The specific aims are (1) to define the biologically relevant antigens of O. volvulus adult worms, microfilariae, and infective larvae, and to relate these antigens to the immune response and to ocular and systemic complications of the disease; and (2) to elucidate the pathogenesis of the corneal lesions in onchocerciasis by defining the mechanisms of the acute and chronic inflammatory response to microfilariae and parasite-derived antigens, and of antibody-dependent complement activation by these organisms. Isolated organisms (adult worms, microfilariae, infective larvae) will be solubilized and subjected to SDS-polyacrylamide gel electrophoresis. Following electrophoretic transfer to nitrocellulose paper, antigens will be identified by immune sera from infected individuals and a peroxidase-conjugated second antibody. Sera from various age groups will be tested to seek evidence for protective antigens. A solid phase Clq binding assay followed by SDS-PAGE and immunoblotting will be used to detect immune complex-bound antigens, using rabbit antisera to O. volvulus antigen. Monoclonal antibodies will be used to isolate and purify specific antigens. cDNA clones to relevant antigens will be produced and identified by in vitro translation. Injection of microfilariae into C4 deficient and cobra venom factor treated guinea pigs will be used to elucidate the role of complement in ocular inflammation due to microfilariae. Induction of collagen synthesis by keratocytes in vitro and migration of fibroblasts towards onchocercal antigens will be used to test these phenomena as contributing to development of sclerosing keratitis. Complement activation by microfilariae will be examined in whole and C4 deficient guinea pig serum. Fractions of C4D GPS will be examined for ability to enhance granulocyte killing of D. immitis microfilariae, as will factor XII deficient and agammaglobulinemic serum. Anti GP Clq will be tested for ability to block enhanced killing. Production of C5a and C3a will be examined in vitro. By combining studies on the role of parasite antigens and the host response to them in the disease process with investigation of the inflammatory response to the parasite, the mechanisms of ocular and systemic disease in O. volvulus infection will be elucidated. In addition, identification of protective antigens which may lead to a vaccine could ultimately result in an effective means of prevention of onchocerciasis, one of the major blinding diseases of man.
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