Endotoxin stimulates hydrogen peroxide detoxifying activity in rat hepatic endothelial cells.
The study aimed to assess the effect of lipopolysaccharide (LPS) in vivo (from Escherichia coli, 2 mg/kg body weight intraperitoneally) on the production and elimination of hydrogen peroxide (H2O2) in rat hepatic endothelial and Kupffer cells. Twenty-two hours after the injection of LPS, hepatic cells were isolated by collagenase and pronase digestion followed by centrifugal elutriation, and cell-associated H2O2 was determined by flow cytometry analysis using 2',7'-dichloroflorescin diacetate (DCF-diacetate). LPS treatment did not alter the basal or phorbol myristate acetate-stimulated levels of H2O2-related fluorescence in endothelial cells; however, it doubled phorbol myristate acetate-stimulated fluorescence in Kupffer cells. Administration of varying concentrations of H202 (range, 10(-7) - 10(-4) mol/L) in vitro caused a significantly delayed increase in fluorescence in endothelial cells from endotoxemic rats as compared with cells from saline-injected animals. The 50% effective concentration of H202 was found at 1.1 x 10(-6) and 8.1 x 10(-6) mol/L on endothelial cells after saline and LPS treatment, respectively. No differences were detected in H2O2-stimulated fluorescence between resting and LPS-stimulated Kupffer cells. Administration of varying glucose concentrations in vitro significantly decreased the H2O2-stimulated fluorescence in endothelial and Kupffer cells from LPS-injected animals. Inhibition of nitric oxide synthase by in vitro administration of NG-monomethyl-L-arginine (L-NNMMA) did not alter the H2O2- or phorbol myristate acetate-stimulated responses in endothelial and Kupffer cells. As shown earlier, LPS stimulates the gene expression of GLUT1 glucose transporter, glucose-6-phosphate dehydrogenase (G6PD), superoxide dismutases, and glutathione peroxidase in hepatic endothelial cells. The present data indicate that the LPS-induced metabolic alterations are accompanied by an increased H2O2-detoxifying capacity in hepatic endothelial cells. This may represent a protective mechanism against exogenous oxidative stress caused by activated hepatic phagocytes during inflammation. Our observations are consistent with primed production of reactive oxygen species (ROS) in LPS-activated Kupffer cells.
Spolarics
Z
Z
Department of Anatomy, Cell Biology, and Injury Sciences, University of Medicine and Dentistry of New Jersey, Newark, NJ, USA.
Stein
D S
DS
Garcia
Z C
ZC
D000818:Animals; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D004731:Endotoxins / Q000494:pharmacology*; D005453:Fluorescence; D005947:Glucose / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000378:metabolism* / Q000494:pharmacology; D007274:Injections, Intraperitoneal; D007728:Kupffer Cells / Q000187:drug effects / Q000502:physiology; D008102:Liver Circulation; D008297:Male; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D051381:Rats; D017207:Rats, Sprague-Dawley; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D019323:omega-N-Methylarginine / Q000494:pharmacology
D004731:Endotoxins; D019323:omega-N-Methylarginine; D006861:Hydrogen Peroxide; D019001:Nitric Oxide Synthase; D005947:Glucose; D013755:Tetradecanoylphorbol Acetate