OPPOSING ROLES OF CD95 (FAS APO-1) AND CD40 IN THE DEATH AND RESCUE OF HUMAN LOW-DENSITY TONSILLAR B-CELLS/

The regulation of B cell death plays roles in the selection of Ag-spec ific B cells in humoral immune responses, controlling B cell homeostas is and perhaps limiting transformation, The present work addresses whe ther CD95 induces tonsillar B tells to undergo apoptosis and, if so, w hether contact-dependent CD40-L:CD40 signaling can rescue tonsillar B cells from CD95-induced apoptosis. CD95 triggering by anti-CD95 mAb (A PO-1) was studied in human tonsillar B cell populations that were sepa rated by density centrifugation into fractions enriched for either low density, CD38(+) B cells or high density, resting B cells. Low densit y tonsillar B cells express CD95 and undergo anti-CD95-mediated apopto sis by analysis of cellular morphology or DNA fragmentation by TUNEL a ssay. The induction of apoptosis in low density tonsillar B cells by a nti-CD95 mAb is inhibited by CD40 signals provided by stably transfect ed CD40-L(+) 293 cells, but not by control transfected 293 cells (expr essing CD8). In addition, the rescuing effect of CD40-L(+) cells is in hibited specifically by anti-CD40-L (mAb 5c8), The counteracting effec ts of CD95 and CD40 signaling were also studied in Ramos 2G6, a homoge neous B cell tumor line of germinal center phenotype that expresses CD 95 and CD40. Similar to the behavior of low density tonsillar B cells, Ramos 2G6 undergoes anti-CD95-mediated apoptosis, which is prevented by CD40-mediated rescue. These data show that CD95 induces apoptosis i n low density tonsillar B cells and that CD40-L:CD40 interactions resc ue low density tonsillar B cells or the B cell tumor Ramos 2G6 from CD 95-induced apoptosis, and suggest roles for CD95 and CD40 in B cell de ath and selection, respectively.