Activated macrophages (M phi s) are important participants in host defense, but their uncontrolled activation leads rapidly to septic shock and death. One mechanism for regulating other dangerous cells in the immune system is programmed cell death, or apoptosis. Monocytes are known to undergo spontaneous apoptosis upon leaving the circulation unless provided with specific survival signals, but mature tissue M phi s are more robust cells, and it was not clear that they could be similarly regulated by apoptosis. We now show that during differentiation monocytes rapidly lose their sensitivity to apoptosis triggered by passive cytokine withdrawal, but they may retain a novel pathway which initiates apoptosis after activation with specific stimuli (zymosan and phorbol esters). Sensitivity to activation-induced apoptosis was developmentally determined, being downregulated by the maturation-promoting cytokine macrophage colony-stimulating factor but stably upregulated by even transient exposure to the proinflammatory cytokine interferon gamma (IFN-gamma). Apoptosis began within 2-4 h of activation, occurred in > 95% of susceptible cells, and in mixed cocultures selectively affected only those M phi s with a history of IFN-gamma priming. Consistent with a possible role for protein kinase C in the signaling pathway leading to cell death, the kinase inhibitor staurosporine was protective against both phorbol ester- and zymosan-induced apoptosis. Our studies describe a novel form of activation-induced M phi apoptosis which is developmentally regulated by two physiologically relevant cytokines. We speculate that apoptosis may serve to restrict the destructive potential of inflammatory M phi s.

SUMMARYHeat-stable antigen (HSA) is a murine differentiating antigen that is expressed on both CD4 ÿ CD8 ÿ double-negative and CD4 CD8 double-positive thymocytes but not CD4 or CD8 single-positive thymocytes. Effects of anti-HSA monoclonal antibody, R13, on thymocyte apoptosis induced by various stimulations were investigated by a single-cell suspension culture system. Immobilized R13 enhanced the CD3-mediated DNA fragmentation and killing of thymocytes but not the dexamethasoneinduced or phorbol myristate acetate-induced killing of thymocytes. Immobilized R13 by itself could not induce thymocyte apoptosis. Soluble R13 enhanced CD3-mediated apoptosis when HSA and T-cell receptor (TCR)/CD3 were co-cross-linked by a cross-reactive secondary antibody. Even without the cross-reactive secondary antibody, soluble R13 enhanced CD3-mediated apoptosis, although a greater than 100-fold increase in the amount of R13 was needed to give a similar enhancement compared with immobilized R13. Neither R13 by itself nor R13 plus secondary antibody induced cytosolic calcium influx, whereas R13 enhanced CD3-mediated cytosolic calcium increase. These results suggest a functional role of HSA in promoting the activation-induced apoptosis of thymocytes and the involvement of HSA in negative selection.