allografts from Mig AS-treated recipients was accompanied by dense collagen deposition and high level expression of Mig and RANTES. CONCLUSIONS: Mig directs T cell infiltration into B6.H-2bm12 skin allografts on C57BL/6 recipients. Delayed T cell and macrophage infiltration and rejection of the grafts following cessation of Mig AS treatment results in rejection that is histologically and molecularly distinct from acute rejection.     

An MHC class II disparity raises the threshold for tolerance induction in pulmonary allografts in miniature swine

OBJECTIVES: The mechanisms and treatment of chronic rejection in pulmonary allotransplantation remain elusive. We have induced robust tolerance to class I-disparate lung allografts in miniature swine using an intensive 12-day course of tacrolimus. Here, we tested whether a tolerant state can be induced in swine receiving fully mismatched lung allografts. METHODS: Orthotopic left lung allografts were performed using MHC class I-disparate (group 1: n = 3) or fully disparate (group 2: n = 6) donors. The recipients received a 12-day postoperative course of tacrolimus (continuous intravenous infusion; target level = 35-50 ng/mL) as their only immunosuppression. RESULTS: All swine in group 1 maintained their grafts long term without developing any lesions of chronic rejection (>497, >432, >451 days). These recipients exhibited donor-specific hyporesponsiveness in cell-mediated lymphocytotoxity (CML) and mixed lymphocyte reaction (MLR) assays. In group 2, five of the six recipients maintained their grafts long term (sacrificed on postoperative days 515, 389, 429, 481, and 438 with viable grafts). Isolated lesions of obliterative bronchiolitis were occasionally seen on biopsy, and donor-specific hyporesponsiveness on assays was consistently observed. The remaining recipient rejected its graft on day 103 with histologic findings of obliterative bronchiolitis. CONCLUSIONS: We report long-term graft acceptance without chronic immunosuppression in five of six recipients across a full MHC disparity, albeit with some evidence of obliterative bronchiolitis. These data suggest that the class II disparity inherent in a fully mismatched transplant increases the requirement for tolerance induction.     

The effects of cyclosporine on the induction of donor class I and class II MHC antigens in heart and kidney allografts in the rat

We have previously reported 5-30-fold increases in the expression of class I and class II major histocompatibility complex (MHC) antigens in rejecting heart and kidney allografts in the DA-to-PVG rat strain combination. We examine here the effects of immunosuppression with cyclosporine on the induction of donor class I and class II MHC antigens in heart and kidney allografts in this strain combination. Immunohistological studies and quantitative absorption analyses using monoclonal antibodies and assay systems specific for donor class I and class II MHC antigens were used throughout. Heart allografts in cyclosporine-treated rats were examined on day 3,5,7,9,11, and 14 after transplantation, and kidney allografts in cyclosporine-treated rats were examined at day 7. In addition, untreated heart and kidney isografts were studied at days 1,3,5, and 7 after grafting. Immunohistological studies on frozen sections showed that cyclosporine-treated heart and kidney allografts showed no induction of class II MHC antigens, in contrast to untreated heart and kidney allografts. Class I MHC antigen induction did occur in spite of cyclosporine-therapy, but at levels lower than those seen in untreated allografts. Moreover, the pattern and degree of class I induction in the cyclosporine-treated allografts resembled very closely those seen in isografts, and so this induction was, in all probability, a consequence of the transplantation procedure rather than of specific immune responses. We also noted, in the cyclosporine-treated heart allografts, that all donor interstitial dendritic cells had disappeared and been replaced by recipient interstitial dendritic cells by the end of the second week after grafting. In addition, there was no reduction in the class II antigen content of kidney allografts treated for 7 days with cyclosporine. The absence of class II antigen induction in allografts where rejection is effectively suppressed with cyclosporine might be of clinical value in the differential diagnosis between rejection and cyclosporine toxicity in renal transplantation, and between active and inactive cellular infiltrates in heart transplantation.     

Does high MHC class II gene expression in normal lungs account for the strong immunogenicity of lung allografts?

Clinical experience has demonstrated that lung allografts are considerably more immunogenic than liver allografts although both organs contain equivalent amounts of lymphoreticular tissue. Northern blot analysis of MHC class II gene expression in various murine organs with I-AB and I-EB gene-specific oligonucleotide probes revealed that MHC class II expression in lungs is semiquantitatively higher than in the liver and other organs with the exception of the spleen. We conclude that this high MHC class II expression strongly suggests that the lymphoreticular tissue in the lungs is in a state of activation. This may be an important reason for the strong immunogenicity of lung allografts.     

