摘要
目的观察甲状腺癌细胞中微小RNA-330(miR-330)对T细胞免疫球蛋白黏蛋白3(TIM-3)的调控作用, 及其对甲状腺癌细胞侵袭能力的影响。方法收集河北医科大学第二医院甲状腺乳腺外科2020年1月到2021年6月30例甲状腺癌及其癌旁组织标本。应用荧光定量聚合酶链反应(PCR)方法检测甲状腺癌组织中miR-330及TIM-3基因的表达量;将miR-330模拟物及阴性对照质粒转染至TPC-1细胞中, 应用双荧光素酶报告基因实验分析TIM-3是否为miR-330的靶基因;应用荧光定量PCR方法检测miR-330的表达对TIM-3基因的影响;随后, 将TIM-3过表达质粒与miR-330模拟物转染至TPC-1细胞中, 应用Transwell细胞侵袭实验分析miR-330和TIM-3表达水平对甲状腺癌细胞侵袭能力的影响, 两组间比较应用t检验。结果荧光定量PCR实验结果表明, miR-330在甲状腺癌组织中的表达水平(0.38±0.07)明显低于癌旁组织(1.01±0.06, t=10.860, P<0.05);TIM-3的表达水平(5.70±0.81)明显高于癌旁组织(1.01±0.04, t=9.923, P<0.05), 相关性分析表明, miR-330与TIM-3两者表达呈负相关(r=-0.492, P<0.05);双荧光素酶报告基因实验结果表明, miR-330模拟物组荧光素酶活性(0.37±0.06)明显低于对照组(1.02±0.07, t=12.460, P<0.05);荧光定量PCR结果表明, 转染miR-330模拟物组TIM-3的表达水平(0.43±0.06)明显低于对照组(1.02±0.02, t=10.080, P<0.05);miR-330模拟物组侵袭细胞数(122.30±18.77)明显低于对照组(285.00±16.00, t=11.420, P<0.05);而与TIM-3过表达质粒共转染细胞后, miR-330模拟物转染组(228.00±12.53), 与对照组(225.00±13.45)比较差异无统计学意义(t=0.286, P>0.05)。结论 miR-330可通过靶向作用于TIM-3基因抑制甲状腺癌细胞侵袭。
Objective To observe the regulatory effect of microRNA(miR)-330 on T-cell immunoglobulin and mucin domain-containing protein 3(TIM-3)in thyroid cancer cells and its effect on the invasive ability of thyroid cancer cells.Methods A total of 30 cases of thyroid cancer and its adjacent tissues were collected from Department of Thyroid and Breast Surgery of the Second Hospital of Hebei Medical University from January 2020 to June 2021.Real-time polymerase chain reaction(PCR)was used to observe the expression levels of miR-330 and TIM-3 genes in thyroid cancer tissues.After transfecting miR-330 mimics and negative control into TPC-1 cells,dual luciferase reporter gene assay was used to detect whether TIM-3 was the target gene of miR-330.Real-time PCR was used to detect the effect of miR-330 mimics on TIM-3 gene.Subsequently,the overexpression plasmid of TIM-3 and miR-330 mimics were co-transfected into TPC-1 cells.The effects of TIM-3 and miR-330 expression levels on the invasion of thyroid cancer cells were observed by invasion assay.T-test was used for comparison between groups.Results The results of real-time PCR experiments showed that the expression level of miR-330 in thyroid cancer tissues(0.38±0.07)was significantly lower than that in adjacent tissues(1.01±0.06,t=10.860,P<0.05).The expression level of TIM-3(5.70±0.81)was significantly higher than that of adjacent tissues(1.01±0.04,t=9.923,P<0.05).Correlation analysis showed that the expression of miR-330 and TIM-3 was negatively correlated(r=-0.492,P<0.05).Dual luciferase reporter gene analysis indicated that the luciferase activity of miR-330 mimics transfection group(0.37±0.06)was significantly lower than that of control group(1.02±0.07,t=12.460,P<0.05).Real-time PCR results showed that the expression level of TIM-3 in the miR-330 mimic transfected group(0.43±0.06)was significantly lower than that in the control group(1.02±0.02,t=10.080,P<0.05).The number of invasive cells in the miR-330 mimic group(122.30±18.77)was significantly less than that in the control group(285.00±16.00,t=11.420,P<0.05).In addition,after co-transfection with the TIM-3 overexpression plasmid,there was no significant difference in the number of invasive cells between the miR-330 mimics group(228.00±12.53)and the negative control group(225.00±13.45,t=0.286,P>0.05).Conclusion MiR-330 could inhibit the invasion of thyroid cancer cells by targeting the TIM-3 gene.
作者
张晓旭
底旺
李清怀
申伟
底罕
冀宏
Zhang Xiaoxu;Di Wang;Li Qinghuai;Shen Wei;Di Han;Ji Hong(Graduate School of Hebei Medical University,Shijiazhuang 050000,China;Department of Thyroid and Breast Surgery,the Second Hospital of Hebei Medical University,Shijiazhuang 050000,China;School of Basic Medicine,Hebei Medical University,Shijiazhuang 050000,China)
出处
《中华实验外科杂志》
CAS
北大核心
2022年第2期254-256,共3页
Chinese Journal of Experimental Surgery
