P22 Effect of hydrogen sulfide on Ca2+ signaling in endothelial and smooth muscle cells

Abstract

Endothelial cells are active participants in inflammatory processes. They are involved in diverse activities including the regulation of leucocyte extravasation, angiogenesis, cytokine production, protease and extracellular matrix synthesis, vasodilation, etc. The small gaseous molecule hydrogen sulfide (H 2 S) is involved in a variety of physiological processes like vascular relaxation, angiogenesis, neurotransmission and inflammation. In the vascular system, ATP-sensitive K + -channels are a major target for H 2 S but over the last few years evidence has accumulated that Na + - and Ca 2+ -permeable channels as well as intracellular Ca 2+ stores are also modulated by H 2 S. In the present study we investigated the effect of H 2 S on Ca 2+ signaling in cultured endothelial and smooth muscle cells with special emphasis given to the role of H 2 S in modulating Ca 2+ entry as well as receptor-mediated release of Ca 2+ from intracellular stores. Experiments were performed with human microvascular endothelial cells (HMEC-1), endothelial cells isolated from porcine aorta, and smooth muscle cells isolated from rat aorta and rat trachea. Mobilization of intracellular Ca 2+ and Ca 2+ entry was monitored by measuring the intracellular free Ca 2+ concentration with FURA-2 in the absence and presence extracellular Ca 2+ , respectively. Activity of endothelial nitric oxide synthase (eNOS) in intact cells was determined as conversion of incorporated l -[ 3 H]-arginine into l -[ 3 H]-citrulline. Incubation of human and porcine endothelial cells with the H 2 S-donor NaHS (100 μM, 10–45 min) markedly diminished the release of Ca 2+ from intracellular stores elicited by receptor agonists such as ATP or histamine. As a consequence, the stimulatory effect of these agonists on endothelial NO formation was strongly diminished, whereas the response to the Ca 2+ ionophore A23187 was barely affected. In contrast to its inhibitory effect on Ca 2+ release, NaHS stimulated Ca 2+ influx from the extracellular space, but the resulting increase in intracellular free Ca 2+ concentration was not sufficient to affect endothelial NO formation. In smooth muscle cells isolated from rat aorta or rat trachea NaHS also blocked receptor-mediated release of Ca 2+ from intracellular stores but in contrast to the results obtained with endothelial cells, no stimulatory effect of NaHS on Ca 2+ entry was observed. H 2 S inhibits the stimulatory effect of Ca 2+ mobilizing agonists on endothelial NO formation by attenuating the release of Ca 2+ from intracellular stores. Inhibition of Ca 2+ release by H 2 S is not a peculiarity of endothelial cells but also occurs in vascular and tracheal smooth muscle cells. These hitherto undescribed effects may be in part responsible for the beneficial effects of H 2 S in sulfur bath therapy.