Comparison of formation of D2/E2-isoprostanes and F2-isoprostanes in vitro and in vivo -: Effects of oxygen tension and glutathione

The isoprostanes (IsoPs) are bioactive prostaglandin-like compounds derived from the free-radical-catalyzed peroxidation of arachidonic acid in vitro and in vivo. IsoPs possessing either an F-type prostane ring (F-2-IsoPs) or D/E-type prostane rings (D2E2-IsoPs) are formed depending on whether IsoP endoperoxide intermediates undergo reduction or isomerization, respectively. Little, however, is known regarding factors influencing the formation of various classes of IsoPs, particularly D-2/E-2-IsoPs. Thus, studies were undertaken to examine the formation of D-2/E-2-IsoPs in relation to F-2-Isops both in vitro and in vivo. In peroxidizing rat liver microsomes, the formation of D-2/E-2-IsoPs increased in a time-and oxygen dependent manner and correlated with F-2-IsoP generation and loss of precursor arachidonic acid, although the absolute amount of D2E2-IsoPs formed exceeded by over B-fold the levels of F-2-IsoPs formed, Surprisingly, however, in liver tissue from rats exposed to an oxidant stress, levels of F-2-IsoPs were up to 10-fold greater than those of D-2/E-2-IsoPs, suggesting that an endogenous process causes IsoP endoperoxide reduction in vivo. Addition of glutathione (GSH) to peroxidizing microsomes at concentrations from 0.01 to 5 mM increased the formation of F-2-IsoPs at the expense of D-2/E-2-IsoPs. Boiling of microsomes did not alter the effect of GSH. Formation of D2E2-IsoPs in liver tissue in vivo was greatly enhanced compared to F-2-IsoPs in rats depleted of GSH. Thus, GSH modulates the formation of different classes of IsoPs in vitro and in vivo. Other thiols, including beta-mercaptoethanol, dithiothreitol, and cysteine, were able to substitute for GSH. These studies indicate that GSH promotes F-2-IsoP formation and diminishes D-2/E-2-IsoP levels in vitro and in vivo by causing reduction of IsoP endoperoxides. (C) 1998 Academic Press.