Th2 cells: orchestrating barrier immunity.
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Ben L Kelly
Publications and source records attributed to Ben L Kelly.
The Leishmania major LACK antigen with an immunodominant epitope at amino acids 156 to 173 is not required for early Th2 development in BALB/c mice.
The Leishmania major LACK antigen contains an immunodominant epitope at amino acids 156 to 173 (LACK(156-173)) that is believed to nucleate the pathological Th2 immune response in susceptible BALB/c mice. To test this hypothesis, we generated L. major parasites that express a mutated LACK that fails to activate Vbeta4/Valpha8 T-cell receptor transgenic T cells specific for this epitope. Although mutant parasites attenuated the expansion of endogenous LACK-specific, interleukin-4 (IL-4)-expressing, CD4 T cells compared to wild-type parasites in vivo, the overall frequency of IL-4 and gamma interferon-secreting lymphocytes was similar to that elicited by wild-type L. major. Mutant parasites demonstrated diminished amastigote viability and delayed lesion development in mice, although parasites could be recovered over 200 days after infection. Complementation with a wild-type lack fusion construct partially rescued these defects, indicating a role for endogenous LACK in parasitism. Mice inoculated with mutant parasites were not protected against subsequent infection with wild-type L. major.
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Leishmania major LACK antigen is required for efficient vertebrate parasitization.
The Leishmania major LACK antigen is a key target of the immune response in susceptible BALB/c mice and remains a viable vaccine candidate for human leishmaniasis. We describe the genomic organization of the four lack genes in the L. major diploid genome together with results of selected lack gene targeting. Parasites containing a single lack gene in either the upstream or downstream locus grew comparably to wild-type promastigotes in vitro, but failed to parasitize BALB/c mice efficiently, even in a T cell-deficient environment. The replication of single copy lack mutants as amastigotes was attenuated in macrophages in vitro, and parasites failed to increase in numbers in immunodeficient mice, despite their persistence over months. Complementation with an additional lack copy was sufficient to induce robust lesion development, which also occurred using parasites with two lack genes. Conversely, attempts to generate lack-null parasites failed, suggesting that LACK is required for parasite viability. These data suggest that LACK is critical for effective mammalian parasitization and thus represents a potential drug target for leishmaniasis.
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Targeted gene deletion in Leishmania major identifies leishmanolysin (GP63) as a virulence factor.
Leishmanolysin, the Leishmania surface metalloproteinase of 63 kDa (GP63) has been described as a parasite virulence factor and is involved in the direct interaction of promastigotes and host macrophage receptors and interaction with the complement cascade. To study the role of leishmanolysin in the pathogenesis and virulence of Leishmania major, targeted gene replacement was used to delete the entire 20 kb region containing all seven leishmanolysin genes (gp63 genes 1-7). The resulting L. major leishmanolysin deficient mutants showed normal development inside the sand fly vector, however, promastigotes recovered from sand flies or from culture showed an increase in sensitivity to complement-mediated lysis and a delay in lesion formation in BALB/c animals. The phenotypic differences could be significantly improved by expression of a cloned leishmanolysin gene. These results demonstrate that leishmanolysin is a vital virulence factor in Leishmania pathogenesis.
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