上调miRNA-145表达对非小细胞肺癌细胞增殖、凋亡及放射敏感性的影响
|
摘要
目的探讨上调miRNA-145表达对非小细胞肺癌细胞增殖、凋亡及放射敏感性的影响。方法收集人正常肺上皮细胞株BEAS-2B和非小细胞肺癌细胞株A549,采用逆转录聚合酶链反应(RT-PCR)检测两种细胞中miRNA-145的相对表达量。以LipofectamineTM2000转染试剂将miRNA-145模拟物和阴性对照质粒转染至A549细胞中,分别作为miRNA-145组和NC组,以只加入转染试剂的A549细胞作为对照组,RT-PCR检测各组细胞中miRNA-145的相对表达量。采用四甲基偶氮唑蓝(MTT)法和流式细胞术(FCM)检测上调miRNA-145表达对细胞增殖和凋亡的影响。给予不同放射剂量X线照射后采用克隆形成实验检测细胞的放射敏感性。结果A549细胞中miRNA-145的相对表达量明显低于正常肺上皮BEAS-2B细胞(P﹤0.01);转染后,miRNA-145组细胞中miRNA-145的相对表达量高于对照组(P﹤0.05);转染后,NC组与对照组细胞中miRNA-145的相对表达量比较,差异无统计学意义(P﹥0.05);miRNA-145组细胞转染24、48、72 h的光密度(OD)值均低于对照组(P﹤0.05);转染48 h后,miRNA-145组细胞的凋亡率高于对照组(P﹤0.05);2、4、6、8 Gy剂量的X线照射后miRNA-145组细胞的存活分数(SF)均低于对照组,放射增敏比(SER)为1.491。结论与正常肺上皮BEAS-2B细胞比较,A549细胞中miRNA-145的相对表达量较低,上调其表达能够抑制A549细胞增殖,促进细胞凋亡,增强放射敏感性。
Objective To investigate the effect of up regulation of miRNA-145 on cell proliferation, apoptosis and radiosensitivity in non-small cell lung cancer. Method The human lung epithelial cell line BEAS-2 B cells and non-small cell lung cancer A549 cells were collected, and the relative expression level of miRNA-145 was detected by quantitative reverse transcription-polymerase chain reaction(RT-PCR). A549 cells were transfected with the miRNA-145 mimic(miRNA-145 group) and negative control plasmid(NC group) using LipofectamineTM2000 transfection reagent, only transfection reagents were added to the control group. The relative expression level of miRNA-145 in above three groups were detected by RT-PCR. Proliferation and apoptosis of A549 cells were also evaluated by methylthiazolyldiphenyl-tetrazolium bromide(MTT) assay and flow cytometry(FCM). The radiosensitivity of the cells was examined by clonogenic assay after various doses of irradiation exposure. Result A549 cells showed significant decreased in miRNA-145 relative expression compared with the normal lung epithelial BEAS-2 B cell(P<0.01). After transfection, the relative expression of miRNA-145 in the miRNA-145 group was significant higher than that in the control group(P<0.05), while there was no significant difference between the NC group and the control group(P>0.05). The optical density(OD) values of miRNA-145 group at 24, 48, 72 h post transfection were lower than those of control group(P<0.05), and the apoptotic rate of nonsmall cell lung cancer cells at 48 h post transfection was higher than that of control group(P<0.05). The cells were irradiated with 2, 4, 6 or 8 Gy X-ray irradiation and the survival fraction(SF) of cells in the miRNA-145 group was lower than that in the control group, and the enhancement ratio(SER) was 1.491. Conclusion Compared with normal lung epithelial BEAS-2 B cells, the relative expression of miRNA-145 is lower in non-small cell lung cancer cells, and up regulation of miRNA-145 can inhibit the proliferation, improve the apoptosis, and enhance the radiosensitivity of A549 cells.
