Biomedical subjects
Publications and source records attributed to C Brautbar.
HLA-A, -B, -C, -DR, and -DQ antigens were determined in 25 Ashkenazi Jews with Paget's disease of bone. HLA-DR2 was more frequent in the Pagetic patients compared with 57 healthy controls of the same ethnic origin. This finding concurs with a previous report and raises the possibility that HLA-DR2 may be associated with Paget's disease of bone, probably by predisposing the bone cells to viral infection.
HLA class-I and class-II antigens were detected in extracts of dry spleen tissue allowed to age from 1 to 17 months, using monomorphic anti HLA monoclonal antibodies and the ELISA technique. Using polymorphic HLA alloantisera samples of spleen tissue could be characterized and differentiated. The potential use of dried tissues and blood stains is discussed.
HLA antigens of two Jewish families with alopecia areata were studied. There was no linkage between the disease and the HLA complex.
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HLA typing of amniotic cells for clinical purposes using the conventional cytotoxicity assay is a laborious and complicated procedure. In the present study we demonstrate that HLA class I typing of the fetus can be determined using the enzyme linked immunosorbent assay (ELISA) either on amniotic cells or soluble free antigens shed by the cells into the culture medium. Our results indicate that the ELISA technique is a sensitive and reliable assay that can be used as an alternative method of HLA class I typing of amniotic cells.
A double antibody ELISA technique is described to detect HLA antigens in extracts of blood stains. The assay involves capture of free HLA determinants using an immobilized monoclonal antibody directed against monomorphic regions of HLA class I and HLA class II antigens. The captured antigens are then detected using alloantisera directed against the polymorphic regions of the captured HLA entities. The technique is able to detect specific HLA-A, B, and DR antigens in extracts prepared from blood smears as well as from dried and freshly thawed lymphocytes. The assay may be of potential use in forensic medicine, particularly in instances where extraction of nucleic acids for fingerprinting is not feasible.
Various immunologic parameters were tested in a family of two patients with common variable immunodeficiency. Both patients had low serum immunoglobulin levels, low peripheral B cell and T4 subclass of lymphocytes, and reversed T4/T8 ratio. One of the patients had excessive suppressor activity. Two of the asymptomatic members of the family (the father and one brother) had also low T4/T8 ratio that was not associated with excessive suppressor activity. No linkage between the disease inheritance to HLA could be observed. A study of the T cell helper activity to an antigen, the response to which is regulated by an HLA-linked gene, suggested a defect in the immune response of the two patients and their asymptomatic brother with immunologic disorders. Treatment with cimetidine of the patient with excessive suppressor activity led to an improvement in his clinical state, reduction in suppressor activity, temporary effect on his proliferative response capacity to mitogens, and an increase in the antigen-specific helper activity.
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The autoimmune dermatologic disease pemphigus vulgaris (PV) is associated with the serotypes HLA-DR4 and HLA-DRw6. Based on nucleotide sequence and oligonucleotide probe analysis of enzymatically amplified DNA encoding HLA-DR beta chain (HLA-DRB) and HLA-DQ beta chain (HLA-DQB; henceforth HLA is omitted from designations), we showed previously that the DR4 susceptibility was associated with the Dw10 DRB1 allele [encoding the mixed lymphocyte culture (MLC)-defined Dw10 specificity]. The DRw6 susceptibility similarly was shown to be associated with a rare DQB allele (DQB1.3), which differed from another nonsusceptible allele by only a valine-to-aspartic acid substitution at position 57. Given the linkage disequilibrium that characterizes HLA haplotypes, it is difficult to assign disease susceptibility to a specific locus rather than to a closely linked gene(s) on the same haplotype. To address this problem, we have analyzed all of the polymorphic loci of the class II HLA region (DRB1, DRB3, DQA, DQB, and DPB) on the DRw6 haplotypes in patients and controls. In 22 PV patients, 4 different DRw6 haplotypes were found that encode the same DQ beta chain (DQB1.3) but contained silent nucleotide differences at the DQB locus as well as coding sequence differences in the DQA and DRB loci. These results, obtained by using a method for allele-specific polymerase chain reaction amplification, strongly support the hypothesis that the allele DQB1.3 confers susceptibility. This DQB allele is correlated with the MLC-defined Dw9 specificity and is associated with two different DRB1 alleles (the common "6A" associated with DRw13 and the rare "6B" associated with DRw14). Since 86% (19 of 22) of DRw6+ patients contain the DQB1.3 allele (vs. 3% of controls), whereas 64% (14 of 22) contain the DRB1 allele 6B (vs. 6% of the controls), we conclude that most of the DRw6 susceptibility to PV can be accounted for by the DQ beta chain.
