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The Journal of Immunology, Vol 153, Issue 6 2506-2514, Copyright © 1994 by American Association of Immunologists
ARTICLES |
CM Walsh, AA Glass, V Chiu and WR Clark
Department of Biology, University of California, Los Angeles 90024.
The murine CTL hybridoma PMMI has been shown by the most sensitive techniques to be devoid of perforin. We thus used PMMI activated with PMA and ionomycin, to investigate possible alternate lytic pathways in CTLs in the absence of perforin. We found that PMMI is equipped with membrane TNF-alpha as a potential lytic mechanism, but TNF-alpha is unlikely to be involved in acute (4 h) lytic reactions. On the other hand, PMMI readily lyses target cells expressing the gene for the Fas Ag, but does not lyse target cells expressing fas antisense DNA. The generation of fas-dependent lysis required protein synthesis in PMMI, but target cell protein synthesis was not required for lysis. Lysis of Fas-positive target cells by PMMI was accompanied by DNA fragmentation, and both lysis and DNA fragmentation were blocked by inhibition of protein synthesis in the effector cell. We find the relative extent and kinetics of fas-dependent lysis and DNA fragmentation indistinguishable from that seen in "classical" CTL lytic assays. Both fas- and perforin- dependent lysis were blocked by inhibitors of poly(ADP) ribosylation. We found very little difference in the sequence of events in target cells lysed by the fas pathway compared with the classical (probably perforin) lytic pathway. Given the widespread distribution of fas, particularly in hematopoietic target cells, caution may be required in interpreting the relationship between parameters such as DNA fragmentation and 51Cr-release solely on the basis of the granule exocytosis model.
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