A Small Dose of Hydrogen Peroxide Enhances Tumor Necrosis Factor-Alpha Toxicity in Inducing Human Vascular Endothelial Cell Apoptosis: Reversal with Propofol

In Brief We designed the present study to test the hypothesis that oxygen free radicals can enhance tumor necrosis factor (TNF)-α cellular toxicity, which might be reversed by propofol, an anesthetic with antioxidant properties, in human vascular endothelial cell line ECV304. Cultured ECV304 were either not treated, treated with 10 μM of hydrogen peroxide (H2O2), treated with TNF-α (40 ng/mL) alone, TNF-α in the presence of 10 μM of H2O2 (H+T), or propofol plus H2O2 for 24 h. Cell viability was measured by lactate dehydrogenate (LDH) assay. Cell apoptosis was assessed by flow cytometry and terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP) nick end-labeling. The antiapoptotic Bcl-2 and pro-apoptotic Bax protein expressions were measured by immunocytochemical analysis. Increases in apoptosis, Bax, lipid peroxidation product malondialdehyde, LDH, and decreases in Bcl-2, superoxide dismutase, and glutathione peroxidase were observed in TNF-α–treated cells. H2O2 10 μM did not cause significant lipid peroxidation (0.75 ± 0.03 nmol/mg of malondialdehyde protein) as compared with control (0.70 ± 0.04 nmol/mg of malondialdehyde protein) (P > 0.05) but further enhanced TNF-α–induced lipid peroxidation, upregulated Bax, and down-regulated Bcl-2 expression and enhanced TNF-α–induced cell apoptosis (P < 0.05). Propofol 50 μM attenuated TNF-α and H2O2-induced cell apoptosis, accompanied by decreases in malondialdehyde and LDH production and restoration of Bcl-2 expression. Propofol exerts protective effects against H2O2-enhanced TNF-α cell toxicity by reducing oxidative injury. IMPLICATIONS: Hydrogen peroxide, at a small concentration (10 μM) that did not cause significant lipid peroxidation per se, significantly enhanced tumor necrosis factor (TNF)-α–induced endothelial cell lipid peroxidation and programmed cell death. Propofol prevented this synergistic effect of hydrogen peroxide and TNF-α and may be beneficial in some clinical circumstances associated with increased oxidative stress.