Objective To investigate the effect of up regulation of miRNA-145 on cell proliferation, apoptosis and radiosensitivity in non-small cell lung cancer. Method The human lung epithelial cell line BEAS-2 B cells and non-small cell lung cancer A549 cells were collected, and the relative expression level of miRNA-145 was detected by quantitative reverse transcription-polymerase chain reaction(RT-PCR). A549 cells were transfected with the miRNA-145 mimic(miRNA-145 group) and negative control plasmid(NC group) using LipofectamineTM2000 transfection reagent, only transfection reagents were added to the control group. The relative expression level of miRNA-145 in above three groups were detected by RT-PCR. Proliferation and apoptosis of A549 cells were also evaluated by methylthiazolyldiphenyl-tetrazolium bromide(MTT) assay and flow cytometry(FCM). The radiosensitivity of the cells was examined by clonogenic assay after various doses of irradiation exposure. Result A549 cells showed significant decreased in miRNA-145 relative expression compared with the normal lung epithelial BEAS-2 B cell(P<0.01). After transfection, the relative expression of miRNA-145 in the miRNA-145 group was significant higher than that in the control group(P<0.05), while there was no significant difference between the NC group and the control group(P>0.05). The optical density(OD) values of miRNA-145 group at 24, 48, 72 h post transfection were lower than those of control group(P<0.05), and the apoptotic rate of nonsmall cell lung cancer cells at 48 h post transfection was higher than that of control group(P<0.05). The cells were irradiated with 2, 4, 6 or 8 Gy X-ray irradiation and the survival fraction(SF) of cells in the miRNA-145 group was lower than that in the control group, and the enhancement ratio(SER) was 1.491. Conclusion Compared with normal lung epithelial BEAS-2 B cells, the relative expression of miRNA-145 is lower in non-small cell lung cancer cells, and up regulation of miRNA-145 can inhibit the proliferation, improve the apoptosis, and enhance the radiosensitivity of A549 cells.
引文
[1]Siegel RL,Miller KD,Jemal A.Cancer Statistics,2017[J]CA Cancer J Clin,2017,67(1):7-30.
[2]Zhou C,Yao LD.Strategies to improve outcomes of pa tients with EGRF-mutant non-small cell lung cancer:re view of the literature[J].J Thorac Oncol,2016,11(2):174186.
[3]刘聪,刘元水,张学德,等.晚期非小细胞肺癌靶向药物临床应用进展[J].山西医药杂志,2016,45(22):2627-2630.
[4]Mei Z,Su T,Ye J,et al.The miR-15 family enhances the ra diosensitivity of breast cancer cells by targeting G2 check points[J].Radiat Res,2015,183(2):196-207.
[5]Yuan W,Xiaoyun H,Haifeng Q,et al.MicroRNA-218 en hances the radiosensitivity of human cervical cancer vi promoting radiation induced apoptosis[J].Int J Med Sci2014,11(7):691-696.
[6]Wang W,Liu J,Wu Q.MiR-205 suppresses autophagy and enhances radiosensitivity of prostate cancer cells by target ing TP53INP1[J].Eur Rev Med Pharmacol Sci,2016,20(1):92-100.
[7]Qiu T,Zhou X,Wang J,et al.MiR-145,miR-33a and miR133b inhibit proliferation,migration,invasion and cell cy cle progression via targeting transcription factor Sp1 in gas tric cancer[J].FEBS Lett,2014,588(7):1168-1177.
[8]贺新伟,林道浙,薛迪新,等.miRNA-145在甲状腺乳头状癌中的表达及功能研究[J].中华内分泌外科杂志,201711(3):224-227;248.
[9]Wei H,Wen-ming C,Jun-bo J.Plasma miRNA-145 as novel biomarker for the diagnosis and radiosensitivity pre diction of human cervical cancer[J].J Int Med Res,2017,45(3):1054-1060.
[10]Cui SY,Wang R,Chen LB.MicroRNA-145:a potent tu mour suppressor that regulates multiple cellular pathway[J].J Cell Mol Med,2014,18(10):1913-1926.
[11]郭文炜,曾志勇,徐驰.miRNA-145与肿瘤相关性研究进展[J].实用医学杂志,2013,29(23):3947-3948.
[12]Lu R,Ji Z,Li X,et al.miRNA-145 functions as tumor sup pressor and targets two oncogenes,ANGPT2 and NEDD9in renal cell carcinoma[J].J Cancer Res Clin Oncol,2014140(3):387-397.
[13]王寰昱,王亚峰,张昆松,等.上调miRNA145对肝癌细胞的作用及其机制研究[J].中国病理生理杂志,2015,31(6):1019-1025.
[14]张伟,刘伟良,王勇,等.miRNA-145在非小细胞肺癌中的表达[J].现代肿瘤医学,2017,25(15):2424-2427.
[15]王春梅,高艳丽,尹丽霞.非小细胞肺癌放射治疗研究进展[J].中华肿瘤防治杂志,2017,24(10):720-724.
