miR-145-5p靶向TAGLN2调控结直肠癌SW620细胞侵袭和迁移的机制探讨

目的:探究微小RNA-145-5p（miR-145-5p）在结直肠癌组织和细胞中的表达情况及其靶向调控肌动蛋白凝胶蛋白2（TAGLN2）对结直肠癌细胞侵袭和迁移能力的影响。方法:采用实时荧光定量PCR（qPCR）技术对48例结直肠癌患者癌组织、配对癌旁组织、结直肠癌细胞株（HCT8、SW620、HCT116、HT-29）及结直肠黏膜细胞FHC中的miR-145-5p和TAGLN2 mRNA表达进行定量分析。将SW620细胞设为空白对照组、miR-145-5p mimics组、mimics-NC组、pcDNA3.1-TAGLN2组、pcDNA3.1-Vector组和miR-145-5p mimics+pcDNA3.1-TAGLN2组，采用qPCR检测miR-145-5p和TAGLN2 mRNA表达，采用Transwell法检测细胞侵袭及迁移能力，采用免疫印迹法（Western blot）检测TAGLN2蛋白及EMT相关蛋白表达，采用双荧光素酶报告实验检测miR-145-5p和TAGLN2间的靶向关系。结果:miR-145-5p在结直肠癌组织中的表达水平显著低于癌旁组织（P<0.05），并与结直肠癌患者的TNM分期和淋巴结转移相关（均P<0.05）；TAGLN2在结直肠癌组织中的表达水平显著高于癌旁组织，并与miR-145-5p表达呈负相关（P<0.05）；miR-145-5p和TAGLN2在结直肠癌HCT8、SW620、HCT116和HT-29细胞中的表达水平显著低于或高于FHC细胞（均P<0.05）。miR-145-5p过表达可降低SW620细胞的侵袭和迁移能力。miR-145-5p靶向调控TAGLN2表达，单独转染TAGLN2阳性质粒可增加SW620细胞的侵袭和迁移能力，与miR-145-5p mimics同时转染后，TAGLN2蛋白、波形蛋白（Vimentin）和神经钙黏素（N-cadherin）表达降低，上皮钙黏素（E-cadherin）表达升高，TAGLN2对SW620细胞侵袭和迁移能力的增强作用被显著抑制。结论:miR-145-5p在结直肠癌中呈低表达状态，其表达水平与结直肠癌患者的TNM分期和淋巴结转移密切相关，miR-145-5p靶向调控TAGLN2抑制结直肠癌细胞的侵袭和迁移能力。

The mechanism of miR-145-5p regulating invasion and migration of colorectal cancer SW620 cells by targeting TAGLN2

Objective:To explore the expression of miR-145-5p in colorectal cancer (CRC),and to investigate the effect of miR-145-5p on the invasion and migration of CRC cells by targeting TAGLN2.Methods:Quantitative real-time PCR (qPCR) was used to detect the expression of miR-145-5p and TAGLN2 mRNA in 48 cases of CRC tissues and corresponding adjacent tissues,CRC cell lines (HCT8,SW620,HCT116,HT-29) and human normal colorectal mucosal cells FHC.The SW620 cells were divided into blank control group,miR-145-5p mimics group,mimics-NC group,pcDNA3.1-TAGLN2 group,pcDNA3.1-Vector group and miR-145-5p mimics+pcDNA3.1-TAGLN2 group.The expression of miR-145-5p and TAGLN2 mRNA in SW620 cells was detected by qPCR.Transwell assay was used to detect the invasion and migration of SW620 cells.The expression of TAGLN2 and EMT-related proteins in SW620 cells was detected by Western blot.The targeting relationship between miR-145-5p and TAGLN2 was detected by dual luciferase reporter assay.Results:The expression of miR-145-5p in CRC tissues was significantly lower than that in adjacent tissues (P<0.05),which was correlated with TNM stage and lymphatic metastasis in CRC patients (all P<0.05).The expression of TAGLN2 in CRC tissues was significantly higher than that in adjacent tissues,and was negatively correlated with the expression of miR-145-5p (P<0.05).The expression of miR-145-5p and TAGLN2 in CRC cell lines（HCT8,SW620,HCT116 and HT-29）were significantly lower or higher than that in FHC cells(all P<0.05).Transfection with miR-145-5p mimics could reduce the invasion and migration of SW620 cells.miR-145-5p targeted the expression of TAGLN2.The up-regulation of TAGLN2 can promote the invasion and migration of SW620 cells.After co-transfection with miR-145-5p mimics,the expression of TAGLN2,Vimentin and N-cadherin was decreased,but the expression of E-cadherin was increased.The positive effects of TAGLN2 overexpression on invasion and migration of SW620 cells were significantly inhibited.Conclusion:The low expression of miR-145-5p in CRC was closely related to TNM stage and lymphatic metastasis of CRC patients.miR-145-5p inhibit the invasion and migration of colorectal cancer cells by targeting TAGLN2.