The objective of our proposal is to study immunologic functions associated with MHC extended haplotypes. We will test two different hypotheses related to their functions: (1) Individuals homozygous for certain MHC haplotypes are associated with nonresponse against HBsAG and nonresponse may be due to a defect of TH1 or Th2 cells. We will use T cell precursor frequency, cell mixture experiments and measurements of cytokines (interferon-gamma, IL-2, IL-4, IL-10 and IL-12) in responders and nonresponders in the presence of HBsAg. We will use HLA identical pairs, related and unrelated. (2) We will test the hypothesis that individuals with low NK activity and low CD56 positive cell number belong to different HLA-B, C complementation groups corresponding to HLA-B, C homozygous and specific HLA-B, C heterozygous combinations. We will produce NK alloreactive clones that react against some but not all cells of a panel of EBV-transformed cell lines or PHA blasts of different genotypes.
Specific aims are: 1. To extend the definition and characterization of complementation groups of NK genes. We will study NK activity an the number of NK(CD16+CD56+) cells in individuals homozygous or heterozygous for each of the following determinants: HLA-B7 (Cw7), HLA-B8 (Cw7), HLA-B44 (Cw4), B44 (Cw 5), HLA- B57 (Cw6), HLA-B35 (Cw4), HLA-B60 (Cw3), HLA-B62 (Cw3) and HLA-B18 (Cw5), HLA-B18 (Cw3) and HLA-B51 (Cwx). II. In order to identify NK specificities and to determine their association with extended haplotypes or with HLA-B(C) alleles in individuals with differences in complotypes we will generate and characterize alloreactive NK clones against [HLA-B8, CS01, DR3], [HLA-B18, F1C30, DR3] [HLA-B7, SC31, DR2], [HLA-B62, SC33, DR4], [HLA-B35, SC31, DR4] homozygous extended haplotypes and against individuals homozygous for HLA-B51 and test their cytotoxic activity against a panel of PHA-activated T cells or EBV-transformed B cell liens. We will also test these clones with transfectants with individual alleles of HLA-C and HLA-B genes. III. We will use HLA-recombinant families (HLA- A/C, B; C/B or B/DR) and homozygous and heterozygous cells of different HLA genotypes to map the gene regulating the expression of the NK target antigens recognized by clones generated in specific aim II and the gene controlling the number and activity of NK cells. Methodologies will use cloning of NK cells, characterization of NK clones by immunofluorescence assay, HLA-C typing by polymerase chain reaction, and transfection of class I negative cells with HLA class I genes and mutagenesis. Identification of a hematopoietic histocompatibility (Hn) genetic system in humans could be useful for matching in one marrow transplantation.
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