Expression of MHC class I and class II antigens in colonic carcinomas

Malignant and non-malignant ('normal') colonic tissues from patients with colonic carcinoma were examined for the expression of MHC class I and class II antigens by immunoenzymatic staining using monoclonal antibodies. The amount of class I antigen as detected by 2 monoclonal antibodies, FMC 16 or W6/32 was clearly diminished in 11 of 14 tumours when compared to the amount present on 'normal' colonic tissue from the same individual. The loss of class I antigen did not correlate with tumour stage or differentiation. The reactivities of FMC 16 and W6/32 with these tissues were not identical, which indicates that the 2 monoclonal antibodies may recognize different epitopes on the HLA class I molecule. Class II antigens were absent from 'normal' colonic epithelium but were present on 20 of 28 tumours, with DR being detected more often than DP, and DQ found only on 4 of 28 tumours. When present, staining for class II antigens was heterogeneous within the tumour, in that all tumour cells did not stain equally. DR and DP antigens were found more often on moderately or poorly differentiated adenocarcinomas and on stage B, C and D tumours in that order of frequency. Thus tumours with a better prognosis were less likely to express DR and DP. The expression of DQ was unrelated to staging or differentiation.     

Expression of MHC class I and class II antigens in primary breast carcinomas and synchronous nodal metastases

Expression of the major histocompatibility (MHC) class I and class II antigens was studied by immunohistochemistry in a series of 70 primary breast carcinomas and in nodal metastases. In particular, the expression of class I (HLA A-B-C) and class II (DP, DQ and DR) molecules was compared in: a) primary breast cancers devoid of nodal metastases (n = 36) and tumors exhibiting metastatic deposits (n = 34) at the time of surgery, and b) primary breast carcinomas and their corresponding synchronous axillary nodal metastases. Reduced or absent HLA A-B-C antigen expression was seen in approximately 54.3% of primary breast carcinomas, whereas a partial or complete induction of class II products was observed in 18.5% (DQ), 30% (DP) or 48.5% (DR) of the same cases. An almost complete overlap of antigen expression was observed in breast tumors in which no metastases were found by histological examination of axillary nodes and in neoplasms showing histologically-diagnosed synchronous metastases. The reactivity for class I and class II antigens in nodal metastases roughly paralleled that exhibited by corresponding primary tumors. A discordant expression was seen in 11 cases (32%) stained for HLA A-B-C and in 8 (24%), 7 (21%) and 6 (18%) cases assayed for DP, DQ and DR products, respectively. When a discordant expression was detected, either decreased or increased staining patterns were observed in metastases. The finding of overlapping MHC antigenic profiles in the majority of primary breast tumors and nodal metastases casts doubts on the hypothesis that loss of MHC antigens can play an important role in the seeding and growth of metastatic breast carcinoma cells.     

Expression of MHC class I and class II antigens in pancreatic adenocarcinomas

The antigens encoded by the major histocompatibility complex (MHC) are cell surface glycoproteins that play a fundamental role in the regulation of the immune response. Anomalous MHC expression in tumor cells has been viewed as an important feature to escape tumor recognition by immune cells. Low or absent MHC class I expression as well as ectopic MHC class II expression have been often observed to correlate with high grade malignancy and metastatic potential in a variety of human cancers. To date, very little investigation of MHC (HLA in man) class I and class II expression inhuman pancreatic cancer has been reported. We investigated this aspect on frozen sections of 8 pancreatic adenocarcinomas and 18 established in vitro cell lines. HLA class I was expressed in all but two cancers whereas de novo HLA class II expression was detected in 3 of 8 cancers. Interestingly, a hierarchy in the expression of the various subsets of HLA class II was found with HLA-DR>-DP>-DQ. Results on cell lines strongly resembled the ones obtained in cancer tissues. However, a peculiar feature was observed in certain cell lines. HLA class II antigens were expressed in only a few cell lines and in some of them a mixed population of positive and negative cells was found. Sorting and cloning of the two populations confirmed the existence of tumor cell clones with stable and distinct HLA class II phenotype. Taken together, these results indicate the cellular heterogeneity of pancreatic cancer cells with regard to the qualitative and quantitative expression of major histocompatibility complex genes, and may provide new insights for a better understanding of the tumor-host relationships in this extremely severe form of neoplasia.     

