發(fā)布時(shí)間：2018-03-12 13:43

  本文選題：骨骼肌衛(wèi)星細(xì)胞　切入點(diǎn)：MRFs　出處：《內(nèi)蒙古農(nóng)業(yè)大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：骨骼肌衛(wèi)星細(xì)胞是一類具有增殖能力和分化潛力的生肌干細(xì)胞,它存在于肌細(xì)胞膜和基膜兩者之間,這類細(xì)胞對(duì)于以后骨骼肌的成長(zhǎng)和再生起著重要的作用。該論文從內(nèi)蒙古錫林郭勒盟采集烏珠穆沁羊的胎兒期(3月齡)及剛出生(0月齡)、中年羊(6月齡)肌肉樣品,采用兩步酶消化法和組織塊法對(duì)骨骼肌衛(wèi)星細(xì)胞進(jìn)行體外提取培養(yǎng);用1%的明膠和4%的多聚甲醛兩種助貼壁劑來檢測(cè)細(xì)胞的貼壁時(shí)間;利用差速貼壁法、peroll密度梯度離心法和改良后的純化方法來進(jìn)行細(xì)胞純化;并用免疫組化鑒定了骨骼肌衛(wèi)星細(xì)胞;用實(shí)時(shí)熒光定量技術(shù)檢測(cè)了骨骼肌細(xì)胞在不同生長(zhǎng)時(shí)期MRFs基因表達(dá)變化。得出如下結(jié)論:以組織塊法分離和改良后的純化方法純化并用1%的明膠作為助貼壁劑得到的細(xì)胞數(shù)目較多且形態(tài)較好、細(xì)胞較純,建立了一套高效穩(wěn)定的骨骼肌衛(wèi)星細(xì)胞分離純化體系。用desmin和myosine抗體對(duì)該細(xì)胞進(jìn)行鑒定,細(xì)胞質(zhì)中的胞漿被染成棕色或者棕黃色,證實(shí)此細(xì)胞為骨骼肌衛(wèi)星細(xì)胞。MRFs家族基因中MyoD和Myf5主要在骨骼肌衛(wèi)星細(xì)胞的增殖期表達(dá),MyoG和Myf6主要在細(xì)胞的分化階段表達(dá)。其中MyoD和Myf5在細(xì)胞增殖期表達(dá)量上升,而到分化期時(shí)表達(dá)量下降;MyoG和Myf6正好相反,在細(xì)胞增殖期表達(dá)量下降,而到分化期時(shí)表達(dá)量上升。
[Abstract]:Skeletal muscle satellite cells are a class of myogenic stem cells with proliferative and differentiation potential, which exist between the muscle cell membrane and the basement membrane. These cells play an important role in the growth and regeneration of skeletal muscle in the future. Two step enzyme digestion method and tissue mass method were used to extract and culture skeletal muscle satellite cells in vitro, and two kinds of adherent agents (1%% gelatin and 4%% paraformaldehyde) were used to detect the adherence time of the cells. The cell was purified by differential adherent density gradient centrifugation and modified purification method, and the skeletal muscle satellite cells were identified by immunohistochemistry. The changes of MRFs gene expression in skeletal muscle cells at different growth stages were detected by real-time fluorescence quantitative technique. The conclusions are as follows: the method of tissue mass separation and purification was used to purify skeletal muscle cells and gelatin 1% was used as adherent agent. To a larger number of cells and a better shape, A set of high efficient and stable isolation and purification system of skeletal muscle satellite cells was established. The cells were identified by desmin and myosine antibodies. The cytoplasm of the cells was stained brown or brown. It was confirmed that the expression of MyoD and Myf5 in skeletal muscle satellite cells. MRFs family genes were mainly expressed in the proliferative phase of skeletal muscle satellite cells. MyoG and Myf6 were mainly expressed in the differentiation phase of skeletal muscle satellite cells, and the expression of MyoD and Myf5 increased in the proliferative phase of the cells. On the contrary, the expression of MyoG and Myf6 decreased at the stage of cell proliferation, but increased at the stage of differentiation.
【學(xué)位授予單位】：內(nèi)蒙古農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：TS251.1

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