Constitutive expression of major histocompatibility complex class II antigens in pulmonary epithelium and endothelium varies among different species

We have observed high constitutive levels of class II antigen expression on porcine and human coronary endothelium, but not on the endothelium of rats and mice. This study examines whether a similar interspecies difference exists in the expression of class II molecules on pulmonary epithelium and endothelium. Lung tissues from na?ve human, porcine, and rodent sources were stained with the monoclonal antibody ISCR3 and examined by light microscopy. Immunoperoxidase staining of class II molecules was observed on human and porcine pulmonary epithelium and endothelium, but was absent in rats and mice. By using an antibody with cross-species reactivity, we demonstrated that na?ve swine pulmonary epithelium and endothelium, unlike those of rodent species, express basal levels of class II antigens in a manner similar to that observed in human lung tissue. These interspecies differences may explain experimental differences observed between murine and large-animal constructs.     

Expression of donor class I major histocompatibility antigens on the vascular endothelium of mouse skin allografts

The expression of a class I MHC antigen on the vascular endothelium of mouse skin allografts was assessed by in vivo uptake of radiolabeled monoclonal anti-class-I antibody in the grafts after i.v. injection into the recipients. Endothelial localization of the bound antibodies was demonstrated via double-labeling immunofluorescence microscopy using factor-VIII-related antigen as a marker for endothelial cells. Treatment of recipients with cyclosporine was accompanied by low levels of class I antigen expression in the grafts, and similarly low levels were measured in grafts carried by nude recipients in the complete absence of rejection. Withdrawal of immunosuppressive therapy was followed by an increased class I antigen expression in the donor skin. An increase was also observed in skin grafts undergoing first-set rejection. We conclude that the expression of class I antigens on the capillary endothelium of mouse skin allografts in vivo is variable and is under influence of the immune status of the recipient.     

Effect of synthetic colloids on major histocompatibility complex class II expression

STUDY OBJECTIVE: Synthetic colloids are used for perioperative fluid management. We hypothesized that their use may be associated with changes in major histocompatibility complex (MHC) class II expression. This could affect patients' morbidity and mortality during clinical intervention. SETTING: University research laboratory. SUBJECTS: Whole blood samples from healthy volunteers. INTERVENTIONS: Whole blood samples from healthy volunteers (n = 6) were incubated with different concentrations of hydroxyethyl starch (HES) from maize and potato (pHES), dextran, and polygelin (gelatine) for 24 hours with or without 100 U/mL human interferon gamma (IFN-gamma; stimulus for MHC class II expression). The expression of human leukocyte antigen (HLA): HLA-DR, HLA-DQ, and HLA-DP was detected simultaneously by a fluoresceinisothiocyanate (FITC)-labeled antibody and analyzed by flow cytometry on lymphocytes and monocytes. MEASUREMENTS AND MAIN RESULTS: Hydroxyethyl starch, pHES, and dextran induced a significant increase in HLA expression. The induction of MHC class II was independent of the structure (50 mg/mL: control, 8.7+/-1.4%; HES, 28 +/- 9.7%; pHES, 29.8 +/- 11.7%; and dextran, 50.2 +/- 8.1%). In contrast, polygelin increased HLA expression only at the highest concentration of gelatine (5 mg/mL, 7.8 +/- 1%; 50 mg/mL, 7.6 +/- 0.8%; 100 mg/mL, 7.3 +/- 1%; 200 mg/mL,16.2 +/- 2.3%). The addition of IFN-gamma decreased HLA expression in presence of highest concentration of HES and dextran. CONCLUSION: In an ex vivo laboratory setting, we demonstrate that high concentrations of plasma expanders are associated with increased HLA expression on lymphocytes and monocytes. However, further in vivo studies are necessary to demonstrate the clinical significance of this observation.     

The relative immunogenicity of corneal epithelium, stroma, and endothelium. The role of major histocompatibility complex antigens

Corneal allografts have been shown to give rise to immune responses, but the role and relative importance of individual corneal cell populations in evoking such responses remain unclear. We dissected ACI (RT1a) rat corneas into separate epithelial, stromal, and endothelial components by a method that yields pure cell populations in tissue culture, and grafted these components separately to groups of fully allogeneic PVG (RT1c) recipients). Grafts of corneal stroma elicited strong cellular cytotoxic immune responses in a cell-mediated lymphocytotoxic assay, but corneal epithelium failed to generate any detectable response. Grafts of corneal endothelium alone, however, evoked a potent cellular cytotoxic response. Using congenic rats, it was found that grafts from PVG.1A (RT1a) donors to PVG (RT1c) recipients (which differ at both the RT1.A and B loci) yielded identical results. However, no corneal component graft from PVG.R1 (RT1rl) donors to PVG recipients (which differ only at RT1.A) generated a detectable immune response. Use of target lymphoblasts from congenic strains established that at least part of all responses detected were directed against class I (RT1.A) major histocompatibility complex antigens. These findings indicate that there is differential immunogenicity of specific corneal tissue components in the rat that may be further influenced by the degree of MHC disparity between donor and recipient.     

