The Protective Effect Of Recombinant Adenovirus(Ad.HO-1) On Liver Cell Against Hypoxia-reoxygenation Injury In Vitro

Hepatic ischemia reperfusion injury (IRI) is a pathophysiologic process, whichoccured in many clinical courses, such as hemorrhage shock, toxic shock, livertransplantation, liver resection and so on. After hypoxia, the injury of blood reflow intoorgans has an important influence on the outcome of the shock/resuscitation and the livergraft survival rate after liver transplantation. Thus, the investigation of hepatic ischemiareperfusion injury will have important meaning to promote the recovery of the organfunction after ischemia.Heme oxygenase-1 (HO-1) is a rate-limiting enzyme, also known as heat shockprotein 32 (hsp 32). It is one of the three HO isoforms that catalyze the degradation ofheme to biliverdin, Fe²⁺, and carbon monoxide (CO), but it is the only one that is inducible.Like other hsp, upregulation of HO-1 may be among the critical cytoprotectivemechanisms that are activated during times of cellular stress such as inflammation,ischemia, hypoxia, hyperoxia, hyperthermia, or radiation. A growing body of evidencesuggests that overexpression of heme oxygenase-1 (HO-1) may protect organs/tissues fromimmune-mediated injury either through prevention of oxidative damage or via a localimmunomodulatory influence on infiltrating inflammatory cells. This protective propertyhas been observed in models of organ transplant rejection. Although HO-1 can be inducedby various stimuli, such as heme and Coprotoporphyrin(CoPP) and so on, exogenousadministration of HO-1 by gene transfer seems a more specific and attractive approach.Objective To construct a recombinant adenovirus vectors (Ad. HO-1), and study itsprotective effect on liver cell through hypoxia-reoxygenation.Methods Recombinant adenovirus (Ad. HO-1) had been constructed with full length human HO-1 gene by Cre-mediated site-specific recombination method. Culturedliver cell were divided into four group: the control group was transfected with vacantrecombinant adenovirus, three group were transfected with Ad. CMV-HSP70 for 24h,48h,72h respectively. The expression of transfected Ad. HO-1 was detected by RT-PCR aftertransfeetion. After the cells in the four groups suffered 4 hours of hypoxia followed by 8hours of reoxygenation, the cell viability was analyzed by MTT method.Results Compared with the control group, the expression of HO-1 were obviouslyincreased after transfection, and the peak level was at 48h after transfection. Afterhypoxia-reoxygenation, the cell viability rates of HO-1 gene transfected groups were29.0%,85.6%,84.6% respectively, all significantly higher than that of control group(2.0%,P＜0.01).Conclusion The over expression of human HO-1 mediated by recombinantadenovirus can protects liver cell against hypoxia-reoxygenation injury in vitro.