miRNA-125a靶向调控SMURF1对结肠癌生物学行为的影响
|
摘要
目的探讨miRNA-125a通过对靶基因SMURF1的调控在结肠癌中的影响。方法 qRT-PCR检测结肠癌患者血清中miRNA-125a和SMURF1表达水平并分析其与结肠癌相关临床指标的相关性;microRNA靶基因数据库预测miRNA-125a靶基因SMURF1并使用荧光素酶报告基因法验证其结合;qRTPCR和Western blot检测miRNA-125a模拟物对SMURF1mRNA及蛋白表达的影响;HE染色检测结肠癌组织中SMURF1的表达;CCK-8法、细胞划痕实验及流式细胞法检测miRNA-125a模拟物和SMURF1对SW480细胞增殖、迁移及周期的影响;构建结肠癌肿瘤异种移植小鼠模型,采用称重及PCNA染色检测miRNA-125a模拟物对小鼠体内肿瘤生长的影响。结果结肠癌患者血清中miRNA-125a表达水平与结肠癌相关临床指标呈负相关性,SMURF1表达水平与结肠癌相关临床指标呈正相关性。结肠癌组织中miRNA-125a的表达水平显著降低,miRNA-125a模拟物可抑制SMURF1的mRNA翻译及蛋白水平。结肠癌肿瘤组织中SMURF1表达水平明显升高,miRNA-125a模拟物可以抑制SW480细胞的增殖、迁移和S期细胞周期的聚集,然而这种抑制效果会因SMURF1表达升高而减弱。miRNA-125a模拟物抑制小鼠体内结肠癌生长及SMURF1的表达。结论 miRNA-125a通过下调SMURF1的表达在结肠癌的发展过程中发挥抑制作用,具有成为结肠癌诊断及治疗靶点的潜力。
Objective To investigate the effect of miRNA-125 a on biological behavior of colon cancer through regulating SMURF1. Methods The expression of miRNA-125 a and SMURF1 in the serum of colon cancer patients were detected by qRT-PCR and their correlation with clinical indicators related to colon cancer was analyzed. The microRNA target gene database predicted the combination of miRNA-125 a target gene and SMURF1 which was also verified by luciferase reporter gene method. qRT-PCR and Western blot were used to detect the effect of miRNA-125 a mimic on the mRNA and protein expression of SMURF1. HE staining was used to detect the expression of SMURF1 in colon cancer tissues; CCK-8 method, cell scratch test and flow cytometry were used to identify the effects of SMURF1 on the proliferation, migration and cycle of SW480 cells. The mouse model of colon cancer xenografts was constructed and the effect of miRNA-125 a mimics on tumor growth in mice was detected by weighing and PCNA staining. Results The expression of miRNA-125 a in the serum of patients with colon cancer was negatively correlated with the clinical indicators of colon cancer. The expression of SMURF1 was positively correlated with the clinical indicators of colon cancer. The expression level of miRNA-125 a in colon cancer tissues was significantly reduced. The miRNA-125 a mimic inhibited SMURF1 mRNA translation and protein levels. The expression of SMURF1 in colon cancer tissues was significantly increased. miRNA-125 a mimics could inhibit the proliferation, migration and cell cycle accumulation of SW480 cells. However, this inhibitory effect could be compensated by the increased expression of SMURF1. miRNA-125 a mimics inhibited the growth of colon cancer and SMURF1 expression in mice. Conclusion miRNA-125 a plays an inhibitory role in the development of colon cancer by down-regulating the expression of SMURF1, and it is a potential target for the diagnosis and treatment of colon cancer.
Objective To investigate the effect of miRNA-125 a on biological behavior of colon cancer through regulating SMURF1. Methods The expression of miRNA-125 a and SMURF1 in the serum of colon cancer patients were detected by qRT-PCR and their correlation with clinical indicators related to colon cancer was analyzed. The microRNA target gene database predicted the combination of miRNA-125 a target gene and SMURF1 which was also verified by luciferase reporter gene method. qRT-PCR and Western blot were used to detect the effect of miRNA-125 a mimic on the mRNA and protein expression of SMURF1. HE staining was used to detect the expression of SMURF1 in colon cancer tissues; CCK-8 method, cell scratch test and flow cytometry were used to identify the effects of SMURF1 on the proliferation, migration and cycle of SW480 cells. The mouse model of colon cancer xenografts was constructed and the effect of miRNA-125 a mimics on tumor growth in mice was detected by weighing and PCNA staining. Results The expression of miRNA-125 a in the serum of patients with colon cancer was negatively correlated with the clinical indicators of colon cancer. The expression of SMURF1 was positively correlated with the clinical indicators of colon cancer. The expression level of miRNA-125 a in colon cancer tissues was significantly reduced. The miRNA-125 a mimic inhibited SMURF1 mRNA translation and protein levels. The expression of SMURF1 in colon cancer tissues was significantly increased. miRNA-125 a mimics could inhibit the proliferation, migration and cell cycle accumulation of SW480 cells. However, this inhibitory effect could be compensated by the increased expression of SMURF1. miRNA-125 a mimics inhibited the growth of colon cancer and SMURF1 expression in mice. Conclusion miRNA-125 a plays an inhibitory role in the development of colon cancer by down-regulating the expression of SMURF1, and it is a potential target for the diagnosis and treatment of colon cancer.