Relative contribution of major histocompatibility complex antigens to the immunogenicity of corneal allografts

The relative contributions made by the major class I (RT1.A) and class II (RT1.B) antigens of the rat major histocompatibility complex (MHC) to the immunogenicity of corneal and skin allografts were investigated using congenic animals. PVG (RT1c) recipients were given skin or heterotopic cornea grafts from congenic PVG.1A (RT1a) or PVG.R1 (RT1r1) donors, which respectively share the entire RT1 complex or only the RT1.A (major class I MHC antigen) region with fully allogeneic ACI (RT1a) rats. Recipient splenocytes were tested at ten days posttransplant for their ability to lyse ACI, PVG.1A, PVG.R1, and PVG target cells in a secondary CML following 6 days in vitro stimulation with irradiated ACI spleen cells. Effector cells from PVG recipients of both RT1.A and B disparate (PVG.1A donor) and RT1.A disparate (PVG.R1) skin or cornea grafts lysed ACI, PVG.1A, and PVG.R1 (but not PVG) targets at levels significantly above controls given syngeneic grafts. However, the level of cytotoxicity against PVG.R1 as well as ACI and PVG.1A allogeneic targets was always significantly higher following PVG.1A grafts than following PVG.R1 grafts, indicating that the addition of a class II MHC antigen difference markedly augmented the immunogenicity of class I MHC antigen disparate cornea and skin grafts. Taken together with other recent evidence confirming the presence of Langerhans cells in the normal rat (and human) cornea, these results suggest that class II MHC-bearing cells make an important contribution to the immunogenicity of corneal allografts.     

Impact of class II major histocompatibility complex antigen expression on the immunogenic potential of isolated rat vascular endothelial cells

We have investigated the immunogenic potential of rat heart vascular endothelial cells by their ability to induce an accelerated rejection of a relevant heart allograft, and related the immunogenic potential to the expression of class II major histocompatibility complex (MHC) antigens on the endothelial cell surface. Only 12% of freshly isolated rat vascular endothelial cells express class II antigens in serum-free medium, and the level of expression is low as judged by immunoperoxidase staining and/or the ability of endothelial cells to bind staphylococci to the cell surface after treatment with monoclonal antibodies to the class II molecule. On the other hand, 99% of the endothelial cells under the same conditions express class I, and the level of expression is high. The class II antigen expression of vascular endothelial cells can be upregulated to more than 98% by recombinant gamma-interferon in vitro--and, concomitantly, the level of expression becomes high, even on the cell surface. Treatment with gamma-interferon did not substantially alter the level of class I expression. The endothelial cells expressing class II antigens weakly, are also weakly immunogenic in vivo: 10(7) endothelial cells are required to reduce the graft survival by 50% of that of the unprimed host. On the contrary, the endothelial cells of the same lineage induced to express class II antigens by gamma-interferon in vitro are highly immunogenic in vivo, as immunogenic as freshly-isolated spleen dendritic cells: only 10(4) endothelial cells are required to induce a 50% reduction of graft survival. These observations demonstrate for the first time that rat vascular endothelial cells are immunogenic in a primary transplantation response in vivo--and, moreover, that the immunogenic capacity of the endothelial cells is directly proportional to the extent of class II MHC antigen expression on the cell surface.     

Expression of class II major histocompatibility complex antigens by bronchial epithelium in rat lung allografts

