Glutathione attenuates ethanol-induced alveolar macrophage oxidative stress and dysfunction by downregulating NADPH oxidases.
Chronic alcohol abuse increases lung oxidative stress and susceptibility to respiratory infections by impairing alveolar macrophage (AM) function. NADPH oxidases (Nox) are major sources of reactive oxygen species in AMs. We hypothesized that treatment with the critical antioxidant glutathione (GSH) attenuates chronic alcohol-induced oxidative stress by downregulating Noxes and restores AM phagocytic function. Bronchoalveolar lavage (BAL) fluid and AMs were isolated from male C57BL/6J mice (8-10 wk) treated ± ethanol in drinking water (20% wt/vol, 12 wk) ± orally gavaged GSH in methylcellulose vehicle (300 mg x kg(-1) x day(-1), during week 12). MH-S cells, a mouse AM cell line, were treated ± ethanol (0.08%, 3 days) ± GSH (500 μM, 3 days or last 1 day of ethanol). BAL and AMs were also isolated from ethanol-fed and control mice ± inoculated airway Klebsiella pneumoniae (200 colony-forming units, 28 h) ± orally gavaged GSH (300 mg/kg, 24 h). GSH levels (HPLC), Nox mRNA (quantitative RT-PCR) and protein levels (Western blot and immunostaining), oxidative stress (2',7'-dichlorofluorescein-diacetate and Amplex Red), and phagocytosis (Staphylococcus aureus internalization) were measured. Chronic alcohol decreased GSH levels, increased Nox expression and activity, enhanced oxidative stress, impaired phagocytic function in AMs in vivo and in vitro, and exacerbated K. pneumonia-induced oxidative stress. Although how oral GSH restored GSH pools in ethanol-fed mice is unknown, oral GSH treatments abrogated the detrimental effects of chronic alcohol exposure and improved AM function. These studies provide GSH as a novel therapeutic approach for attenuating alcohol-induced derangements in AM Nox expression, oxidative stress, dysfunction, and risk for pneumonia.
Dept. of Pediatrics, Division of Neonatal-Perinatal Medicine, Emory Univ., 2015 Uppergate Dr., Atlanta, GA 30322. lbrow03@emory.edu.
Samantha M
SM
Yeligar
Frank L
FL
Harris
C Michael
CM
Hart
Lou Ann S
LA
Brown
D000437:Alcoholism / Q000276:immunology* / Q000378:metabolism; D000818:Animals; D000975:Antioxidants / Q000378:metabolism* / Q000494:pharmacology; D001992:Bronchoalveolar Lavage Fluid / Q000276:immunology; D002460:Cell Line; D002492:Central Nervous System Depressants / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects / Q000276:immunology; D004789:Enzyme Activation / Q000187:drug effects / Q000276:immunology; D000431:Ethanol / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism* / Q000494:pharmacology; D007710:Klebsiella Infections / Q000276:immunology / Q000378:metabolism; D007711:Klebsiella pneumoniae / Q000276:immunology; D016676:Macrophages, Alveolar / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D000074624:NADPH Oxidase 1; D000074662:NADPH Oxidase 2; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects / Q000276:immunology; D010750:Phosphoproteins / Q000378:metabolism
D000975:Antioxidants; D002492:Central Nervous System Depressants; D008562:Membrane Glycoproteins; D010750:Phosphoproteins; C079789:neutrophil cytosol factor 67K; D000431:Ethanol; D009247:NADH, NADPH Oxidoreductases; C488842:Cybb protein, mouse; D000074624:NADPH Oxidase 1; D000074662:NADPH Oxidase 2; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C422943:Nox4 protein, mouse; C060505:neutrophil cytosolic factor 1; D005978:Glutathione
alveolar macrophage; glutathione; nicotinamide adenine dinucleotide phosphate oxidases; oxidative stress; phagocytic function