引文
[1]Yong FL,Law CW,Wang CW.Potentiality of a triple microRNAclassifier:miR-193a-3p,miR-23a and miR-338-5p for early detection of colorectal cancer[J].BMC Cancer,2013,13:280.
[2]Zhou Y,Wan G,Spizzo R,et al.miR-203 induces oxaliplatin resistance in colorectal cancer cells by negatively regulating ATMkinase[J].Mol Oncol,2014,8(1):83-92.
[3]Nie J,Liu L,Xing G,et al.CKIP-1 acts as a colonic tumor suppressor by repressing oncogenic Smurf1 synthesis and promoting Smurf1 autodegradation[J].Oncogene,2014,33(28):3677-87.
[4]范忠义,焦顺昌.Mir-125a与相关疾病关系的研究进展[J].解放军医学院学报,2015,36(11):1152-5.[Fan ZY,Jiao SC.Advances in miR-125a and its relative diseases[J].Jie Fang Jun Yi Xue Yuan Xue Bao,2015,36(11):1152-5.]
[5]Yu W,Chai H.Inhibition of BAMBI reduces the viability and motility of colon cancer via activating TGF-β/Smad pathway in vitro and in vivo[J].Oncol Lett,2017,14(4):4793-9.
[6]Al-Haidari AA,Syk I,Thorlacius H.MiR-155-5p positively regulates CCL17-induced colon cancer cell migration by targeting RhoA[J].Oncotarget,2017,8(9):14887-96.
[7]Yang C,Wang MH,Zhou JD,et al.Upregulation of miR-542-3p inhibits the growth and invasion of human colon cancer cells through PI3K/AKT/survivin signaling[J].Oncol Rep,2017,38(6):3545-53.
[8]罗吉,罗燕,李勇敏,等.健脾消癌方对结肠癌TGF-β/lncRNA-ATB/miR-200a信号通路的影响[J].中国实验方剂学杂志,2018,24(6):161-6.[Luo J,Luo Y,Li YM,et al.Effect of Jianpi Xiaoai Prescription on TGF-β/lncRNA-ATB/miR-200a Signal Pathway in Colorectal Cancer[J].Zhongguo Shi Yan Fang Ji Xue Za Zhi,2018,24(6):161-6.]
[9]Pagliuca A,Valvo C,Fabrizi E,et al.Analysis of the combined action of miR-143 and miR-145 on oncogenic pathways in colorectal cancer cells reveals a coordinate program of gene repression[J].Oncogene,2013,32(40):4806-13.
[10]龙建辰,赵泉,杜晓倩,等.miRNA-101通过靶向CDK8调控结肠癌细胞侵袭和Wnt/β-catenin信号通路活化[J].肿瘤防治研究,2018,45(9):666-71.[Long JC,Zhao Q,Du XQ,et al.miR-NA-101 regulates colon cancer cell invasion and Wnt/β-catenin pathway activation through targeting CDK8[J].Zhong Liu Fang Zhi Yan Jiu,2018,45(9):666-71.]
[11]Kristina S,Kjetil B,Weum AT,et al.Clinical relevance of microRNA miR-21,miR-31,miR-92a,miR-101,miR-106a and miR-145 in colorectal cancer[J].BMC Cancer,2012,12:505.
[12]Pichler M,Winter E,Ress AL,et al.miR-181a is associated with poor clinical outcome in patients with colorectal cancer treated with EGFR inhibitor[J].J Clin Pathol,2014,67(3):198-203.
[13]To KK,Leung WW,Ng SS.A novel miR-203-DNMT3bABCG2 regulatory pathway predisposing colorectal cancer development[J].Mol Carcinog,2016,56(2):464-77.
[14]Lyu H,Huang J,Edgerton SM,et al.Increased erbB3 promotes erbB2/neu-driven mammary tumor proliferation and co-targeting of erbB2/erbB3 receptors exhibits potent inhibitory effects on breast cancer cells[J].Int J Clin Exp Pathol,2015,8(6):6143-56.
