微小RNA-21通过调控肌球家族蛋白1对食管鳞癌迁移和侵袭的作用

Effects of microRNA-21 on migration and invasion of esophageal squamous cell carcinoma by regulating tropomyosin 1

目的 观察微小RNA-21（miRNA-21）通过调控肌球家族蛋白1（TPM1）对食管鳞癌侵袭转移的影响.方法 40例食管鳞癌组织及对应癌旁组织标本,聚合酶链反应（PCR）检测miRNA-21表达;随机抽取其中8例,PCR检测miRNA-21表达,Western blot检测TPM1表达.对食管癌细胞株EC109转染,利用反义miRNA-21核苷酸下调miRNA-21、miRNA-21核苷酸前体上调miRNA-21,分别以无义核苷酸转染作为对照组,检测miRNA-21和TPM1表达水平变化,并进行细胞迁移、侵袭功能实验.利用反义miRNA-21核苷酸和TPM1小干扰RNA作共转染,反义miRNA-21核苷酸和TPM1对照双链无义RNA共转染作为对照组,检测miRNA-21和TPM1表达水平变化,并进行细胞迁移、侵袭功能实验.结果 40例食管鳞癌组织中miRNA-21相对表达量为4.02 ±0.12,癌旁组织为0.60±0.11;其中随机8例患者癌组织中TPM1相对表达量为0.09±0.06,癌旁组织为0.87±0.34,与其miRNA-21表达呈负相关.在EC109中miRNA-21表达量为2.55±0.11;与对照组比较,下调miRNA-21表达后（0.30±0.12/2.35±0.32,P＜0.05）,TPM1表达上调（0.74±0.21/0.14±0.05,P＜0.05）,迁移、侵袭能力减弱（P＜0.05）;上调miRNA-21表达后（6.73±0.55/2.45±0.33,P＜0.05）,TPM1表达缺失（0.01 ±0.00/0.17±0.03,P＜0.05）,迁移、侵袭能力增强（P＜0.05）.共转染处理后,实验组较对照组TPM1 mRNA表达下降（0.15±0.03/3.55±1.25,P＜0.05）,miRNA-21表达被抑制（0.24±0.03/0.26±0.04,P＞0.05）,TPM1蛋白出现表达缺失（0.01 ±0.01/0.80 ±0.11,P＜0.05）,EC109侵袭、迁移能力增强（P＜0.05）.结论 在食管鳞癌中,miRNA-21可能通过抑制TPM1表达介导肿瘤侵袭与转移.

Objective To determine the effect of microRNA-21 （miRNA-21） on the migration and invasion of esophageal squamous cell carcinoma （ESCC） by targeting tropomyosin 1 （TPM1）.Methods The expression of miRNA-21 in 40 clinical ESCC samples and their adjacent normal epithelial samples was detected by polymerase chain reaction （PCR）,and the expression of TPM1 in 8 random pairs of these samples by Western blotting.EC109 cells were transfected with the anti-miRNA-21 oligonucleotide to down-regulate the expression of miRNA-21,and with the miRNA-21 mimics to up-regulate the expression of miRNA-21.The expression of miRNA-21 expression and TPM1 protein expression in these models was detected,and the ability of migration and invasion was examined.EC109 cells were co-transfected with TPM1-siRNA and anti-miRNA-21 oligonucleotide as the control group,and those cotransfected the TPM1-scramble and the anti-miRNA-21 oligonucleotide as the negative control group.The expression changes of miRNA-21 and TPM1 were observed,and the ability of cell migration and invasion was examined.Results The expression of miRNA-21 in these 40 ESCC samples was 4.02 ± 0.12,and 0.60 ± 0.11 in their adjacent normal epithelial samples.In these 8 random matched samples,the expression of TPM1 was 0.09 ± 0.06 in cancer samples,and 0.87 ± 0.34 in their adjacent samples,meanwhile negatively correlated with the miRNA-21 expression.The expression of miRNA-21 in EC109 cells was 2.55 ± 0.11.As compared with negative control group,depression of miRNA-21 （0.30 ± 0.12/2.35 ±0.32,P〈0.05） could restore the expression of TPM1 （0.74±0.21/0.14 ±0.05,P〈 0.05） in EC109 cells and promote migration and invasion （P〈 0.05）,while over-expression of miRNA-21 （6.73 ± 0.55/2.45 ± 0.33,P 〈 0.05） could delete the expression of TPM1 （0.01 ± 0.00/ 0.17 ± 0.03,P 〈 0.05） and relegate the migration and invasion capability （P 〈 0.05） of EC109s.In co-transfection models,as compared with the negative control group,the expression of TPM1 in control group decreased at mRNA level （0.15± 0.03/3.55 ± 1.25,P 〈 0.05）,the expression of miRNA-21 was down-regulated （0.24 ± 0.03/0.26 ± 0.04,P 〉 0.05） and the expression of TPM1 was suppressed （0.01 ±0.01/0.80 ±0.11,P 〈0.05）.Meanwhile EC109 cells in control group still had a high migration and invasion capability （P 〈 0.05）.Conclusion miRNA-21 could influence the migration and invasion of ESCC by regulating TPM1.