环状RNA-EIF3I对肝细胞癌生长转移的影响及机制研究

Effect and mechanism of circ-EIF3I on the growth and metastasis of hepatocellular carcinoma

目的探讨环状RNA-EIF3I(circ-EIF3I)对肝细胞癌(HCC)生长转移的影响及其可能作用机制。方法选取2014年5月至2015年10月河北医科大学第四医院收治的39例HCC患者为研究对象,其中男性29例,女性10例,年龄(62.2±5.6)岁。术中获取部分HCC组织与癌旁组织,用荧光定量聚合酶链反应或蛋白质印迹法分别检测二者circ-EIF3I和磷酸甘油酸激酶1(PGK1)的表达量;利用小分子RNA干扰和基因过表达实验调节其在Hep3B、Huh-7肝癌细胞中的表达,然后以噻唑兰细胞活力实验检测细胞增殖能力,Transwell实验检测细胞迁移及侵袭能力,最后以双荧光素酶报告实验检测靶向关系。结果与癌旁组织比较,HCC组织中circ-EIF3I[(4.32±0.62)比(1.24±0.59)]和PGK1[(2.69±0.19)比(1.00±0.07)]的相对表达量升高,差异有统计学意义(均P<0.05)。与阴性对照组比较,circ-EIF3I小分子干扰RNA组细胞活力降低[Hep3B:(55.3±7.5)%比(100.0±9.2)%;Huh-7:(42.7±6.0)%比(100.0±5.6)%],迁移细胞数减少[Hep3B:(71.0±10.0)比(130.0±15.0)个;Huh-7:(50.0±8.5)比(125.0±10.0)个],侵袭细胞数亦减少[Hep3B:(52.0±7.0)比(105.0±13.0)个;Huh-7:(60.0±8.0)比(144.0±11.0)个],差异均有统计学意义(均P<0.05)。双荧光素酶报告实验证实circ-EIF3I可靶向结合miR-149-5p/miR-1271-5p,miR-149-5p/miR-1271-5p可靶向结合PGK1;PGK1过表达可明显逆转敲低circ-EIF3I对肝癌细胞增殖、迁移及侵袭的作用。结论敲低circ-EIF3I可通过调控miR-149-5p/miR-1271-5p/PGK1分子轴从而抑制HCC细胞增殖、迁移及侵袭。

Objective To investigate the function of circ-EIF3I on the growth and metastasis of hepatocellular carcinoma(HCC)and its possible mechanism.Methods A total of 39 HCC patients admitted to the Fourth Hospital of Hebei Medical University from May 2014 to October 2015 were selected as the study subjects,including 29 males and 10 females,aged(62.2±5.6)years old.Part of HCC tissues and adjacent tissues were obtained during the surgical operation.Fluorescence quantitative PCR was used to detect the expression of circ-EIF3I,and Western blotting was used to detect phosphoglycerate kinase 1(PGK1)in HCC and adjacent tissues,respectively.Small molecule RNA interference and gene overexpression experiments were used to adjust its expression in Hep3B and Huh-7 HCC cell lines,and then MTT cell viability test was used to detect cell proliferation ability.Transwell assaya was used to detect cell migration and invasion ability.Finally,the dual luciferase report experiment was used to detect the targeting relationship between circ-EIF3I and miR-149-5p/miR-1271-5p,and the targeting relationship between miR-149-5p/miR-1271-5p and PGK1.Results Compared with adjacent tissues,the relative expression of circ-EIF3I[(4.32±0.62)vs.(1.24±0.59)]and PGK1[(2.69±0.19)vs.(1.00±0.07)]in HCC tissues from 39 cases were increased,and the differences were statistically significant(all P<0.05).Compared with the negative control group,the cell viability of the circ-EIF3I small molecule interfering RNA group was reduced[Hep3B:(55.3±7.5)%vs.(100.0±9.2)%;Huh-7:(42.7±6.0)%vs.(100.0±5.6)%],the number of migrating cells was decreased[Hep3B:(71.0±10.0)vs.(130.0±15.0);Huh-7:(50.0±8.5)vs.(125.0±10.0)],the number of invasive cells also was decreased[Hep3B:(52.0±7.0)vs.(105.0±13.0);Huh-7:(60.0±8.0)vs.(144.0±11.0)],the difference was statistically significant(all P<0.05).The dual luciferase report experiment confirmed that circ-EIF3I could target miR-149-5p/miR-1271-5p,and miR-149-5p/miR-1271-5p can target PGK1.Over expression of PGK1 could significantly reverse the effects of knockdown of circ-EIF3I on the proliferation,migration and invasion of HCC cells.Conclusion Knockdown of circ-EIF3I could inhibit the proliferation,migration,invasion of HCC cells by regulating the miR-149-5p/miR-1271-5p/PGK1 molecular axis.