Role of the life span determinant P66(shcA) in ethanol-induced liver damage.
Mice lacking the 66 kDa isoform of the adapter molecule shcA (p66(shcA)) display increased resistance to oxidative stress and delayed aging. In cultured cell lines, p66 promotes formation of Reactive Oxygen Species (ROS) in mitochondria, and apoptotic cell death in response to a variety of pro-oxidant noxious stimuli. As mitochondrial ROS and oxidative cell damage are clearly involved in alcohol-induced pathology, we hypothesized that p66 may also have a role in ethanol. In vivo, changes observed in p66+/+ mice after 6-week exposure to ethanol in the drinking water, including elevated serum alanine aminotransferase (ALT), liver swelling and evident liver steatosis, were significantly attenuated in p66-/- mutant mice. Biochemical analysis of liver tissues revealed induction of the p66 protein by ethanol, whereas p66-deficient livers responded to alcohol with a significant upregulation of the mitochondrial antioxidant enzyme MnSOD, nearly absent in control mice. Evidence of an inverse correlation between expression level of p66 and protection from alcohol-induced oxidative stress was also confirmed in vitro in primary hepatocytes and in HepG2-E47 cells, an ethanol-responsive hepatoma cell line. In fact, MnSOD upregulation by exposure to ethanol in vitro was much more pronounced in p66KO versus wild-type isolated liver cells, and blunted in HepG2 cells overexpressing p66shc. p66 overexpression also prevented the activation of a luciferase reporter gene controlled by the SOD2 promoter, indicating that p66 repression of MnSOD operates at a transcriptional level. Finally, p66 generated ROS in HepG2 cells and potentiated oxidative stress and mitochondrial depolarization by ethanol. Taken together, the above observations clearly indicate a role for p66 in alcohol-induced cell damage, likely via a cell-autonomous mechanism involving reduced expression of antioxidant defenses and mitochondrial dysfunction.
Koch
Osvaldo R
OR
Department of Pathology, Faculty of Medicine University of Buenos Aires, Buenos Aires, Argentina.
Fusco
Salvatore
S
Ranieri
Sofia Chiatamone
SC
Maulucci
Giuseppe
G
Palozza
Paola
P
Larocca
Luigi Maria
LM
Cravero
Amerys A M
AA
Farre'
Stella M
SM
De Spirito
Marco
M
Galeotti
Tommaso
T
Pani
Giovambattista
G
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000502:physiology*; D000410:Alanine Transaminase / Q000097:blood; D000818:Animals; D006528:Carcinoma, Hepatocellular; D045744:Cell Line, Tumor; D000431:Ethanol / Q000494:pharmacology*; D005235:Fatty Liver, Alcoholic / Q000378:metabolism / Q000473:pathology; D005786:Gene Expression Regulation; D022781:Hepatocytes / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008113:Liver Neoplasms; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008930:Mitochondria, Liver / Q000378:metabolism; D020033:Protein Isoforms / Q000235:genetics / Q000502:physiology; D017382:Reactive Oxygen Species / Q000378:metabolism; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D013482:Superoxide Dismutase / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D020033:Protein Isoforms; D017382:Reactive Oxygen Species; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; C528034:Shc1 protein, mouse; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D000431:Ethanol; D013482:Superoxide Dismutase; D000410:Alanine Transaminase