ucn-1028-c and Necrosis

Necrosis, as opposed to apoptosis, is recognized as a nonspecific cell death that induces tissue inflammation and is preceded by cell edema. In non-neuronal cells, the latter has been explained by defective outward pumping of Na(+) caused by metabolic depletion or by increased Na(+) influx via membrane transporters. Here we describe a novel mechanism of swelling and necrosis; namely the influx of Na(+) through oxidative stress-activated nonselective cation channels. Exposure of liver epithelial Clone 9 cells to the free-radical donors calphostin C or menadione induced the rapid activation of an approximately 16-pS nonselective cation channel (NSCC). Blockage of this conductance with flufenamic acid protected the cells against swelling, calcium overload, and necrosis. Protection was also achieved by Gd(3+), an inhibitor of stretch-activated cation channels, or by isosmotic replacement of extracellular Na(+) with N-methyl-D-glucamine. It is proposed that NSCCs, which are ubiquitous although largely inactive in healthy cells, become activated under severe oxidative stress. The ensuing influx of Na(+) initiates a positive feedback of metabolic and electrolytic disturbances leading cells to their necrotic demise.

Topics: Animals; Calcium; Calcium Channel Blockers; Cations; Cells, Cultured; Electric Conductivity; Extracellular Space; Free Radicals; Gadolinium; Ion Channels; Liver; Meglumine; Naphthalenes; Necrosis; ortho-Aminobenzoates; Oxidative Stress; Rats; Sodium; Vitamin K

The present study is focused on the role of oxidative stress in the induction of either necrosis or apoptosis by eicosapentaenoic acid (EPA) in the lymphoma cell lines Raji and Ramos, respectively. To investigate the different death modes induced by EPA, we assessed the importance of some antioxidants and reactive oxygen species in the two cell lines. We observed that different antioxidants counteracted the necrotic effect of EPA on Raji cells to a different extent, and that vitamin E counteracted EPA-induced accumulation of superoxide anion in this cell line. On the contrary, no effects of antioxidants were observed on development of apoptosis induced by EPA in Ramos cells, and vitamin E did not counteract EPA-induced accumulation of superoxide anions in Ramos cells. Moreover, apoptosis was partly inhibited by transcription inhibitors (actinomycin D) and protein synthesis inhibitors (cycloheximide), suggesting dependency upon new protein synthesis prior to apoptosis. Kinase inhibitors (staurosporin and calphostin C) did not alter the EPA-induced apoptosis. The observed cellular accumulation of superoxide anion following EPA incubation may be important for induction of necrosis in Raji cells. In contrast, none of the other investigated parameters indicated a role of oxidative stress promoted by EPA in the induction of apoptosis in Ramos cells.

Topics: Antioxidants; Apoptosis; Eicosapentaenoic Acid; Enzyme Inhibitors; Glutathione Peroxidase; Humans; Lymphoma; Naphthalenes; Necrosis; Protein Kinase C; Protein Synthesis Inhibitors; Superoxides; Transcription, Genetic; Tumor Cells, Cultured; Vitamin E

Protein kinase C (PKC) has been implicated in the cardioprotective effects of ischemic preconditioning in rabbits, but whether it plays a role in rats is unknown. We tested this preconditioning PKC theory by assessing whether the inhibition of PKC with calphostin C, a potent and specific inhibitor of PKC, can block the preconditioning effects in this model. Four groups of rats were studied: 1) control + vehicle, 2) control + calphostin C, 3) preconditioning + vehicle, and 4) preconditioning + calphostin C. All rats underwent 90 min of coronary occlusion followed by 4 h of reperfusion; in addition, preconditioned rats underwent three 3-min episodes of ischemia and 5 min of reperfusion before the 90 min of ischemia. Two injections of vehicle or calphostin C (0.1 mg/kg) were administered in intravenous boluses 29 min and 3 min before the 90-min coronary occlusion, i.e., one dose was given 5 min before preconditioning, and another dose was given between preconditioning and the sustained 90 min of ischemia in preconditioned rats. After 4 h of reperfusion, the area at risk (AR) was delineated by dye injection and area of necrosis was assessed by triphenyltetrazolium chloride staining. The electrocardiogram was recorded for the incidence of ventricular tachycardia (VT) and ventricular fibrillation. AR was similar in all four groups. In the nonpreconditioned control rats receiving vehicle, myocardial infarct size expressed as a percentage of the AR averaged 45.7 +/- 1.7%. Pretreatment with calphostin C had no effect on infarct size (48.9 +/- 3.4%) in nonpreconditioned control rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Analysis of Variance; Animals; Arrhythmias, Cardiac; Blood Pressure; Drug Administration Schedule; Female; Heart Rate; Myocardial Ischemia; Myocardial Reperfusion; Myocardium; Naphthalenes; Necrosis; Polycyclic Compounds; Premedication; Protein Kinase C; Rabbits; Rats; Rats, Sprague-Dawley; Reference Values; Time Factors