Analysis of the role of MHC class II presentation in the stimulation of cytotoxic T lymphocytes by antigens targeted into the exogenous antigen-MHC class I presentation pathway

By conjugation of proteins to beads, Ags can be selectively targeted into the MHC class I pathway of phagocytes in vivo and can stimulate CTL responses. Because phagocytes also present particulate Ag on MHC class II molecules, we examined whether these Ags stimulated concomitant CD4 T cell immunity. Although the priming of CD4 T cells with soluble OVA required adjuvants, particulate Ag was stimulatory when injected in saline. We next examined whether CD4 T cell responses played a role in the generation of CTL to particulate Ag. At low concentrations of Ag, OVA primed CTLs in wild-type mice but not in MHC class II-deficient animals, indicating that MHC class II presentation of Ag was essential for CTL generation. These data both support a model where CD4 T cells collaborate with CTLs as part of a three-cell interaction and identify a phagocyte as the third cell in this reaction. Interestingly, injection of higher concentrations of the same Ag primed equivalent CTL responses in both wild-type and MHC class II-deficient mice. These results indicate that a key variable in determining whether CTL generation is helper cell dependent or independent is the dose of immunogen. This may explain in part why CTL responses to abundant Ags, such as viruses, tend to be helper independent, while responses to less abundant Ags, such as minor histocompatibility Ags, require T helper cells. In addition, these results also point to the potential of using particulate Ags to prime or boost responses in settings with CD4 immunodeficiency.     

Human cytomegalovirus-infected cells have unstable assembly of major histocompatibility complex class I complexes and are resistant to lysis by cytotoxic T lymphocytes

Viruses which cause persistence in the naturally infected host are predicted to have evolved immune evasion mechanisms. Human cytomegalovirus (HCMV) causes significant morbidity and mortality in immunocompromised patients yet persists without clinical manifestations in seropositive individuals who have normal immune function. We report that HCMV infection in vitro impairs major histocompatibility complex class I (MHC-I) assembly accompanied by resistance to killing by cytotoxic CD8+ T lymphocytes. Pulse-chase metabolic labelling experiments show that MHC-I complexes continue to be assembled by both uninfected and HCMV-infected cells. However, MHC-I molecules are unstable in HCMV-infected cells and are rapidly broken down. Endoglycosidase H treatment of immunoprecipitates indicates that the breakdown of MHC-I complexes in HCMV-infected cells occurs primarily in a pre-Golgi compartment. Interference with normal MHC-I assembly and expression, if relevant in vivo, may have implications for the restriction of the diversity of the CD8+ cytotoxic T lymphocyte repertoire directed against HCMV antigens and may be an important mechanism of viral persistence.     

Pretransplant cytotoxic donor T-cell activity specific to patient HLA class I antigens correlating with mortality after unrelated BMT

Unrelated bone marrow transplantation (BMT) is associated with increased post-transplant complication rates, partly because more transplantation antigens are mismatched than in HLA-identical related BMT. We have shown previously that the cytotoxic T-lymphocyte precursor (CTLp) test performed before transplantation specifically detects HLA class I mismatches demonstrating its usefulness for the identification of new HLA class I alleles. In this study we analysed the clinical relevance of the CTLp test in 41 patients who underwent unrelated BMT between 1990 and 1994. All patient-donor pairs were HLA-A, -B, -DR compatible as defined by AB-serology and oligotyping for DR1-14. The host-reactive CTLp test was performed using previously frozen peripheral blood mononuclear cells (PBMC) as stimulators and PHA blasts as target cells. We found 10 CTLp-positive and 31 CTLp-negative patient-donor pairs. Between the two groups there were no significant differences for age, diagnosis, sex, preconditioning and GvHD prophylaxis. The clinical results for the CTLp positive and the CTLp negative transplants were: severe acute GvHD III-IV 67% and 26% (P = 0.0315), transplant-related mortality 60% and 26% (P = 0.0085), and patient survival at 3.5 years 10% and 54% (P = 0.0006). Seven patient-donor pairs were mismatched for HLA-DR and/or -DQ subtypes. Only one of these seven class II mismatched pairs had a positive CTLp test. In the remaining nine CTLp positive pairs the CTL reactivity was directed against HLA-A, -B or -C antigens, revealing a statistically significant (P < 0.005) correlation between the CTLp frequency and HLA class I matching. In conclusion, the CTLp test helped to select optimally matched bone marrow donors and was particularly useful in association with high resolution oligotyping for DR- and DQ-subtypes for precise matching of both classes of HLA antigens.     

