本文關(guān)鍵詞： miR-98 運(yùn)動(dòng)性骨骼肌損傷 E2F5 ID1　出處：《上海體育學(xué)院》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：研究目的:隨著人們觀念的改變和大眾健身的普及,運(yùn)動(dòng)性骨骼肌損傷已經(jīng)不僅存在于競技體育,也成為困擾大眾健身的常見問題。損傷后的修復(fù)會(huì)出現(xiàn)一系列問題,如:瘢痕、纖維化、疼痛等。相關(guān)研究發(fā)現(xiàn):miR-98可以調(diào)控骨骼肌衛(wèi)星細(xì)胞的分化,使其成為骨骼肌損傷修復(fù)的又一靶點(diǎn)。本實(shí)驗(yàn)通過研究下坡跑大鼠股直肌損傷修復(fù)過程中miR-98的作用,為運(yùn)動(dòng)性骨骼肌損傷的治療提供新的理論基礎(chǔ)和實(shí)驗(yàn)依據(jù)。研究方法:72只兩月齡健康雄性SD大鼠,隨機(jī)分為9組,每組8只。分組:安靜對照組(C)、運(yùn)動(dòng)后0 h組、運(yùn)動(dòng)后6 h組、運(yùn)動(dòng)后12h組、運(yùn)動(dòng)后24 h組、運(yùn)動(dòng)后48 h組、運(yùn)動(dòng)后72 h組、運(yùn)動(dòng)后1 W組、運(yùn)動(dòng)后2 W組。利用下坡跑建立運(yùn)動(dòng)性骨骼肌損傷實(shí)驗(yàn)?zāi)Ｐ?運(yùn)動(dòng)方案參數(shù):跑臺(tái)速度為16m/min,坡度為-16°,運(yùn)動(dòng)時(shí)長為120min。用10%的水合氯醛腹腔注射麻醉,通過下腔靜脈取血,離心取血清后保存在-80℃C冰箱;取血后迅速取股直肌,肌肉標(biāo)本迅速放入用液氮冷卻的異戊烷中冷凍10s左右;完全冷凍后放于干冰上,待異戊烷完全揮發(fā)后,迅速用錫箔紙包上放入超低溫冰箱儲(chǔ)存。所取組織樣本進(jìn)行以下實(shí)驗(yàn):1)股直肌進(jìn)行冰凍切片HE染色(橫切)進(jìn)行形態(tài)學(xué)觀察;2)采用比色法和ELISA試劑盒分別對血清中CK和CK-MM的變化進(jìn)行檢測;3)采用RT-PCR對股直肌中miR-98、E2F5、ID1 mRNA水平進(jìn)行檢測;4)采用Western-Blot實(shí)驗(yàn)對股直肌中E2F5、ID1蛋白表達(dá)情況進(jìn)行檢測。研究結(jié)果:1.HE染色結(jié)果:對照組結(jié)果顯示:肌纖維成多邊形,大小均一,排列緊密規(guī)則,肌細(xì)胞核均勻的分布在細(xì)胞膜下;運(yùn)動(dòng)后Oh、6h、12h、24h、48h、72h組切片顯示:肌纖維排列散亂,細(xì)胞間隙變大,形狀不規(guī)則,呈腫脹狀態(tài);運(yùn)動(dòng)后1w和2w組切片顯示:肌纖維形態(tài)和對照組相似,趨于正常。2.血清 CK 和 CK-MM:與C組相比,運(yùn)動(dòng)后Oh組血清CK的含量明顯升高,具有顯著性差異(P0.05);運(yùn)動(dòng)后6h組血清CK含量幾乎持平;運(yùn)動(dòng)后12h、24h、48h組血清CK含量稍有升高,無顯著性差異;運(yùn)動(dòng)后72h、1W、2W組血清CK含量稍微降低,但無顯著性差異。運(yùn)動(dòng)后大鼠血清CK-MMELISA結(jié)果顯示:與C組相比,運(yùn)動(dòng)后Oh、6h、12h組血清中CK-MM的含量增加,無顯著性差異;運(yùn)動(dòng)后24h血清中CK-MM的含量顯著升高,具有顯著性差異(P0.05);運(yùn)動(dòng)后48h和72h血清中含量上升,無顯著性差異;運(yùn)動(dòng)后1W和2W后,血清CK-MM的含量已經(jīng)基本恢復(fù)。3.運(yùn)動(dòng)后股直肌中miR-98、E2F5和ID1 mRNA水平變化:與C組相比,運(yùn)動(dòng)后Oh和6h組miR-98 mRNA的含量稍有升高,無顯著性差異;運(yùn)動(dòng)后12h組miR-98 mRNA含量下降,無顯著性差異;運(yùn)動(dòng)后24h、48h和72h組miR-98 mRNA含量明顯下降,具有顯著性差異(P0.05);運(yùn)動(dòng)后1W組miR-98 mRNA含量稍降低,無顯著性差異;運(yùn)動(dòng)后W組miR-98 mRNA含量稍升高,無顯著性差異。與C組相比,運(yùn)動(dòng)后Oh組E2F5mRNA含量升高,無顯著性差異;運(yùn)動(dòng)后6h、12h和24h組E2F5 mRNA含量明顯升高,具有顯著性差異(P0.05);運(yùn)動(dòng)后48h和72h組E2F5 mRNA含量上升,無顯著性差異;運(yùn)動(dòng)后1W和2W組E2F5 mRNA含量稍有降低,無顯著性差異。與C組ID1 mRNA含量相比,運(yùn)動(dòng)后Oh和6h組ID1 mRNA含量增加,無顯著性差異;運(yùn)動(dòng)后12h組ID1 mRNA含量降低,無顯著性差異;運(yùn)動(dòng)后24h、48h和72h組ID1 mRNA含量明顯下降,具有顯著性差異(P0.05);運(yùn)動(dòng)后1W和2W組ID1 mRNA含量降低,但無顯著性差異。4.運(yùn)動(dòng)后各組股直肌中E2F5、ID1蛋白含量的變化:與C組相比,運(yùn)動(dòng)后0h、6h、12h、24h和48h組E2F5蛋白含量明顯升高,具有顯著性差異(P0.05);運(yùn)動(dòng)后72h、1W和2W組E2F5蛋白含量稍有升高,但無顯著性差異。與C組相比,運(yùn)動(dòng)后0h、6h和12h組股直肌中ID1含量下降,但無顯著性差異;運(yùn)動(dòng)后24h和48h組股直肌中ID1含量明顯下降,具有顯著性差異(P0.05);運(yùn)動(dòng)后72h、1W和2W組股直肌中ID1含量降低,但無顯著性差異。研究結(jié)論:1.一次長時(shí)間離心運(yùn)動(dòng)可以成功建立運(yùn)動(dòng)性骨骼肌損傷實(shí)驗(yàn)?zāi)Ｐ?2.運(yùn)動(dòng)性骨骼肌損傷時(shí)miR-98含量下降,促進(jìn)骨骼肌損傷的修復(fù)。
[Abstract]:Objective: with the popularity of the people idea change and the public fitness, exercise induced skeletal muscle injury has not only exist in the sports, has become a common problem of public health. The repair of the injured will appear a series of problems, such as scarring, fibrosis, pain and so on. The study found that miR-98 can regulate differentiation of skeletal muscle satellite cells, which has become a target of skeletal muscle injury. In this experiment, miR-98 ran downhill rectus femoris of rats during the injury and repair effect through research, to provide theoretical and experimental basis for the new treatment of exercise-induced skeletal muscle injury. Methods: 72 of 2 month old healthy