球体形成实验联合药物筛选建立耐顺铂胰腺癌干细胞株-论文.pdf

《球体形成实验联合药物筛选建立耐顺铂胰腺癌干细胞株-论文.pdf》由会员上传分享，免费在线阅读，更多相关内容在应用文档-天天文库。

1、第26卷肝胆胰外科杂志第1期·实验研究·球体形成实验联合药物筛选建立耐顺铂胰腺癌干细胞株杨德君，傅红兵，卫子然，徐嘉鹏，朱振新，蔡清萍(第二军医大学附属长征医院胃肠外科，上海200003)[摘要]目的探索建立顺铂耐药人胰腺癌干细胞株，并进行初步鉴定。方法通过球体形成实验联合顺铂药物筛选建立耐顺铂胰腺癌干细胞株；MTT法计算耐药指数；免疫荧光半定量检测OCT-4、SOX-2表达水平，检测干细胞标记物；免疫荧光半定量检测MUC-1表达水平，检测细胞分化能力；通过流式细胞仪测量侧群细胞和表面特异抗原标记(CD44／CD24)检测肿瘤干细胞含量。结果耐顺铂干细胞株的耐药指数是亲代株PANC—l的7

2、1．5倍；干细胞标记物的免疫荧光检测发现，耐药株中OCT-4、SOX-2、MUC-1均有不同程度的表达；耐药干细胞株及亲代株SP细胞比例分别为(25．1士1．1)％和(7．8土0．9)％，差异有统计学意义(t=6．89，JD<0．01)；耐药株及亲代株中CD44／CD24表型的细胞亚群分别占(20．8土1．3)％和(2．4土0．8)％，差异有统计学意义(t=16．13，尸<0．01)。结论球体形成实验联合顺铂药物筛选能建立耐顺铂胰腺癌细胞株，该细胞株能高效富集胰腺癌肿瘤干细胞。[关键词]胰腺癌；肿瘤干细胞；球体形成实验；耐药；筛选[中图分类号]R392．12[文献标识码]A[文章编号]l0

3、07—1954(2014)一0038—05Establishmentofcisplatin-resistantpancreaticcancerstemcellsthroughsphere-formationas-sayanddrugscreeningYANGDe-jun，FUHong—bing，WEIZi-ran，eta1．DepartmentofGastrointes—tinalSurgery，ChangzhengHospitalAffiliatedtotheSecondMilitaryMedicalUniversity，Shanghai200003，ChinaObjectiveToes

4、tablishcisplatin·-resistantpancreaticcancerstemcellsfromPANC·-1and~den·-tifythecancerstemcellpreliminarily．MethodsCisplatin—resistantpancreaticcancerstemcellswereobtainedthroughsphere—formationassayanddrugscreeninginculturemediumfor4weeks．ResistanceindexofthecellswasmeasuredbyMTT．Proteinexpression

5、sofOCT-4andSOX一2weredeterminedbyimmunofluorescence．CellulardifferentiativecapacitywasdetectedbasedontheexpressionofMUC一1．SidepopulationanalysisandCD44+／CD24cellssubsetsexplorationwereperformedbyflowcytometrytoexaminecancerstemcellsproportion．ResultsCisplatin—resistantcancercellswereobtainedwhichwe

6、re71．5timesincreasedindrugresistancerelativetoparentalcells．Tovaryingdegree，resistantcellhadexpressionsofOCT-4，SOX一2andMUC一1．FlowcytometryanalysisshowedthatSPcellproportioninresistantcellwas(25．1±1．1)％，whereasinparentalPANC一1was(7．8土0．9)％，thedifferencewasstatisticallysignificant(f=6．89，P<0．01)；CD4

7、4+／CD24cellswas(20．8士1．3)％inresistantcell，whereas(2．4土0．8)％inPANC—l，therewassignificantdifferencebetweenthem(t=16．13，P<0．01)．ConclusionTocombinesphere—formationassaywithdrugscreeningcanestablishdrug—resistancepan