An acidic modification of the cytoplasmic domain contributes to the charge heterogeneity of the MHC class I antigens

Pulmonary atresia with intact ventricular septum (PAIVS) is a rare congenital cardiac anomaly that has been classified into two types: one is a more frequent type having dysplasia of tricuspid valve (TV) with a small annulus, underdeveloped right ventricle (RV) with a hypoplastic cavity and a hypertrophic wall; the other type has severe dysplasia of TV and dilatation of RV, right atrium (RA), and right atrioventricular junction with thinning of the RV wall. We performed a morphologic study on 11 autopsied hearts with PAIVS, giving particular emphasis to the variation of morphology of the TV. We could classify these hearts into 3 groups according to the degree of right ventricular development. In the first group of 7 cases (type I), the RVs were underdeveloped. Thick leaflets, restricted valve apparatus with short chordae, and small annuli were characteristics of the TV. In the second group of 3 cases (type II), the RVs showed marked enlargement of the cavity and thinning of the wall. The TV showed redundant, dysplastic, sail-like anterior leaflets, and the downward displacement of septal leaflet and/or posterior leaflet, which are the findings frequently observed in Ebstein's malformation. The RVs were dilated and with partially unguarded tricuspid orifice. The septal leaflet of the TV was dysplastic and, in two cases, the septal leaflet showed chordal structure at the upper surface facing the RA, which is a peculiar finding that has not been described in the literature. The remaining case was a heart with a moderately developed RV (type III). The TV showed mildly dysplastic appearance and we classify this as a separate type, because we could expect the best surgical results in this type. This type had optimal size of RV and the mildest degree of dysplasia of TV. In PAIVS, the morphology of TV correlates well with the type of the right ventricular development.     

De novo expression of MHC class I and class II antigens on endomyocardial biopsies from patients with inflammatory heart disease and rejection following heart transplantation

Inflammation of the heart muscle is caused either by infection (i.e. coxsackie virus) resulting in myocarditis or by rejection following heart transplantation. These processes induce activation of the immune system. We examined endomyocardial biopsies from patients with myocarditis, perimyocarditis and rejection following heart transplantation and compared these to biopsies from patients with coronary artery disease. The biopsies were examined immunohistologically with specific monoclonal antibodies against class I and class II molecules of the major histocompatibility complex (MHC). MHC class I antigens on the normally negative myocytes were evident in myocarditis (38%) and in rejection after heart transplantation (68%). In the interstitium there was an increase of both MHC class I and class II antigens. MHC class II antigens, however, were never seen on myocytes. MHC class I antigens are required for the action of CD 8 positive cytotoxic T cells. Therefore myocytes which express MHC class I antigens are susceptible to cytotoxic effects of the immune system. MHC class II antigens are essential to T helper cells. By cytokine release, activated T helper cells play a central role in the initiation, regulation and mediation of an immune response in myocarditis and rejection following heart transplantation.     

