Amnion membrane epithelial cells express class I HLA and contain class I HLA mRNA

Amnion epithelial cells in membranes from term deliveries, which have been reported not to express histocompatibility Ag, were evaluated for HLA by using an avidin-biotin immunoperoxidase staining system and for class I HLA mRNA by Northern blotting and in situ hybridization. There were three major findings from these studies. 1) Amnion cells frequently expressed class I HLA. Three mAb to monomorphic determinants of class I HLA were used: 61D2, PA2.6, and W6/32. 61D2 identified 1 of 8 fresh amnion membranes as class I positive whereas PA2.6 identified 4/8 and W6/32 identified 5/8. 2) Amnion cells contained class I HLA mRNA. RNA extracted from amnion membranes hybridized to a class I HLA probe (pHLA1.1) in Northern blotting. In situ hybridization procedures with pHLA1.1 showed that essentially all amnion cells contained class I HLA mRNA. 3) Levels of class I HLA mRNA in amnion cells could be modulated. Exposure of amnion explants to medium containing IFN-gamma enhanced levels of class I HLA mRNA in amnion cells, whereas epidermal growth factor diminished those levels. The results suggest that amnion cells transcribe class I HLA genes and are capable of synthesizing class I H chains but that expression may be modulated by extrinsic regulatory molecules.     

HLA antibody responses in HLA class I transgenic mice

In a previous report we described how cross-immunizations of pairs of transgenic mice expressing different HLA class I antigens led to the production of antibodies directed exclusively at polymorphic epitopes. This was ascribed to self-tolerance of HLA that prevents immune responses to monomorphic epitopes and focuses responses on polymorphic ones. In the present report we extend our findings and demonstrate that immunizations of class I transgenic mice with HLA transfected mouse fibrosarcoma as well as with human lymphoblastoid cells also preferentially yield antibodies to polymorphic epitopes. This was the case whether or not immunizations were carried out across locus barriers [e.g., Tg (HLA-A *0201) or Tg (HLA-Cw*0301) transgenic mice immunized with HLA-B27 transfectants] or within the same locus [e.g., Tg (HLA-B*1302) transgenic mice immunized with HLA-B27 transfectants or B27-expressing lympho-blastoid cell]. Use of an extended immunization protocol with four or more booster injections favored antibodies of IgG isotype with affinities high enough to lyse normal peripheral blood lymphocytes (PBLs) in complement-dependent cytotoxicity assays and to immunoprecipitate HLA antigens. The specificities covered by the monoclonal antibodies (mAbs) could be either broad or narrow, depending on the genetic distance of the HLA antigens or alleles involved. For instance, a Tg(HLA-B*1302) transgenic mouse immunized with B27 produced both broad B7/B27-specific antibodies, Bw4-specific antibodies, and one antibody reacting with all B alleles except B13 and with some C alleles. On the other hand, a Tg(HLA-B*1302) transgenic mouse immunized with Bw47 transfectants responded narrowly with an antibody to Bw60 and Bw47. Thus it appears that by choosing appropriate recipient mice and closely related or more distant HLA antigens, antibodies of a programmed specificity can be generated. Address correspondence and offprint requests to: U. Hämmerling.     

HLA class I allele promiscuity revisited

The peptide repertoire presented on human leukocyte antigen (HLA) class I molecules is largely determined by the structure of the peptide binding groove. It is expected that the molecules having similar grooves (i.e., belonging to the same supertype) might present similar/overlapping peptides. However, the extent of promiscuity among HLA class I ligands remains controversial: while in many studies T cell responses are detected against epitopes presented by alternative molecules across HLA class I supertypes and loci, peptide elution studies report minute overlaps between the peptide repertoires of even related HLA molecules. To get more insight into the promiscuous peptide binding by HLA molecules, we analyzed the HLA peptide binding data from the large epitope repository, Immune Epitope Database (IEDB), and further performed in silico analysis to estimate the promiscuity at the population level. Both analyses suggest that an unexpectedly large fraction of HLA ligands (>50%) bind two or more HLA molecules, often across supertype or even loci. These results suggest that different HLA class I molecules can nevertheless present largely overlapping peptide sets, and that “functional” HLA polymorphism on individual and population level is probably much lower than previously anticipated.     

