本文選題：nestin　切入點(diǎn)：Cdk5　出處：《重慶醫(yī)科大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：目的:通過觀察電針與跑臺(tái)訓(xùn)練聯(lián)合作用于大鼠急性骨骼肌損傷修復(fù)過程中對(duì)nestin、Cdk5和ε-n ACh R表達(dá)的影響,探討電針聯(lián)合跑臺(tái)訓(xùn)練對(duì)于骨骼肌損傷及促進(jìn)神經(jīng)肌肉接頭修復(fù)的可能機(jī)制。方法:將60只成年SD雄性大鼠,設(shè)置為正常組和干預(yù)組和自然恢復(fù)組,采用隨機(jī)數(shù)字表法隨機(jī)抽取6只作為正常組,剩余54只大鼠作為自然恢復(fù)組和干預(yù)組。通過自制打擊器制備右下肢的腓腸肌急性鈍挫傷模型。將隨機(jī)抽取的48只建模成功大鼠按照隨機(jī)數(shù)字表法分為2組(每組n=24):自然恢復(fù)組和干預(yù)組。干預(yù)組在造模成功48h后,按照跑臺(tái)計(jì)劃訓(xùn)練表干預(yù),跑臺(tái)結(jié)束后即刻選取阿是穴、足三里穴位進(jìn)行電針干預(yù),每日一次,連續(xù)干預(yù)5天后休息2天。每組分別于造模后7天、14天、21天、28天四個(gè)時(shí)間點(diǎn)分別隨機(jī)選取6只大鼠進(jìn)行檢測(cè)。采用HE染色觀察損傷局部組織形態(tài)變化;采用Western blot、免疫熒光和RT-PCR觀察nestin、Cdk5以及ε-n ACh R蛋白及m RNA的表達(dá)水平。結(jié)果:1、HE染色顯示:正常組肌纖維形態(tài)完整,肌細(xì)胞大小規(guī)則,排列整齊;自然恢復(fù)組、干預(yù)組與正常組相比,均出現(xiàn)不同程度的肌纖維斷裂、變性,壞死;干預(yù)組與自然恢復(fù)組相比,肌細(xì)胞結(jié)構(gòu)損傷后恢復(fù)更迅速,肌組織內(nèi)的瘀血、出血灶和水腫減輕更快,新生肌細(xì)胞更多,整體肌肉組織形態(tài)恢復(fù)更好。2、免疫熒光顯示:Nestin與Cdk5免疫熒光雙標(biāo)記染色:正常組中,nestin與Cdk5無(wú)共表達(dá);在自然恢復(fù)組和干預(yù)組中,nestin與Cdk5熒光共表達(dá)逐漸降低,干預(yù)組中共表達(dá)低于自然恢復(fù)組。3、Western blot檢測(cè)結(jié)果:自然恢復(fù)組和干預(yù)組中,nestin蛋白在7天,14天,21天表達(dá)量均高于正常組(P0.05),且干預(yù)組表達(dá)量高于自然恢復(fù)組(P0.05),28天各組相比無(wú)明顯差異(P0.05);Cdk5蛋白在7天,14天,21天表達(dá)量均比正常組高(P0.05),干預(yù)組明顯低于自然恢復(fù)組(P0.05),在28天各組無(wú)明顯差異(P0.05);ε-n ACh R蛋白表達(dá)逐漸升高,各個(gè)時(shí)間點(diǎn)干預(yù)組表達(dá)量高于自然恢復(fù)組(P0.05),7天、14天、21天干預(yù)組和自然恢復(fù)組中表達(dá)量均低于正常組(P0.05),28天干預(yù)組表達(dá)量高于自然恢復(fù)組(P0.05),與正常組表達(dá)量無(wú)明顯差異(P0.05)。4、RT-PCR檢測(cè)結(jié)果:自然恢復(fù)組和干預(yù)組中,nestin m RNA在7天,14天均高于正常組(P0.05),且呈現(xiàn)遞減趨勢(shì),在21天,28天干預(yù)組和自然恢復(fù)組中nestin m RNA表達(dá)量與正常組相比無(wú)統(tǒng)計(jì)學(xué)意義(P0.05);Cdk5 m RNA在7天,14天,21天干預(yù)組與自然恢復(fù)組表達(dá)量均比正常組高(P0.05),各時(shí)間點(diǎn)的干預(yù)組明顯低于自然恢復(fù)組(P0.05),28天各組中Cdk5m RNA表達(dá)量與正常組相比無(wú)統(tǒng)計(jì)學(xué)意義(P0.05);ε-ACh R m RNA的表達(dá)在四個(gè)時(shí)間點(diǎn)逐漸升高,7天、14天、21天干預(yù)組和自然恢復(fù)組中表達(dá)量均低于正常組(P0.05),干預(yù)組表達(dá)量高于自然恢復(fù)組(P0.05),28天干預(yù)組表達(dá)量高于自然恢復(fù)組(P0.05),與正常組表達(dá)量無(wú)統(tǒng)計(jì)學(xué)差異(P0.05)。結(jié)論:大鼠骨骼肌急性損傷后,電針聯(lián)合運(yùn)動(dòng)訓(xùn)練在一定程度上促進(jìn)神經(jīng)肌肉接頭的修復(fù),其作用機(jī)制可能是通過抑制nestin和Cdk5蛋白在肌細(xì)胞膜上的共表達(dá),促進(jìn)了ε-n ACh R與nestin的表達(dá),從而促進(jìn)神經(jīng)肌接頭的再生。
[Abstract]:Objective: To observe the effects of electroacupuncture combined with the treadmill training in repairing rat acute skeletal muscle injury of nestin, expression of Cdk5 and R ACh e -n, electroacupuncture combined with treadmill training on skeletal muscle injury and promote the mechanism of neuromuscular junction repair. Methods: 60 adult male SD rats that is set to normal group and intervention group and spontaneous recovery group were randomly selected 6 rats as normal group, the remaining 54 rats as the natural recovery group and intervention group. The acute gastrocnemius blunt contusion model was prepared to attack right lower limb system. 48 rats were randomly selected were randomly divided into 2 groups (n=24 each): natural recovery group and intervention group. The intervention group in rats after 48h, in accordance with the treadmill treadmill training schedule intervention plan, immediately after the selection of Ashi point, Zusanli acupoint for acupuncture The intervention, once a day, continuous intervention after 5 days of rest for 2 days. Each group after modeling for 7 days, 14 days, 21 days, 28 days four time points were randomly selected 6 rats were detected. HE staining was used to observe the morphological changes of local tissue injury; by Western blot, immunofluorescence and RT-PCR. Nestin, Cdk5 and the expression level of R protein and m ACh e -n RNA. Results: 1. HE staining: normal group muscle fiber morphology, muscle cell size rule, neatly arranged; natural recovery group, the intervention group compared with normal group, there were different degrees of muscle fiber fracture, degeneration, necrosis; intervention group and natural recovery group, injury of muscle cell structure recovered more quickly, muscle tissue congestion, hemorrhage and edema in neonatal myocytes faster, more whole muscle morphology recovery better.2, immunofluorescence showed that Nestin and Cdk5 double immunofluorescence staining: normal 緇勪腑,nestin涓嶤dk5鏃犲叡琛ㄨ揪;鍦ㄨ嚜鐒舵仮澶嶇粍鍜屽共棰勭粍涓，

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