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1、農(nóng)村小學(xué)小班化識(shí)字研究近年來(lái)，隨著國(guó)家計(jì)劃生育政策的不斷完作者：周永列，呂亞萍，胡惟孝，邱蓮女，王文松，劉建棟，吳建國(guó)【摘要】本研究探討四嗪二甲酰胺誘導(dǎo)白血病細(xì)胞株SHI1凋亡作用的機(jī)制。以不同濃度的ZGDHu1在體外培養(yǎng)SHI1細(xì)胞，用細(xì)胞形態(tài)學(xué)、DNA凝膠電泳、DNA含量及細(xì)胞周期分析AnnexinV/PI雙標(biāo)記和Hoechst33258熒光染色等分析細(xì)胞凋亡。用Rhl23/PI測(cè)定線粒體跨膜電位，用流式細(xì)胞術(shù)檢測(cè)ZGDHu1誘導(dǎo)SHI1細(xì)胞凋亡過(guò)程中bcl2、bax、p53、Fas蛋白和線粒體膜蛋白的表
2、達(dá)變化。用RTPCR觀察經(jīng)ZGDHu1作用后SHI1細(xì)胞bcl2、bax、p53基因表達(dá)改變；同時(shí)利用實(shí)時(shí)熒光PCR定量檢測(cè)端粒酶hTERTmRNA表迗的變化。結(jié)果表明：ZGDHu1能誘導(dǎo)SHI1細(xì)胞凋亡，呈作用時(shí)間和劑量的量效關(guān)系，出現(xiàn)典型的細(xì)胞形態(tài)改變，DNA片段化，亞G1峰檢出并顯著增加；AnnexinV/PI和Hoechst33258熒光染色后細(xì)胞發(fā)生凋亡的特征性改變。SHI1細(xì)胞經(jīng)不同濃度的ZGDHu1體外培養(yǎng)后，bcl2基因和蛋白有所下調(diào)，但主要是bax基因和蛋白上調(diào)，導(dǎo)致bax/bcl2比值明顯
3、增高，p53基因和蛋白與Fas蛋白表達(dá)也上調(diào)，均呈劑量依賴性。隨ZGDHu1作用濃度的增加，AWm下降，而線粒體膜蛋白表迗顯著升高，端粒酶hTERTmRNA的表達(dá)顯著下調(diào)。結(jié)論：ZGDHu1通過(guò)上調(diào)p53基因、bax基因和bax/bcl2比值，使線粒體跨膜電位顯著下降的線粒體通路是ZGDHu1誘導(dǎo)細(xì)胞凋亡的途徑之一，F(xiàn)as也參與其中，端粒酶有可能是其抗腫瘤的作用靶點(diǎn)?！娟P(guān)鍵詞】四嗪二甲酰胺細(xì)胞株SHI1細(xì)胞凋亡線粒體端粒酶AbstractTheaimofstudywastoinvestigatethemech
4、anismofN,Nfdi(mmethylphenyl)3,6dimethyl1,4dihydro1,2,4,5tetrazine1,4dicarboamide(ZGDHu1)inducingapoptosisinSHI1humanleukemiacellline.DifferentconcentrationsofZGDHulanddifferenttimesofculturewereusedtotreatSHIlcells;theapoptosisof’SHI1cellswasanalyzedbymorph
5、ology,DNAagarosegelelectrophoresis,DNAcontentdetection,AnnexinV/PlandHoechst332581abelingmethod,themitochondrialtransmembrancepotentialweremeasuredbydihydrorhodaminl23,andexpressionsofbe12,bax,Fas,p53andmitochondrialmembraneprotEinwereanalyzedbyflowcytometr
6、y,whilethebcl2,baxandp53genewereanalyzedbyRTPCR.ThetranscriptionallevelofhTERTmRNAwasmeasuredbyrealtimefluorescencequantitativeRTPCR.TheresultsshowedthatafterexposuretoZGDHu1,SHIlcellswereinducedtoapoptosisinatimeanddoesdependentmanner.SHIlcellapoposiswasco
7、mfirmedbytypicalcellmorphology,DNAfragmentHoechst33258andAnnation,subexinGlphase,/PIlabelingetc.Theexpressionofbax,bax/bcl2,p53andFasgenesignificantlyincreasedandbcl2slightlydecreased.ZGDHulcouldincreasedtheexpressionofmitochondrialmembraneprotEIninadosedep
8、endentmannerwhile△^mreduced.TheexpressionofhTERTmRNAsignificantlydecreased.ItisconcludedthatZGDHulcanupregulatetheexpressionofp5ThemitochondrialpacentofmitochondriaentialmaybeoneoftingapoptosisbyZ3,bax
