NITRIC OXIDE-MEDIATED APOPTOSIS IN MURINE PERITONEAL-MACROPHAGES

Nitric oxide (NO) synthase, the enzyme responsible for the generation of the cytotoxic compound NO from L-arginine, is induced in macrophage s during activation. Previous work demonstrated that the cytotoxicity of NO extends to the macrophages that produce it, because the activity of NO synthase in these cells correlates inversely with their life sp an in culture. Data presented here demonstrate that the NO-dependent d eath of murine peritoneal macrophages activated in vitro with IFN-gamm a and LPS is mediated through apoptosis. Evidence in this direction wa s provided by microscopic examination of the cells, which revealed the presence of nuclear and cytoplasmic alterations characteristic of apo ptosis, and by the specific pattern of internucleosomal DNA fragmentat ion detected by electrophoresis. That these alterations resulted from the production of NO was confirmed by the preventive effects of cell a ctivation in L-arginine-restricted medium or in medium containing an i nhibitor of NO synthase, N(G)-monomethy L-arginine, and more directly by the induction of apoptosis by exposure of the cells to authentic NO gas. Additional results demonstrated that glucose starvation, the inh ibition of the tricarboxylic acid cycle with fluorocitrate or of glyco lysis with iodoacetate, but not the suppression of the electron transp ort chain wit h potassium cyanide, also induced macrophage apoptosis. The potential role of metabolic inhibition as a mechanism for NO-media ted apoptosis, as well as the relationship of these findings with even ts occurring in wounds and other sites of macrophage infiltration are discussed.