In vitro induction of naive cytotoxic T lymphocytes with complexes of peptide and recombinant MHC class I molecules coated onto beads: role of TCR/ligand density

We previously reported that complexes of peptide with soluble single-chain recombinant MHC (SC-MHC) class I molecules are able to induce cytotoxic T lymphocytes (CTL) in vitro in a murine system with an efficiency comparable to that observed with peptide-pulsed dendritic cells as antigen-presenting cells. In this report, we have assessed the capacity of preformed peptide/SC-Kd complexes in monomeric or dimeric form as well as of peptide/SC-Kd-loaded beads to generate in vitro specific CTL responses from naive DBA/2 spleen cells. Peptide/SC-Kd-coated beads were consistently more efficient. We evaluated the role of costimulatory molecules, using monoclonal antibodies anti-CD80 or anti-CD86. In addition, the capacity of peptide/SC-Kd-coated beads to generate a CTL response from purified naive CD8+ T cells was ascertained. Taken together, the results indicate that, under our conditions, CTL priming does not require the participation of co-stimulatory molecules and is the consequence of a direct interaction between the cognate TCR on peptide-specific CTL precursors and the peptide/SC-Kd-loaded beads. Titration of the amount of preformed complexes of SC-Kd and peptide 170-179 of HLA-CW3 that need to be coated onto the beads to prime CTL precursors shows an activation threshold which can be calculated to be between 25000 and 50000 complexes. In effect, in cultures stimulated with specific peptide CW3/SC-Kd complexes representing less than 50% occupancy of the total (10(5)) complexes on the beads, no peptide-specific cytolytic activity was observed. These results suggest that the efficiency of the primary CTL induction depends on the density of specific peptide/SC-Kd complexes present on the beads.     

Tumors with reduced expression of a cytotoxic T lymphocyte recognized antigen lack immunogenicity but retain sensitivity to lysis by cytotoxic T lymphocytes

A murine solid tumor was transfected to express various levels of an allogeneic major histocompatibility complex class I gene (K216), in order to test the effect of the level of antigen expression on immunogenicity and sensitivity to lysis by cytotoxic T lymphocytes (CTL). The growth rates of clones of tumor cells expressing different levels of the transfected gene were similar in vitro and in nude mice. Although all tumor cells, including cells freshly isolated from growing tumors, were equally sensitive to lysis by specific CTL, only tumor cells expressing the highest level of the K216 antigen stimulated CTL and were rejected by normal mice. In contrast, tumor cells expressing lower levels of antigen failed to immunize for CTL and grew progressively in normal mice, despite retaining expression of the transfected gene and remaining fully sensitive to CTL-mediated lysis; thus, the threshold of antigen needed to stimulate CTL responses was considerably higher than that needed to lyse tumor cells. Reduction of K216 antigen expression from 100-fold to 40-fold above background, impaired significantly the ability of the tumor cells to induce a K216-specific immune response, while tumor cells expressing K216 at levels 2-fold above background were as susceptible to CTL-mediated lysis as tumor cells expressing 50-fold more antigen. The important implication of these findings is that some tumors occurring in nature may not be immunogenic but nevertheless express antigens which are potential targets for immune therapy.     

The susceptibility to natural killer cell-mediated lysis of HLA class I-positive melanomas reflects the expression of insufficient amounts of different HLA class I alleles

NK cells selectively lyse tumor cells which do not express one or more MHC class I alleles. The ability to discriminate between self normal or tumor cells is due to the expression of MHC class I-specific killer inhibitory receptors (KIR). In the present study we analyzed melanoma cell lines which were highly susceptible to NK cell-mediated lysis in spite of the expression of a complete set of HLA class I alleles. Quantitative analysis of the HLA class I expression using allele-specific monoclonal antibodies (mAb) revealed a down-regulation of all HLA class I molecules. Treatment of melanoma cells with IFN-gamma resulted in up-regulation of all HLA class I alleles that was paralleled by the acquisition of resistance to lysis. That resistance to lysis reflected the up-regulation of HLA class I molecules was revealed by the finding that mAb-mediated masking of either KIR or their HLA class I ligands completely restored the melanoma cell lysis. These results were obtained by the use of selected NK cell clones derived either from allogeneic or autologous donors. In addition, similar results were obtained using in vitro expanded autologous NK cell populations. Our data indicate that NK cells can lyse not only melanoma cells which have lost the expression of one or more HLA class I alleles but also cells expressing a decreased amount of class I molecules.     

