GNB2L1在宫颈癌中的表达及与患者预后的相关性分析

Expression of GNB2L1 in cervical cancer and its correlation with prognosis

目的研究鸟嘌呤核苷酸结合蛋白β多肽2样蛋白1(GNB2L1)在宫颈癌组织中的表达情况与临床病理学特征及预后的相关性。方法收集2010年6月—2013年3月新疆医科大学第一附属医院妇科收治的132例宫颈鳞癌(CSCC)患者、56例宫颈上皮内瘤变(CIN)患者宫颈组织及32例无宫颈疾病患者正常宫颈(NC)组织,采用免疫组化、qRT-PCR、Western blot方法检测CSCC、CIN、NC组织中GNB2L1和mRNA表达水平,分析与临床病理学特征及预后的相关性。基因测序检测CSCC组织中GNB2L1基因拷贝数的变化。Western blot、qRT-PCR检测GNB2L1在宫颈癌细胞C33a、SiHa及正常宫颈上皮细胞H8中的表达情况。STRING数据库构建基因调控网络;RNA干扰调控技术下调子宫颈癌SiHa和C33a细胞中GNB2L1基因表达后,BODIPY染色检测SiHa、C33a细胞内脂质含量的变化。Kaplan-Meier和logrank试验分析GNB2L1蛋白的表达水平与患者总生存期的相关性。结果与CIN和NC组织相比,CSCC组织中GNB2L1的mRNA与蛋白质表达水平较高,差异具有统计学意义(P<0.01),且GNB2L1过表达与细胞分化程度(P=0.001)、临床分期(P=0.017)、淋巴结转移(P=0.049)密切相关。与GNB2L1低表达患者相比,GNB2L1高表达患者的总生存时间更短(P=0.002)。与NC组织(215100±7276)copies/ng相比,CSCC组织(297200±10815)copies/ng中的GNB2L1 DNA拷贝数显著升高(P=0.009)。qRT-PCR与Western blot结果显示,与H8细胞比较,C33a、SiHa细胞中GNB2L1的mRNA及蛋白表达水平均明显升高(P<0.01)。下调GNB2L1表达后,BODIPY检测结果显示,与正常对照组和阴性对照组比较,实验组细胞内脂质含量明显减少。结论GNB2L1在宫颈癌组织中高表达可能是CSCC患者独立的预后因素,过表达GNB2L1可能促进肿瘤的进展,有望成为CSCC患者潜在的预后生物标志物。

Objective To investigate the correlation between the expression of GNB2L1 in cervical cancer tissues and the characteristics with prognosis of clinical pathology.Methods The cervical tissues of 132 patients with cervical squamous cell carcinoma(CSCC)and 56 patients with cervical intraepithelial neoplasia(CIN)and 32 patients without cervical diseases were collected in the Department of Gynecology of the hospital,and the expression levels of GNB2L1 and mRNA in CSCC,CIN and NC tissues were detected by immunohistochemistry,qRT-PCR and Western blot methods,and the correlation with clinical pathological features and prognosis were analyzed.Gene sequencing detected changes in the number of copies of GNB2L1 gene in cervical squamous cell carcinoma tissue.Western blot and qRTPCR measured the expression of GNB2L1 in cervical cancer cells C33a,SiHa,and normal cervical endothelial cells H8.The STRING database constructs a gene regulation network,and the expression of GNB2L1 gene in SiHa and C33a cells of cervical cancer was down-regulated by RNA interference regulation technology,and BODIPY staining detected the change of lipid content in SiHa and C33a cells.Kaplan-meier analysis and Logrank test were used to analyze the correlation between GNB2L1 protein expression level,overall survival and progression-free survival.Results Compared with CIN and NC tissues,GNB2L1 mRNA and protein were significantly overexpressed in CSCC tissues,and GNB2L1 overexpression was closely related to pathological grading(P=0.001),clinical stage(P=0.017),and lymph node metastasis(P=0.049).Patients with high GNB2L1 expression had a shorter overall survival time(P=0.002)compared with the patients with low expression of GNB2L1.The number of GNB2L1 DNA copies in CSCC organizations(297200±9815.061)were significantly higher(P=0.009)compared with NC tissues(195100±5676.392).The results of qRT-PCR and Western blotting showed that GNB2L1 mRNA and protein expression levels in C33a and SiHa cells were significantly increased(P<0.001)compared with H8 cells.After down-regulating GNB2L1 expression,the intracellular lipid content in shGNB2L1 group was significantly reduced compared with the normal control group and the sh-NON group.Conclusion The high expression of GNB2L1 in cervical cancer tissues may be an independent prognostic factor for CSCC patients.Overexpression of GNB2L1 may contribute to tumor progression.Therefore,it may be a potential prognostic biomarker for CSCC patients.