miRNA-145对结肠癌细胞体外增殖、侵袭、凋亡的影响
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摘要
目的:探讨miRNA-145对结肠癌细胞体外增殖、侵袭、凋亡的影响。方法:采用脂质体转染法转染miRNA-145拟似物、阻遏物和阴性对照物至HCT116细胞,采用荧光实时定量PCR法检测细胞内miRNA-145表达,采用MTT法检测细胞增殖活性,采用Transwell小室法检测HCT116细胞侵袭,采用Annexin V-FITC/PI双染法检测转染后HCT116细胞凋亡情况。结果:转染后24 h、48 h、72 h时拟似物转染组细胞增殖活性显著低于空白对照组、阴性对照组和阻遏物转染组(P<0. 05),空白对照组和阴性对照组细胞增殖活性差异无统计学意义(P>0. 05),阻遏物转染组细胞增殖活性显著高于空白对照组、阴性对照组和拟似物转染组(P<0. 05)。转染24 h、48 h、72 h时拟似物转染组miRNA-145表达显著高于空白对照组、阴性对照组和阻遏物转染组,阻遏物转染组miRNA-145表达显著低于拟似物转染组、空白对照组和阴性对照组,差异均有统计学意义(P<0. 05);空白对照组和阴性对照组miRNA-145表达水平差异均无统计学意义(P>0. 05)。Transwell小室法检测结果显示,拟似物转染组穿膜细胞数显著少于空白对照组、阴性对照组和阻遏物转染组,阻遏物转染组穿膜细胞数显著多于空白对照组、阴性对照组和拟似物转染组,差异均有统计学意义(P<0. 05);空白对照组和阴性对照组穿膜细胞数相比较差异均无统计学意义(P>0. 05)。Annexin V-FITC/PI双染法检查结果显示,拟似物转染组细胞凋亡率显著高于空白对照组、阴性对照组和阻遏物转染组,阻遏物转染组细胞凋亡率显著低于空白对照组、阴性对照组和拟似物转染组,差异均有统计学意义(P<0. 05);空白对照组和阴性对照组细胞凋亡率相比较差异无统计学意义(P>0. 05)。结论:miRNA-145表达异常是影响HCT116细胞增殖、侵袭、凋亡重要因素,为新的基因靶点的药物开发提供了思路。
Objective: To study the effect of miRNA-145 on the proliferation,invasion and apoptosis of colorectal cancer cells in vitro. Methods: The miRNA-145 mimetic,repressor and negative control substance were transfected into HCT116 cells by liposome transfection. The expression of miRNA-145 in cells was detected by real-time fluorescence quantitative PCR. The proliferative activity of cells was detected by MTT assay. The invasion of HCT116 cells was detected by Transwell migration assay,and the apoptosis of HCT116 cells was detected by Annexin V-FITC/PI double staining. Results: At 24 h,48 h and 72 h after transfection,cell proliferation activity in mimetic transfection group was significantly lower than that in blank control group,negative control group and repressor transfection group( P<0. 05). There was no significant difference in proliferative activity between blank control group and negative control group( P>0. 05),and repressor transfection group had significantly higher proliferative activity than blank control group,negative control group and repressor transfection group( P< 0. 05). The expression of miRNA-145 in mimetic transfection group was significantly higher than that in blank control group,negative control group and repressor transfection group,and the expression of miRNA-145 in repressor transfection group was significantly lower than that in mimetic transfection group,blank control group and negative control group,the difference was statistically significant( P<0. 05),but there was no significant difference in expression level between blank control group and negative control group( P<0. 05). The detection results of Transwell migration method showed that the number of transmembrane cells of the mimetic transfection group was significantly less than that in blank control group,negative control group and repressor transfection group,the number of transmembrane cells was more in repressor transfection group than in blank control group,negative control group and mimetic transfection group,the differences were statistically significant( P<0. 05). There was no significant difference in the number of membrane cells between the blank control group and the negative control group( P>0. 05). The Annexin V-FITC/PI double staining showed that the apoptosis rate in mimetic transfection group was significantly higher than that of blank control group,negative control group and repressor transfection group. The apoptosis rate in repressor transfection group was significantly lower in blank control group,negative control group and mimetic transfection group,the differences were statistically significant( P< 0. 05). There was no significant difference in cell apoptosis rate between blank control group and negative control group( P>0. 05). Conclusion: The abnormal expression of miRNA-145 is an important factor in the proliferation,invasion and apoptosis of HCT116 cells,which provides a way for the development of new genes as a target for the development of drugs.