Development and antigen specificity of CD8+ cytotoxic T lymphocytes in beta 2-microglobulin-negative, MHC class I-deficient mice in response to immunization with tumor cells

beta 2-Microglobulin knockout mice (beta 2-m-/-) with MHC class I expression deficiency are able to develop functional TCR(+)-alpha beta, CD8+ CTLs in response to tumor cell injection. The i.p. injection of beta 2-m-/- mice with tumor results in the massive accumulation of highly lytic CD8+ CTLs in the peritoneum and causes the local recruitment of CD8+ T cells into lymph nodes and spleens of immune animals. The accumulation of CD8+ CTLs in peritoneum is accompanied by the rejection of tumor cells and the survival of animals. The deficiency in MHC class I expression in beta 2-m/- mice is reflected in the delayed tumor rejection and CD8+ cell accumulation during the primary anti-tumor response in comparison with normal mice. The secondary response, however, is identical in normal and MHC class I-deficient mice. The rejection of tumor cells appears to be MHC class I directed because no rejection of tumors, no accumulation of CD8+ CTLs, and no survival of animals were observed when syngeneic tumor cells were used for injection with the notable exception of anti-minor Ag response. The Ag specificity of CD8+ CTLs in beta 2-m-/- mice is demonstrated using a panel of tumor target cells and class I transfectants. Although no substantial differences were found in the number and specificity of peritoneal CD8+ CTLs in beta 2-m-/- and normal mice using tumor rejection studies, the analysis of TCR-V beta phenotype using the panel of mAbs revealed the reduction in proportion of TCR-V beta 5 and TCR-V beta 6 used by CD8+ cell population from beta 2-m-/- mice. Development of lytic and H-2-directed CD8+ cells in regional lymph nodes was also observed after footpad immunization of beta 2-m-/- mice with TNP-labeled C57BL/6 splenocytes, suggesting anti-minor Ag reaction.     

Interaction of murine MHC class I molecules with tapasin and TAP enhances peptide loading and involves the heavy chain alpha3 domain

In human cells the association of MHC class I molecules with TAP is thought to be mediated by a third protein termed tapasin. We now show that tapasin is present in murine TAP-class I complexes as well. Furthermore, we demonstrate that a mutant H-2Dd molecule that does not interact with TAP due to a Glu to Lys mutation at residue 222 of the H chain (Dd(E222K)) also fails to bind to tapasin. This finding supports the view that tapasin bridges the association between class I and TAP and implicates residue 222 as a site of contact with tapasin. The inability of Dd(E222K) to interact with tapasin and TAP results in impaired peptide loading within the endoplasmic reticulum. However, significant acquisition of peptides can still be detected as assessed by the decay kinetics of cell surface Dd(E222K) molecules and by the finding that prolonged viral infection accumulates sufficient target structures to stimulate T cells at 50% the level observed with wild-type Dd. Thus, although interaction with tapasin and TAP enhances peptide loading, it is not essential. Finally, a cohort of Dd(E222K) molecules decays more rapidly on the cell surface compared with wild-type Dd molecules but much more slowly than peptide-deficient molecules. This suggests that some of the peptides obtained in the absence of an interaction with tapasin and TAP are suboptimal, suggesting a peptide-editing function for tapasin/TAP in addition to their role in enhancing peptide loading.     

1,25-dihydroxycholecalciferol enhances the expression of MHC class II antigens and intercellular adhesion molecule-1 by human renal tubular epithelial cells

