INHIBITION OF DNA-SYNTHESIS AND IL-2 BIOACTIVITY IN MLR BY SPLENIC PREGNANCY-ASSOCIATED NATURAL SUPPRESSOR CELLS INVOLVES THE PRODUCTION OFA TGF-BETA-1-LIKE MOLECULE AND A 2ND DISTINCT INHIBITORY FACTOR

Natural suppressor cells exhibiting a double-negative, immature T cell phenotype have been identified in maternal spleen during syngeneic mu rine pregnancy. In the present study, splenic pregnancy-associated nat ural suppressor (SPANS) cells are shown to express alpha/beta T cell r eceptors. SPANS cell-mediated inhibition of DNA synthesis by spleen ce lls responding in mixed lymphocyte reactions (MLR) is associated with a reduction in interleukin (IL)-2 bioactivity beginning after 96 h of culture. Although culture supernatants from suppressed MLR exhibit dim inished ability to support the growth of IL-2-dependent CTLL-2 cells, SPANS cells themselves are unable to inhibit IL-2-driven CTLL-2 prolif eration, suggesting that SPANS cells down-regulate IL-2 synthesis in M LR. IL-2 utilization in MLR is also inhibited by SPANS cells, since th e addition of exogenous IL-2 fails to relieve the inhibitory effect of SPANS cells on lymphoproliferative responses in MLR. Flow cytofluorom etric analysis reveals that MLR performed in the presence of SPANS cel ls contain normal percentages of CD4 and IL-2 receptor-bearing spleen cells. Thus, SPANS cells do not inhibit cellular proliferation in MLR by selectively interfering with clonal expansion of IL-2-producing hel per T cells or by down-regulating IL-2 receptor expression. We have de termined that SPANS cells inhibit DNA synthesis in MLR via the product ion of a transforming growth factor (TGF)-beta1-like suppressor factor , since cellular proliferation in MLR is restored to normal levels in the presence of anti-TGF-beta1 neutralizing antibody. However, IL-2 bi oactivity in these cultures remains low in comparison to control MLR, suggesting the presence of a second distinct suppressor factor. Althou gh the identity of this second inhibitory molecule has yet to be deter mined, neutralizing antibody studies have ruled out IL-10.