Effect Of Exercise Precondition On Myocardial Apoptsis By TNF-? After Exhaustive Exercise Injury In Rats

Objective:Exhaustive exercise can induce myocardial injury, which even resulted in sudden death,such events are not uncommon. So it is one of the most important causes faced by the sports medicine and military medicine and clinical medicine. Previous studies focused on the treatment of myocardial injury after exhaustive, There is currently a lack of practical and effective prevention measures. Recent people have recognized ischemic preconditioning induced cardioprotection by repeated transient ischemia, which make people from to direct cell protection research to the endogenous protective mechanism research. Exercise preconditioning which is as a special form of ischemia preconditioning induced self protection though exercise. It could improve the ability of myocardial ischemia and hypoxia tolerance. Recent studies have shown that myocardial apoptosis is cell biology based after exhausted myocardial injury. Exhaustive exercise increased the ratio of Bax / Bcl-2, promoted the expression of Fas protein, which decided to myocardial cells into the execution phase of apoptosis, and then triggered a series of complex cellular signal transduction pathways such as TNF-??NF-KB?Fas-L, ultimately leads to the cell apoptosis. Cysteinyl aspartate specific proteinase paly an important role. EP has a protective effect on the myocardium, but it is not clear whether EP exerts protective effects by inhibiting the apoptosis of cardiomyocytes and its effect on the intracellular signal transduction pathways. This study intends to establish exercise preconditioning and exhaustive exercise model, researching effects of exercise preconditioning on TNF-alpha and Caspase expression. Explore the protective effect of exercise preconditioning cardioprotection relevant whether if TNF-alpha mediated apoptosis pathway or not, further understanding of exercise preconditioning on myocardial protective effect of molecular mechanism. At the same time it can be for exhaustive exercise induced myocardial injury prevention measures to provide new experimental evidence and measures. Methods:1. Forty-eight male Sprague Dawley rats were randomly divided into four groups(n=12 in each group): sedentary control group, exhausted group, short-term EP exhausted group, long-term EP exhausted group. The short-term EP underwent 3 days of intermittent swimming exercise, and the long-term exercise preconditioning group 3 weeks some as above. Except for sedentary control group, the exhausted exercise group and EP group underwent exhausted exercise. Rats were killed 30 minutes after exhausted exercise(Thomas exhausted standardization), besides sedentary control group which were killed in resting state during the same period.2. The myocardial tissue was taken as pathological section, and the microstructure was observed under light microscope after HE staining.3. The content of serum ischemia modified albumin(IMA), creatine kinase isoenzyme(CK-MB) and cardiac troponin I(c Tn I) was detected by ELISA.4. The myocardial cell apoptosis was measured by the method of terminal-deoxynucleotidyl transferase mediated d-UTP nick end labeling(TUNEL).5. Using fluorescence quantitative polymerase chain reaction(PCR) to detect the gene expression of TNF-??Caspase-8?Caspase-9?Caspase-3 in myocardial in every group.6. Western Blot was used to detect the protein expression of TNF-??Caspase-8?Caspase-9?Caspase-3 in myocardial in every group. Results:1. Histomorphology of myocardium samples observed by light microscopy: cardiac sarcomeres of control group rats were arranged in neat rows, the density was uniform and there was no organelle edema. Myocardium of Group Exhausted suffered a high degree of edema, and muscle fibers rupture.In Groups of, Short-EP and Long-EP group part of myocardium edema, furthermore, Long-EP is more close to the control group.2. The comparision of IMA ?CK-MB?c Tn I in rats serum of different groups(1) Compared with the sedentary control group, exhaustive group, short-term EP and long-term EP serum IMA and CK-MB levels were significantly increased(P < 0.05), in addition,exhaustive group and short-EP group, serum c Tn I level was significantly increased(P<0.05), but long-term EP group had no significant difference.(2) Compared with the exhaustive group, long-term EP group, the serum levels of IMA, CK-MB, c Tn I content decreased significantly, there is significant difference(P< 0.05), and short-term EP group serum CK-MB and c Tn I decreased significantly(P < 0.05), meanwhile IMA decreased is not obvious.3. Apoptosis cell of cardiomyocyte tissue Apoptotic nuclei were stained brown and the normal nuclei were blue on TUNEL. Results revealed that the control group rats myocardial tissue has a few scattered in apoptosis; exhaustive group massive apoptosis of rats myocardial tissue was significantly higher than that of the control group(P < 0.05); Short-EP and Long-EP apoptosis was significantly decreased(P < 0.05).4. Expression of TNF-??Caspase-8?Caspase-9?Caspase-3 m RNA in myocardial of different groups(1)Compared with control group, except Caspase-9 of the long-term EP, the TNF-??Caspase-8?Caspase-9?Caspase-3 m RNA of exhausted group, Short-EP and Long-EP group are significant higher(P<0.05).(2)Compared with exhausted group, except Caspase-3 of the Short-EP, TNF-??Caspase-8?Caspase-9?Caspase-3m RNA of Short-EP and Long-EP is significant decrease(P < 0.05). Furthermore Long-EP group are decreased more significantly(P<0.05).5. The expression of TNF-??Caspase-8?Caspase-9?Caspase-3 protein in myocardial tissue of different groups(1)Compared with the control group, expression of TNF-??Caspase-8?Caspase-9?Caspase-3 protein of exhausted group and Short-EP is significant higher(P<0.05),but there was no significant difference with Long-EP group.(2)Compared with exhausted group, there are no significant difference in TNF-??Caspase-8?Caspase-9?Caspase-3 protein of Short-EP group, but Long-EP is significant decrease(P<0.05). Conclusion:1. Exhaustive exercise can induce myocardial injury, exercise preconditioning can reduce myocardial injury after exhaustive exercise.2. Cardiomyocyte apoptosis is part of myocardial injury induced by exhaustive exercise, When the body is exhausted, the activation of TNF-? mediated apoptosis signaling pathway and regulation of Caspase protease induces apoptosis of cardiac muscle cells.3. Exercise preconditioning inhibits myocardial apoptosis by restraining TNF-? mediated apoptosis pathway, then reduce myocardial apoptosis. It plays a important role in myocardial protection.4. Short-EP and long-EP can inhibit the apoptosis in different degrees, and The Long-EP heart protection effect is stronger than the Short-EP.