Aberrant colonic expression of MHC class II antigens in Hirschsprung's disease.

The major histocompatibility complex (MHC) Class I and II cell surface antigens responsible for the recognition of self vs non-self were studied in patients with documented Hirschsprung's disease. Monoclonal antibodies reactive with monomorphic determinants of human lymphocyte antigen (HLA)-A,B,C (Class I) and HLA-DR (Class II) were used to demonstrate immunohistochemically the expression of MHC antigens in 27 biopsy specimens from a variety of colorectal disorders. The rectal specimens examined from patients with Hirschsprung's disease showed an unexpected, marked elevation of Class II antigens with abnormal localization in the mucosa and lamina propria. This ectopic expression was not seen in any portion of the small or large bowel of patients who did not have Hirschsprung's disease. Furthermore, proximal normal colon of children with Hirschsprung's disease failed to show increased expression of Class II antigen. In an attempt to better define the effector arm at a cellular level, the distribution of helper T cells (CD4+), cytotoxic/suppressor T cells (CD8+) and natural killer cells (NK; CD16+) was examined in 5 cases. In Hirschsprung's disease, rectal infiltration of CD8+ and CD16+ cells was found, but not CD4+ cells. Ectopic expression of Class II antigen with increased numbers of rectal T cells and NK cells suggested that an early immunologic event may be causal in Hirschsprung's disease.     

Delayed rejection of MHC class II-disparate skin allografts in mice treated with farnesyltransferase inhibitors

Farnesyltransferase inhibitors (FTIs), developed as anti-cancer drugs, have the potential to modulate immune responses without causing nonspecific immune suppression. We have investigated the possibility that FTIs, by affecting T cell cytokine secretion, can attenuate alloreactive immune responses. The effects of FTIs on murine alloreactive T cells were determined both in vitro, by measuring cytokine secretion or cell proliferation in mixed lymphocyte cultures, and in vivo, by performing skin allografts from H-2(bm12) mice to MHC class II-disparate B6 mice. We found that two different FTIs, ABT-100 and L-744,832, blocked secretion of IFN-gamma, IL-2, IL-4, and TNF-alpha from na?ve T cells in vitro. ABT-100 and L-744,832 blocked cytokine production from both CD4(+) and CD8(+) na?ve T cells stimulated with CD3 and CD28 antibodies, but only if the cells were pretreated with the FTIs for 48 h. Proliferation of alloreactive T cells in mixed lymphocyte cultures was blocked by either FTI. We also found that the proliferation of enriched T cells stimulated with IL-2 was blocked by ABT-100 treatment. In mice with an MHC class II-disparate skin graft, rejection of primary allografts was significantly delayed by treatment with either ABT-100 or L-744,832. Secondary rejection in mice previously primed to the alloantigen was found to be unaffected by L-744,832 treatment. We have shown that FTIs can block T cell cytokine secretion and attenuate alloreactive immune responses. The ability of FTIs to block secretion of cytokines, including IFN-gamma and IL-4, from na?ve T cells provides a likely biological mechanism for the specific suppression of class II MHC-mediated allorejection. These results demonstrate that FTIs may have useful immunomodulatory activity due to their ability to delay priming to alloantigens.     

Expression of MHC class II antigens on human glioma cells modulated by transfection with genes encoding these antigens.

Four glioma cell lines not expressing human leukocyte antigen (HLA)-DR or DQ did so after transfection with genes encoding HLA-DR and DQ. The glioma cells had enhanced ability to stimulate allogeneic and autologous responding lymphocytes in the mixed lymphocyte response (MLR). Glioma cells expressing only HLA-DR had weaker MLR enhancement than those expressing both HLA-DR and DQ or only HLA-DQ. Stimulation by transformed glioma cells increased the killing activity of responding lymphocytes against autologous glioma cells 2.0 times. Both HLA-DR and DQ are important in MLR, and the immunogenicity of glioma cells might be increased by transfection with genes encoding these antigens.     