Objective: To study the effect of miRNA-145 on the proliferation,invasion and apoptosis of colorectal cancer cells in vitro. Methods: The miRNA-145 mimetic,repressor and negative control substance were transfected into HCT116 cells by liposome transfection. The expression of miRNA-145 in cells was detected by real-time fluorescence quantitative PCR. The proliferative activity of cells was detected by MTT assay. The invasion of HCT116 cells was detected by Transwell migration assay,and the apoptosis of HCT116 cells was detected by Annexin V-FITC/PI double staining. Results: At 24 h,48 h and 72 h after transfection,cell proliferation activity in mimetic transfection group was significantly lower than that in blank control group,negative control group and repressor transfection group( P<0. 05). There was no significant difference in proliferative activity between blank control group and negative control group( P>0. 05),and repressor transfection group had significantly higher proliferative activity than blank control group,negative control group and repressor transfection group( P< 0. 05). The expression of miRNA-145 in mimetic transfection group was significantly higher than that in blank control group,negative control group and repressor transfection group,and the expression of miRNA-145 in repressor transfection group was significantly lower than that in mimetic transfection group,blank control group and negative control group,the difference was statistically significant( P<0. 05),but there was no significant difference in expression level between blank control group and negative control group( P<0. 05). The detection results of Transwell migration method showed that the number of transmembrane cells of the mimetic transfection group was significantly less than that in blank control group,negative control group and repressor transfection group,the number of transmembrane cells was more in repressor transfection group than in blank control group,negative control group and mimetic transfection group,the differences were statistically significant( P<0. 05). There was no significant difference in the number of membrane cells between the blank control group and the negative control group( P>0. 05). The Annexin V-FITC/PI double staining showed that the apoptosis rate in mimetic transfection group was significantly higher than that of blank control group,negative control group and repressor transfection group. The apoptosis rate in repressor transfection group was significantly lower in blank control group,negative control group and mimetic transfection group,the differences were statistically significant( P< 0. 05). There was no significant difference in cell apoptosis rate between blank control group and negative control group( P>0. 05). Conclusion: The abnormal expression of miRNA-145 is an important factor in the proliferation,invasion and apoptosis of HCT116 cells,which provides a way for the development of new genes as a target for the development of drugs.
引文
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[9]邢晓芳,李子禹.miR-143和miR-145在胃癌中的表达及功能研究[J].中华胃肠外科杂志,2015,18(1):50-53.Xing XF,Li ZY.Expression and function of miR-143 and miR-145in gastric cancer[J].Chin J Gastrointest Surg,2015,18(1):50-53.
[10]张定富,吴秋芳,戈长征.miRNA-145在喉癌中的表达及临床意义[J].中国临床研究,2017,30(1):48-50.Zhang DF,Wu QF,Ge CZ.Expression and clinical significance of miRNA-145 in laryngeal carcinoma[J].Chin J Clin Res,2017,30(1):48-50.
[11]Doberstein K,Steinmeyer N,Hartemtz AK,et al.MicroRNA-145targets the metalloprotease ADAM17 and is suppressed in renal cell carcinoma patients[J].Neoplasia,2013,15(2):218-230.
[12]喻金梅,安云婷,戴红春.miRNA-145在不同宫颈组织中的表达意义[J].实用癌症杂志,2016,31(11):1747-1748,1753.Yu JM,An YT,Dai HC.Significance of miRNA-145 expression in different cervix tissues[J].Pract J Cancer,2016,31(11):1747-1748,1753.
[13]强勇,杨楠,张雷,等.非小细胞肺癌患者血清中miRNA-145的表达和意义[J].医学研究生学报,2016,29(1):62-65.Qiang Y,Yang N,Zhang L,et al.Expression and significance of miRNA-145 in serum of patients with non-small cell lung cancer[J].J Med Postgrad,2016,29(1):62-65.
[14]王丹.miRNA-145抑制肺癌A549细胞增殖活性的机制[J].江苏大学学报(医学版),2015,25(4):311-316.Wang D.MiRNA-145 inhibits the proliferation activity of A549cells in lung cancer[J].J Jiangsu Univ(Med Edi),2015,25(4):311-316.
[15]赵一奇,焦峰,闵波,等.贲门癌miRNA-145的表达[J].现代肿瘤医学,2016,24(6):891-893.Zhao YQ,Jiao F,Min B,et al.The expression of miRNA-145 in cardia cancer[J].J Mod Oncol,2016,24(6):891-893.
