Role Of Nrf2in MSCs Regulating Oxidative Stress In Infracted Myocardium

Objective:To investigate the inhibitory effect of transplanted MSCs on the oxidative stress in rat myocardium,and evaluate the function of Nrf2/ARE pathway in the inhibitory effect.Method:108rats are randomized into3groups:Sham group、myocardial infarction grouop (MI group) and MSCs transplantation after myocardial infarction group (MSCs group). Each group is divided into6subgroups:2hours after MI subgroup (2h subgroup),8h subgroup,24h subgroup,7d subgroup,14d subgroup and28d subgroup. Rats are executed at6time points, the oxidative stress of myocardium is detected and the expression levels of Nrf2and HO-1gene and protein are detected by RT-PCR and Western blotting. Rats are executed28days after myocardial infarction and myocardial morphological changes are observed by HE staining and Masson staining. Survival and differentiation of MSCs are detected by immunofluorescence technic.Results:1.HE staining28days after infarction shows that myocardial cells in sham operation groups are in neat order, while the myocardial cells in the control groups have been replaced by fiber cicatrical tissue. The MSCs groups area of infarction fibre cicatrical tissue has obviously decreased, compared with the control group. The Masson staining result shows that myocardial tissue in sham operation groups is stained red without blue collagenous fibre. In the control groups, collagenous fibre is more clearly formed. The blue collagenous fibre in MSCs group has obviously decreased, compared with the control group.2. After observation under fluorescence microscope,it is found out that DAPI-marked stem cells still exist28days after infarction. Immunofluorescence detection for myocardial troponin shows that troponin is found not only in myocardial cells, but also in DAPI-marked MSCs endochylema.3. Oxidative stress level:SOD level, GSH level, the Capacity of restraining hydroxyl radical and MDA level had no significant changes in Sham operation groups. There were significant reduces in SOD level, GSH level and the Capacity of restraining hydroxyl radical2hours after myocardial infarction(P<0.05) compared with Sham operation groups,while MDA level increased dramatically(P<.0.05). SOD level, GSH level and the Capacity of restraining hydroxyl radical kept on increasing from2hours to24hours after myocardial infarction and the peak was reached at24hours.MDA level increased from2hours to24hours after myocardial infarction and the peak was reached at24hours. SOD level in MI groups were still lower than in Sham operation groups(P<0.05),while GSH level in the two groups had no significant differences(P>0.05). the Capacity of restraining hydroxyl radical and MDA level in MI groups were significant higher than Sham operation groups(P<0.05). SOD level, GSH level and the Capacity of restraining hydroxyl radical reduced in7days and14days and then increased in28days. SOD level, GSH level and the Capacity of restraining hydroxyl radical in MI groups in28days were significantly lower than in Sham operation groups(P<0.05),while MDA level kept on decreasing from7days to28days.The lowest MDA level which was reached at28days in MI groups was still significantly higher than in Sham operation groups(P<0.05). SOD level, GSH level, the Capacity of restraining hydroxyl radical and MDA level in each time point in MSCs groups had similar changes with MI groups.But SOD level, GSH level and the Capacity of restraining hydroxyl radical in MSCs groups were higher than in MI groups(P<0.05) in each time point while MDA level in MI groups were significantly lower than in MSCs groups(P<0.05) in each time point.4. PCR and Western blotting analyse:Nrf2and HO-1expressing levels began to rise after myocardial infarction and the peaks ware reached at24hours. The levels in7d groups and14d groups were much lower than24h groups, but still higher than Sham groups.In28d the expressing levels came to another peak. Nrf2and HO-1expressing levels were higher in MSCs groups than in MI groups except in2h.Conclusion:1. Transplanted MSCs can decrease oxidative stress in infracted myocardium.2. MSCs can increase Nrf2and HO-1expression in both gene and protein levels and the inhibitory function of MSCs to oxidative stress may be mediated by Nrf2/ARE pathway.