PURPOSE: Beside its role in calcium and phosphorus metabolism, 1,25-dihydroxycholecalciferol (1,25-D3) exerts multiple effects on cytokine and major histocompatibility complex (MHC) class II expression in monocytes and lymphocytes. In different renal diseases tubular epithelial cells express MHC class II molecules and cell adhesion molecules,, such as intracellular adhesion molecule-1 (ICAM-1). Therefore, modulation of MHC class II and ICAM-1 expression in renal tubular epithelial cells by 1,25-D3 may be relevant to lymphocyte adhesion to tubular epithelial cells and immune mediated renal injury. However, the expression of MHC class II antigens and cellular adhesion molecules by renal tubular epithelial cells in response to 1,25-D3 has not been investigated. MATERIALS AND METHODS: We generated human renal tubular epithelial cells and SV40 transfected tubular epithelial cells to investigate immune modulation of 1,25 on renal epithelial cells. Major histocompatibility complex class II molecules and ICAM-1 were detected by a specific enzyme linked immunoassay. RESULTS: We found a dose-dependent increase of both constitutive and induced MHC class II and ICAM-1 expression in tubular epithelial cells stimulated with 1,25-D3. Dose-dependent stimulation of MHC class II and ICAM-1 expression was not restricted to primary human renal tubular epithelial cells but was also detected in SV40 transfected cells. CONCLUSIONS: Expression of MHC class II and ICAM-1 is crucial for antigen presentation by and lymphocyte adhesion to renal tubular epithelial cells. Modulatory effects of 1,25-D3 on immune accessory function of renal tubular epithelial cells may be of clinical significance in renal diseases.     

Regulation of MHC class I heterodimer stability and interaction with TAP by tapasin

Major histocompatibility complex (MHC) class I molecules are heterodimers of a class I heavy chain and beta 2-microglobulin that bind peptides supplied by the MHC region-encoded transporters associated with antigen processing (TAP). Peptide binding by class I heterodimers is necessary for their maturation into stable complexes and is dependent on their physical association with TAP. In human mutant 721.220 cells, however, a novel genetic defect causes the failure of class I heterodimers to associate with TAP. This deficiency correlates with lack of expression of a glycoprotein, tapasin (TAP-associated glycoprotein), which has been found in association with class I heterodimers and TAP. Employing a transcomplementation analysis, we obtained evidence co-localizing the genetic defect of mutant 220 cells and the structural or a regulatory gene controlling the expression of tapasin on the short arm of chromosome 6, which includes the MHC. Expression of tapasin and the normal interaction of class I heterodimers with TAP are concomitantly restored, indicating the probable function of tapasin as a physical link between these complexes. In further support of this model, the absence of tapasin in mutant 220 cells correlates with reduced class I heterodimer stability, suggesting that tapasin may stabilize class I heterodimers and thereby enhance their association with TAP. These results further implicate tapasin in a mechanism that promotes peptide binding by class I heterodimers through their interaction with TAP.     

Fc epsilon receptor I on dendritic cells delivers IgE-bound multivalent antigens into a cathepsin S-dependent pathway of MHC class II presentation

In this study, we elucidate the Fc epsilon RI-mediated Ag uptake and presentation mechanisms of dendritic cells (DC). We found that Fc epsilon RI-bound IgE, after polyvalent but not after monovalent ligation, is efficiently internalized into acidic, proteolytic compartments, degraded, and delivered into organelles containing MHC class II, HLA-DM, and lysosomal proteins. To follow the fate of the fragmented ligand, we sought to interfere with invariant chain (Ii) degradation, a process critical for peptide loading of nascent MHC class II molecules. We found DC to express cathepsin (Cat) S, a cysteine protease involved in Ii processing by B cells. Exposure of DC to a specific, active-site inhibitor of Cat S resulted in the loss of anti-Cat S immunoreactivity, led to the appearance of an N-terminal Ii remnant, and decreased the export of newly synthesized MHC class II to the DC surface. Furthermore, inactivation of Cat S as well as blockade of protein neosynthesis by cycloheximide strongly reduced IgE/Fc epsilon RI-mediated Ag presentation by DC. Thus, multimeric ligands of Fc epsilon RI, instead of being delivered into a recycling MHC class H pathway, are channeled efficiently into MIIC (MHC class II compartment)-like organelles of DC, in which Cat S-dependent Ii processing and peptide loading of newly synthesized MHC class II molecules occur. This IgE/Fc epsilon RI-dependent signaling pathway in DC may be a particularly effective route for immunization and a promising target for interfering with the early steps of allergen presentation.     

