Sites of inhibition of mitochondrial electron transport in macrophage-injured neoplastic cells.

Abstract

Previous work has shown that injury of neoplastic cells by cytotoxic macrophages (CM) in cell culture is accompanied by inhibition of mitochondrial respiration. We have investigated the nature of this inhibition by studying mitochondrial respiration in CM-injured leukemia L1210 cells permeabilized with digitonin. CM-induced injury affects the mitochondrial respiratory chain proper. Complex I (NADH-coenzyme Q reductase) and complex II (succinatecoenzyme Q reductase) are markedly inhibited. In addition a minor inhibition of cytochrome oxidase was found. Electron transport from c~-glycerophosphate through the respiratory chain to oxygen is unaffected and permeabilized CM-injured L1210 cells oxidizing this substrate exhibit acceptor control. However, glycerophosphate shuttle activity was found not to occur within CM-injured or uninjured L1210 cells in culture hence, c~-glycerophosphate is apparently unavailable for mitochondrial oxidation in the intact cell. It is concluded that the failure of respiration of intact neoplastic cells injured by CM is caused by the nearly complete inhibition of complexes I and II of the mitochondrial electron transport chain. The time courses of CMinduced electron transport inhibition and arrest of L1210 cell division are examined and the possible relationship between these phenomena is discussed.

Inhibition of mitochondrial respiratory chain complex I by TNF results in cytochrome c release, membrane permeability transition, and apoptosis

“…Oxygen consumption was measured with a Clark oxygen electrode (model 5300: Yellow Spring Instrument Co., Yellow Spring, OH, USA) as described (Granger and Lehninger, 1982). Cells were permibilized with 0.0025% digitonin in respiration medium (0.25 M sucrose, 20 mM HEPES, pH 7.2, 2 mM KH 2 PO 4 and 1 mM EGTA), washed and then suspended at 3610 7 cells/ml in respiration medium.…”

Nitrogen oxide‐induced autoprotection in isolated rat hepatocytes

“…SSDI 0014-5793(95)01115-3 Effective permeabilization was determined by Trypan blue staining. Measurement of mitochondrial respiration was initiated by adding permeabilized hepatocytes into respiration medium supplemented with 1 mM ADP and 10 mM fl-hydroxybutyrate, 2.5 mM MgC12 and 0.7% BSA and appropriate substrate/inhibitor as described [14].…”

Nitric Oxide, Mitochondria, and Cell Death

“…However, after several hours of exposure to NO an irreversible inhibition develops, probably due to conversion of NO to RNS that inhibit respiration at multiple sites (15). One of the more rapid effects is an inactivation of complex I, possibly due to S-nitrosylation of the complex (15,24), followed by inhibition of aconitase and complex II, possibly due to removal of iron from iron-sulphur centers (25)(26)(27). Peroxynitrite can inhibit complex I, complex II, cytochrome oxidase, the ATP synthase, aconitase, Mn-SOD, creatine kinase, and probably many other proteins (2,10,28).…”