MHC class II antigens on canine bronchoalveolar cells

To evaluate the expression of MHC (major histocompatibility complex) antigens on canine bronchoalveolar cells (BAC), bronchoalveolar lavages (BAL) were performed in mongrel and German shepherd dogs. MHC class II antigens on canine BAC and peripheral blood mononuclear cells (PBMC) were detected by monoclonal antibodies (mAbs) B1F6, 7.5.10.1 and Q5/13 recognising canine MHC class II antigens, using cytofluorometry. These mAbs reacted with more than 20% of BAC and PBMC in both breeds of dog. The percentage of MHC class II positive cells in BAC were lower than those in PBMC. There was no significant difference in the percentages of MHC class II positive BAC and PBMC in mongrel and German shepherd dogs. To further identify the expression of MHC class II antigens on BAC, the cells were separated into adherent and nonadherent cells by petri dish adherence. The percentages of MHC class II positive cells in adherent and non-adherent cell populations were similar. Nearly half the lymphocytes in normal BAC were T cells detected by mAbs F3-20-7 and 1A1; B cells were scarce and represented less than 10% of nonadherent cells. Immunoprecipitation by anti-MHC class II mAbs, and SDS-polyacrylamide gel electrophoresis (SDS-PAGE) revealed MHC class II-like molecules on canine BAC and PBMC. After stimulation with phytohaemagglutinin (PHA), the percentages of class II positive cells in BAC and PBMC were significantly increased. Thus, these anti-MHC class II mAbs may prove to be of advantage in experiments designed to evaluate the changes in class II antigen expression on canine BAC during the course of immune response in the lung, as in pulmonary allograft rejection.     

Techniques used to define human MHC antigens: monoclonals, class I and class II biochemistry

A variety of straightforward biochemical techniques have been applied to the analysis of HLA molecules. These techniques centre largely on two components: monoclonal antibodies (mAbs) and gel analysis systems. Although only the basic applications are discussed here, the techniques have broad applications. Monoclonal-gel systems may be used for the characterisation of polymorphisms in HLA class I alpha chains or class II alpha and beta chains. This requires the use of mAbs which are locus-specific and monomorphic. Alternatively, B lymphoblastoid cell lines (B-LCL) whose HLA antigen mobilities are known in advance may be used for screening uncharacterised monoclonals for locus specificity.     

Equine T lymphocytes express MHC class II antigens

Six anti-HLA class II mouse monoclonal antibodies (mAbs) were used in conjunction with a rat monoclonal antibody raised against horse lymphocytes to define class II major histocompatibility complex (MHC) molecules in the horse. By utilizing an ELISA assay and complement dependent lymphocytotoxicity assay, five out of the six anti-HLA class II antibodies and the rat anti-horse monoclonal antibody were found to react with a high percentage of peripheral blood lymphocytes. Flow cytometry demonstrated a variable antigen density on peripheral blood lymphocytes and clear evidence for expression by lymphocytes that carried no detectable surface immunoglobulin. None of the antibodies reacted with equine platelets. The mAbs immunoprecipitated an antigenic complex of Mr 29,000-33,000 from horse lymphocytes. It appears that the distribution of MHC class II antigens in the horse is different from that in man but is similar to that in the dog, since MHC class II antigens are expressed on resting peripheral blood lymphocytes which lack membrane-bound immunoglobulins. Correlations between the distribution of MHC class II antigens on lymphocyte subpopulations and their role in immunological phenomena may contribute to our understanding of the functional properties of these molecules.     

Signal transduction via MHC class II antigens on B lymphocytes

The role of the MHC class II antigens in the activation of resting human B lymphocytes (B-Go) was examined with respect to both early and late events in the activation process. The (Ca2+)i induced by anti-IgM was enhanced in the presence of, or following pre-incubation with, an anti-MHC class II DR antibody (D1.12). Pre-incubation with a sepharose conjugated antibody (Seph.-D1.12) augmented the proliferation of B-Go in response to a sub-optimal concentration of anti-IgM. The 2D PAGE profile of B-Go differed from that of in vivo activated B lymphocytes. The 2D PAGE profile of B-Go activated by Seph.-D1.12 was not identical to the profile of B-Go activated by either anti-IgM or PMA. These data suggest that the activation of B-Go via the class II antigens shares part of the pathway of anti-IgM induced activation but does not follow an identical pathway.     

