顺铂在喉肿瘤细胞转化为干细胞样肿瘤细胞作用的初步探讨

Exploration of the role of cisplatin on transformation of laryngeal tumor cells to stem-like cancer cells

目的:探讨喉癌Hep-2细胞株经流式分选后的非侧群细胞经化疗药物顺铂诱导转变为干细胞样肿瘤细胞的可能机制。方法:喉癌Hep-2细胞株经流式细胞仪荧光活化细胞分选系统分选后,获得的非侧群分别加入头颈部常用化疗药物——顺铂(实验组)和生理盐水(对照组)培养48h,再次上流式细胞仪检测两组侧群细胞比例;并通过RT-PCR及Western blot分别在mRNA和蛋白水平检测两组细胞的干细胞样标记β-catenin、notch-1的基因和蛋白的表达情况。结果:实验组非侧群细胞经顺铂(终浓度1μg/ml)作用48h后,侧群细胞比例为(17.16±0.18)%,对照组则为(10.05±1.20)%,两组差异有统计学意义(t=5.844,P<0.01)。荧光定量RT-PCR示经顺铂处理实验组的β-catenin、notch-1mRNA较对照组显著增多;Western blot示实验组细胞中β-catenin、notch-1蛋白表达量较对照组显著增加(t=5.155,P=0.031;t=5.977,P=0.004)。结论:非侧群细胞经化疗药物顺铂诱导后可分化为干细胞样肿瘤细胞。其可能机制是通过wnt/β-catenin、notch转导通路异常诱导非侧群细胞转化为干细胞样肿瘤细胞。

Objective：To explore the possibility mechanism of non-side population cells（NSP）of Hep-2be induced into stem-like cancer cells by chemotherapy drug--cisplatin.Method：Hep-2cell lines were sorted by fluorescence-actived cell sorting.The acquired NSP cells in trail group were co-cultured with cisplatin for more than 48 hours,while the control group with normal saline（NS）.Then identified the percentage of the side population（SP）cells by flow cytometer.Theβ-catenin,notch-1mRNA in trial and control group were detected using quantitative realtime PCR,and theβ-catenin,notch-1protein in two groups were compared by Western blot.Result：The percentage of side population cells in two groups were（17.16±0.18）%,（10.05±1.20）%,respectively.There was significant difference between two groups（t=5.844,P〈0.01）.The expression ofβ-catenin,notch-1was higher in trail group by qRT-PCR;the protein levels ofβ-catenin,notch-1was found to inceased in the trail group by Western blot（t=5.155,P=0.031;t=5.977,P=0.004）.Statistical analysis showed significant difference between two groups（P〈0.05）.Conclusion：NSP cells can be differentiated into stem-like cancer cells after being treating with cisplatin.The supposed mechanism is maybe through wnt/β-catenin,notch signaling transduction pathway abnormalities.