| Objective. To study the injuries of heart induced by Chronic Obstructive Pulmonary Disease (COPD) in old rats; 2. To Observe the oxidative stress modulation mechanisms of SIRT1 during this process; 3. To explore how resveratrol and SIRT1 exert antioxidant, antiapoptotic effects, which protect the heart against the adverse effects of COPD induced by cigarette smoking exposure and LPS.Methods. In vivo experiments, SD rats at 3-month-old and 22-month-old were divided into five groups:the young control group (12), the young COPD group (12), the old control group (12), the old COPD group (11), the old COPD group treated with resveratrol (COPD+Res) group (12). ALL of the COPD groups (n= 35) were exposed to the smoke of 8 commercial cigarettes (11 mg tar and 0.8 mg nicotine/cigarette) each day for 8 weeks and instillation given of LPS 200μg/200μl twice, whereas the control group was not exposed to cigarette smoking. To assess the heart protective effects of resveratrol, the COPD+Res group of rats was pretreated with resveratrol 25 mg-kg-1·day-1for 8 weeks and then exposed to the above-described cigarette smoking protocol. The changes of weight, heart weight, LVEF,LVEDD, RVP, PAP, LVEDP, SOD, MDA,8-OHdG, collagen, cadiocyte apoptosis were detected. The expression of SIRT1 were evalulated by Taqman real-time RT-PCR and Western Blot. The expression of p-FOXO3a were evalulated by Western blot. In vitro Experiments 1. H9C2 cells were divided into control group (C group), solvent control group (DMSO group), cigarette smoking extraction stimulus group (CSE group), CSE group with Res treatment group (CSE+Res group) and Res group.25μg/ml CSE intervened to CSE group from 3-72h and Res group was pre treated with 20μmol/L Res 30min before. Res group was treated with the equal amount of Res.2. Cell proliferation was investigated by MTT method and the CSE concentration was determined. Specific fluorescent probe CM-H2DCFDA and fluorescent micro-plate method was employed for the determination of ROS expression. Mitochondrial membrane potential was measured by fluorescence spectrophotometer and the level of apoptosis was measured by activity of caspase 3. RT-PCR was employed for the determination of the expression of SIRT1 mRNA.The protein of SIRT1 and p-FOXO3a expressions were observed by Weston Blot.Results Part 1 1.The changes in histopathology of lung in rats of the two COPD groups were similary to those in COPD pantients.2.Compared with old rats in control group, the obvious increase were found in HW/BW, thickness of left ventricle, diameter of cadiocyte, collagen, LVEDD, PAP, LVEDP,8-OHdG and MDA in rats of old COPD group (P<0.01), and significantly decrease were found in SOD (P<0.01). It was found more hypertrophy changes in the HE-stained myocardial cells of old COPD group than that of old control group, while we also found the myocardial fibrosis significantly increased by masson staining. Part 2 In vivo experiments 1. Compared with rats in old control group, the expression of SIRT1mRNA and protein decreased in rats of old COPD group (P<0.05), the expression of SIRT1 staining by immunofluorescence decreased, and the expression of p-FOXO3a protein decreased (P<0.05).2. Compared with rats in old COPD group, the expression of SIRT1mRNA and protein increased in old COPD+Res group (P<0.05), the expression of SIRT1 staining by immunefluorescence increased, and the expression of p-FOXO3a protein increased (P<0.05).3.Compared with rats in old COPD group, the obvious decrease were found in thickness of left ventricle, diameter of cadiocyte, collagen, LVEDD, PAP, LVEDP, 8-OHdG and MDA of old COPD+Res group (P<0.05),and significantly increase were found in SOD (P<0.01).4.Compared with old control group, the apoptosis index of cardiac muscle increased in old COPD group (P<0.05).The apoptosis index decreased in old COPD+Res group as opposed to old COPD group (P<0.05). In vitro experiments 1. The expression of ROS increased in H9C2 cells of CSE group against the cells of C group (P<0.05), whereas and the expression of SIRT1 and p-FOXO3a decreased (P<0.05), the activity of caspase-3 increased(P<0.05)in CSE group; 2.The expression of ROS decreased in CSE+Res group as compared with the H9C2 cells of CSE group (P<0.05), the activity of caspase-3 also was found decreased (P< 0.05), and the expression of SIRT1 and p-FOXO3a increased (P<0.05). Part 3 In vivo experiments 1.The more severe damage of chondriosome was found in old COPD group than that of old control group, while the damage of chondriosome significantly lessen in Res treated group. In vitro experiments 1.The mitochondria membrane potential drop (P<0.05) in CSE group, whereas can be increased by Res (P<0.05)Conclusion 1. The left ventricular remodeling and heart dysfunction can be induced by oxidative stress in old COPD rat.2. SIRT1 may be regulate the oxidative stress in old COPD rat by the effect on ROS production of mitochondria in the cardiac.3. p-FOXO3a may also involved in the oxidative stress regulation in the heart injury induced by COPD with SIRT1.4. The myocardial protection of resveratrol against COPD related stress may be by inducing expression of SIRT1. Rresveratrol may be considered as anti-oxidative stress therapy for the heart injury in old COPD rat.
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