[2]刘娜,李文强,高成伟.冬凌草甲素通过下调NF-κB/COX-2表达抑制结肠癌Lo Vo细胞增殖[J].山西医科大学学报,2017,48(3):251-255.Liu N,Li WQ,Gao CW.Oridonin inhibits the proliferation of colon cancer Lo Vo cells by down-regulation of NF-kappa B/COX-2 expression[J].J Shanxi Med Univ,2017,48(3):251-255.
[3]连建安,姜斌骅,傅永清.结肠癌细胞OPN和HIF-1α基因表达水平与结肠癌恶性生物学行为的关系[J].中国卫生检验杂志,2017,27(21):3140-3141,3144.Lian JN,Jiang BH,Fu YQ.The relationship between the gene expression level of OPN and HIF-1 alpha in colon cancer cells and the malignant biological behavior of colon cancer[J].Chin J Health Lab Technol,2017,27(21):3140-3141,3144.
[4]银瑞,董新伟,王铮,等.肺腺癌相关微小RNA的筛选及其功能研究[J].中华实验外科杂志,2016,33(1):115-117.Yin R,Dong XW,Wang Z,et al.Screening and functional study of small RNA associated with lung adenocarcinoma[J].Chin J Exp Surg,2016,33(1):115-117.
[5]郭莹,瞿文,王一浩,等.microRNA在自身免疫性疾病中的研究进展[J].中国免疫学杂志,2016,32(8):1237-1240,1244.Guo Y,Qu W,Wang YH,et al.microRNA research progress in autoimmune diseases[J].Chin J Immunol,2016,32(8):1237-1240,1244.
[6]Xu Q,Liu LZ,Qian X,et al.miR-145,a new regulator of the DNAfragmentation factor-45(DEF-45)-mediated apoptotic network[J].Mol Cancer,2010,9(2):11-16.
[7]Yan X,Chen X,Liang H,et al.miR-143 and miR-145synergistically regulate ERBB3 to suppress cell proliferation and invasion in breast cancer[J].Mol Cancer,2014,24(13):220.
[8]郭斌,郭枫,郭永连,等.miRNA-145在前列腺癌中的表达及临床意义[J].中国老年学杂志,2017,37(19):4824-4825.Guo B,Guo F,Guo YL,et al.Expression and clinical significance of MiRNA-145 in prostate cancer[J].Chin J Gerontol,2017,37(19):4824-4825.
[9]邢晓芳,李子禹.miR-143和miR-145在胃癌中的表达及功能研究[J].中华胃肠外科杂志,2015,18(1):50-53.Xing XF,Li ZY.Expression and function of miR-143 and miR-145in gastric cancer[J].Chin J Gastrointest Surg,2015,18(1):50-53.
[10]张定富,吴秋芳,戈长征.miRNA-145在喉癌中的表达及临床意义[J].中国临床研究,2017,30(1):48-50.Zhang DF,Wu QF,Ge CZ.Expression and clinical significance of miRNA-145 in laryngeal carcinoma[J].Chin J Clin Res,2017,30(1):48-50.
[11]Doberstein K,Steinmeyer N,Hartemtz AK,et al.MicroRNA-145targets the metalloprotease ADAM17 and is suppressed in renal cell carcinoma patients[J].Neoplasia,2013,15(2):218-230.
[12]喻金梅,安云婷,戴红春.miRNA-145在不同宫颈组织中的表达意义[J].实用癌症杂志,2016,31(11):1747-1748,1753.Yu JM,An YT,Dai HC.Significance of miRNA-145 expression in different cervix tissues[J].Pract J Cancer,2016,31(11):1747-1748,1753.
[13]强勇,杨楠,张雷,等.非小细胞肺癌患者血清中miRNA-145的表达和意义[J].医学研究生学报,2016,29(1):62-65.Qiang Y,Yang N,Zhang L,et al.Expression and significance of miRNA-145 in serum of patients with non-small cell lung cancer[J].J Med Postgrad,2016,29(1):62-65.
[14]王丹.miRNA-145抑制肺癌A549细胞增殖活性的机制[J].江苏大学学报(医学版),2015,25(4):311-316.Wang D.MiRNA-145 inhibits the proliferation activity of A549cells in lung cancer[J].J Jiangsu Univ(Med Edi),2015,25(4):311-316.
[15]赵一奇,焦峰,闵波,等.贲门癌miRNA-145的表达[J].现代肿瘤医学,2016,24(6):891-893.Zhao YQ,Jiao F,Min B,et al.The expression of miRNA-145 in cardia cancer[J].J Mod Oncol,2016,24(6):891-893.