摘要
目的:探究含硬化蛋白域蛋白1(SOSTDC1)对宫颈癌细胞恶性生物学行为的调控及其分子机制。方法:收集2020年8月至2022年5月间在福建省肿瘤医院活检或手术切除的53例宫颈癌组织和相应的癌旁组织标本,免疫组化法检测SOSTDC1蛋白在宫颈癌组织及相应癌旁组织中的表达,qPCR法检测正常宫颈细胞、宫颈癌细胞中SOSTDC1 mRNA表达;将SOSTDC1过表达慢病毒(OE-sostdc1)和对照空病毒(NC)感染宫颈癌细胞SiHa及CaSki,将其分为SiHa-OE-sostdc1、SiHa-NC、CaSki-OE-sostdc1、CaSki-NC组,采用WST-1法、细胞集落形成实验、Transwell实验和WB法检测转染各组SiHa及CaSki细胞的增殖、集落形成、迁移和侵袭能力和BMP、Wnt/β-catenin信号途径相关蛋白及上皮-间充质转化(EMT)相关蛋白的表达。用DNA甲基化酶抑制剂5-氮杂2'-脱氧胞苷(5'-Aza-CdR)处理宫颈癌细胞后采用qPCR和WB法检测SOSTDC1 m RNA及蛋白的表达变化,用甲基化特异性PCR(MSP)检测5例配对宫颈癌组织与癌旁组织中SOSTDC1基因启动子区甲基化水平,同时qPCR检测其SOSTDC1 mRNA水平。结果:与癌旁组织比较,SOSTDC1蛋白在宫颈癌组织中呈低表达(P<0.01),且与淋巴结转移与FIGO分期有关联(均P<0.05);与正常宫颈HUCEC细胞比较,SOSTDC1 mRNA在宫颈癌C33A、HeLa、SiHa、CaSki细胞中均呈低表达(均P<0.01)。过表达SOSTDC1显著抑制SiHa及CaSki细胞的增殖、迁移和侵袭能力(均P<0.05)。WB法结果检测显示,过表达SOSTDC1显著抑制SiHa及CaSki细胞中磷酸化Smad、Dvl2/3、β-catenin、VIM、N-cadherin、Snail蛋白的表达(均P<0.05),5'-Aza-CdR处理后的SiHa及CaSki细胞中SOSTDC1 mRNA和蛋白水平均显著增加(均P<0.05),MSP检测结果显示,相较于癌旁组织,宫颈癌组织中SOSTDC1基因启动子区呈高度甲基化,且SOSTDC1 mRNA水平降低(P<0.01)。结论:SOSTDC1在宫颈癌组织中呈低表达且与肿瘤的恶性进展关联,其表达下调与其基因启动子区高度甲基化有关,过表达SO
Objective:To investigate the role of sclerostin domain-containing protein 1(SOSTDC1)in regulating malignant biological behaviors of cervical cancer(CC)cells and its molecular mechanism through in vitro experiments.Methods:Fifty-three cervical cancer tissues and corresponding paracancerous tissues were collected from biopsy or surgical resection at Fujian Cancer Hospital between August 2020 and May 2022,immunohistochemistry was used to investigate SOSTDC1 expression in CC and adjacent cervical tissue specimens.qPCR was performed to detect SOSTDC1 mRNA expression levels in normal cervical tissues and CC cells.CC cells SiHa and CaSki transfected with SOSTDC1 over-expression lentiviruses(OE-sostdc1)and negative control(NC)viruses were divided into SiHa-OE-sostdc1 group,SiHa-NC group,CaSki-OE-sostdc1 group and CaSki-NC group.WST-1,colony formation,and Transwell assays were used to detect the proliferative,colony-formative,migratory and invasive abilities of SiHa and CaSki cells in all the groups.The expressions of proteins related to BMP,Wnt/β-catenin signaling pathways,and epithelial mesenchymal transition(EMT)were detected by Western blotting.qPCR and WB arrays were performed on CC cells treated with the DNA methylation inhibitor 5-aza-2'-deoxycytidine(5'-Aza-CdR)to detect the changes in the expressions of SOSTDC1 mRNA and proteins.Methylation-specific PCR(MSP)assay was performed to analyze the level of gene promoter methylation of SOSTDC1 in 5 pairs of CC and para-cancerous tissue samples,while qPCR was used to detect SOSTDC1 mRNA levels in the tissues.Results:The expression of SOSTDC1 protein was significantly reduced in CC tissues compared with para-cancerous tissues(P<0.01),and low SOSTDC1 expression was found to be correlated with lymph node metastasis and FIGO staging(all P<0.05).The expression levels of SOSTDC1 mRNA were significantly reduced in C33A,HeLa,SiHa and CaSki cells compared with normal cervical HUCEC cells.Over-expression of SOSTDC1 significantly inhibited the proliferative,migratory,and invasive
作者
林万松
郭爱华
陈淑萍
汪阳
黄天莹
冯梅
叶韵斌
LIN Wansong;GUO Aihua;CHEN Shuping;WANG Yang;HUANG Tianying;FENG Mei;YE Yunbin(Laboratory of Immuno-Oncology,Clinical Oncology School of Fujian Medical University,Fujian Cancer Hospital,Fuzhou 350014,Fujian,China;Department of Gynecologic Oncology,Clinical Oncology School of Fujian Medical University,Fujian Cancer Hospital,Fuzhou 350014,Fujian,China;Fujian Provincial Key Laboratory of Translational Cancer Medicine,Fuzhou 350014,Fujian,China;School of Basic Medical Sciences,Fujian Medical University,Fuzhou 350122,Fujian,China)
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
北大核心
2023年第6期473-481,共9页
Chinese Journal of Cancer Biotherapy
基金
福建省医学创新课题基金(No.2020CXA013)
福建省科技创新联合资金项目(No.2018Y9104)
福建省卫生健康中青年骨干人才培养项目(No.2017-ZQN-14)。
