Onchocerciasis is a leading cause of blindness in the world today and the major cause of blindness in areas with severe onchocerciasis is sclerosing keratitis. Studies to elucidate pathogenesis of onchocercal sclerosing keratitis have been impeded by lack of a suitable animal model as well as unavailability of sufficient parasite material. In our laboratory, we have successfully developed an animal model in guinea pigs in which sclerosing keratitis, clinically and histologically similar to that in human onchocerciasis, is induced in presensitized guinea pigs by intrastromal injection of soluble antigens. The availability of defined antigens produced by recombinant DNA techniques in Project 2 (utilizing reagents from Projects 1 and 3 and purified in quantity in this Project will make it possible to study whether in the guinea pig model specific defined antigens are relevant in the elicitation of sclerosing keratitis. Further, examination of synthetic or enzymatically derived peptides/polypeptides from the pathogenic antigens will allow identification of the pathogenic epitopes of these antigens. Using a mouse model, detailed histologic and immunohistologic comparison will be carried out at different stages during progress of the disease. The availability of specific antibodies directed against mononuclear cell subpopulations in the mouse model provides a unique approach to define potential immunopathologic mechanisms involved in vivo in response to each defined antigen of interest from Project 2. This will provide new information on immunologic mechanisms likely to be involved in the pathogenesis of the ocular disease. As an additional step, most promising pathogenic peptide antigens will be tested in inbred strains of mice for induction of sclerosing keratitis. This will allow dissection of whether MHC haplotype determines susceptibility to sclerosing keratitis induced by a specific epitope.
The specific aim of the present proposal are: 1) to identify recombinant antigens that induce sclerosing keratitis in the guinea pig model. This work will start with two different recombinant antigens, RAL-1 and RAL-2; 2) To define pathogenic epitopes of specific recombinant antigens capable of inducing sclerosing keratitis in the guinea pig model, and 3) To perform detailed histological and immunohistological study of the cornea representing different stages of sclerosing keratitis using different defined antigens in a mouse model. This will elucidate the precise nature of the in vivo immune response to defined antigens of interest.
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