Heat shock induces IkappaB-alpha and prevents stress-induced endothelial cell apoptosis

S. L. DeMeester, T. G. Buchman, Y. Qiu, A. K. Jacob, K. Dunnigan, R. S. Hotchkiss, I. Karl and J. P. Cobb
Department of Surgery, Washington University School of Medicine, St Louis, Mo, USA.

OBJECTIVE: To determine whether prior heat shock would attenuate endothelial cell apoptosis and whether any effect of preemptive heat shock is mediated through a nuclear factor kappa B and inhibitor kappa B alpha mechanism. DESIGN: A randomized, controlled in vitro study. SETTING: A laboratory in a large, academic medical center. INTERVENTIONS: Cultured primary porcine endothelial cells were treated with increasing doses of sodium arsenite (40-160 micromol/L), after which the interval until subsequent apoptotic (lipopolysaccharide-arsenite) challenge was varied (4-16 hours). The degree of cell death and apoptosis were determined using neutral red uptake and staining with annexin V and propidium iodide, respectively. Inducible heat shock protein 70 and inhibitor kappa B alpha levels in treated cells were determined by Western blot analysis. Lipopolysaccharide-induced nuclear factor kappa B activity was assessed using an electrophoretic mobility shift assay. RESULTS: Prior arsenite treatment decreased cell death by apoptosis in a time- and dose-dependent manner. Specifically, a higher sodium arsenite concentration and shorter intervals afforded better protection (P=.01, 160 micromol/L at 4 hours). Protection against apoptosis correlated with increased heat shock protein 70 and inhibitor kappa B alpha levels and decreased nuclear factor kappa B binding activity. CONCLUSIONS: Arsenite, an inducer of the heat shock response, decreased stress-induced endothelial cell apoptosis. The mechanism of this protection may include decreased nuclear factor kappa B activity or increased inducible heat shock protein 70 levels. Heat shock protein 70 may serve as a molecular marker to determine not only the phenotypic state of the cell but also the durability of protection afforded by heat shock. These data support the hypothesis that stress-induced changes in transcription factor activity and protein expression can regulate the induction of apoptosis.