B7-CTLA4 interaction enhances both production of antitumor cytotoxic T lymphocytes and resistance to tumor challenge

Expression of B7-family costimulatory molecules CD80 (B7-1) and CD86 (B7-2) on tumor cells enhances host immunity. However, the role of the two B7 receptors, CD28 and CTLA4 (CD152), on T cells in antitumor immune response has not been clearly elucidated. Based on the effects of anti-CD28 and anti-CTLA4 mAbs on T cell response, it was proposed that CD28-B7 interaction promotes antitumor immunity, whereas B7-CTLA4 interaction down-regulates it. A critical test for the hypothesis is whether selective engagement of CTLA4 receptors by their natural ligands CD80 and CD86 enhances or reduces antitumor immunity. Here we used tumors expressing wild-type and mutant CD80, as well as mice with targeted mutation of CD28, to address this issue. We report that in syngeneic wild-type mice, B7W (W88>A), a CD80 mutant that has lost binding to CD28 but retained binding to CTLA4, can enhance the induction of antitumor cytotoxic T lymphocytes (CTL); B7Y (Y201>A), which binds neither CD28 nor CTLA4, fails to do so. Consistent with these observations, B7W-transfected J558 plasmocytoma and EL4 thymoma grow significantly more slowly than those transfected with either vector alone or with B7Y. Optimal tumor rejection requires wild-type CD80. Moreover, expression of a high level of CD80 on thymoma EL4 cells conveys immunity in mice with a targeted mutation of CD28 gene. Taken together, our results demonstrate that B7-CTLA4 interaction enhances production of antitumor CTL and resistance to tumor challenge and that optimal enhancement of antitumor immunity by CD80 requires its engagement of both CD28 and CTLA4.     

Involvement of MHC class I molecule and ICAM-1 in the enhancement of adhesion and cytotoxic susceptibility to immune effector cells of tumor cells transfected with the interleukin (IL)-2, IL-4 or IL-6 gene

To investigate the molecular and cellular mechanisms involved in the reduced tumorigenicity and increased immunogenicity of interleukin-2 (IL-2)-, IL-4- or IL-6-gene-transfected B16 melanoma vaccine, we have analyzed the functional and phenotypic properties of these genetically engineered melanoma cells in the present study. The cytokine-gene-transfected B16 melanoma cells showed stronger adhesion to the lymphokine-activated killer (LAK) cells or cytotoxic T lymphocytes (CTL), and higher sensitivity to cytotoxicity of LAK cells or CTL. Using fluorescence-activated cell sorting analysis, we found that both MHC class I and ICAM-1 expression were increased after IL-2, IL-4 or IL-6 gene transfection. The increased level of MHC class I and ICAM-1 expression seems to be responsible for the high sensitivity of these gene-transfected B16 cells to LAK or CTL cytotoxicity because anti-(MHC class I) or anti-ICAM-1 mAb could inhibit the adhesion and cytotoxicity increment simultaneously. The CTL induction was partly inhibited by anti-ICAM-1 mAb and was completely blocked by anti-MHC class I mAb. These results suggested that the decreased tumorigenicity of IL-2-, IL- 4-, and IL-6-gene-transfected B16 melanoma cells may be partly due to the increased sensitivity to effector cell cytotoxicity mediated by increased expression of ICAM-1 or MHC class I molecules on the tumor cell surface after cytokine gene transfection.     

Adoptively transferred CD4+ lymphocytes from CD8 -/- mice are sufficient to mediate the rejection of MHC class II or class I disparate skin grafts