male SD rats were randomly divided into 9 groups, 8 rats in each group. Group: normal control group (C), 0 h after exercise group, 6 h after exercise group, 12h after exercise group, exercise group after 24 h, 48 h after exercise group, exercise group after 72 h, 1 W after exercise group, W after exercise 2 group. By running the establishment of exercise-induced skeletal muscle injury experimental model of downhill exercise program parameters: treadmill speed of 16m/min, the slope is -16 degrees, the movement for a long time by intraperitoneal injection with 10% chloral hydrate 120min. anesthesia, the inferior vena cava blood, serum were stored in the -80 C C refrigerator; blood samples taken immediately after the rectus femoris muscle specimens, quickly put frozen in isopentane cooled in liquid nitrogen is about 10s; after completely frozen on dry ice, to isopentane completely volatilized quickly using the foil bag into the ultra low temperature freezer storage. The tissue samples were the following experiments: 1) of rectus femoris HE staining (cross) were observed; 2) by colorimetric method and ELISA kit respectively on the change of CK and CK-MM in serum were detected by RT-PCR E2F5; 3) of miR-98, ID1 were detected in the rectus femoris, mRNA level; 4) using Western-Blot experiments of rectus femoris E2F5, ID1 protein expression were detected. Results: the results of 1.HE staining: the control group showed that muscle fiber into polygon, uniform size, closely arranged rules, muscle cells evenly distributed in the cell membrane; Oh after exercise, 6h, 12h, 24h, 48h, 72h group section showed that the muscle fibers were scattered and the cell gap becomes larger, irregular shape, showed swelling of the state; and 1W after exercise group 2W section showed that muscle fiber morphology and similar to the control group, serum CK and CK-MM: became normal in.2. compared with C group, the content of serum CK in Oh group increased significantly after exercise, with a significant difference (P0.05); after exercise 6h serum CK was almost unchanged after exercise; 12h, 24h, 48h group, serum CK content increased slightly, no significant difference; after exercise 72h, 1W, 2W group of serum CK content decreased slightly, but there was no significant difference. The results show CK-MMELISA in the serum of rats after exercise: compared with C group, after exercise Oh, 6h, 1 Increase the content of CK-MM in serum of 2H group, no significant difference; the content of CK-MM 24h in serum after exercise was significantly increased, with significant difference (P0.05); after exercise the content of 48h and 72h in serum increased, no significant difference; after exercise 1W and 2W, the content of serum CK-MM has returned to.3. after the rectus femoris miR-98, E2F5 and ID1 change mRNA level: compared with C group, the content of Oh and 6h after exercise group miR-98 mRNA increased slightly, no significant difference; after exercise group 12h miR-98 mRNA decreased, no significant difference; after exercise 24h, 48h and 72h group miR-98 mRNA was significantly decreased, with significant difference (P0.05); 1W after exercise group miR-98 mRNA content decreased slightly, no significant difference; W group miR-98 mRNA content increased slightly after exercise, there was no significant difference. Compared with the C group, Oh group increased the content of E2F5mRNA after exercise, there was no significant difference; after exercise 6h, 12h and 24h group E2F5 mRNA鍚噺鏄庢樉鍗囬珮,鍏鋒湁鏄捐憲鎬у樊寮，

本文編號(hào)：1549461