本文選題：仙臺(tái)病毒Tianjin株 + MDA-MB-231細(xì)胞�。� 參考：《天津醫(yī)科大學(xué)》2015年碩士論文

【摘要】：目的:近年來(lái),利用病毒治療腫瘤取得了巨大進(jìn)展。仙臺(tái)病毒Tianjin株是仙臺(tái)病毒新基因型。本論文擬采用紫外線滅活仙臺(tái)病毒Tianjin株(以下簡(jiǎn)稱UV-Tianjin)感染人乳腺癌MDA-MB-231細(xì)胞和人宮頸癌HeLa細(xì)胞,通過檢測(cè)細(xì)胞的凋亡情況及凋亡發(fā)生的可能機(jī)制來(lái)評(píng)價(jià)UV-Tianjin的抗腫瘤作用,以期為病毒抗腫瘤作用機(jī)制研究及開發(fā)新的治療乳腺癌和宮頸癌的生物制劑提供參考依據(jù)。方法:用不同滴度UV-Tianjin分別感染人乳腺癌細(xì)胞MDA-MB-231和人宮頸癌細(xì)胞HeLa,24小時(shí)后,(1)MTT法檢測(cè)不同滴度UV-Tianjin(MOI:10,20,40,80,160,320)對(duì)MDA-MB-231細(xì)胞和人正常乳腺上皮細(xì)胞MCF 10A,或HeLa細(xì)胞增殖的影響;(2)Hoechst染色熒光顯微鏡觀察UV-Tianjin(MOI:40,80)感染后MDA-MB-231細(xì)胞或HeLa細(xì)胞的形態(tài)學(xué)改變;(3)AnnexinⅤ-FITC/PI標(biāo)記流式細(xì)胞儀檢測(cè)UV-Tianjin(MOI:20,40,80)誘導(dǎo)MDA-MB-231細(xì)胞或HeLa細(xì)胞產(chǎn)生凋亡的凋亡率;(4)分光光度法檢測(cè)UV-Tianjin(MOI:20,40,80)感染后MDA-MB-231細(xì)胞或HeLa細(xì)胞中Caspase-3、-8和-9的活性變化;(5)JC-10染色流式細(xì)胞儀檢測(cè)UV-Tianjin(MOI:5,20,80)感染后MDA-MB-231細(xì)胞或UV-Tianjin(MOI:20,40,80)感染后HeLa細(xì)胞線粒體膜電位的變化;(6)Western blot檢測(cè)UV-Tianjin(MOI:20,80)感染后MDA-MB-231細(xì)胞中Bcl-2、Bax、細(xì)胞色素c、caspase-9、Fas、FasL、caspase-8和caspase-3蛋白表達(dá)水平或UV-Tianjin(MOI:20,40,80)感染后HeLa細(xì)胞中細(xì)胞色素c、APAF-1、caspase-9、Fas、FasL、FADD、caspase-8和caspase-3蛋白表達(dá)水平。結(jié)果:(1)MTT檢測(cè)結(jié)果顯示UV-Tianjin能夠抑制MDA-MB-231細(xì)胞或HeLa細(xì)胞的增殖,且呈劑量依賴性;而對(duì)正常乳腺上皮細(xì)胞MCF 10A無(wú)明顯抑制作用;(2)Hoechst染色結(jié)果顯示滅活病毒組MDA-MB-231細(xì)胞或HeLa細(xì)胞均發(fā)生了明顯的形態(tài)學(xué)變化:細(xì)胞核致密濃染,呈高亮的藍(lán)色熒光,且具有劑量依賴性趨勢(shì);(3)AnnexinⅤ-FITC/PI標(biāo)記流式細(xì)胞儀檢測(cè)結(jié)果顯示滅活病毒組MDA-MB-231細(xì)胞或HeLa細(xì)胞的凋亡率呈劑量依賴性升高(P0.01);(4)分光光度法檢測(cè)結(jié)果顯示滅活病毒組MDA-MB-231細(xì)胞或HeLa細(xì)胞中caspase-3,-8和-9的活性增加且呈劑量依賴性(P0.05);(5)JC-10染色流式細(xì)胞儀分析顯示滅活病毒組MDA-MB-231細(xì)胞或HeLa細(xì)胞線粒體膜電位呈劑量依賴性降低,且與細(xì)胞對(duì)照組比較均有統(tǒng)計(jì)學(xué)差異(P0.01);(6)Western blot結(jié)果顯示高劑量滅活病毒組MDA-MB-231細(xì)胞中Bcl-2、Bax、細(xì)胞色素c、caspase-9、Fas、FasL、caspase-8和caspase-3蛋白表達(dá)水平或HeLa細(xì)胞中細(xì)胞色素c、APAF-1、caspase-9、Fas、FasL、FADD、caspase-8和caspase-3蛋白表達(dá)水平與對(duì)照組比較均有顯著差異(P0.05)。結(jié)論:滅活仙臺(tái)病毒Tianjin株能夠抑制人乳腺癌MDA-MB-231細(xì)胞和人宮頸癌HeLa細(xì)胞的增殖,誘導(dǎo)MDA-MB-231細(xì)胞和HeLa細(xì)胞發(fā)生凋亡,且呈劑量依賴性,其凋亡發(fā)生的機(jī)制可能與線粒體內(nèi)源性途徑和死亡受體外源性途徑相關(guān)。
[Abstract]:Objective: in recent years, great progress has been made in the treatment of tumor with virus. Sendai virus Tianjin strain is a new genotype of Sendai virus. In this paper, ultraviolet inactivated Sendai virus Tianjin strain (UV-Tianjin) was used to infect human breast cancer MDA-MB-231 cells and human cervical cancer HeLa cells. The anti-tumor effect of UV-Tianjin was evaluated by detecting the apoptosis of the cells and the possible mechanism of apoptosis. The aim of this study was to provide a reference for the study of the antitumor mechanism of the virus and the development of new biological agents for the treatment of breast and cervical cancer. Methods: the effects of different titers of UV-Tianjin on the proliferation of MDA-MB-231 cells and normal breast epithelial cells (MCF 10A or HeLa cells) were detected by MTT assay after 24 hours after infection with human breast cancer cell line MDA-MB-231 and human cervical cancer cell line HeLa. Morphologic changes of MDA-MB-231 cells or HeLa cells after UV-Tianjin MOI: 4080) the apoptosis rate of MDA-MB-231 cells or HeLa cells induced by UV-Tianjin: MOI: 4040 ~ 80) was detected by flow cytometry with annexexin 鈪，

本文編號(hào)：1964398