人同源盒基因1对宫颈癌细胞增殖、迁移及侵袭的影响

Effect of OTX1 on proliferation, migration and invasion of cervical cancer cells

摘要

目的 探讨人同源盒基因1(OTX1)在宫颈癌组织中的表达情况,探究其对宫颈癌细胞增殖、迁移及侵袭能力的影响。方法 回顾性收集2020年5月至2021年12月在湖北省妇幼保健院行子宫切除术的51例女性宫颈癌患者与35例子宫良性病变患者的宫颈组织.免疫组化及RT-qPCR实验用于检测OTX1在临床收集的宫颈癌组织及正常宫颈上皮组织中的表达情况;RT-qPCR及western blot实验用于检测OTX1在宫颈癌细胞株及正常宫颈细胞株中的表达情况;在宫颈癌细胞株C-33A及CaSki中敲低/过表达OTX1后,采用western blot检测MMP-2、MMP-9及TIMP-2的蛋白表达变化;通过MTT法、细胞划痕实验及Transwell侵袭实验检测OTX1对宫颈癌细胞活性、迁移及侵袭能力的影响.使用GraphPad Prism 7.0以及ImageJ进行数据处理与分析。结果 OTX1在宫颈癌组织中的表达显著高于正常宫颈组织(t＝-11.979、-7.859,P＜0.05),并与临床上宫颈癌的病理特征密切相关,在体积大、FIGO分期级别高、分化程度低、有淋巴结转移的宫颈癌组织中高表达占比高(χ²＝6.920、4.230、11.230、6.890,P＜0.05);OTX1在宫颈癌细胞株中的表达显著高于正常宫颈细胞株(t值介于-47.334~-13.301之间,P＜0.05);在宫颈癌细胞株中敲低OTX1,显著抑制了宫颈癌细胞的活性以及增殖、迁移及侵袭的能力(t值介于6.131~19.854之间,P＜0.05);在宫颈癌细胞株中过表达OTX1,促进了宫颈癌细胞的活性以及增殖、迁移及侵袭的能力(t值介于-25.147~-4.290之间,P＜0.05)。结论 OTX1在宫颈癌组织及细胞中高表达,且与宫颈癌细胞的增殖、迁移、侵袭密切相关.提示OTX1可能是治疗宫颈癌的新靶点,并为宫颈癌的研究提供了新方向.

Abstract

Objective To investigate expression of orthodenticle homeobox 1 (OTX1) in cervical cancer tissues and its effects on capabilities in proliferation, migration, and invasion of the cervical cancer cells. Methods The cervical tissues samples including 51 cases of cervical cancer tissues of patients who underwent hysterectomy and 35 cases of normal cervical epithelial tissues of patients with benign cervical lesions who underwent hysterectomy in Hubei Provincial Maternal and Child Health Hospital over a period from May 2020 to December 2021 were collected retrospectively. Immunohistochemistry and real time quantitative reverse transcription polymerase chain reaction (RT-qPCR) were conducted to detect expression of OTX1 in the cervical cancer tissues and the normal cervical epithelial tissues clinically collected. RT-qPCR and western blot assay were employed to detect expression of OTX1 in the cervical cancer cell lines and the normal cervical cell lines. After knockdown/overexpression of OTX1 in the cervical cancer cell lines C-33A and CaSki, western blot method was used to detect changes in expression of proteins MMP-2, MMP-9 and TIMP-2. Colorimetric methyl thiazolyl tetrazolium (MTT) assay, cell scratch assay and Transwell invasion assay were employed to investigate effects of OTX1 on activity, migration and invasion capabilities of the cervical cancer cells. GraphPad Prism 7.0 and ImageJ software were used to conduct statistical analysis of the data. Results The expression of OTX1 in the cervical cancer tissues was significantly higher than that in the normal cervical tissues (in immunohistochemistry: t=-11.979; in RT-qPCR: t=-7.859, both P＜0.05), and was closely associated with clinopathological features of the cervical cancers: the expression of OTX1 was highly expressed in those cervical cancers with larger tumor size, lower differentiation degree and having lymphatic metastasis (χ ²=6.920, 4.230, 11.230 and 6.890 respectively, all P＜0.05). The expressions of OTX1 in the cervical cancer cell lines C-33A, SiHa, CaSki and ME-180 were significantly higher than those in the normal cervical cells (H8) (t values ranged from -47.334 to -13.301, all P＜0.05). Knockdown OTX1 significantly inhibited activity, proliferation and migration capabilities of the cervical cancer cell lines (t values ranged from 6.131 to 19.854, all P＜0.05), while overexpression of OTX1 enhanced activity, proliferation and migration capabilities of the cervical cancer cell lines (t values ranged from -25.147 to -4.290, all P＜0.05). Conclusion OTX1 is highly expressed in human cervical cancer tissues and cells, and is closely related to proliferation, migration and invasion of the cervical cancer cells. This suggests that OTX1 may be a novel target for treatment of cervical cancer and provide a new direction for study on cervical cancer.

关键词

人同源盒基因1 / 宫颈癌 / 细胞 / 增殖 / 迁移 / 侵袭

Key words

OTX1 / cervical cancer / cell / proliferation / migration / invasion

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基金资助

湖北省卫生健康委员会联合基金项目(WJ2019H188)

病毒学国家重点实验室开放研究基金项目(2021KF005)