Biochemical analysis of class I and class II MHC antigens in cynomolgus macaques by one-dimensional isoelectric focusing

To obtain a better estimate of major histocompatibility complex (MHC) polymorphism in the Old World macaque, Macaca fascicularis, class I and class II MHC proteins from 42 animals were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and one-dimensional isoelectric focusing (1D-IEF). The panel represented both related and unrelated animals with a total of at least 30 serologically distinct haplotypes. Cells were sequentially immunoprecipitated with monoclonal antibody (mAb) W6/32 for class I and with mAb L243 for class II molecules, followed by neuraminidase treatment. Both sets of immunoprecipitates yielded 5-7 major bands on IEF. All bands present in offspring were present in at least 1 parent. Siblings which were serologically identical for class I and which were non-stimulatory in mixed lymphocyte culture (MLC) yielded identical IEF patterns for both class I and class II; in other sibling pairs which were serologically identical for class I antigens (Ag), IEF produced convincing evidence that the siblings were indeed nonidentical, or helped to verify that recombination had occurred within the MHC. Specific bands were found which correlated with class I specificities A8, A24, and B25 previously defined by serology. Comparison of serological and biochemical data will broaden our understanding of the MHC in Macaca fascicularis and will increase the potential use of this species in transplantation research, as a model of disease, and for comparative studies.     

Presentation of cytosolic antigens via MHC class II molecules

Major histocompatibility (MHC) class II molecules function to present antigenic peptides to CD4 T lymphocytes. The pathways by which these molecules present exogenous antigens have been extensively studied. However by contrast, far less is known about the processing and trafficking of cytosolic antigens, which can also serve as an alternative source of ligands for MHC class II molecules. Self-proteins, tumor antigens, as well as viral proteins found within the cytosol of cells, can be presented via MHC class II molecules, resulting in the activation of specific CD4 T cells. Studies have begun to reveal unique steps as well as some similarities in the pathways for cytosolic and exogenous antigen presentation. Recent developments in this area are summarized here.     

Dog MHC class I antigens contain beta 2-microglobulin

Detergent-solubilized DLA class I antigens have been isolated by immunoprecipitation from peripheral blood lymphocytes with an anti-beta 2-microglobulin serum and an alloantiserum. Amino acid composition and NH2-terminal amino acid sequence analysis confirmed that the DLA light chain is identical to dog urinary beta 2-microglobulin, and that the DLA heavy chain shows homology to the major histocompatibility complex class I heavy chains of other species.     

Distribution of class I and class III MHC antigens in the Tarasco Amerindians

Class I and class III major histocompatibility complex (MHC) antigen frequencies were analyzed in 130 haplotypes from 33 families belonging to a group of Amerindians culturally and linguistically isolated for more than 12 centuries in Mexico: the Tarascos. The most frequent antigens in this ethnic group of the HLA-A locus are: A2 (gf 0.353), A24 (gf = 0.223), A31 (gf = 0.184), and A28 (gf = 0.161); and the most frequent of the HLA-B locus are: B35 (gf = 0.230), B39 (gf = 0.192), B15 (gf = 0.146), and B5 (gf = 0.123). On the other hand, class III antigens demonstrated relatively high frequencies of the SC31 (frequency = 0.561), SC01 (frequency = 0.076), and SC42 (frequency = 0.069) complotypes. Also important was the relatively high frequency of the HLA-B27 antigen (gf 0.061) and the SC33 complotype (frequency = 0.046), which are either absent or found infrequently in other Amerindian groups. Analysis of MHC haplotypes revealed that four of them have relatively high frequencies, these were the following: [B39;SC31] (11.6%), [B35;SC31] (11.6%), [B15;SC31] (8.0%), and [B5;SC31] (5.8%). Other MHC haplotypes had frequencies lower than 5.0%. The decreased frequency of BF alleles other than BF*S and the presence of the SC33 and SC32 complotypes suggest long time preservation from genetic admixture. This information withstands the basis for population genetic analysis and disease association studies in Mexican mestizos.     