[16]Florczuk M,Szpechcinski A,Chorostowska-Wynimko JmiRNAs as biomarkers and therapeutic targets in non small cell lung cancer:current perspectives[J].Target On col,2017,12(2):179-200.
[17]Ye C,Sun NX,Ma Y,et al.MicroRNA-145 contributes to enhancing radiosensitivity of cervical cancer cells[J]FEBS Lett,2015,589(6):702-709.
[18]Yan S,Li X,Jin Q,et al.MicroRNA-145 sensitizes cervi cal cancer cells to low-dose irradiation by downregulating OCT4 expression[J].Exp Ther Med,2016,12(5):31303136.
[19]Gong P,Zhang T,He D,et al.MicroRNA-145 modulate tumor sensitivity to radiation in prostate cancer[J].Radia Res,2015,184(6):630-638.
[2]Zhou C,Yao LD.Strategies to improve outcomes of pa tients with EGRF-mutant non-small cell lung cancer:re view of the literature[J].J Thorac Oncol,2016,11(2):174186.
[3]刘聪,刘元水,张学德,等.晚期非小细胞肺癌靶向药物临床应用进展[J].山西医药杂志,2016,45(22):2627-2630.
[4]Mei Z,Su T,Ye J,et al.The miR-15 family enhances the ra diosensitivity of breast cancer cells by targeting G2 check points[J].Radiat Res,2015,183(2):196-207.
[5]Yuan W,Xiaoyun H,Haifeng Q,et al.MicroRNA-218 en hances the radiosensitivity of human cervical cancer vi promoting radiation induced apoptosis[J].Int J Med Sci2014,11(7):691-696.
[6]Wang W,Liu J,Wu Q.MiR-205 suppresses autophagy and enhances radiosensitivity of prostate cancer cells by target ing TP53INP1[J].Eur Rev Med Pharmacol Sci,2016,20(1):92-100.
[7]Qiu T,Zhou X,Wang J,et al.MiR-145,miR-33a and miR133b inhibit proliferation,migration,invasion and cell cy cle progression via targeting transcription factor Sp1 in gas tric cancer[J].FEBS Lett,2014,588(7):1168-1177.
[8]贺新伟,林道浙,薛迪新,等.miRNA-145在甲状腺乳头状癌中的表达及功能研究[J].中华内分泌外科杂志,201711(3):224-227;248.
[9]Wei H,Wen-ming C,Jun-bo J.Plasma miRNA-145 as novel biomarker for the diagnosis and radiosensitivity pre diction of human cervical cancer[J].J Int Med Res,2017,45(3):1054-1060.
[10]Cui SY,Wang R,Chen LB.MicroRNA-145:a potent tu mour suppressor that regulates multiple cellular pathway[J].J Cell Mol Med,2014,18(10):1913-1926.
[11]郭文炜,曾志勇,徐驰.miRNA-145与肿瘤相关性研究进展[J].实用医学杂志,2013,29(23):3947-3948.
[12]Lu R,Ji Z,Li X,et al.miRNA-145 functions as tumor sup pressor and targets two oncogenes,ANGPT2 and NEDD9in renal cell carcinoma[J].J Cancer Res Clin Oncol,2014140(3):387-397.
[13]王寰昱,王亚峰,张昆松,等.上调miRNA145对肝癌细胞的作用及其机制研究[J].中国病理生理杂志,2015,31(6):1019-1025.
[14]张伟,刘伟良,王勇,等.miRNA-145在非小细胞肺癌中的表达[J].现代肿瘤医学,2017,25(15):2424-2427.
[15]王春梅,高艳丽,尹丽霞.非小细胞肺癌放射治疗研究进展[J].中华肿瘤防治杂志,2017,24(10):720-724.
[16]Florczuk M,Szpechcinski A,Chorostowska-Wynimko JmiRNAs as biomarkers and therapeutic targets in non small cell lung cancer:current perspectives[J].Target On col,2017,12(2):179-200.
[17]Ye C,Sun NX,Ma Y,et al.MicroRNA-145 contributes to enhancing radiosensitivity of cervical cancer cells[J]FEBS Lett,2015,589(6):702-709.
[18]Yan S,Li X,Jin Q,et al.MicroRNA-145 sensitizes cervi cal cancer cells to low-dose irradiation by downregulating OCT4 expression[J].Exp Ther Med,2016,12(5):31303136.
[19]Gong P,Zhang T,He D,et al.MicroRNA-145 modulate tumor sensitivity to radiation in prostate cancer[J].Radia Res,2015,184(6):630-638.