miR-155-5p mimic靶向HIF-1α对非小细胞肺癌A549细胞生长和运动能力的调节作用和机制

The regulatory effect and mechanism of miR-155-5p mimic targeting HIF-1α on the growth and motility of non-small cell lung cancer A549 cells

目的探讨miR-155-5p mimic靶向缺氧诱导因子-1α(HIF-1α)对非小细胞肺癌A549细胞生长和运动能力的调节作用和机制。方法取对数期生长的A549细胞,将未经转染的细胞设为Control组,miR-155 mimic及HIF-1α分别转染至A549细胞,设为miR-155 mimic组和HIF-1α组;再将两者共同转染于A549细胞,设为mimic+HIF-1α组。采用荧光素酶报告基因检测miR-375与SLC7A11的靶向关系,利用RT-PCR检测各组细胞miR-155-5p及HIF-1αmRNA表达水平,Edu染色检测细胞增殖,划痕实验检测细胞迁移,Transwell小室实验检测细胞侵袭,Western blot检测细胞中各蛋白表达。结果 HIF-1α是miR-155-5p mimic的靶向基因。与Control组相比,miR-155 mimic组HIF-1αmRNA水平、HIF-1α和Ki67蛋白表达量及增殖细胞数、侵袭细胞数及划痕愈合率均降低(P<0.05),HIF-1α组HIF-1αmRNA水平、HIF-1α和Ki67蛋白表达量及增殖细胞数、侵袭细胞数及划痕愈合率均升高(P<0.05)。与HIF-1α组相比,mimic+HIF-1α组HIF-1αmRNA水平、HIF-1α和Ki67蛋白表达量及增殖细胞数、侵袭细胞数及划痕愈合率均降低(P<0.05)。与Control组相比,miR-155 mimic组N-cadherin、VEGF、VEGFR2、P38蛋白表达量降低,E-cadherin蛋白表达量升高(P<0.05),HIF-1α组N-cadherin、VEGF、VEGFR2、P38蛋白表达量升高,E-cadherin蛋白表达量降低(P<0.05)。与HIF-1α组相比,mimic+HIF-1α组N-cadherin、VEGF、VEGFR2、P38蛋白表达量降低,E-cadherin蛋白表达量升高(P<0.05)。结论 miR-155-5p可通过靶向抑制HIF-1α表达抑制非小细胞肺癌A549细胞增殖、侵袭及迁移,其作用机制可能与抑制上皮间质转化及VEGF/P38通路相关。

Objective To investigate the regulatory effect and mechanism of miR-155-5 p mimic targeting hypoxia inducible factor 1α(HIF-1α) on the growth and motility of non-small cell lung cancer A549 cells. Methods The untransfected A549 cells growing in logarithmic phase ells were set as control group. miR-155 mimic and HIF-1α were transfected into A549 cells respectively, which were divided into Mir-155 mimic group and HIF-1α group. Then they were co transfected into A549 cells and set up as mimic+HIF-1α group. The targeting relationship between Mir 375 and SLC7 A11 was detected by luciferase reporter gene. The mRNA expression levels of Mir-155-5 p and HIF-1α were detected by RT PCR. The cell proliferation was detected by Edu staining. The cell migration was detected by scratch test. The cell invasion was detected by Transwell chamber test. The protein expression was detected by Western blot. Results HIF-1α is the target gene of Mir-155-5 p mimic. Compared with control group, HIF-1α mRNA level, HIF-1α and Ki67 protein expression, proliferative cell number, invasive cell number and scratch healing rate were all decreased in miR-155 group(P<0.05), while HIF-1 α mRNA level, HIF-1α and Ki67 protein expression, proliferative cell number, invasive cell number and scratch healing rate were all increased in HIF-1α group(P<0.05). Compared with HIF-1α group, HIF-1α mRNA level, HIF-1α and Ki67 protein expression, proliferative cell number, invasive cell number and scratch healing rate were all decreased in mic+HIF-1α group(P<0.05). Compared with the control group, the expression of N cadherin, VEGF, VEGFR2, p38 protein in miR-155-mic group decreased, E cadherin protein increased(P<0.05), N cadherin, VEGF, VEGFR2, p38 protein in HIF 1 α group increased, E cadherin protein decreased(P<0.05). Compared with HIF-1α group, the expression of N cadherin, VEGF, VEGFR2 and p38 protein in mic+HIF-1α group decreased, while the expression of E cadherin protein increased(P<0.05). Conclusion miR-155-5 p can inhibit the proliferation, invasion and migration of non-small cell lung cancer A549 cells by targeting the expression of HIF-1α. Its mechanism may be related to the inhibition of epithelial stromal transformation and VEGF/p38 pathway.