沉默atf5對(duì)卵巢癌細(xì)胞增殖、凋亡及化療敏感性的影響

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1、青島大學(xué)碩士學(xué)位論文沉默ATF5對(duì)卵巢癌細(xì)胞增殖、凋亡及化療敏感性的影響姓名：齊亞妮申請(qǐng)學(xué)位級(jí)別：碩士專業(yè)：婦產(chǎn)科學(xué)指導(dǎo)教師：陳愛平20120610摘要目的l探討顯性負(fù)抑制人轉(zhuǎn)錄激活因子5(Activatingtranscriptionfactor5，ATF5)對(duì)卵巢癌細(xì)胞體外增殖、凋亡及順鉑敏感性的影響。方法。1．采用RT-PCR和Wegem．blot檢測(cè)SKOV-3、A2780、OVCAR8卵巢癌細(xì)胞系中ATF5的表達(dá)情況，篩選出ATF5高表達(dá)細(xì)胞系。2．采用脂質(zhì)體介導(dǎo)法將真核表達(dá)質(zhì)粒pLeGFP．C1-NTAzip-ATF5和

2、pLeGFP-C1分別轉(zhuǎn)染SKOV-3細(xì)胞，同時(shí)設(shè)空白對(duì)照組。3．采用MTT及AnnexinV／7．AAD雙染法流式細(xì)胞術(shù)檢測(cè)轉(zhuǎn)染后不同時(shí)間點(diǎn)各組細(xì)胞的增殖活性變化及早期凋亡情況。4．采用M1vr檢測(cè)不同濃度的順鉑作用24h后上述三組細(xì)胞的增殖情況。5．采用流式細(xì)胞術(shù)檢測(cè)4umol／L順鉑作用Oh、12h、24h、48h后各組細(xì)胞的凋亡。6．采用Western．blot檢測(cè)顯性負(fù)抑制ATF5聯(lián)合順鉑作用后，下游基因bel-2蛋白表達(dá)情況。結(jié)果：1．ATF5在SKOV-3細(xì)胞中的表達(dá)明顯高于其它兩株卵巢癌細(xì)胞，表達(dá)差異有統(tǒng)計(jì)學(xué)意義(P

3、

4、指導(dǎo)教師陳愛平教授關(guān)鍵詞：ATFS；上皮性卵巢癌；細(xì)胞增殖；細(xì)胞凋亡；化療敏感性AbstractObjectiveToinvestigatetheeffectofATF5genesilencedbydominantnegativeongrowth，apoptosisandchemosensitivityofovariancarcinomacells．Method1．TheexpressionofATF5indifferentovariancarcinomacelllinesweredetectedbyRT-PCRandWestern

5、blot．2．ThepLeGFP-C1-NTAzip-ATF5andpLeGFP·ClrecombinantplasmidswererespectivelytransfectedintoSKOV-3ceilswithLipofectamineTM2000reagent．3．MTTandFlowcytometrywasrespectivelyusedtoevaluatetheproliferationandapoptosisratesofcells，whichwasinterfered誦tllthefunctionofATF5．4．T

6、hecellswereexposedtocisplatinatdifferentconcentrationsfor24hours．TheproliferationwasassayedbyMTT．5．Aftertransfection24hours，thecellsweretreatedwith4umol／Lcisplatinatdifferenthours(Oh，12h，24h，48h)，flowcytometryWasusedtodetectapoptosis．6．Theexpressionofbcl-2proteinwasdet

7、ectedbywesternblot．Results1．TheexpressionofATF5inSKOV-3WaSthehighestamongthethreekindsofovariancancercelllines，theexpressiondifferenceWasstatisticallysignificant妒<0．05)．2．TherecombinantplasmidCanefficientlyandspecificallyinhibittheexpressionofATF5gene．Lossofthefunction

8、ofATF5ledtoapoptosisofSKOV-3cells，especiallyintheearlystage．3．Comparedwiththenonspecificandcontrolgroup，thegrowthinhi