HLA class I nucleotide sequences, 1992

The HLA class I sequences included in this compilation are taken from publications listed in the papers: Nomenclature for factors of the HLA system, 1991 (Bodmer et al. 1992); Nomenclature for factors of the HLA system, 1990 (Bodmer et al. 1991); and Nomenclature for factors of the HLA system, 1989 (Bodmer et al. 1990). Due to the increased number of sequences we have only included sequences for exons 2, 3, and 4 in this compilation. Where discrepancies have arisen between reported sequences, the original authors have been contacted where possible, and necessary amendments to published sequences have been incorporated into this alignment. Future sequencing may identify errors in this list and we would welcome any evidence that helps to maintain the accuracy of this compilation. In the sequence alignments, identify between nucleotides is indicated by a hyphen (-). An unavailable sequence is indicated by a period (.). Gaps in the sequence are inserted to maintain the alignment between different alleles showing variation in amino acid number. *** DIRECT SUPPORT *** A4903038 00002     

HLA class I nucleotide sequences, 1991

The HLA class I sequences included in this compilation are taken from publications listed in the accompanying paper, Nomenclature for factors of the HLA system, 1990 (Bodmer et al. 1991) and Nomeclature for factors of the HLA system, 1989 (Bodmer et al. 1990). Where discrepancies have arisen between reported sequences the original authors have been contavted where possible, and necessary amendments to published sequences have been incorporated into this alignment. Future sequencing may identify errors in this list and we would welcome any evidence that helps to maintain the accuracy of this compilation. In the sequence alignments identify between residues is indicated by a hyphen (-). Unavailable sequence is indicated by a period (.). Gaps in the sequence are inserted to maintain the alignment between different alleles showing variation in amino acid number.     

Biochemical complexity of serum HLA class I molecules

Human serum was found to contain a variety of class I-like molecules by Western blotting with anti-class I heavy chain reagents: major bands usually are observed around M _r 44 000, 40 000, and 35 000–37 000. HLA-A24-positive individuals are distinguished by higher serum levels of M _r 44 000 and 40 000 class I-like molecules than those found in HLA-A24-negative individuals. The M _r 44 000 serum molecules are probably intact class I molecules that have been shed from the cell membrane, because they contain both a transmembrane segment (TM), as deduced from detergent-binding experiments, and a cytoplasmic tail (CT), as inferred from reactivity with an antipeptide serum specific for the cytoplasmic domain of class I antigens (RaCT). The M _r 35 000 and 37 000 molecules contain neither a TM nor a CT region and therefore are probably proteolytic breakdown products of cellular and/or serum M _r 44 000 molecules, although the existence of Q10-like molecules in man cannot be ruled out. The M _r 40 000 molecules do not contain a TM region. M _r 40 000 molecules reactive with the RaCT serum were found in the minority (2/13) of sera tested. We conclude that alternative splicing resulting in a precise excision of the TM exon plays a minor role in the generation of serum HLA class I antigens.Abbreviations used in this paper B2m beta-2 microglobulin - BSA bovine serum albumin - EBV-BLCL Epstein-Barr virus-transformed B lymphoblastoid cell line - mAb monoclonal antibody - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis - TCA trichloroacetic acid     

Soluble HLA class I and HLA class II antigens in the blood of HIV infected patients

The levels of HLA, class I, antigen and HLA-DR antigen in the blood sera of HIV-infected persons were determined by the enzyme immunoassay. The levels of soluble antigens HLR-DR and of HLA, class I, in serum samples containing only antibodies to HIV were elevated, respectively, 1.8- and 1.3-fold in comparison with the norm. The level of soluble HLA-DR antigen in samples containing the markers of other infections (HBsAg, antibodies to hepatitis C virus, anti-IgM antibodies to cytomegalovirus) was significantly higher in comparison with samples containing only antibodies to HIV and serum samples from healthy donors (p < 0.05). The concentration of soluble HLA antigen, class I, in the samples containing antibodies to HIV and markers of other infections was also elevated, but the statistically authentic increase in comparison with the normal level was observed only in the presence of the markers of HIV infection, hepatitis C and cytomegalovirus infection.     

