Schistosomiasis, a debilitating disease caused by Schistosoma mansoni, is estimated to affect 200-300 million people worldwide. Clinical and experimental evidence supports the notion that natural immunity does exist in the infected host, indicating that it should be possible to develop an effective vaccine against schistosomiasis. Studies on immune response modulation during infection or following vaccination using attenuated parasites in mice and rat models indicate that CD4+ T- lymphocytes play a major role in protective immunity. Foreign protein molecules are taken up by antigen-presenting cells such as macrophages and B-lymphocytes and are processed within these cells by unfolding and proteolysis to short peptide fragments. These fragments are then displayed on the cell surface, bound to MHC class II molecules. Binding of T-cell receptor to these peptides in the context of MHC molecules triggers a cascade of T-helper (Th) cell immune responses. Elegant methodological developments have made it possible to elute these peptides from purified MHC molecules by acid extraction and to analyze them by microsequencing. The goal of this proposal is to isolate and identify important T-cell epitopes of schistosome origin using this novel methodological approach.
The Specific Aims of this proposal are: 1) Isolation and identification of MHC class II bound S. mansoni peptides. Eluted peptides will be sequenced and large quantities of appropriate peptides will be synthesized for further studies. 2) To elicit immune response in mice using synthetic peptides. Synthetic peptides, constructed on the basis of positive proliferation response and Th1-type cytokine secretions in the T-cell screening assay sequence homology with S. mansoni antigen, will be used to address this aim of the proposal and 3) To study in vivo protective response in Balb/c, C57BL/6, SJL and CBA/J mice following immunization with synthetic peptides. We had previously presented data showing that peptides eluted from MHC class II molecules of A20 cell line pulsed with NP40 extract of schistosomule stimulate a proliferative response in bulk lymph node cells harvested from vaccinated BALB/c as well as C57BL/6 mice. During the period October 1997- June 1998 of pilot funding, we have successfully separated peptide fractions eluted from class II molecules by reverse phase HPLC. Further, we have tested the ability of individual fractions to stimulate antigen-specific T cell proliferation and cytokine secretion response. Individual fractions have been analysed by our collaborator Dr. John Coligan, NIH and are being sequenced by Dr. Donald Hunt, University of Virginia who has joined this project as a collaborator, using microcapillary HPLC-electrospray ionization with triple quadrupole-tandem mass spectrometry. We are therefore confident that this project we have submitted is a feasible one and that it can be completed successfully.