Activation by interferon-gamma of expression of ICAM-1 and MHC class II antigens in tumour cells from colorectal carcinomas

Aims-To determine whether lack of MHC class II antigen and intercellular adhesion molecule-1 (ICAM-1) expression in some tumours is due to the inability of the tumour cells to respond to the cytokine interferon-gamma (IFN-gamma), an important activator of these surface molecules.Methods-Cells from 40 colorectal tumours which did not constitutively express class II MHC antigens or ICAM-1 were kept in short term culture after disaggregation for a few days to two weeks without significant loss of viability. These were treated with IFN-gamma. Expression of class II MHC antigens and ICAM-1 was determined using immunohistological techniques.Results-There was clear induction in vitro of both MHC class II antigens and ICAM-1 in cells from eight of the tumours, with between 50 and 80% of the tumour cells in the cultures staining positively. The staining was apparent within 24 hours, appeared maximal at about three days, and declined thereafter. There were no obvious differences in cell morphology or viability between the cultures which were inducible and those which were not, nor were there obvious differences between the tumours from which they were derived.Conclusions-Expression of MHC class II antigens and ICAM-1 may be induced by IFN-gamma in a small proportion of colorectal tumours which do not constitutively express these antigens, showing that only a minority of tumours are capable of responding to this cytokine.     

Identification of woodchuck class I MHC antigens using monoclonal antibodies

Abstract: Two class I major histocompatibility complex (MHC) proteins with molecular masses of 43- and 39-kDa were identified in the cell surface membranes of normal woodchucks using a newly developed anti-woodchuck class I monoclonal antibody (mAb) B1b.B9 and immuno-blotting. B1b.B9 was generated by immunizing mice with viable wood-chuck peripheral blood mononuclear cells and was selected for anti-class I MHC reactivity using a cellular enzyme–linked immunoassay, indirect immunofluorescence on tissue sections and flow cytofluorimetry. The distribution pattern of class I MHC antigen on woodchuck lymphoid cells was found to be similar to that reported in other species. Also, the antigen expression on normal woodchuck hepatocytes was comparable to that observed on normal human liver parenchymal cells; thus, the antigen was not detected on hepatocytes by staining of liver tissue sections, but was found by indirect immunofluorescence staining of isolated liver cells. Western blot analysis of the plasma membranes from normal woodchuck hepatocytes revealed the presence of a single species of class I MHC heavy chain protein with a molecular mass of 43-kDa, whereas splenocyte plasma membranes showed intense expression of a 43-kDa species, as well as the presence of a 39-kDa protein. The 39- and 43-kDa proteins were extracted with Triton X-114 to the hydrophobic protein phase, suggesting that they both contain a hydrophobic transmembrane domain. The data obtained indicate that the B1b.B9 identifies a nonpolymorphic epitope of woodchuck class I MHC heavy chains, providing an important reagent for the study of the pathogenesis of hepatitis B virus infection in a woodchuck model.     

Processing of bacterial antigens for peptide presentation on MHC class I molecules

Summary: Professional antigen-presenting cells (pAPC) can process and present exogenous antigens on major histocompatibility complex class 1 (MHC-I) molecules. This unusual pathway for antigen presentation may represent a physiologically important step in the course of priming and tolerance induction of CD8⁺ T cells. In addition, it may play an important role in immunological surveillance for pathogens that survive in vacuolar compartments in APC. The goal of the present review is to discuss recent studies on the processing of bacterial-derived antigens for presentation on MHC-1 molecules. The antigen presentation emphasized will include bacteria that remain confined in vacuolar compartments. This is in contrast to antigens derived from bacteria that have intrinsic properties allowing translocation across membranes and access into the classical MHC-I presentation pathway In particular, presentation of bacterial antigens by dendritic cells (DC) will be emphasized, and MHC-I presentation of antigens derived from apoptotic cells, particularly cells induced to undergo apoptosis by microbial infection, will be presented. Finally, some special aspects of the interaction between bacteria and DC will be discussed as ii relates to DC maturation, antigen presentation and T-cell stimulation.