Stimulation of prostaglandin synthesis in cultured liver cells by CCl4

Abstract

The contribution of hepatocytes to liver prostaglandin synthesis is controversial, partly because hepatocytes of varying purity have been studied. In this study, prostaglandin synthesis was examined in conventional and ricin-purified rat hepatocytes that were incubated with gaseous carbon tetrachloride as a model stimulus for lipid peroxidation and prostaglandin synthesis. Hepatocytes that were incubated for 2 hours with 1 ml, of liquid CCl4/5 L gas volume showed no evidence of cell death or injury. However, twice this volume of CCl4 caused total cell death. Enzyme immunoassay (EIA) failed to detect prostaglandin E2 (PGE2) synthesis in hepatocyte cultures after the first day in culture, under a variety of cell culture conditions. Conventional hepatocyte cultures, but not ricin-purified hepatocytes, synthesized thromboxane B2 (approximately 300 pg/mg protein) and prostaglandin D2 (PGD2) (range, 1,000-6,000 pg/mg protein). Conventional hepatocyte cultures released prostaglandin F2alpha (PGF2alpha) immunoreactivity (ranging from approximately 900 to 1,800 pg/mg protein). Ricin-purified hepatocyte cultures made at least half as much PGF2 alpha immunoreactivity as did corresponding conventional hepatocytes. However, cyclooxygenase inhibitors did not inhibit the portion of PGF2 alpha immunoreactivity made by ricin-purified hepatocytes. PGF2 alpha immunoreactivity released by ricin-purified hepatocytes cochromatographed with PGF2 alpha, and probably represents F2-isoprostanes resulting from lipid peroxidation in hepatocytes. F2-isoprostane release was detected by immunoassay. Conventional cultures of rat hepatocytes contain Kupffer and endothelial cells, which can synthesize significant amounts of cyclooxygenase products. Highly purified hepatocytes do not produce cyclooxygenase products, even with a maximal stimulus to lipid peroxidation. CCl4 causes the release of F2-isoprostanes from hepatocytes with or without observable cell injury, as detected by Trypan blue exclusion or lactate dehydrogenase (LDH) release.