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Mice vaccinated by injection with tumor cells expressing the Mycobacterium leprae gene for hsp65 acquire a remarkably high degree of protection against challenge with Mycobacterium tuberculosis. We used limiting-dilution analysis to assess the frequency of CD4+ CD8- and CD4- CD8+ splenocytes responding to mycobacterial hsp65 in such vaccinated mice. Cells of both phenotypes were present at very high and equal frequencies (approximately 1:100). Vaccination with live Mycobacterium bovis BCG also increased the frequencies of both phenotypes of hsp65-reactive cells equally (to approximately 1:2,500), whereas vaccination procedures that were not protective, with either dead BCG, hsp65 protein in incomplete Freund's adjuvant, or hsp65 mixed with tumor cells, resulted in preferential increase in CD4+ CD8- cells. Twelve CD4+ CD8- and twelve CD4- CD8+ hsp65-responsive T-cell clones were obtained and characterized. All showed conventional antigen recognition via major histocompatibility complex class II and class I pathways but differed in secretion of gamma interferon and interleukin 4 and cytotoxicity. In tests of antimycobacterial activity against M. tuberculosis, both in infected macrophages in vitro and by adoptive transfer of protection with T-cell clones injected into irradiated mice, the most effective clones were the most cytotoxic and secretion of gamma interferon made only a secondary contribution.

infection and immunity

Characterization of T cells that confer a high degree of protective immunity against tuberculosis in mice after vaccination with tumor cells expressing mycobacterial hsp65