Altered intragraft immune responses and improved renal function in MHC class II-deficient mouse kidney allografts

BACKGROUND: During renal allograft rejection, expression of MHC class II antigens is up-regulated on the parenchymal cells of the kidney. This up-regulation of MHC class II proteins may stimulate the intragraft alloimmune response by promoting their recognition by recipient CD4+ T cells. In previous studies, absence of donor MHC class II antigens did not affect skin graft survival, but resulted in prolonged survival of cardiac allografts. METHODS: To further explore the role of MHC class II antigens in kidney graft rejection, we performed vascularized kidney transplants using donor kidneys from A(beta)b-deficient mice that lack MHC class II expression. RESULTS: At 4 weeks after transplant, GFR was substantially depressed in control allografts (2.18+/-0.46 ml/min/kg) compared to nonrejecting isografts (7.98+/-1.62 ml/min/kg; P<0.01), but significantly higher in class II- allografts (4.38+/-0.60 ml/min/kg; P<0.05). Despite the improvement in renal function, class II- allograft demonstrated histologic features of acute rejection, not unlike control allografts. However, morphometric analysis at 1 week after transplantation demonstrated significantly fewer CD4+ T cells infiltrating class II- allografts (12.8+/-1.2 cells/mm2) compared to controls (25.5+/-2.6 cells/mm2; P=0.0007). Finally, the intragraft profile of cytokines was altered in class II- allografts, with significantly reduced expression of Th2 cytokine mRNA compared to controls. CONCLUSIONS: These results support a role of MHC class II antigens in the kidney regulating immune cells within the graft. Further, effector pathways triggered by class II antigens promote renal injury during rejection.     

Vulnerability of allografts to rejection by MHC class II-restricted T-cell receptor transgenic mice

BACKGROUND: Examination of the in vivo activation and function of CD4+ T cells in response to allografts may advance our understanding of the rejection process. We analyzed the capacity of transgenic class II-restricted CD4 T cells to reject skin, cardiac, and islet transplants. METHODS: TS1 mice possess a high frequency of CD4+ T cells specific for the immunodominant epitope of the viral hemagglutinin (HA) protein. We analyzed the kinetics of rejection of skin, heart, and islet grafts by na?ve and sensitized TS1 mice and by adoptively transferred TS1 lymphocytes. RESULTS: Rejection of heart transplants was more rapid than skin grafts (mean survival time, 12.9 vs. 26.6 days), and islet grafts survived indefinitely in TS1 mice. These findings may be partly attributable to the supranormal frequency of HA-reactive cells in TS1 mice. In support of this, we found that adoptive transfer of 5 x 10(5) TS1 lymphocytes to Balb/c hosts effected consistent rejection of HA-bearing skin transplants, whereas a significantly greater number (3 x 10(6)) was required for heart transplant rejection. The in vivo proliferative response of HA-specific T cells to heart and skin was found to be robust and predominantly localized to the draining lymph nodes. CONCLUSION: We developed a model of allograft rejection in which the responding T cells and relevant graft antigen are specifically defined. Adoptive transfer of carboxy-fluorescein succinimidyl ester-labeled transgenic T cells allowed us to visualize a robust proliferative response in vivo to heart and skin allografts, which in both cases was localized to regional lymph nodes.     

Scintigraphic imaging of MHC class II antigen induction in mouse kidney allografts: a new approach to noninvasive detection of early rejection

Mice with kidney transplants were investigated to determine whether early kidney allograft rejection could be detected by radioimmune scintigraphy targeting major histocompatibility complex (MHC) class II antigens induced on donor organ cells. Allografts from C3H/He (H2^k) donors were transplanted into BALB/c (H2^d) recipients. Each mouse was injected intravenously with 100 Ci of ¹²³I-labeled anti-MHC class II monoclonal antibody (mAb; Y17, anti-IE^k) 16 h before scintigraphy. After imaging, mice were sacrificed for tissue counting and histopathological examination. Radiotracer uptake in the nontreated allografts increased starting on the 3rd day after transplantation, peaked at around the 6th day, and then gradually decreased. Rejecting allografts with only focal perivascular mononuclear cell infiltration could be identified by scintigraphy. However, allografted mice without evidence of rejection and isografted mice did not show an increase in radiotracer uptake. Rejecting BALB/c kidney transplanted into C3H/He mice did not show an increase in Y17 mAb uptake, suggesting that class II antigens induced on donor kidneys are solely responsible for the mAb uptake in positive scintigrams of rejecting allografts. Five allografted mice were treated with anti-CD3 mAb and cyclosporin starting 3–9 days after transplantation. Radiotracer uptake decreased after 4 weeks of treatment and increased 2 weeks after the cessation of immunosuppressive treatment, reflecting suppression and recurrence of rejection, as determined by histological examination. These changes could be followed scintigraphically. We conclude that changes in class II antigen expression can be assessed by the ¹²³I-labeled anti-MHC class II antigen mAb and that it is a sensitive and noninvasive method for detecting kidney allograft rejection.