本文選題：股直肌 + MyoD　； 參考：《上海體育學院》2016年碩士論文

【摘要】：研究目的通過對大鼠進行一次性下坡跑訓練,建立運動性骨骼肌損傷動物模型。結合股直肌的組織形態(tài)學變化,同時檢測血清CK和CK-MM的變化,探究在運動性骨骼肌損傷修復過程中Myo D和Myogenin的表達變化情況。研究方法選取72只兩月齡健康雄性SD大鼠為實驗對象,將其隨機分為9組,每組8只。分組情況如下:a.安靜對照組(C);b.運動后即刻組(T0h);c.運動后6小時組(T6h);d.運動后12小時組(T12h);e.運動后24小時組(T24h);f.運動后48小時組(T48h);g.運動后72小時組(T72h);h.運動后1周組(T1w);i.運動后2周組(T2w)。結合Armstrong離心運動誘導運動性骨骼肌損傷實驗模型以及課題組之前的運動實驗方案,確定運動組的運動方案。運動方案具體參數(shù)如下:跑臺速度為16m/min,坡度為-16°,運動時長為120min。對照組不進行下坡跑運動直接取材,運動訓練各組分別在運動后0小時、6小時、12小時、24小時、48小時、72小時、1周、2周進行取材。所有實驗對象均使用10%的水合氯醛腹腔注射進行麻醉。通過腹主動脈取血,離心取血清后保存在超低溫冰箱待用;此外,迅速分離出股直肌,所取肌肉標本利用液氮冷卻的異戊烷進行快速冷凍,之后置于干冰上待異戊烷揮發(fā)完全后于超低溫冰箱保存?zhèn)溆谩Ｋ〗M織樣本進行以下各項實驗:1)采用股直肌冰凍切片HE染色(橫切)進行組織形態(tài)學分析;2)采用比色法和ELISA試劑盒分別對血清CK和CK-MM水平進行檢測;3)采用RT-PCR實驗對Myo D、Myogenin m RNA水平進行檢測;4)采用Western-Blot實驗對Myo D、Myogenin蛋白表達情況進行檢測。研究結果1.HE染色結果:對照組組織切片染色結果顯示肌纖維成多邊形,大小均勻,排列緊密規(guī)則,肌細胞核可見于肌膜邊緣;運動后不同時間組取材的組織切片結果與對照組的結果相比顯示,肌纖維排列不緊密,間隙變大,形狀不規(guī)則,呈腫脹狀態(tài)。運動后6h組股直肌橫切片就發(fā)現(xiàn)有腫脹的肌纖維,運動后12h組到運動后48h組肌纖維腫脹持續(xù)加重,并出現(xiàn)核集聚現(xiàn)象;運動后48組和運動后72h組的切片顯示肌纖維排列最為散亂,間隙最大,核外溢最多。運動后1w組和運動后2w組的切片顯示肌纖維分布趨于正常狀態(tài),肌纖維緊密排列。2.血清CK和CK-MM:血清CK結果顯示該指標在運動后0h組、運動后6h組、運動后72h組及運動后1w組與對照組相比有統(tǒng)計學顯著性差異。并且該指標在運動后0h組到運動后48h組表達水平在不斷增高。血清CK-MM的結果顯示該指標在運動后24h組、運動后48h組與對照組相比具有統(tǒng)計學顯著性差異。并且該指標在運動后0h組到運動后24h組表達水平在不斷增高,之后呈下降趨勢。3.Myo D、Myogenin m RNA的表達變化:RT-PCR實驗結果顯示,Myo D m RNA表達水平在運動后12h組、運動后48h組、運動后72h組以及運動后1w組與對照組相比有統(tǒng)計學顯著性差異(p0.05);Myogenin m RNA表達水平在運動后12h組、運動后24h組及運動后48h組與對照組相比有統(tǒng)計學顯著性差異(p0.05);Id1 m RNA表達水平在運動后6h組和運動后2w組與對照組相比有統(tǒng)計學顯著性差異(p0.05)。4.Myo D、Myogenin蛋白的表達變化:一次性下坡跑運動后,Myogenin蛋白的Western blot結果顯示,運動后6h組、運動后12h組和對照組相比有統(tǒng)計學顯著性差異(p0.05),同樣是在運動后即刻開始下降,之后在運動后24h組達到最大值,后繼續(xù)下降至運動前水平。一次性下坡跑運動后,Myo D蛋白的Western blot結果顯示,運動后0h組、運動后6h組、運動后12h組和運動后1w組與對照組有統(tǒng)計學顯著性差異(p0.05),并且在運動后即刻呈下降的趨勢,在運動后24h組達到峰值后又繼續(xù)下降至對照組水平。研究結論1.一次性長時間下坡跑運動可以成功誘導運動性骨骼肌損傷實驗模型;2.結合血清CK和CK-MM的變化特點,血清CK可以用作運動性骨骼肌損傷評價的敏感指標,血清CK-MM作為輔助評價運動性骨骼肌損傷更準確;3.在一次性下坡跑誘導的運動性骨骼肌損傷中Myo D和Myogenin在轉錄水平存在時相性差異。
[Abstract]:The purpose of this study was to establish an animal model of athletic skeletal muscle injury by one-off downhill running training in rats. In combination with the histomorphological changes of the rectus femoris, the changes of serum CK and CK-MM were detected at the same time, and the changes of Myo D and Myogenin during the repair of sports skeletal muscle injury were investigated. The study method selected 72 2 month old. Healthy male SD rats were randomly divided into 9 groups, each group was divided into 8 rats in each group as follows: A. quiet control group (C), immediate group after exercise (T0h), 6 hour group after C. exercise (T6h), 12 hours after D. exercise (T12h), 24 hours after E. exercise (T24h), 48 hours after F. exercise, 72 hours after exercise; 1 weeks post exercise group 1W); 2 weeks after I. movement (T2w). Combined with the experimental model of motor skeletal muscle injury induced by Armstrong centrifugation and the program before the task group, the motion scheme was determined. The specific parameters of the exercise program are as follows: the speed of the runway is 16m/min, the slope is -16 degrees, and the long movement time is that the 120min. control group does not carry out the downhill running straight. All the groups were taken 0 hours, 6 hours, 12 hours, 24 hours, 48 hours, 48 hours, 72 hours, 1 weeks and 2 weeks after the exercise. All the subjects were anesthetized by intraperitoneal injection of chloral hydrate of 10%. The blood was taken through the abdominal aorta and the cryogenic refrigerator was preserved after the centrifugation. In addition, the rectus femoris muscle was separated quickly, The muscle specimens were frozen with liquid nitrogen cooled isopentane and then placed on dry ice to be preserved in ultra low temperature refrigerator after volatilization of isopentane. The samples taken as follows: 1) histomorphology analysis