Resveratrol attenuates hypoxic injury in a primary hepatocyte model of hemorrhagic shock and resuscitation.
BACKGROUND
Oxidative stress following hemorrhagic shock and resuscitation (HSR) is regulated, in part, by inflammatory and apoptotic mediators such as necrosis factor κB (NF-κB) and p53. Sirtuin 1 (Sirt-1) is a metabolic intermediary that regulates stress responses by suppressing NF-κB and p53 activity. Resveratrol is a naturally occurring polyphenolic antioxidant and Sirt-1 agonist. The aim of this study was to determine whether resveratrol protects hepatocytes following HSR or hypoxia.


METHODS
In vivo, HSR was achieved in male rats by arterial blood withdrawal to 30 ± 2 mm Hg for 1 hour before resuscitation with or without resveratrol (Res, 30 mg/kg). Hepatic tissue was stained and scored for necrosis, interleukin 6, and Sirt-1 expression. In vitro, primary rat hepatocytes were subjected to 8 hours of hypoxia without or with Res (100 µM). Cells were analyzed immediately or after 6 hours of normoxia, for survival and markers of injury (lactate dehydrogenase assay, lipid peroxidation, and mitochondrial integrity). Cell lysates were collected for cytochrome c analysis and immunoprecipitated using antibodies against NF-κB (p65) or p53.


RESULTS
In vivo, animals subject to HSR exhibited increased expression of markers of hepatocyte damage compared with those sham operated, concomitant with lower Sirt-1 expression. In vitro, hypoxia followed by normoxia resulted in increased cell death, an effect that was blunted by Res. Analysis of cell and mitochondrial function demonstrated that Res inhibited the detrimental effects of hypoxia in isolated hepatocytes.


CONCLUSIONS
Resveratrol prevents cell death in HSR and exerts a protective effect on the mitochondria in a hepatocyte model of hypoxic injury-reoxygenation possibly via Sirt-1 modulation of p53 and NF-κB activity.
From the FH "Sammy" Ross Trauma Center, Department of Surgery, Carolinas Medical Center, Charlotte, North Carolina.
Rebecca D
RD
Powell
Jacob H
JH
Swet
Kenneth L
KL
Kennedy
Toan T
TT
Huynh
Iain H
IH
McKillop
Susan L
SL
Evans
D000818:Animals; D015153:Blotting, Western; D016923:Cell Death / Q000187:drug effects; D015687:Cell Hypoxia / Q000187:drug effects; D002470:Cell Survival; D004195:Disease Models, Animal; D004797:Enzyme-Linked Immunosorbent Assay; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D015850:Interleukin-6 / Q000032:analysis / Q000378:metabolism; D008297:Male; D008930:Mitochondria, Liver / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects*; D011897:Random Allocation; D051381:Rats; D017207:Rats, Sprague-Dawley; D012016:Reference Values; D012151:Resuscitation / Q000379:methods*; D000077185:Resveratrol; D012771:Shock, Hemorrhagic / Q000401:mortality / Q000503:physiopathology / Q000628:therapy*; D056564:Sirtuin 1 / Q000187:drug effects / Q000378:metabolism; D013267:Stilbenes / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D015850:Interleukin-6; D016328:NF-kappa B; D013267:Stilbenes; D014409:Tumor Necrosis Factor-alpha; D056564:Sirtuin 1; D000077185:Resveratrol