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We describe here a simple procedure, by which HLA class II antigens can be accurately and reliably identified in those patients where there is minimal or absent expression of HLA-DR,DQw antigens on B cells, or when the total number of leukocytes recovered from the patients do not permit reliable typing. Ficoll-Hypaque-separated peripheral blood mononuclear leukocytes, fresh or cryopreserved, were activated by PHA and then propagated in IL-2-containing medium until enough cells for typing were obtained (usually 7-14 days). At this stage, the cultured cells were shown to be primarily T cells (greater than 90% CD3+). Since the activated T cells propagate in the presence of IL-2, even a small number (10(4] of fresh or cryopreserved patients' cells suffice for this protocol. To date we have been able to successfully HLA-DR,DQw type 34/34 bone marrow transplantation candidates and 12/12 long-term dialysis patients, who were untypable using fresh cells. HLA-DR,DQw antigens on activated T cells from normal individuals were identical to those found on their uncultured B cells. In addition, class I antigens that were undetectable on the uncultured cells of one patient could be identified on activated T cells. The HLA antigens identified on the patients' activated T cells were confirmed by phenotypic analysis of cells from family members.
Pairs of BL-derived cell lines and in vitro EBV-transformed LCLs, derived from the same patient, were compared for the expression of MHC class-I antigenic determinants as shown both by monoclonal antibody (MAb) binding and by sensitivity to HLA-specific CTL clones. BL lines expressed all polymorphic determinants tested at a lower level than the corresponding LCLs, as indicated by the binding of the MAbs AUF 5.13 (anti-HLA A3,A11), GS 142.1 (anti-HLA A1), GS 114.1 (anti-HLA A24), GSP 35.1 (anti-HLA A2,A28), GSP 55.1 (anti-HLA A25,A32), TER MA32 (anti-HLA A32), GSP 145.2 (anti-HLA B27), B27 M.1 (anti-HLA B27,B7) and GSP 8.1 (anti-HLA B8). The difference was most pronounced for HLA A11 and least for B27,A1 and B8 with intermediate differences for the other specificities. The BL lines were also less sensitive to lysis by HLA-specific CTL clones directed to the same and to additional antigens. The polymorphic determinants detected by the AUF 5.13 and GSP 35.1. MAbs were expressed at a lower level in resting T and B cells compared to mitogen- and EBV-induced blasts. An analogous change in the expression of polymorphic determinants was observed in EBV-converted sublines of originally EBV-negative BLs that have become more "LCL-like" after conversion. The appearance of B-cell activation markers was paralleled by the up-regulation of both the serologically defined and the CTL-target epitopes. The findings suggest that the low expression of HLA determinants on the BL cells is a phenotypic vestige of the normal BL precursor.
The inheritance of particular alleles of major histocompatibility complex class II genes increases the risk for various human autoimmune diseases; however, only a small percentage of individuals having an allele associated with susceptibility develop disease. The identification of allelic variants more precisely correlated with disease susceptibility would greatly facilitate clinical screening and diagnosis. Oligonucleotide-primed gene amplification in vitro was used to determine the nucleotide sequence of a class II variant found almost exclusively in patients with the autoimmune skin disease pemphigus vulgaris. In addition to clinical implications, the disease-restricted distribution of this variant should provide insight into the molecular mechanisms underlying associations between diseases and HLA-class II genes.
