TPX2在舌鳞状细胞癌组织中的表达及其对Cal27细胞增殖和凋亡的影响

Investigation of the expression of TPX2 in tongue squamous cell carcinoma and its effect on Cal27 cell proliferation and apoptosis

目的研究Xklp2靶蛋白(targeting protein for xenopus kinesin-like protein2,TPX2)在舌鳞状细胞癌组织中的表达及意义,并探讨其在舌鳞癌细胞株Cal27增殖和凋亡过程中的作用。方法收集新鲜的舌鳞状细胞癌组织及其配对癌旁组织30例于液氮中保存,采用荧光定量PCR、免疫印迹western blot方法检测TPX2在舌鳞状细胞癌癌组织和癌旁组织中mRNA、蛋白水平的表达情况,分析TPX2表达水平与临床病理参数的相关性;进一步用小干扰RNA(small interfering RNA,siRNA)敲低舌鳞状细胞癌细胞株Cal27细胞TPX2的表达量,以MTT法、流式细胞术和Western blot法分别检测细胞增殖、凋亡以及凋亡相关蛋白(cleaved caspase 3、caspase 3)表达水平的变化。结果在转录水平,TPX2在30例舌鳞状细胞癌患者癌组织中呈高表达状态,23例患者的癌组织TPX2的表达水平高于其对应的癌旁组织(t=3.254,P=0.002 9);在蛋白水平,TPX2在30例舌鳞状细胞癌患者癌组织中呈高表达状态,20例患者的癌组织TPX2的表达水平高于其对应的癌旁组织(t=2.862,P=0.007 7),且TPX2的高表达与舌鳞状细胞癌的T分期、淋巴结转移具有相关性;转染TPX2 siRNA 48 h可以抑制Cal27细胞增殖能力,增加凋亡细胞比例(F=342.9,P<0.000 1),同时上调Cleaved caspase 3(F=46.98,P=0.001 4)、Caspase 3(F=33.35,P=0.002 7)相关凋亡蛋白的表达。结论TPX2在舌鳞状细胞癌组织中呈高表达,干扰Cal27细胞中TPX2的表达可以抑制细胞增殖,诱导细胞凋亡,提示TPX2可能是舌鳞状细胞癌基因治疗的新靶点之一。

Objective To investigate the expression of targeting protein for xenopus kinesin-like protein2（TPX2）in tongue squamous cell carcinoma（TSCC） tissues and explore its effect on cell proliferation and apoptosis. Methods30 cases of TSCC tissues, paired normal tissues were collected in Dongguan People＇s Hospital during 2013-2016. The m RNA and protein expression level of TPX2 was determined by qRT-PCR and western blot, respectively and analyzed The correlation of TPX2 expression level and clinic opathological parameters. Cal27 cell was transfected with si RNAs to knockdown the expression of TPX2, then cell proliferation, cell apoptosis and related proteins（cleaved caspase 3 and caspase 3） were detected by MTT assay, flow cytometry and western blot. Results TPX2 was highly expressed in（t=3.254, P=0.002 9） tumor tissues at m RNA level compared to adjacent normal parts. In protein level, TPX2 was highly expressed（66.7%） in tumor tissues, TPX2 expression level of 20 case was higher than the corresponding tissue adjacent to carcinoma（20/30, t=2.862, P=2.862）, and high expression of TPX2 was related to T staging, lymph node metastasis of tongue cancer. Knockdown TPX2 effectively reduced cell proliferation, increased apoptosis rate（F=342.9, P 0.000 1） and upregulated the expression of apoptosis-related proteins cleaved caspase 3（F=46.98,P=0.001 4） and caspase 3（F=33.35, P=0.002 7）. Conclusion Overexpression of TPX2 was found in TSCC tissues.Silencing of TPX2 might inhibit cell proliferation and promote cell apoptosis. TPX2 could be a new target for gene therapy of TSCC.