Recent studies revealed that CD4+ cells initiate allograft rejection through direct recognition of allogeneic MHC class II Ags and indirect recognition of MHC peptides processed by self APCs. Both pathways were shown to help CD8+ cells that eventually lysed allogeneic MHC class I-presenting targets. There was little evidence, however, that CD4+ cells are sufficient for graft rejection. We studied skin graft rejection by CD8-deficient (CD8 -/-) mice. We showed that BALB/cJ(H-2d) CD8 -/- mice could reject allogeneic skin from C57BL/6J(H-2b) mice deficient in MHC class I or in MHC class II Ags. To understand the role of CD4+ cells in this process, we isolated them from CD8 -/- mice and transferred them to BALB/cJ nude mice that had been grafted with allogeneic skin (H-2b) from animals deficient in MHC class I or MHC class II. Nude mice injected with CD4+ cells rejected MHC class II and, albeit more slowly, MHC class I disparate skins. We showed in vitro evidence that CD4+ cells were not cytotoxic toward MHC class I or MHC class II disparate targets and that they recognized MHC class I allogeneic targets through indirect recognition. CD4+ cells produced Th1 cytokines, but not IL-4, following stimulation with allogeneic cells. Furthermore, intragraft TNF-alpha was elevated in skin grafted onto nude mice reconstituted with CD4+ cells compared with nonreconstituted mice. This suggests that MHC class II- or MHC class I-guided CD4+ cells alone are sufficient to induce rejection by the generation of cytokine-induced lesions.     

Alterations in TCR-MHC contacts subsequent to cross-recognition of class I MHC and singly substituted peptide variants

Vesicular stomatitis virus (VSV) elicits H-2Kb-restricted CTLs specific for the immunodominant VSV octapeptide RGYVYQGL. To study the structural features important for interaction between the TCR beta-chain and the peptide/MHC complex, we immunized TCR alpha-chain transgenic mice with the VSV peptide and raised a panel of anti-VSV CTL clones with identical TCR alpha-chains. Consistent with our previous analysis of uncloned populations of primary CTLs, the anti-VSV CTL clones were all Vbeta13+ and expressed TCR beta-chains with highly homologous complementarity-determining region 3 (CDR3) loops. Although the clones expressed similar TCRs, they differed in their ability to cross-react with VSV peptide variants singly substituted at TCR contact positions 4 and 6. These findings allowed us to identify short stretches of amino acids in the C-terminal region of the CDR3beta loop that, when altered, modify the cross-reaction capability of the TCR to position 4 and position 6 variant peptides. To further probe the structural correlates of biologic cross-reactivity, we used cross-reactive CTL clones and cell lines expressing point mutations in H-2Kb to investigate the effect of single amino acid changes in the peptide on the pattern of recognition of the TCR for the peptide/MHC complex. Single conservative substitutions in the peptide were sufficient to alter the recognition contacts between a cross-reactive TCR and the MHC molecule, supporting the idea that the TCR can make overall structural adjustments in MHC contacts to accommodate single amino acid changes in the peptide.     

Cytomegalovirus infection enhances the neointima formation in rat aortic allografts: effect of major histocompatibility complex class I and class II antigen differences

BACKGROUND: The development of chronic rejection has emerged as a major cause of long-term graft failure. Previous studies have demonstrated that cytomegalovirus (CMV) infection is associated with an increased incidence of chronic allograft rejection in renal, cardiac, and aortic allografts. This study was designed to investigate the effects of the major histocompatibility complex (MHC) class I or class II mismatches on CMV-enhanced chronic rejection. METHODS: Aortic transplantation was performed between different inbred rat strain combinations; the Lewis to RP combination was class I-mismatched and Wag/Rij to RP class II-mismatched. At 7, 28, and 90 days after transplantation, the intensity of chronic rejection in mismatched grafts with or without CMV infection was evaluated using histological and immunohistological analysis. RESULTS: The results of this study demonstrated that CMV infection led to an increased influx of monocytes/ macrophages in class I-mismatched grafts at 1 week after transplantation and enhanced infiltration of T lymphocytes in class II-mismatched grafts at 4 weeks. Although more vascular lesions were observed in the class II-mismatched combinations, an intensified neointima formation by CMV infection was observed only in the MHC class I-mismatched allografts. CONCLUSIONS: CMV infection may increase neointima formation of allografts when an MHC class I disparity between donor and recipient is present. This may be associated with the increased perivascular influx of monocytes/macrophages observed in CMV-infected animals early after transplantation.