Pathogen-driven selection and worldwide HLA class I diversity

The human leukocyte antigen (HLA; known as MHC in other vertebrates) plays a central role in the recognition and presentation of antigens to the immune system and represents the most polymorphic gene cluster in the human genome [1]. Pathogen-driven balancing selection (PDBS) has been previously hypothesized to explain the remarkable polymorphism in the HLA complex, but there is, as yet, no direct support for this hypothesis [2 and 3]. A straightforward prediction coming out of the PDBS hypothesis is that populations from areas with high pathogen diversity should have increased HLA diversity in relation to their average genomic diversity. We tested this prediction by using HLA class I genetic diversity from 61 human populations. Our results show that human colonization history explains a substantial proportion of HLA genetic diversity worldwide. However, between-population variation at the HLA class I genes is also positively correlated with local pathogen richness (notably for the HLA B gene), thus providing support for the PDBS hypothesis. The proportion of variations explained by pathogen richness is higher for the HLA B gene than for the HLA A and HLA C genes. This is in good agreement with both previous immunological and genetic data suggesting that HLA B could be under a higher selective pressure from pathogens.     

HLA class I and class II polymorphism in three Sicilian populations

Two human leukocyte antigen (HLA) class I loci (HLA-A and HLA-B) and one class II locus (HLA-DR) were typed at the DNA level in the Sicilian population. Study participants were of Sicilian origin (183 for class I loci and 260 for class II loci) and live in three towns, chosen on the basis of geographic position and different historical events. These towns are Sciacca (southwest Sicily, located at sea level, conquered by Arabs in A.D. 814), Piana degli Albanesi (northwest Sicily, 720 m above sea level, has maintained religious, cultural, and linguistic peculiarities traced to Albanian settlement in 1488), and Troina (northeast Sicily, 1120 m above sea level, known as the first settlement of Normans). The assumptions underlying the study of genetic structure, based on HLA allele polymorphism, are that these three towns are located in areas that can be distinguished according to historical criteria and that they are likely to have contributed to cultural and probably genetic differences. As such, the high frequency of some alleles in Sciacca and Troina seems to be correlated with Greek, Phoenician, North African, and Arab influence. In accordance with different human settlements in Sicily, we found that the HLA allele frequencies support the existence of genetic differentiation between the western and eastern sides of Sicily. This separation is attributed to Greek colonization in the east and to Phoenician-Carthaginian-Arab influence in the west. Moreover, the comparisons of all allele frequencies between Mediterranean and AfrIcan populations show the same trend, highlighting in some cases European origin and in other cases non-European origin.     

HLA class I polymorphism in the Albanian population

The HLA class I polymorphism was studied in a sample of the Albanian population. Ninety-three unrelated healthy Albanians were typed for HLA-A, -B and -Cw antigens by standard microlyphocytotoxicity test. The antigens with the highest frequencies were: HLA-A2 (34.4%), A3 (14.5%) and A1 (12.4%); B51 (19.3%), B35 (12.4%) and B18 (10.2%); Cw4 (16.2%), Cw7 (16.2%) and Cw6 (10.8%). The HLA haplotypes with high frequency in Albanians included A2-B51 (4.3%), A2-B18 (2.4%), A2-B35 (2.4%), Cw4-B35 (7.6%), and Cw7-B18 (6.5%), which are not significantly different from the other neighboring populations. Low frequency of HLA-A1-B8 haplotype (1.1%) is noted in the Albanian population. The frequency of HLA-B27 antigen (1.1%) is one of the lowest frequencies observed in Caucasians. Such results are important in studies of HLA-A1-B8, HLA-B27 and disease associations. These findings should be also useful in understanding the origin of Albanians, representing a base for future studies about HLA polymorphism in the Albanian population.     

Association of HLA class I antigens and HLA class II alleles with vitiligo in a Turkish population

As is the case with many other autoimmune diseases, there is an association between vitiligo and HLA complex. HLA subtypes vary with racial/ethnic background. The purpose of this study was to determine which HLA class I antigens and HLA class II alleles are associated with Turkish vitiligo patients. Forty-one patients with vitiligo and 61 healthy control subjects were typed for HLA class II alleles. Thirty-three out of 41 patients with vitiligo and 100 healthy transplant donors were typed for HLA class I antigens. HLA DNA typing was performed by polymerase chain reaction/sequence specific primer method for class II. HLA typing for class I was performed by serological method. The frequency of HLA DRB1*03 was 0.6340 in patients compared to 0.2950 in controls (P = 0.0014). The frequency of HLA DRB1*04 was found to be 0.6830 in patients compared to 0.2950 in controls (P = 0.00026). The allele HLA DRB1*07 was present in 0.390 of patients compared to 0.0820 of the controls (P = 0.0004). A preventive antigen for the manifestation of vitiligo has not been identified in this study. Our findings suggest that DRB1*03, DRB1*04 and DRB1*07 alleles are genetic markers for general susceptibility to vitiligo in a Turkish population.     

