| Exercise-induced Fatigue is a temporary decline in exercise capacity after a long period of exercise, and the body physiological and mental processes can not sustain their level of functioning and maintenance of a specific predetermined exercise intensity, and body organs and system function are out of tune, leading to a state of decline in exercise capacity.With the rapid development in competitive sports nowadays, competition becomes increasingly fierce, and many of the items are at or near the world record level of human limits. In order to achieve good results, the load that athletes bear are becoming more and more high in training. So the body can not adapt to cause exercise-induced fatigue. Elimination and relief exercise-induced fatigue effectively and timely is essential for improving the results of training for athletes. Sports medical community pays more and more attention on the research on Traditional Chinese medicine in improving exercise capacity and delay fatigue. In this study, exercise-induced fatigue rats were exposed to Traditional Chinese medicine, and we observed its effect on the expression of serum testosterone, blood urea nitrogen, lactic acid and17(3-HSD in exercise-induced fatigue rats. Our study will lay the theoretical and experimental basis for further exploring the mechanism of Traditional Chinese medicine anti-fatigue.Objective:We used biological chemistry, immunohistochemistry, RT-PCR and Western Blotting method, and we detected the content of the serum urea nitrogen, lactic acid and testosterone and the expression of testicular tissue17β-HSD to explore the mechanism that heshouwuyin relieve exercise-induced fatigue.Method: 1.The choice of experimental animals:We used Seventy-five male SD(Sprague-Dawley, SD) rats who were all Eight-week-old. They body weight180-200g.2. Establishment animal models of exercise fatigue:The swimming pail was round, and the wall was smooth, and the diameter was50cm, and the depth was70cm. The water temperature was30℃±2℃. We made the rats to swim adaptively at the first week. The rats swam for30min at the first time, and then the time increased by40%daily until the rats could swim for2hours at the first weekend. The adaptive weight loading swimming training began at the second week. The rats burdened with0.5%of their body weight for the first time,and the weight increased by1%、2%、3%、4%daily. The rats swam for2hours each time.The load increased to5%of their body weight at the sixth time, and the swimming time was significantly shorter, so we chose4%of their body weigh as the ultimate load. And we did swimming exhaustion experiments (The swimming exhaustion standard was that the rats still could not return to the surface from the water after10seconds by sinking). The swimming exhaustion experiments did one time each day,and lasted for42days.3. Animal grouping:The rats were randomly divided into5groups, normal control group (N group), Heshouwuyin administered normal group (HA group), model control group (M group), Heshouwuyin treated group (HT group), and Heshouwuyin beforehand-prevention group (HP group),each group15animals.N group and HA group rats did not swim, and lavaged the same volume of Sodium Chloride and Heshouwuyin(20g/kg/d). M group, HT group, and E group did swimming training every afternoon to copy exercise-induced fatigue model. HP group was fed with Heshouwuyin (20g/kg/d) every morning for the prevention and swam in the afternoon for42days. We collectd about1.5ml the medial canthal orbital vein blood. After42days since we setted up the models, we collectd about1.5ml the medial canthal orbital vein blood. The blood was centrifugated,and we drew on the supernatant. We detectd the concentration of blood urea nitrogen, blood lactate and serum testosterone. Then we judged the modeling is successful or not. The judging standard is as following:Compared with quiet control group, the concentration of blood urea nitrogen and blood lactate increased significantly in model group rats which indicatd that the modeling was successful. After the success of the model, M group, HT group was given Heshouwuyin (20g/kg/d) for treatment and M group was given Sodium Chloride for42days. N group, HA group and the HP group rats was supplied with eating and drinking freely.4. specimen collection and treatmentAfter the end of the experiment, Rats in each group were anesthetize with10%chloral Hydrate. We collectd about1.5ml the medial canthal orbital vein blood, the blood was centrifugated for20minutes at a speed of1000r/min, and then we drew on the supernatant, and the supernatant was keep at-80℃refrigerator. Then we detectd the concentration of serum testosterone, blood urea nitrogen and blood lactic acid.Next from left ventricle to the aorta, and then we perfused approximately250ml Sodium Chloride fast which was treated with DEPC until the flowing liquid was clear. Then we took the left testicular tissue quickly.We kept the tissue in liquid nitrogen for30minutes, and took the tissue to-80℃refrigerator for keep to spare for the Western blotting, RT-PCR detection. The right testicular tissue was fixed with4%paraformaldehyde by perfusion, and the tissue was embedded in paraffin in order to spare for immunohistochemistry.Results:1. Changes of body body weight of rats:Compared with quiet control group, the rats in model group lost weight significantly (P<0.01). The rats in Heshouwuyin treatment group put on weight significantly compared with the model group(P<0.05). Compared with treatment group, there was no significant difference in body weight in prevention group (P>0.05).2. The concentration of blood urea nitrogen: Compared with quiet control group,the concentration of blood urea nitrogen in model control group increased significantly (P<0.01). Compared with model control group the concentration of blood urea nitrogen in Heshouwuyin treatment group become significantly lower(P<0.05). Compared with treatment group,there was no significant difference in the concentration of blood urea nitrogen in prevention group (P>0.05).3. The concentration of blood lactic acid:Compared with quiet control group,the concentration of blood lactic acid in model control group increased significantly (P<0.01). Compared with model control group the concentration of blood lactic acid in Heshouwuyin treatment group become significantly lower(P<0.05). Compared with treatment group, there was no significant difference in the concentration of blood lactic acid in prevention group (P>0.05).4. The concentration of serum testosterone:Compared with quiet control group,the concentration of testosterone in model control group decreased significantly (P<0.01). Compared with model control group the concentration of serum testosterone in Heshouwuyin treatment group increased significantly (P<0.05). Compared with treatment group,there was no significant difference in the concentration of serum testosterone in prevention group (P>0.05).5. The expression of17β-HSD:Immunohistochemical staining showed that17β-HSD positive signal located in the cytoplasm and nuclus in Leydig cells. The positive expression were brown particles. Immunohistochemistry, Western blotting and RT-PCR tests results are as follows:The expression of rat testis decreased significantly in the model group compared with the normal group (P<0.01). The expression of rat testis increased significantly in the treatment group and prevention group compared with the model group(P<0.05). There was no significant difference in the expression of rat testis in prevention group compared with treatment group (P>0.05).Conclusion 1. The rat model of exercise-induced fatigue is successful.2. Heshouwuyin can increase the concentration of serum testosterone.3. Heshouwuyin can significantly reduce the concentration of serum urea nitrogen and lactic acid of exercise-induced fatigue rats.4. Heshouwuyin can increase the expression of17P-HSD in testicular tissue of exercise-induced fatigue rats. |
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