CD49f维持宫颈癌干细胞特性的作用研究

Role of CD49f in the maintenance of cervical cancer stem cell characteristics

目的研究整合素家族蛋白成员整合素α6,即CD49f在维持宫颈癌干细胞特性中的作用。方法采用生物信息学分析肿瘤基因组图谱(The Cancer Genome Atlas,TCGA)和基因表达汇编(Gene Expression Omnibus,GEO)数据库中CD49f转录本水平在宫颈癌组织和正常宫颈组织中的表达差异。在宫颈癌细胞系HeLa,SiHa,C33A,HT-3和CaSki中,采用免疫细胞化学方法检测CD49f蛋白的表达。采用流式细胞计数仪检测和分选获得CD49f^high和CD49f^low的两群细胞并进行MTT实验检测细胞活力。检测两群细胞对化疗药顺铂和5-氟尿嘧啶的敏感性。体外肿瘤球形成实验和体外分化培养检测两群细胞的自我更新能力和分化能力的差异。结果数据库结果显示CD49f转录本水平在宫颈癌组织中的表达高于正常宫颈组织(P<0.05)。CD49f蛋白在宫颈癌5个细胞系中均有不同程度的表达且定位于细胞膜。流式细胞计数仪分选获得CD49f^high和CD49f^low的两群细胞,MTT结果显示两群细胞的体外增殖能力差异无统计学意义(P>0.05),而肿瘤干细胞特性包括较强的肿瘤球形成能力、多项分化能力和对传统化疗药物的抵抗性,在CD49f^high的宫颈癌细胞中显著高于CD49f^low的细胞群,差异有统计学意义(P<0.05)。结论CD49f在宫颈癌组织中表达增高,能够显著促进宫颈癌干细胞的自我更新能力、抑制分化和化疗药敏感性等特性,提示其可能作为宫颈癌干细胞诱发的复发转移和不良预后的生物标志物。

Objective To investigate the role of a member of integrins family proteins integrinα6,CD49f in the maintenance of cervical cancer stem cell characteristics.Methods Bioinformatics was used to analyze the expression of CD49f transcript in cervical can-cer and normal cervical tissues in The Cancer Genome Atlas(TCGA)and Gene Expression Omnibus(GEO)databases.Immunocytochemistry was used to detect the expression of CD49f protein in cervical cancer cell lines HeLa,SiHa,C33A,HT-3 and CaSki.The posi-tive rate of CD49f in cervical cancer cells was detected by flow cytometry,and two subpopulation of CD49f^high and CD49f^low cells were isolated.Cell viability of the two subpopulation cells and the sensitivity to chemotherapeutic drugs cisplatin and 5-fluorouracil were evaluated by MTT assay.The tumorsphere formation and differentiation culture assays were performed to detect the abilities of self-renewal and differentiation.Results The database showed that the expression of CD49f in cervical cancer was significantly higher than that in normal cervical tissues(P<0.05).In cervical cancer cell lines,CD49f protein was located in the cell membrane and showed different expression level in five cancer cell lines.The proliferation abilities of CD49f^high and CD49f^low cells were not significantly different in vitro(P>0.05).The abilities of chemotherapy resistance,tumorsphere formation and anti-differentiation were significantly higher in CD49f^high cells those in CD49f^low cells(P<0.05).Conclusion CD49f shows high expression in cervical cancer and can significantly promote the self-renewal of cervical cancer stem cell,and inhibit the ability of differentiation and chemotherapy sensitivity,suggesting that CD49f may be a biomarker of poor prognosis of cervical cancer.