本文關鍵詞：人剪切修復基因XPD抑制肝癌HepG2細胞增殖遷移及對自噬水平調(diào)控的實驗研究　出處：《南昌大學》2015年碩士論文　論文類型：學位論文

  更多相關文章： 肝癌 XPD 增殖 凋亡 遷移 自噬

【摘要】：目的:探究上調(diào)肝癌細胞株HepG2中XPD基因的表達后對腫瘤細胞增殖凋亡、遷移以及自噬水平變化的影響。方法:采用脂質(zhì)體作為載體介質(zhì)將重組質(zhì)粒pEGFP-N2/XPD和空載質(zhì)粒pEGFP-N2瞬時轉(zhuǎn)染入人肝癌細胞株HepG2細胞內(nèi),實驗分為四組1)空白對照組、2)脂質(zhì)體組、3)pEGFP-N2組、4)pEGFP-N2/XPD組,使用熒光顯微鏡觀察綠色熒光表達情況;MTT法觀察腫瘤細胞的增殖能力;流式細胞儀檢測腫瘤細胞凋亡情況;體外平板克隆形成實驗觀察腫瘤細胞體外克隆集落形成情況;劃痕愈合實驗及Transwell實驗檢測腫瘤細胞遷移能力的變化。Western blotting檢測HepG2細胞XPD及自噬相關蛋白LC3、P62表達量的變化。結(jié)果:熒光顯微鏡觀察轉(zhuǎn)染空載質(zhì)粒pEGFP-N2和重組質(zhì)粒pEGFPN2/XPD的細胞可見綠色熒光,轉(zhuǎn)染有效。上調(diào)XPD的表達后,結(jié)果顯示與對照組比較,實驗組細胞增殖能力減弱(p0.01),細胞凋亡率增加(p0.01),體外克隆形成數(shù)量減少(p0.01),遷移能力減弱(p0.05),并且上調(diào)LC3蛋白表達(p0.01),而隨之P62的表達水平減少(p0.01)。結(jié)論:過表達XPD基因能顯著抑制肝癌細胞增殖促進其凋亡,抑制體外克隆形成,并且降低腫瘤細胞的遷移能力,同時調(diào)控肝癌細胞自噬水平的活化。
[Abstract]:Objective: to investigate the proliferation and apoptosis of hepatoma cells after up-regulating the expression of XPD gene in hepatoma cell line HepG2. Effects of migration and changes in autophagy levels. Methods:. The recombinant plasmid pEGFP-N2/XPD and the empty plasmid pEGFP-N2 were transiently transfected into human hepatoma cell line HepG2 cells using liposome as carrier medium. The experiment was divided into four groups (1) blank control group (2)) liposome group (pEGFP-N2) and pEGFP-N _ 2 / XPD group. The expression of green fluorescence was observed by fluorescence microscope. The proliferative ability of tumor cells was observed by MTT method. Apoptosis of tumor cells was detected by flow cytometry. The colony formation of tumor cells in vitro was observed by plate clone formation in vitro. Effect of scratch healing test and Transwell assay on the migration ability of tumor cells. XPD and autophagy associated protein LC3 in HepG2 cells were detected by blotting. Results: green fluorescence was observed in the cells transfected with empty plasmid pEGFP-N2 and recombinant plasmid pEGFPN2/XPD. After up-regulating the expression of XPD, the results showed that compared with the control group, the proliferation ability of the experimental group decreased p0.01a, and the apoptosis rate increased (p0.01). The number of clone formation in vitro decreased p0.01a, the migration capacity decreased, and the expression of LC3 protein was upregulated (p0.01). Conclusion: overexpression of XPD gene can significantly inhibit the proliferation of hepatoma cells and promote its apoptosis, and inhibit the formation of in vitro cloning. It also reduces the migration ability of tumor cells and regulates the activation of autophagy of hepatoma cells.
【學位授予單位】：南昌大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：R735.7

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