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1、338Labeledlmmunoassays＆ClinMed，Apr．2015，Vo1．22，No．4·方法研究·膀胱癌T24順鉑耐藥細(xì)胞系的建立及生物學(xué)特性趙熳，張曼，馬萍(1．徐州醫(yī)學(xué)院醫(yī)學(xué)技術(shù)學(xué)院，江蘇徐州221004；2．首都醫(yī)科大學(xué)附屬北京世紀(jì)壇醫(yī)院，北京100038)摘要：目的采用濃度梯度法誘導(dǎo)建立人膀胱癌順鉑耐藥細(xì)胞系(T24／DDP)，并觀察其生物學(xué)特性。方法利用人膀胱癌細(xì)胞株T24，采用順鉑(DDP)作為誘導(dǎo)劑在體外建立人膀胱癌順鉑耐藥細(xì)胞模型T24／DDP(0．6mL)。通過倒置相差顯微鏡下觀察
2、T24、T24／DDP細(xì)胞形態(tài)，MTF法檢測細(xì)胞多藥耐藥性，生長曲線檢測細(xì)胞的增殖能力，流式細(xì)胞術(shù)測細(xì)胞的周期，qRT-PCR檢測耐藥基因MRP(multidrugresistance—associatedprotein)的表達(dá)。結(jié)果經(jīng)過順鉑14個月的誘導(dǎo)，成功建立了T24／DDP。倒置相差顯微鏡下觀察與T24細(xì)胞相比較，T24／DDP排列不規(guī)則，形態(tài)出現(xiàn)多形性，出現(xiàn)巨細(xì)胞；與T24相比較，T24／DDP對順鉑、阿霉素、長春新堿、甲氨蝶呤均產(chǎn)生不同程度的交叉耐藥，比T24細(xì)胞倍增時間明顯延長，細(xì)胞周期中G1期細(xì)胞增
3、多，G2、S期細(xì)胞減少。qRT—PCR結(jié)果顯示，耐藥細(xì)胞MRP基因的表達(dá)明顯高于親本細(xì)胞。結(jié)論人膀胱癌耐順鉑細(xì)胞株T24／DDP具有多藥耐藥性，可以作為研究膀胱癌細(xì)胞多藥耐藥的良好實(shí)驗(yàn)?zāi)Ｐ汀ｊP(guān)鍵詞：人膀胱癌細(xì)胞；順鉑；多藥耐藥EstablishmentofCisplatin-multidrugResistanceCellLineofHumanBladderCancerT24andItsBiologicalCharacteristicsZHA0Man。ZHANGManI，．MAPing(1．CollegeofMedi
4、calTechnique，XuzhouMedicalCollege，Xuzhou221004，China；2．BeijingShijitanHospital，CapitalMedicalUniversity，Beijing100038，China)Abstract：ObjectiveToestablishacisplatinresistantbladdercancercelllineandtoanalysisitsbiologicalcharacteristics．MethodsDrugconcentrationg
5、radientincreasingmethodwasadoptedtoinducedrugresistance．T24andT24／DDPwereobservedunderphasecontrastmicroscope．MultidrugresistanceOfthetwocelllineswasdeterminedbyMT-F．Cellproliferationwasevaluatedaccordingtothegrowthcurve．Flowcytometrywasperformedtodeterminecyc
6、lesofthetwocelllines．Theexpressionofmultidrugresistancerelatedgenemuhidrugresistance—associatedprotein(MRP)wasdetectedbyqRT—PCR．ResultsAfter14months，cisplatin—resistanthumanbladdercancercelllineT24／DDPwassuccessfullyestablishedfromitsparentalcelllineT24．Twokin
7、dsofcellsweresignificantlydifferentfromeachotherbyamicroscope．T24／DDPcellwas9．51timesmoreresistanttoDDPthanT24．T24／DDPcelllineexhibitedthecharacteristicsofcross—resistancetocisplatin，adriamycin，methotrexate，vincristine．ThecelldoublingtimeofT24／DDPwaslongerthan
8、parentalcelllineT24．ComparedwithT24，theproportionofG2andSphasecellsinT24／DDPdecreasedsignificantly，whilethatofG1phasecellsincreasedsignificantly．Theexpressionofdrug-resistancerelat
