Alcohol increases hepatic oxygen consumption by affecting communication between Kupffer cells and parenchymal cells

This dissertation was designed to more fully characterize the mechanisms of elevated O{dollar}sb2{dollar} metabolism in response to alcohol in order to develop more effective preventive and therapeutic strategies for alcoholic liver injury.; The effect of conditioned media from Kupffer cells isolated from rats chronically exposed to ethanol on O{dollar}sb2{dollar} uptake of normal parenchymal cells was studied to evaluate the possibility that cell to cell communication is involved in this mechanism. The results showed that O{dollar}sb2{dollar} uptake was increased more than 30% by this conditioned media. Nisoldipine and indomethacin blocked this stimulation, suggesting the involvement of eicosanoids. Prostaglandin E{dollar}sb2{dollar} (PGE{dollar}sb2){dollar} added directly to parenchymal cells increased O{dollar}sb2{dollar} uptake nearly 60% and PGE{dollar}sb2{dollar} levels were elevated twofold. The addition of endotoxin to cultured cells caused a similar phenomenon. Thus, Kupffer cells are activated by ethanol treatment to release PGE{dollar}sb2{dollar} which stimulates O{dollar}sb2{dollar} uptake in parenchymal cells, possibly by mechanisms involving bacterial endotoxin.; To characterize subtypes of PGE{dollar}sb2{dollar} receptors and signal transduction pathways involved in elevated O{dollar}sb2{dollar} uptake, 17-phenyl-omega-trinor-PGE{dollar}sb2{dollar} (17-PGE{dollar}sb2),{dollar} an EP{dollar}sb1{dollar}-specific agonist, and 11-deoxy PGE{dollar}sb1,{dollar} (11-PGE{dollar}sb1),{dollar} an EP{dollar}sb2{dollar}-specific agonist, were added to isolated parenchymal cells and O{dollar}sb2{dollar} uptake was measured. 11-PGE{dollar}sb1{dollar} stimulated O{dollar}sb2{dollar} uptake in a dose-dependent manner; however, 17-PGE{dollar}sb2{dollar} had no effect. PGE{dollar}sb2{dollar} stimulated a dose-dependent increase in cAMP formation which correlated well with increased O{dollar}sb2{dollar} uptake in isolated parenchymal cells. Dibutyryl cAMP also caused a dose-dependent increase in O{dollar}sb2{dollar} uptake, mimicking the effect of PGE{dollar}sb2.{dollar} The PGE{dollar}sb2{dollar}-stimulated increase in O{dollar}sb2{dollar} uptake was blocked by H-89, a protein kinase A inhibitor, antimycin A, which inhibits the respiratory chain between cytochrome b and cytochrome c, and potassium cyanide, an inhibitor of mitochondrial cytochrome oxidase. Incubation of mitochondria with the purified catalytic subunit of the cAMP-dependent protein kinase significantly increased both state 3 rates of O{dollar}sb2{dollar} uptake and the respiratory control ratio. Both processes were prevented by incubation with the PKA inhibitory peptide, PKI. Thus, the ability of PGE{dollar}sb2{dollar} to stimulate O{dollar}sb2{dollar} uptake is mediated via the EP{dollar}sb2{dollar} subclass of receptors; increased cAMP activates cAMP-dependent protein kinase. It is proposed that phosphorylation of mitochondrial components in turn stimulates mitochondrial O{dollar}sb2{dollar} uptake.; In liver perfusion studies, PGE{dollar}sb2{dollar} infusion increased O{dollar}sb2{dollar} uptake at normal flow rates; however, the increase was nearly twice as large at high flow rates. Concomitantly, glucose output increased rapidly while glycolysis decreased. The PGE{dollar}sb2{dollar}-induced increase in O{dollar}sb2{dollar} uptake is oxygen dependent. High flow rates also increased basal rates of O{dollar}sb2{dollar} uptake. The increase in O{dollar}sb2{dollar} uptake due to high flow rates was prevented by inactivation of Kupffer cells with GdCl{dollar}sb3.{dollar} Conditioned medium from Kupffer cells incubated at high oxygen tension (75% O{dollar}sb2){dollar} stimulated parenchymal cell O{dollar}sb2{dollar} uptake over 30% and elevated PGE{dollar}sb2{dollar} production about two-fold compared to cultures exposed to 21% O{dollar}sb2.{dollar} These effects were blocked completely by both indomethacin and nisoldipine.