摘要
目的:探讨LncRNA DLG1-AS1对甲状腺乳头状癌(PTC)细胞恶性化生长和转移的影响,并对其可能的分子机制进行研究。方法:体外培养PTC细胞(TPC1、KTC-1、B-CPAP、HTori-3)和正常甲状腺上皮细胞Nthy-ori3-1,采用实时荧光定量聚合酶链反应(qRT-PCR)检测细胞中DLG1-AS1和miR-203表达差异。将B-CPAP细胞分为空白对照组、阴性对照组(转染pcDNA3.1空载体和阴性对照序列)、shDLG1-AS1组(转染pcDNA3.1-shDLG1-AS1)、shDLG1-AS1+miR-203抑制物组(转染pcDNA3.1-shDLG1-AS1和miR-203抑制物)。通过CCK-8法和Transwell小室法检测细胞增殖、迁移和侵袭活性。蛋白质印迹法检测上皮间质转化(EMT)相关蛋白表达。双荧光素酶报告分析DLG1-AS1、miR-203、ZEB2之间的靶关系。结果:经qRT-PCR法检测,与Nthy-ori3-1细胞相比,TPC1、KTC-1、B-CPAP、HTori-3细胞中DLG1-AS1相对表达量均升高,同时miR-203相对表达量均降低(均P<0.05),选择DLG1-AS1相对表达量最高的B-CPAP细胞进行后续实验。与空白对照组和阴性对照组相比,shDLG1-AS1组细胞增殖、迁移、侵袭活性降低,ZEB2、N-cadherin、vimentin、β-catenin蛋白表达下调,同时E-cadherin表达上调(均P<0.05)。与shDLG1-AS1组相比,shDLG1-AS1+miR-203 inhibitor组细胞增殖、迁移、侵袭活性被逆转,ZEB2、N-cadherin、vimentin、β-catenin蛋白表达上调,同时E-cadherin表达下调(均P<0.05)。经双荧光素酶报告证实,DLG1-AS1可以直接调控miR-203,而ZEB2则是miR-203的下游靶基因。结论:LncRNA DLG1-AS1通过与miR-203竞争结合,消除miR-203对靶基因ZEB2表达的抑制,促进PTC细胞发生EMT,这可能是PTC恶性化生长和转移的重要机制。
Objective:To investigate the effect of LncRNA DLG1-AS1 on the proliferation and metastasis of papillary thyroid carcinoma(PTC)cells,and further to explore its mechanism.Methods:PTC cells TPC1,KTC-1,B-CPAP,HTori-3 and normal follicular cell Nthy-ori3-1 were used to detect DLG1-AS1 and miR-203 expressions by real time quantitative polymerase chain reaction(qRT-PCR)method.B-CPAP cells were divided into blank group,NC group(transfected pcDNA3.1-vector and negative control inhibitor),shDLG1-AS1 group(transfected pcdna 3.1-shDLG1-AS1),shDLG1-AS1+miR-203 inhibitor group(transfected pcdna3.1-shDLG1-AS1 and miR-203 inhibitor).The proliferation and invasion,migration activities of cells were detected by CCK-8 assay and transwell method.Western blotting was used to detect the relative expression of epithelial-mesenchymal transition(EMT)related proteins.The luciferase report was used to analyze the target relationship between DLG1-AS1,miR-203,and ZEB2.Results:qRT-PCR results showed compared with Nthy-ori3-1 cells,the relative expression of DLG1-AS1 in TPC1,KTC-1,B-CPAP and HTori-3 cells was increased,while the relative expression of miR-203 was decreased(P<0.05).B-CPAP cells with the highest relative expression of DLG1-AS1 were selected for follow-up study.Compared with blank group and NC group,the proliferation,migration and invasion of shDLG1-AS1 group cells were decreased,while the expression of ZEB2,N-cadherin,vimentin,β-catenin protein was down-regulated,and the expression of E-cadherin was up-regulated(P<0.05).Compared with shDLG1-AS1 group,shDLG1-AS1+miR-203 inhibitor group showed a reversal of cell proliferation,migration and invasion,while the expression of ZEB2,N-cadherin,vimentin,β-catenin protein was up-regulated and the expression of E-cadherin was down-regulated(P<0.05).DLG1-AS1 could directly regulate miR-203,and ZEB2 was the downstream target gene of miR-203.Conclusion:By combining with miR-203,LncRNA DLG1-AS1 eliminates the inhibition of miR-203 to ZEB2 expression,promotes the EMT of PTC cells,which may be
作者
冯立新
翟传夫
谭清玉
FENG Lixin;ZHAI Chuanfu;TAN Qingyu(Department of General Surgery, Linyi Hospital of Traditional Chinese Medicine, Linyi 276000, China)
出处
《东南大学学报（医学版）》
CAS
2021年第2期133-141,共9页
Journal of Southeast University(Medical Science Edition)
基金
山东医药卫生科技发展计划面上项目(2018WS391)。
