| Objective: The myocardial ischemia-reperfusion injury (IR) is a main interferen -ce in the reperfusion therapy for acute myocardial infarction patients, and ischemic preconditioning (IPC) can obviously reduce the injury of cardiac muscle. The protection of small heat shock proteins to myocardium from ischemic preconditioning in adult rats has already been known, but some studies showed that this protection had not been found in aged rats. Previous studies had shown that the small heat shock proteins (sHSPs) protect the myocardium from I/R-induced damage. To learn the relationship between the expression of sHSPs and aging of rats during IPC, the present study was designed to investigate the expression of alpha-B crystallin (αBC)and HSP 27 in the myocardium during IPC both in mRNA level and protein level in adult and young rats to explain the essence and regulation of descending adaptability and ascending sensitivity to injury in aged heart. Material and methods:1) Adult male (8-20 week old) and aged male (20-24 months old) SD rats were randomly divided into two groups: controlled group (Ctrl) and ischemic precondi -tioned group (IPC). In the Ctrl group, rats' breast were opened with continuous perfusing for 60 minutes, while in the IPC group, rats were treated with ischemia (5 minutes each) 3 times, reperfused for 5minutes. After 3 times of ischemia and reperfusion anterior wall of left ventricle treated with ischemia and posterior wall Of left ventricle treated with non-ischemia were taken in 0min, 5min, 15min, 45min and 60min, and there were 4 rats in each group. Then these samples were put into liquid nitrogen for 5 minute and frozen into -70℃ refrigerator.2) The distribution of HSP27 and αBC proteins in soluble endochylema andinsoluble organelle: the heart tissues were placed into split buffers, and then the soluble protein and in soluble protein were collected and detected via western blot.3) The location of HSP27 and αBC proteins in heart cells: the fresh defrosted heart tissues were marked with HSP27 and αBC proteins via immunofluorescence methods, and then observed under laser confocal microscopy.4) mRNA expression of HSP27 and αBC proteins in different time with ischemia treatment: the mRNA expression of HSP27 and αBC proteins was detected via semi-quantitative RT-PCR.Results:1) RT-PCR showed that the expression of alpha-B crystalline and HSP27 mRNA and protein did not change both in aging and adult rats, but the expression of αBC and HSP27 mRNA increased after about 15 min ischemia treatment both in adult and aged rats.2) Western blot demonstrated that the expression of αBC and HSP27 protein did not change both in aging and adult rats, the expression of alpha-B crystalline protein increased a little after ischemia treatment in anterior wall of left ventricle in aged rats compared with adult rats. The ability of translocation of a αBC and HSP27 protein in anterior wall of left ventricle from the cytosol to sites of the myofibrillar system reduced in aging rats compared with adult rats, and the abilty of translocation of the two proteins in posterior wall of left ventricle treated with non-ischemia had no changes.3) Immunohistology indicated that after 15min ischemia pretreatment, HSP27 protein shifted to Z line and I band, while αBC protein shifted to Z line and meanwhile gathered at intercalated discs in adult rats compared with aged rats. Conclusion:1) It was demonstrated that the aged rats still had ability of genetic transcription and protein synthesis of αBC and HSP27 during ischemic preconditioning.2) The translocation ability of αBC and HSP27 in aging rat reduced afterischemia preconditioning, which might explain why aging rat heart partially loss the protection of IP. |
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