was carried out by freezing section of the rectus femoris HE staining (crosscutting); 2) the colorimetric method and the ELISA Kit were used respectively. Serum CK and CK-MM levels were detected; 3) RT-PCR test was used to detect Myo D, Myogenin m RNA level; 4) Myo D, and the expression of Myogenin protein was detected by Western-Blot experiment. The cell nucleus can be seen at the edge of the musculi membrane. The tissue section results from different time groups after exercise show that the muscle fibers are not arranged closely, the gap becomes larger, the shape is irregular and swollen. The swelling of the muscle fiber is found in the transverse section of the rectus femoris in group 6h after exercise, and the muscle fiber swelling in group 12h after exercise to 48h group after exercise. After exercise, the 48 groups and 72h groups after exercise showed that the muscle fibers were most scattered, the space was the most scattered, the gap was the largest, and the nuclear spillover was the most. The slices of 1W and 2W groups after exercise showed that the distribution of muscle fibers tended to be normal, and the muscle fibers were closely arranged in.2. serum CK and CK-MM: serum CK results showed that the index was in transport. After exercise, group 0h, group 6h, 72h group after exercise and 1W group after exercise were significantly different from those in control group, and the expression level of 48h group was increasing in 0h group after exercise after exercise. The result of serum CK-MM showed that the index was in 24h group after exercise, and there was a statistically significant difference between the 48h group and the control group after exercise. After exercise, the expression level of the 24h group was increasing in group 0h and after exercise, then the decline trend of.3.Myo D, Myogenin m RNA expression changes: the RT-PCR experimental results showed that Myo D m RNA expression level after exercise, after exercise group, after exercise and after exercise, there were statistical significance compared with the control group. Difference (P0.05); Myogenin m RNA expression level in group 12h after exercise, 24h group after exercise and after exercise 48h group compared with control group, there was significant difference (P0.05), Id1 m RNA expression level in 6h group and after exercise after exercise compared with the control group compared with the control group has statistical significant difference. After the downhill running, the Western blot results of Myogenin protein showed that there was a significant difference between the 6h group after exercise and the 12h group after exercise (P0.05), and the same was immediately after the exercise. After the exercise, the 24h group reached the maximum value, then continued down to the pre exercise level. Myo D eggs after the one-off downhill running. The results of white Western blot showed that after exercise, group 0h, group 6h after exercise, 12h group after exercise and 1W group after exercise were significantly different from those in control group (P0.05), and the trend was declining immediately after exercise. After the peak of exercise, the group reached the peak value and continued to decline to the control group. Conclusion 1. a long time downhill running movement was carried out. The experimental model of motor skeletal muscle injury can be successfully induced. 2. in combination with the changes of serum CK and CK-MM, serum CK can be used as a sensitive index for evaluating the damage of athletic skeletal muscle. Serum CK-MM is more accurate as a auxiliary evaluation of motor skeletal muscle injury; 3. Myo D and Myogenin in the injury of skeletal muscle induced by one-off downhill running. The phase difference exists at the transcriptional level.
【學位授予單位】：上海體育學院
【學位級別】：碩士
【學位授予年份】：2016
【分類號】：R87

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