miR-119a靶向作用FHIT对膀胱肿瘤细胞生物学行为的影响

The effects of miR-119a targeting FHIT on the biological beharior of bladder tumor cells

目的探讨miR-119a靶向作用FHIT对膀胱肿瘤细胞生物学行为的影响。进一步明确膀胱癌发生、发展过程中的分子作用机制,以及寻找膀胱癌生物治疗新作用靶点提供理论依据。方法将miR-119a作为研究的目的基因,采用TargetScan靶基因预测软件,预测miR-119a可能靶向调控FHIT基因。选取125例膀胱癌患者的膀胱癌组织及癌旁正常组织,实时定量PCR(Real-time PCR)用于检验膀胱癌组织及癌旁正常组织miR-119a及FHIT的表达量并进行对比。设计合成并构建miR-119a慢病毒表达载体,并成功转染至人膀胱癌T24和5637细胞中。然后基于目前常用的体外实验方法,如CCK-8、周期、平板克隆、Transwell及细胞划痕实验等实验,对比分析实验组(即上调miR-119a组)和对照组(即未上调miR-119a组)在细胞增殖、迁移、侵袭等过程的差异性。结果膀胱癌组织中的miR-119a表达量(11.330±0.106)明显比正常组织中(0.365±0.076)的高,差异有统计学意义(P<0.05);但FHIT的表达量较低(0.596±0.125 vs 1.203±0.212)。上调miR-119a组中5637细胞和T24细胞的miR-119a表达量均有所增加,与对照组相比差异有统计学意义(P<0.05);CCK-8实验及平板克隆实验与对照组相比,上调组的细胞吸光度(OD)值及细胞集落数量明显增加,差异有统计学意义(P<0.05)。细胞周期实验,上调miR-119a后停留在分裂期的膀胱肿瘤细胞明显增加(T24:64.325±5.214 vs 55.315±4.215)(5637:53.103±3.124 vs 46.324±4.452),差异有统计学意义(P<0.05);Transwell实验及细胞划痕实验,与对组照组相比,上调膀胱肿瘤细胞穿膜细胞的数量及爬行速率升高明显,差异有统计学意义(P<0.05)。结论 miR-119a在膀胱癌组织中表达量显著上升,上调miR-119a可能与靶向抑制FHIT的表达来促进膀胱肿瘤细胞增值与迁移效应有关。

Objective To explore the effects of microRNA-119a(miR-119a) on the proliferation and invasiveness of bladder cancer cells. To further clarify the molecular mechanism in the development of bladder cancer, and to provide theoretical basis for the search of new biological therapeutic targets for bladder cancer. Methods Using miR-119a as the target gene, the association between miR-119a and FHIT gene was detected by TargetScan target gene prediction software. Bladder cancer tissues and adjacent normal tissues from 125 bladder cancer patients were prepared for this study. Real-time quantitative PCR was used to examine the expression of miR-119a and FHIT in bladder cancer tissues and normal tissues. Then, the expression vector of miR-119a lentivirus was designed and successfully transfected into T24 and 5637 cells. CCK-8, cycle, plate cloning, Transwell and cell scratch experiments were performed to detect cell proliferation effect and invasion effect. Results The expression of miR-119a in bladder cancer tissues was significantly higher(11.330±0.106) than that in normal tissues(0.365±0.076), and the difference was significant(P<0.05). But the expression of FHIT was lower(0.596±0.125 vs 1.203±0.212) in bladder cancer tissues. By upregulating the expression miR-119a, the expression of miR-119a in T24 and 5637 cells was increased, which was significantly different from the negative control group(P<0.05). Compared with the control group, the cell OD value and cell colony number were significantly increased in the up-regulation group(P<0.05). After upregulation of miR-119a, bladder cancer cells were significantly increased(T24: 64.325±5.214 vs 55.315±4.215)(5637: 53.103±3.124 vs 46.324±4.452), with statistically significant differences(P<0.05). Compared with the miR-119a upregulation group, the number of Transwell cells and the crawl rate were decreased, and there was a significant difference in the number of cell migration between the upregulation group and the negative control group(P<0.05). Conclusion The expression level of miR-119a was significantly increased in bladder cancer tissues. Up-regulation miR-119a might be related the inhibition of FHIT expression, and promote proliferation and migration effect on bladder cancer cells.