Narcolepsy is a very rare disease among Israeli Jews with a frequency of 7/3 X 10(6). An investigation of the association of narcolepsy with the human leukocyte antigen system was conducted in Israeli Jews at the serologic and genomic levels. The human leukocyte antigen class I and class II antigen typing of 7 clinically diagnosed narcoleptics, 3 individuals suffering from sleep disorders other than narcolepsy, and 11 healthy matched controls revealed that all narcoleptic patients (100%) investigated in the present study carried the HLA-DR2 haplotype, whereas patients with other sleep disorders did not. The HLA-B7 and DR2 occurred jointly in 57% (4/7) of the narcoleptic patients, as compared to 2% in randomly selected Israeli healthy controls. Restriction fragment length polymorphism analysis was performed with several restriction enzymes and three cDNA probes for DQ alpha, DQ beta, and DR beta genes on genomic DNAs obtained from narcoleptics and patients with other sleep disorders, matched controls, and 3 homozygous typing cells representing the DR2 subtypes Dw2, Dw12, and DwAZH. The restriction fragment length polymorphism analysis showed that all narcoleptics (7 of 7) shared virtually identical restriction fragment length polymorphisms with one of the homozygous typing cells (GSO), which defines DR2,Dw2. The frequency of the DR2,Dw2 haplotype in the healthy Israeli population is 3.2%. Other non-narcoleptic patients did not share these restriction fragment length polymorphisms. These findings indicate that narcolepsy is associated worldwide with the HLA-DR2,Dw2 haplotype.
The amount of HLA class-I antigens was determined on the surface of enriched populations of B and T lymphocytes obtained from human peripheral blood, lymph nodes and spleens. The assays were performed using monoclonal antibodies that recognize different determinants on HLA class-I antigens and utilizing a simple and sensitive enzyme-linked immunosorbent assay (ELISA). Our results indicate that B lymphocytes obtained from human peripheral blood, lymph nodes and spleens express at least twice as many HLA class-I antigens as T lymphocytes obtained from the same organs.
The autoimmune dermatologic disease pemphigus vulgaris (PV) is associated with the HLA serotypes DR4 and DRw6. Susceptibility to PV could be conferred either by sequences shared between the DR4 and DRw6 haplotypes or by different sequences in these haplotypes. We have examined the distribution of DR and DQ beta-chain and DQ alpha-chain alleles in PV patients and in control subjects by hybridization with oligonucleotide probes and sequence analysis of in vitro amplified DNA. Ninety percent (34/38) of the DR4 haplotypes in patients contain a specific DR beta I sequence present in 36% (16 of 44) of DR4 controls (P = 0.001). This sequence is also found in DRw6 haplotypes. However, it is present in only 25% (6 of 24) of DRw6 patients. The results of our analysis indicate that predisposition to PV is conferred by different sequences in DR4 and DRw6 haplotypes. The DR4 susceptibility is highly associated with the Dw10 DR beta I allele, implicating the polymorphic residues in the third hypervariable region. The DRw6 susceptibility is strongly associated with a rare DQ beta allele (DQB1.3). This allele differs from a common DQ beta allele (DQB1.1) only by a valine----aspartic acid substitution at position 57.
Human decidual antigen presenting cells (DAPCs) exposed to fetal cells in vitro induce generation of suppressor T cells among a population of peripheral blood lymphocytes. Human lymphocyte antigen (HLA)-class II positive antigen presenting cells were isolated from early normal human decidual tissue and from peripheral blood (PAPCs) by adhering Ficoll-Paque separated cell suspensions to fibronectin. In contrast to PAPCs, DAPCs pulsed with fetal antigens induced a radio-sensitive, Leu 1,2-positive T suppressor cell population. A nylon wool adherent B cell population is required during the in vitro induction of the suppressor cells. These suppressor cells impair primary mitogen and mixed lymphocyte culture (MLC) responses, generation of anti-trinitrophenyl (TNP) cytotoxic T lymphocytes, and antibody response of autologous and allogeneic lymphocytes. Only intact viable embryonic cells can effectively confer upon DAPCs the ability to induce T suppressor cells. The T suppressor cell induction by DAPCs primed with fetal antigens is restricted by the major histocompatibility complex. Our results show that the HLA-DR molecules are the most prominent restriction elements.