Variations in expression of class II major histocompatibility complex antigens on bronchial epithelial cells and vascular endothelium were investigated in normal rat lungs and allografted lungs during acute rejection and after cyclosporine (CsA) treatment. BN (RT1n) left lungs were transplanted into LEW (RT1l) recipients. Lungs were excised during acute rejection in untreated rats on postoperative days 1 through 5, and after CsA treatment (25 mg/kg on days 2 and 3) on days 5 and 100. Cryostat sections were examined for class II antigen expression with an immunoperoxidase technique, using various monoclonal antibodies. In the normal lung, class II antigens were not expressed by epithelial or endothelial cells. In the allografts, induction of class II antigens closely correlated with the rejection process: on day 2, the ciliated bronchial epithelium was locally positive; it became uniformly positive with increasing cellular peribronchial infiltration on days 3 and 4. CsA treatment prevented class II antigen expression to a certain extent, leaving the bronchial epithelium weakly positive at 100 days. Endothelial cells were invariably negative for class II antigens in all allografted lungs. The class II antigens expressed on the bronchial epithelial cells were of graft origin, except for recipient-type class II molecules found on the ciliated surface in CsA-treated animals. We conclude that expression of class II antigens by bronchial epithelium is the result of a bronchus-directed rejection process, and hypothesize that such a rejection process may have caused bronchiolitis obliterans in several of the patients with combined heart-lung transplants. Important is the observation that class II molecules can be present on the membranes of cells that do not themselves produce these antigens.     

Lack of correlation between the induction of donor class I and class II major histocompatibility complex antigens and graft rejection

The induction of donor major histocompatibility complex (MHC) antigens on nonrejected and rejected rat renal allografts was compared at various times after transplantation in two strain combinations, DA-to-PVG and LEW-to-DA. Graft rejection was prevented by preoperative donor-specific blood transfusion (DST). Quantitative absorption analysis and immunohistology were performed using monoclonal antibodies specific for donor class I and class II MHC antigens. A significant increase in the expression of donor MHC antigens, both class I and class II, was demonstrated on nonrejected as well as rejected kidneys after transplantation. A kinetic analysis showed that induction of donor class I antigens was accelerated on the nonrejected grafts, and by day 5 the nonrejected kidneys showed increased expression of class I antigen when compared with the rejected grafts (a 37- vs. a 25-fold increase in expression). Increased expression of donor class I antigens persisted on the nonrejected grafts and was still detectable on long-term-surviving kidneys, 50 days after transplantation. The magnitude of class II antigen induction was similar on both rejected and nonrejected grafts (8-fold by 5 days after transplantation). Immunohistology demonstrated that class I and class II antigens were induced on identical structures in the kidney in both situations. In particular the vessel endothelia, which do not express class II antigens in normal kidney, become strongly positive in both rejected and nonrejected grafts 5 days after transplantation. Although renal allograft rejection is completely suppressed in rats given a single donor-specific blood transfusion before transplantation, graft survival cannot be explained by the lack of induction of donor MHC antigens. Donor MHC antigens are induced on these nonrejected kidney grafts, and therefore they could act as target molecules for the effector cells that mediate graft destruction. Thus the induction of donor MHC antigens on tissue allografts should not be considered as indicative of a rejection response resulting in graft destruction.     

Presentation of donor major histocompatibility complex class II antigens by dna vaccination prolongs heart allograft survival

BACKGROUND: Donor major histocompatibility complex (MHC) antigens play an important role in both allograft rejection and tolerance. With the use of several animal models, it has been shown that presentation of donor antigens before transplantation can lead to allograft tolerance. Vaccination of animals with a DNA plasmid encoding an antigen enables highly efficient expression of the protein in vivo. METHODS: In this study, we used DNA vaccination delivered through intramuscular, intraperitoneal, or intravenous routes to indirectly present donor antigens and to determine the effect in the modulation of the allograft response. LEW.1A recipients of a LEW.1W heart allograft were treated before grafting by vaccination with a plasmid encoding the donor RT1.D MHC class II or RT1.A class I molecules. RESULTS: Only anti-MHC II vaccination significantly prolonged allograft survival compared with untreated rats. We observed a significant prolongation of heart allograft survival with the intramuscular route of injection, but surprisingly we found the intravenous and intraperitoneal routes to be the best. CONCLUSION: After transplantation the anti-donor cellular response was significantly decreased in vaccinated rats. This was accompanied by a significant reduction in interferon-gamma mRNA expression in the grafted hearts and T helper 1-type alloantibody production, indicating that the vaccination modifies the alloresponse against the grafts.     

