Hydrogen peroxide induces intracellular calcium oscillations in human aortic endothelial cells

Background-Because the vascular endothelium is exposed to oxidant stress resulting from ischemia/reperfusion and from the products of polymorphonuclear leukocytes or monocytes, studies were performed to examine the effect of hydrogen peroxide (1 mu mol/L to 10 mmol/L) on endothelial Ca2+ signaling, Methods and Results-At low concentrations (1 to 10 mu mol/L), hydrogen peroxide did not affect intracellular Ca2+ concentration in subconfluent, indo 1-loaded human aortic endothelial monolayers, At a concentration of 100 mu mol/L hydrogen peroxide, intracellular free Ca2+ gradually increased from 125.3 +/- 6.8 to 286.3 +/- 19.9 nmol/L over 4.2 +/- 0.9 minutes before repetitive Ca2+ oscillations were observed, consisting of an initial large, transient spike of approximate to 1 mu mol/L followed by several spikes of decreasing amplitudes at a frequency of 0.7 +/- 0.1 min(-1) over 12.0 +/- 1.1 minutes, After these oscillations, intracellular Ca2+ reached a plateau of 543.4 +/- 64.0 nmol/L, which was maintained above baseline levels for > 5 minutes and then partially reversible on washout of hydrogen peroxide in most monolayers, Intracellular Ca2+ oscillations were typically observed when monolayers were exposed to 100 to 500 mu mol/L hydrogen peroxide. Higher concentrations of hydrogen peroxide (1 and 10 mmol/L) increased intracellular Ca2+ but only rarely (2 of 6 monolayers at 1 mmol/L) or never (at 10 mmol/L) stimulated intracellular Ca2+ oscillations. Removal of Ca2+ from the buffer either before hydrogen peroxide stimulation or during an established response did not block intracellular Ca2+ oscillations in response to 100 mu mol/L hydrogen peroxide, but prior depletion of an intracellular Ca2+ store with either caffeine, histamine, or thapsigargin abolished Ca2+ oscillations. Conclusions-Hydrogen peroxide induces concentration-dependent intracellular Ca2+ oscillations in human endothelial cells, which results from release of an endoplasmic reticulum Ca2+ store. Because oxidant production appears to occur In the micromolar range in the postischemic/anoxic endothelium and is associated with impaired endothelium-dependent relaxation, the effects of micromolar concentrations of hydrogen peroxide on endothelial Ca2+ signaling described in the present study maybe important in the pathogenesis of postischemic endothelial dysfunction.