[15]Nie J,Liu L,Xing G,et al.CKIP-1 acts as a colonic tumor suppressor by repressing oncogenic Smurf1 synthesis and promoting Smurf1 autodegradation[J].Oncogene,2014,33(28):3677-87.
[16]Liu J,Chen Y,Huang Q,et al.IRAK2 counterbalances oncogenic Smurf1 in colon cancer cells by dictating ER stress[J].Cell Signal,2018,48:69-80.
[2]Zhou Y,Wan G,Spizzo R,et al.miR-203 induces oxaliplatin resistance in colorectal cancer cells by negatively regulating ATMkinase[J].Mol Oncol,2014,8(1):83-92.
[3]Nie J,Liu L,Xing G,et al.CKIP-1 acts as a colonic tumor suppressor by repressing oncogenic Smurf1 synthesis and promoting Smurf1 autodegradation[J].Oncogene,2014,33(28):3677-87.
[4]范忠义,焦顺昌.Mir-125a与相关疾病关系的研究进展[J].解放军医学院学报,2015,36(11):1152-5.[Fan ZY,Jiao SC.Advances in miR-125a and its relative diseases[J].Jie Fang Jun Yi Xue Yuan Xue Bao,2015,36(11):1152-5.]
[5]Yu W,Chai H.Inhibition of BAMBI reduces the viability and motility of colon cancer via activating TGF-β/Smad pathway in vitro and in vivo[J].Oncol Lett,2017,14(4):4793-9.
[6]Al-Haidari AA,Syk I,Thorlacius H.MiR-155-5p positively regulates CCL17-induced colon cancer cell migration by targeting RhoA[J].Oncotarget,2017,8(9):14887-96.
[7]Yang C,Wang MH,Zhou JD,et al.Upregulation of miR-542-3p inhibits the growth and invasion of human colon cancer cells through PI3K/AKT/survivin signaling[J].Oncol Rep,2017,38(6):3545-53.
[8]罗吉,罗燕,李勇敏,等.健脾消癌方对结肠癌TGF-β/lncRNA-ATB/miR-200a信号通路的影响[J].中国实验方剂学杂志,2018,24(6):161-6.[Luo J,Luo Y,Li YM,et al.Effect of Jianpi Xiaoai Prescription on TGF-β/lncRNA-ATB/miR-200a Signal Pathway in Colorectal Cancer[J].Zhongguo Shi Yan Fang Ji Xue Za Zhi,2018,24(6):161-6.]
[9]Pagliuca A,Valvo C,Fabrizi E,et al.Analysis of the combined action of miR-143 and miR-145 on oncogenic pathways in colorectal cancer cells reveals a coordinate program of gene repression[J].Oncogene,2013,32(40):4806-13.
[10]龙建辰,赵泉,杜晓倩,等.miRNA-101通过靶向CDK8调控结肠癌细胞侵袭和Wnt/β-catenin信号通路活化[J].肿瘤防治研究,2018,45(9):666-71.[Long JC,Zhao Q,Du XQ,et al.miR-NA-101 regulates colon cancer cell invasion and Wnt/β-catenin pathway activation through targeting CDK8[J].Zhong Liu Fang Zhi Yan Jiu,2018,45(9):666-71.]
[11]Kristina S,Kjetil B,Weum AT,et al.Clinical relevance of microRNA miR-21,miR-31,miR-92a,miR-101,miR-106a and miR-145 in colorectal cancer[J].BMC Cancer,2012,12:505.
[12]Pichler M,Winter E,Ress AL,et al.miR-181a is associated with poor clinical outcome in patients with colorectal cancer treated with EGFR inhibitor[J].J Clin Pathol,2014,67(3):198-203.
[13]To KK,Leung WW,Ng SS.A novel miR-203-DNMT3bABCG2 regulatory pathway predisposing colorectal cancer development[J].Mol Carcinog,2016,56(2):464-77.
[14]Lyu H,Huang J,Edgerton SM,et al.Increased erbB3 promotes erbB2/neu-driven mammary tumor proliferation and co-targeting of erbB2/erbB3 receptors exhibits potent inhibitory effects on breast cancer cells[J].Int J Clin Exp Pathol,2015,8(6):6143-56.
[15]Nie J,Liu L,Xing G,et al.CKIP-1 acts as a colonic tumor suppressor by repressing oncogenic Smurf1 synthesis and promoting Smurf1 autodegradation[J].Oncogene,2014,33(28):3677-87.
[16]Liu J,Chen Y,Huang Q,et al.IRAK2 counterbalances oncogenic Smurf1 in colon cancer cells by dictating ER stress[J].Cell Signal,2018,48:69-80.