Increased expression of class I major histocompatibility complex antigens on hepatocytes in rejecting human liver allografts

We studied hepatocellular expression of major histocompatibility (MHC) antigens in 43 serial liver transplant biopsies from 22 patients (42 percutaneous, 1 autopsy specimen), 4 normal liver biopsies, and 8 percutaneous biopsies of diseased livers from non-liver-transplant patients. Frozen tissue sections were stained by an indirect immunofluorescence technique using monoclonal antibodies (MCAb) that recognize nonpolymorphic human class I or class II MHC determinants. Ethidium bromide was used to stain nuclei and rhodamine-conjugated anti-basement-membrane antibodies to delineate epithelial and vascular structures. HLA-DR antigens recognized by MCAb OKIa1 and I2 were not detected on hepatocytes but were detected on the bile duct epithelium in 7 of 27 transplant biopsies, including 5 with acute rejection and 1 with chronic liver disease that later progressed to chronic rejection. HLA-A, B, C antigens recognized by MCAb 34/28 intensely stained cells lining the liver sinusoids but were negative on hepatocytes in 4 normal liver biopsies and 7 of 8 non-transplant biopsies. Expression of class I MHC antigens on hepatocyte membranes was increased in 17 of 21 (81%) biopsies from patients with acute rejection, in 4 of 4 with chronic transplant liver disease, but in only 3 of 18 (17%) biopsies from patients with no rejection (chi square = 8.62, P less than 0.01). Our observations demonstrate increased expression of MHC class I antigens in association with acute rejection in human orthotopic liver transplantation. Histologic resolution of the rejection episode is generally followed by a decrease in hepatocyte class I antigen expression. Further analysis of this response may have value in assessing the severity of the rejection and effectiveness of treatment.     

In vivo suppression of major histocompatibility complex class II expression on porcine vascular endothelial cells by an HMG-CoA reductase inhibitor

BACKGROUND: Vascular endothelial cells of man and pig, but not rodents, strongly express major histocompatibility complex (MHC) class II antigens in vivo, probably via the inducible promoter IV of the class II transactivator. There is abundant in vitro evidence that MHC class II positive vascular endothelial cells can activate T cells. Peripheral antigen presentation by endothelial cells is potentially important for organ-specific immunity, for allograft rejection, and possibly for immune responsiveness in general. Given the reported effects of statins on promoter IV of the class II transactivator, we evaluated in vivo expression of MHC class II antigens in pigs treated with atorvastatin calcium. METHODS: Pigs were given 3 mg/kg/day of atorvastatin orally daily for 16 days, and then killed 24 hr after the last dose. Heart, kidney, and liver were removed for immunohistological and quantitative absorption analysis. RESULTS: Double-labeling studies using immunofluorescence on frozen section for Factor VIII and MHC class II showed a marked suppression of MHC class II on vascular endothelial cells in all 4 treated pigs, in comparison with untreated pigs. This was confirmed using immunoperoxidase techniques on frozen sections. Quantitative absorption analysis showed up to 25-fold reduction in MHC class II expression. CONCLUSIONS: Statins substantially suppress endothelial cell MHC class II expression in vivo. This is likely to inhibit organ-specific immune responses, and possibly also general immune responsiveness. In a transplantation setting, in addition to other regulatory effects on the recipients immune system, statins might reduce the long-term capacity of the donor organ to activate rejection mechanisms.     

Expression of class I and class II major histocompatibility antigens in normal and transplanted human heart

The expression of monomorphic determinants of class I and class II histocompatibility antigens in human heart before and after cardiac transplantation was examined using monoclonal antibodies and an immunoperoxidase technique. Thirty-five biopsy specimens were examined. Seven were from nontransplanted hearts and 28 were from patients who had received orthotopic cardiac transplantation 5-270 days previously. In normal hearts no class I was found on the surface of the myocardium, but intercalated discs were occasionally stained and interstitial structures (blood vessels and discrete mononuclear cells) strongly expressed the antigen. In contrast, of the 28 specimens from transplanted patients, 24 expressed class I antigen strongly on the outer surface and intercalated discs of the myocardium as well as on the interstitial structures. Class II antigen was not found on the myocardium of normal heart. It was present on small capillaries, the endothelia of some large capillaries, and discrete mononuclear cells. In 20 of 25 samples from transplant patients there was increased expression of class II antigen on interstitial structures but not on myocardium. The increased expression of class I antigen on the myocardium following transplantation may make it a potential target for cytotoxic T cells. The relevance of increased expression of class I and class II antigens in response to transplantation is discussed.     

Non specific increased expression of class I major histocompatibility complex (MHC) antigens on rat liver grafts

Major histocompatibility complex (MHC) antigens play a major role in the rejection reaction and their increased expression may increase the host response to the foreign graft [1]. Several clinical [2–5] and experimental studies [6, 7] have demonstrated increased expression of MHC antigens on the different cell components of liver allografts during rejection. However modified expression of MHC antigens may also occur in certain liver diseases [8–10], after cholestasis [11] or on a regenerating liver [11]. In this experimental study in inbred rats, we compared the expression of MHC antigens on liver cells during rejection and non-immunological situations (cholestasis, cytolysis, regeneration).