摘要
目的 观察微小RNA-135a（miRNA-135a）对人胰腺癌细胞株Bxpc-3增殖及凋亡的影响.方法 噻唑蓝（MTT）实验检测5-氟尿嘧啶（5-Fu）对Bxpc-3细胞的抑制率,选择合适的5-Fu浓度进行后续实验；流式细胞术检测5-Fu作用后Bxpc-3细胞的凋亡率；实时定量荧光逆转录聚合酶链反应（FQ-PCR）检测5 Fu作用后Bxpc-3细胞miRNA-135a的含量变化.设计miRNA-135a模拟物及反义寡核苷酸链,分别转染Bxpc-3细胞,并转染空病毒组作为对照,RT-qPCR检测各转染组miRNA-135a表达量；MTT法实验检测转染后各组Bxpc-3细胞的增殖率；流式细胞术检测5-Fu作用后各转染组Bxpc-3细胞的凋亡率.结果 5-Fu对胰腺癌细胞Bxpc-3的抑制有浓度和时间依赖性,随着药物浓度增大及作用时间延长,5-Fu对Bxpc-3抑制率增高.经5-Fu处理3、6、12h后miRNA-135a表达量是对照组的（0.45±0.02）、（0.60±0.03）、（0.73±0.02）倍,差异均有统计学意义（P＜0.05）.转染miRNA-135a模拟物组较空病毒miRNA-135a表达量升高（9.73±0.83）倍,差异有统计学意义（P＜0.05）；转染miRNA-135a反义寡核苷酸链组是空病毒组miRNA-135a表达量的（0.26±0.01）倍,差异有统计学意义（P＜0.05）.转染miRNA-135a模拟物组较转染空病毒组细胞增殖率明显升高,相反转染miRNA135反义寡核苷酸链组较转染空病毒组细胞增殖率明显降低.24h后,转染miRNA-135a模拟物组Bxpc-3细胞凋亡率[（12.03±1.07）％]较空病毒组[（20.61±1.53）％]明显降低,差异有统计学意义（P＜0.05）；转染miRNA-135a反义寡核苷酸链组Bxpc-3细胞凋亡率[（38.20±2.64）％]较空病毒组的（20.61±1.53）％明显升高,差异有统计学意义（P＜0.05）.结论 5-Fu作用于Bxpc-3细胞后miRNA-135a表达量降低；上调miRNA-135a表达,Bxpc-3细胞的增殖速率升高,Bxpc-3细胞的凋亡率降低；下调miRNA-135a的表达,Bxpc-3细胞增殖率降低,Bxpc-3细胞的凋亡率增高.
Objective To investigate the influence of miRNA-135a on the proliferation and apoptosis of the pancreatic cancer cells.Methods Methyl thiazol tetrazolium （MTT） was used to detect the inhibitory effect of 5-fluorouracil （5-Fu） on Bxpc-3 cells and the the appropriate concentration of 5-Fu was chosen in the following experiment.The apoptosis of Bxpc-3 cells was detected by using flow cytometry.Real-time fluorescent quantitative polymerase chain reaction （FQ-PCR） was used to detect the mRNA expression level of miRNA-135a after the Bxpc-3 cells were treated with 5-Fu.The mimic and anti-miRNA oligonuclotide （AMO） were designed and transfected into Bxpc-3 cells,and the Bxpc-3 cells transfected with vehicle served as control group.FQ-PCR was used to detect the expression of miRNA-135a and the proliferation rate was detected by using MTT assay.Flow cytometry was used to detect the apoptosis of transfected Bxpc-3 cells after treatment with 5-Fu.Results The 5-Fu inhibited Bxpc-3 cells in a concentration-and time-dependent manner.With the increase of concentration and prolongation of time,the inhibition rate of 5-Fu was elevated.The miRNA-135a level was reduced to （0.45 ±0.02）,（0.60 ±0.03）and （0.73 ± 0.02） times after treatment with 5-Fu （P 〈 0.05 for all）.After transfection the mRNA levels of miRNA-13a was elevated （9.73 ±0.83） times in the mimic groups and reduced to （0.26 ±0.01） times in the AMO groups as compared with the vehicle groups （P 〈 0.05 for all）.The proliferation of Bxpc-3 cells was elevated in the mimic group and reduced in the AMO group as compared with the vehicle group.The apoptosis of Bxpc-3 cells,which was （12.03 1.07） %,was reduced significantantly in the mimic groups as compared with the vehicle groups [（20.61 ± 1.53） %] （P 〈 0.05）,while the apoptosis was elevated in the AMO groups [（38.20 ±2.64）%] （P 〈0.05）.Conclusion The level of miRNA-135a was lower in Bxpe-3 cells treated with 5-Fu.The up-regulated expression of miRNA-135
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2014年第1期28-31,共4页
Chinese Journal of Experimental Surgery
基金
国家自然科学基金资助项目（30972928、31140078）