Conservation of HLA class I private epitopes in macaques

Fifty mouse monoclonal antibodies (mAb) specific for HLA class I epitopes were compared for their reactivity against two closely related nonhuman primate species, pigtailed macaques (Macaca nemestrina, Mn) and longtailed macaques (M. fascicularis, Mfl), which diverged from the hominoids 23–40 million years ago. An analysis of Nei's genetic identity (I) and distance (D) based on reactivity of all class I-specific mAb showed, as expected, that the macaques are more closely related to each other (1=0.959) than to man (I=0.782 for Mn and 0.859 for Mfl). However, there were clear differences in genetic similarity with respect to certain epitopes. Macaques were most different from each other and from man in expression of heterologous epitopes recognized by the mouse that are not polymorphic among humans. In contrast, the most polymorphic epitopes unique to single HLA alleles, so-called private epitopes, were present in all the species, and neither macaque species could be distinguished from humans, suggesting that certain class I private epitopes may be highly conserved in evolution.[/p]Abbreviations used in this paper D Nei's index of genetic distance - I Nei's index of genetic identity - mAb monoclonal antibodies - Mfl Macaca fascicularis - MHC major histocompatibility complex - Mn Macaca nemestrina - PF phenotypic frequency     

c-myc down-regulates class I HLA expression in human melanomas

Expression of class I HLA antigen has been shown to be reduced in a number of human tumours. Here we show that in a panel of 11 melanoma cell lines with variable class I HLA expression an inverse correlation exists between the mRNA levels of c-myc and class I HLA. This suggests that high expression of the c-myc oncogene might inhibit the class I HLA expression. To test this hypothesis a melanoma cell line with a low c-myc and high class I HLA mRNA expression was transfected with a c-myc expression vector. All clones expressing the transfected c-myc gene show reduced class I HLA mRNA and beta 2-microglobulin mRNA expression. Reduced class I HLA mRNA levels result in a lowered class I protein expression on the cell surface. Treatment with gamma-interferon fully restores the class I HLA and beta 2-microglobulin expression in these cells. This effect is preceded by a transient decrease of the c-myc mRNA level. These results show that the class I HLA expression is modulated by the level of c-myc expression, thus opening up the possibility that high expression of this oncogene influences the interaction of melanoma cells with the immune system.     

Lack of HLA class I and class II antigens on human preimplantation embryos

The expression of HLA Ag on polyploid early human embryonic stages, obtained in an in vitro fertilization and embryo transfer program, was investigated by indirect immunofluorescence tests using a panel of mAb. Neither HLA class I Ag, beta 2-microglobulin, nor HLA class II molecules could be detected on blastomers. The zona pellucida also lacked these Ag, but granulosa cells expressed HLA class I Ag, beta 2-microglobulin, and HLA class II Ag. These results make it likely that the absence of HLA Ag is one of the mechanisms involved in protecting the implanting embryo from rejection by the immunocompetent mother.     

Signal transduction pathways in the induction of HLA class I antigen expression on Huh 6 cells by interferon-gamma.

This study investigated the intracellular signal transduction regulating the appearance of HLA class I antigens on Huh 6 cells induced by interferon-gamma. The expression was blocked by a protein kinase C inhibitor, H-7, but not by a calmodulin antagonist, W-7, nor by a protein kinase A inhibitor, H-8, at low dose. The antigen expression was induced by a direct activator of protein kinase C, phorbol myristate acetate, but not by calcium ionophore A23187 nor an analog of cAMP, dbcAMP. Therefore, we concluded that protein kinase C is involved in the expression of HLA class I antigens on Huh 6 cells induced by interferon-gamma but Ca(2+)-calmodulin and cAMP are not.