【摘要】：目的:建立Beclin 1過(guò)表達(dá)和低表達(dá)MCF-7細(xì)胞模型,檢測(cè)4Gy照射后細(xì)胞自噬和凋亡的變化,探討B(tài)eclin 1的分子調(diào)控作用機(jī)制。方法:實(shí)驗(yàn)分為MCF-7組、MCF-7+4Gy組、MCF-7-Beclin1+4Gy組和MCF-7-Belin1RNAi+4 Gy組。利用分子生物學(xué)方法構(gòu)建Beclin 1過(guò)表達(dá)載體pcDNA3.1-Beclin 1,并建立Beclin 1過(guò)表達(dá)和低表達(dá)細(xì)胞模型。細(xì)胞經(jīng)4 Gy照射后,采用MDC染色熒光顯微鏡觀察自噬細(xì)胞百分比,AnnexinⅤ-FITC和PI染色流式細(xì)胞術(shù)檢測(cè)凋亡細(xì)胞百分比,Western blotting法檢測(cè)Beclin 1、P53、Bcl-2和Bax蛋白表達(dá)。結(jié)果:與MCF-7組比較,MCF-7+4Gy組、MCF-7-Beclin 1+4Gy組和MCF-7-Beclin 1RNAi+4Gy組自噬和凋亡細(xì)胞百分比均明顯升高(P0.05或P0.01),以MCF-7-Beclin 1+4Gy組升高最明顯,且高于MCF-7+4Gy組(P0.05);各組壞死細(xì)胞百分比無(wú)明顯差異。4Gy照射后,MCF-7組和MCF-7-Beclin 1組細(xì)胞中Beclin 1、P53和Bax蛋白表達(dá)水平均升高,Bcl-2蛋白表達(dá)水平降低,且以MCF-7-Beclin 1組最為明顯。結(jié)論:成功建立Beclin 1過(guò)表達(dá)和低表達(dá)MCF-7細(xì)胞模型,電離輻射能夠誘導(dǎo)其細(xì)胞自噬和凋亡,且對(duì)Beclin 1過(guò)表達(dá)細(xì)胞作用更明顯。Beclin1通過(guò)激活P53,進(jìn)而抑制Bcl-2和激活Bax,從而形成以P53為中心的自噬與凋亡分子調(diào)控機(jī)制。
[Abstract]:Aim: to establish Beclin 1 overexpression and low expression MCF-7 cell model, to detect the changes of autophagy and apoptosis after 4Gy irradiation, and to explore the molecular regulatory mechanism of Beclin 1. Methods: the experiment was divided into MCF-7 group, MCF-7 4Gy group, MCF-7-Beclin1 4Gy group and MCF-7-Belin1RNAi 4 Gy group. Beclin 1 overexpression vector pcDNA3.1-Beclin 1 was constructed by molecular biological method, and Beclin 1 overexpression and low expression cell models were established. After 4 Gy irradiation, the percentage of autophagy cells was observed by MDC staining fluorescence microscope, and the percentage of apoptotic cells was detected by Annexin 鈪，

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