肾癌细胞株786-0侧群细胞生物学特性与XIAP mRNA表达的研究

目的 肿瘤干细胞在肿瘤复发和耐药中起着关键作用,侧群(side population,SP)细胞中富含肿瘤干细胞.本研究探讨肾癌786-0细胞株中SP细胞的生物学特性及凋亡抑制蛋白XIAP在SP中的表达.方法 制备肾癌786-0细胞株单细胞悬液,Hoechst33342染色,流式细胞荧光激活技术分选出肾癌786-0细胞系中的SP细胞和非侧群(non-SP,NSP)细胞2个亚群,MTT法绘制细胞生长曲线,比较SP细胞和NSP细胞的增殖能力;无血清悬浮培养观察两组亚群细胞肿瘤球形成能力,流式细胞仪检测两组亚群细胞体外分化能力;实时荧光定量PCR法检测SP细胞、NSP细胞及未经分选的肾癌786-0细胞XIAP mRNA的表达情况.结果 流式细胞分选结果显示,肾癌786-0细胞株中SP细胞所占比例为(2.9±0.1)％.细胞生长曲线结果显示,SP细胞增殖速度快,与NSP细胞增殖能力比较,差异有统计学意义,t=4.32,P=0.033.SP细胞在无血清培养基中的肿瘤球形成能力强于NSP细胞;无血清培养2周后SP细胞亚组SP细胞所占比例下降为(18.1±0.3)％,提示SP细胞可以分化为NSP细胞.XIAP mRNA在SP细胞中的表达量为10.74±0.25,与NSP细胞亚组及未经分选的786-0细胞比较差异有统计学意义,F=61.02,P=0.012.结论 肾癌786-0细胞株中存在具有干细胞特性的SP细胞,高表达凋亡抑制蛋白XIAP,SP细胞可作为肾癌分子靶向治疗研究的切入点.

Biological characteristics and XIAP mRNA expression of side population cells in renal carcinoma cell line 786-0

OBJECTIVE Cancer stem cells play a key role in tumor resistance and relapse.Side population (SP) cells are rich in cancer stem cells.This study was to isolate the SP cells from renal carcinoma cell line 786-0 and analysis the biological characteristics and XIAP mRNA expression of side population cells.METHODS SP and non-SP cells were sorted from renal carcinoma cell line 786-0 by fluorescence-activated cell sorting (FACS) using Hoechst 33342 staining,the capacity of proliferation of both subgroups was tested by MTT method and both subgroups were cultivated to survey the capacity to form into suspended tumor cell spheres.The differentiation ability of both subgroups was determined by FACS.Real-time PCR was used to evaluate the XIAP mRNA expression levels of XIAP mRNA in the SP cells,non-SP cells and non-sorted renal carcinoma cell line 786-0.RESULTS The proportion of SP cells in renal carcinoma cell line 786-0 was (2.9±0.1)％.Cell growth curve indicated that proliferative capacity of the SP cells was better than that of non-SP cells (t=4.32,P=0.033).The capacity to form into suspended tumor cell spheres of the SP cells in serum-free medium was better than that of non-SP cells.After two weeks,the proportion of SP cells in SP subgroups fell to (18.1± 0.3) ％,which revealed the SP cells can differentiate into NSP cells.The expression levels of XIAP mRNA in the SP cells (10.74±0.25) was significantly higher than that in non-SP cells and non-sorted renal carcinoma cell line 786 0 (F1=61.02,P=0.012).CONCLUSIONS The results of this study indicate the existence of cancer stem-1ike SP cells in renal carcinoma cell line 786-0.The expression levels of XIAP are higher in SP cells.SP cells may provide new insights for molecular target therapy of renal cell carcinoma.