Prostanoid release by Kupffer cells upon hypoxia-reoxygenation: role of pHi and Cai2+

Primary cultures of rat Kupffer cells liberated significant amounts of prostaglandin (PG) D2, PGE2, and thromboxane (measured as thromboxane B2) when exposed to reoxygenation after 4 h of hypoxia. After a delayed onset, prostanoids were released at high rates for at least 8 h and after that time 700 pmol PGD2, 280 pmol PGE2, and 200 pmol thromboxane per 10(6) cells had been liberated. Unlike prostanoid release, leukotriene B4 production in reoxygenated cell cultures was only twice as much as in aerobic controls. Superoxide dismutase and catalase had no effect on PGD2, PGE2, and thromboxane production, indicating that prostanoid formation was independent of reactive oxygen species generated extracellularly and of cell injury. On the other hand, diphenyliodonium, as well as amiloride, blocked hypoxia-reoxygenation-induced PGD2, PGE2, and thromboxane release. The elevated prostanoid synthesis was preceded by increases in intracellular pH (from 7.23 to 7.38) and in intracellular Ca2+ (from 55 nM to a maximum level of 807 nM). These observations suggest a participation of NADPH oxidase and a related Na(+)-H+ exchange in the enhanced prostanoid synthesis, probably through the induction of an increased intracellular Ca2+ concentration.