This project on human immunogenetics of the major histocompatibility complex is an integrated investigation of immunological, genetic, and biochemical aspects of the HLA complex and closely linked genes. The overall objectives for this project are to obtain a better understanding of the genetic factors responsible for allograft reactions, to identify the immunological mechanisms involved in such reactions, and to understand the relationship between the HLA complex and genetic factors predisposing to diseases. The studies are divided into four projects. (1) Cell surface phenotype and function of human alloactivated lymphocytes. This project utilizes the techniques of T-lymphocyte cloning for the analysis of lymphocyte diversity. (2) In vitro production by human B lymphocytes of human monoclonal antibodies detecting cell surface antibodies is being investigated. (3) Serological, biochemical, and genetic analysis of the human equivalent of the thymus-leukemia (TL) antigen is undertaken and compared with the murine counterparts. (4) The relationship between a disease-causing gene and HLA is being investigated using congenital adrenal hyperplasia due to 21-hydroxylase deficiency as a model. This project utilizes molecular genetic tools for the analysis of the genes in the HLA region. The major emphasis in the studies of alloreactivity during the last year has been the relationship between T lymphocyte differentiation antigens and MHC gene products. These studies have involved allocytotoxic, human T-cell clones. The conclusions are based upon the combined analysis of biochemical studies of MHC antigens using murine anti-HLA monoclonal antibodies, analysis of T-lymphocyte differentiation antigens with monoclonal antibodies and blocking studies of allocytotoxicity with these two sets of antibodies. The investigations have revealed a distinct but complex pattern. It has been demonstrated that multiple HLA-D-related class II antigens all carry allotypic determinants which can be recognized by cytotoxic T cells. Other studies have involved the investigation of T cell activation via the cell surface antigens T3 and T11. The relationship between these two T cell activation pathways is presently under investigation. We have achieved major progress in this project on human immunogenetics in the molecular genetic analysis of congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OH-Def.). During the last year, we have cloned and sequenced a full length cDNA for adrenal 21-hydroxylase, a cytochrome P450. This cDNA has been used to map two 21-OH-genes within the murine H-2 complex. These genes are located within the S-Region of H-2 and very close to H-2D. The same cDNA probe has been used to characterize the two 21-OH genes within the HLA complex. At least one of these two genes are deleted in the HLA-Bw47 positive HLA haplotypes found in some patients with the severe, salt-wasting form of 21-OH-Def. These studies indicate that CAH due to 21-OH-Def. is a defect in the structural gene for adrenal cytochrome P450 21-hydroxylase. (CS)
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