Abstract
We are interested in understanding the effector pathways used by host T cells to inhibit engraftment following allogeneic HCT. Previous work in MHC matched models found that mice deficient in either perforin or fasl continue to mediate resistance, however, the numbers of BMCs necessary to override the host barrier is less than in cytotoxically normal mice. These findings suggest both cytotoxic pathways are likely to contribute to the barrier. Notably, in the combined absence of both pathways, resistance is virtually absent. In contrast, animals which contain memory CD8 T cells can resist large numbers of BMCs regardless of the absence of both perforin and fasl pathways. The present studies examined the use of these pathways by T cell populations with respect to the inhibition of progenitor cell (PC)activity (CFU-IL3) from BMC ex-vivo to determine the relative contributions of perforin and/or fasL in inhibition mediated by effector cells derived from naive vs. memory cell populations. Effector cells were generated from unsensitized BALB/c and BALB/c-perf−/− mice as well as mice immunized against H2b alloantigens. Primary effectors from unsensitized mice and memory effectors were generated following 5 and 2–3 day MLC cultures, respectively. Effector cells were co-cultured up to 48 hrs. with BMC and then plated with rmIL-3 to elicit CFU formation. Effector cells from unsensitized BALB/c-wt mice inhibited allogeneic B6-wt CFU (>80%) as well as B6-lpr CFU (>70%) formation. Syngeneic BM was not inhibited. These data demonstrate that the perforin dependent pathway is sufficient for inhibition, consistent with in vivo resistance observed in FasL defective recipients. Effector cells generated from unsensitized BALB/c-perf−/− also effectively inhibited B6-wt CFU (>80%), consistent with a fas mediated pathway. However, in contrast to effectors generated from unsensitized BALB/C-wt mice, effector cells generated from BALB/c-perf−/−did not inhibit CFU from B6-lpr BM, suggesting that: 1) both perforin and fasl effector pathways can be utilized to inhibit CFU formation ex-vivo and, 2) in the absence of both pathways, these primary effector cells could not mediate such inhibition. As anticipated, memory effector cells from BALB/c-wt mice exhibited the same pattern as the "naive" effectors (i.e.>90% inhibition) against both B6-wt and B6-lpr BM. This finding is consistent with inhibition by perforin and/or additional effector pathways. However, memory effector cells from cytotoxically deficient BALB/c-perf−/− mice failed to mediate the same pattern of inhibition as "naive" effectors from this strain. In contrast to the inability of primary effectors to inhibit B6-lpr CFU activity, the memory effectors efficiently (i.e.>90% inhibition) inhibited CFU generation from B6-lpr BM. These findings demonstrate that unlike effector cells generated from naive T cells, memory effectors can use a non-perforin/fasL dependent pathway to inhibit allogeneic PC activity. The results are consistent with the inhibition of engraftment in vivo following conditioning and transplant into cytotoxically double (perforin + fasL) defective mice and support the notion that memory - but not primary effectors utilize a non-perforin/fasl pathway to inhibit allogeneic CFU PC function. Current experiments are examining the kinetics of CFU inhibition and precise characterization of the effector T cells.
An intense form of homeostatic proliferation of naive CD8+ cells driven by IL-2
