MiR-155和MiR-21协同靶向SOCS6促进肺癌发生、发展的机制研究

发布时间：2019-07-08 15:25

【摘要】：研究背景:肺癌是所有癌症中发病率和死亡率最高的癌症。miR-155和miR-21在肺癌的发生、发展过程中起着重要的作用,然而miR-155和miR-21是如何促进肺癌的发生、发展机制仍不明确。研究目的:探讨miR-155和miR-21通过直接靶向SOCS6促进肺癌发生、发展;协同miR-155/21对肺癌发生、发展的促进作用优于单一 miR-155或miR-21对肺癌的促进作用,从而为临床找到一种可能更加有效的治疗肺癌的手段。材料和方法:(1)临床标本中首先检测了 80对肺癌患者癌与癌旁组织的miR-155、miR-21以及SOCS6的表达情况,并分析之间的相互关系;(2)体内实验选取细胞系A549和H1299作为肺癌的细胞株,分别在A549和H1299细胞系中抑制或过表达miR-155、miR-21以及miR-155/21,检测各组中基因SOCS6的mRNA及蛋白的表达情况;观察各组肿瘤细胞的增殖、活性、凋亡情况;(3)体外实验将转染miR-155、miR-21和miR-155/21的抑制剂A549细胞注射到裸鼠背部皮下,观察各组裸鼠后背肿瘤的成瘤以及基因SOCS6在肿瘤组织中的表达情况。结果:(1)临床标本实验,检测了 80对肺癌患者癌与癌旁组织,其中癌组织miR-155和miR-21的表达明显高于癌旁组织,同时SOCS6的表达明显低于癌旁组织,miR-155和miR-21与SOCS6的表达成反向关系;随着肿瘤分期进展,miR-155和miR-21的表达逐渐升高,而SOCS6的表达逐渐减低。(2)体外实验,促进或抑制miR-155、miR-21和miR-155/miR-21表达,均能抑制或促进SOCS6 mRNA及蛋白的表达;但协同促进或抑制miR-155/miR-21时,对SOCS6的抑制或促进效果优于单一miR-155或miR-21对SOCS6的作用效果;当过表达野生型miR-155、miR-21时,SOCS6-3'-UTR荧光活性明显减低,过表达突变型miR-155、miR-21,SOCS6-3'-UTR 荧光活性未见变化,证实 miR-155、miR-21 是通过直接靶向SOCS6的3'-UTR而降低其表达的。(3)体外实验,抑制miR-155、miR-21和miR-155/miR-21可以促进A549和H1299细胞的凋亡,同时也抑制了 A549和H1299细胞的增殖比率(细胞数目、活性、克隆数),但协同抑制miR-155/miR-21的效果无论在促进A549和H1299细胞的凋亡,还是在抑制A549和H1299增殖比率均优于单一抑制miR-155或miR-21的效果。(4)体内实验,裸鼠后背注射miR-155/21抑制剂组的肿瘤大小明显低于单纯注射miR-155或miR-21抑制剂组;免疫组化示:miR-155/21抑制剂组的SOCS6表达明显高于单一 miR-155或miR-21抑制剂组SOCS6的表达。结论:(1)临床患者标本、体外和体内实验证实,miR-155和miR-21是促进肺癌发生、发展的关键因子;(2) miR-155和miR-21是通过直接靶向SOCS6促进肺癌发生、发展;(3)协同抑制miR-155/miR-21对肺癌的抑制效果优于单一 miR-155或miR-21对肺癌的抑制效果。
文内图片：

图片说明：图２邋：邋ｍｉＲ－１５５、ｍｉＲ－２１和ｍｉＲ－２１／１５５的调控网络分析：（Ａ）邋ｍｉＲ－２１的调控网络；逡逑
[Abstract]:Background: lung cancer has the highest incidence and mortality among all cancers. MIR-155 and miR-21 play an important role in the occurrence and development of lung cancer. However, how miR-155 and miR-21 promote the occurrence of lung cancer, the development mechanism is still unclear. Objective: to investigate the effect of miR-155 and miR-21 on the occurrence and development of lung cancer by directly targeting SOCS6, and the promoting effect of miR-155/21 on the occurrence and development of lung cancer is better than that of single miR-155 or miR-21 on lung cancer, so as to find a more effective method for clinical treatment of lung cancer. Materials and methods: (1) the expression of miR-155,miR-21 and SOCS6 in 80 pairs of lung cancer and paracancerous tissues were detected in clinical specimens, and the relationship between them was analyzed. (2) in vivo, cell lines A549 and H1299 were selected as lung cancer cell lines to inhibit or overexpress miR-155,miR-21 in A549 and H1299 cell lines, and miR-155/21, was used to detect the expression of SOCS6 mRNA and protein in each group, and the proliferation, activity and apoptosis of tumor cells in each group were observed. (3) miR-155,miR-21 and miR-155/21 inhibitor A549 cells were injected subcutaneously into the back of nude mice in vitro to observe the tumorigenesis of tumor and the expression of gene SOCS6 in tumor tissue of nude mice in each group. Results: (1) 80 pairs of lung cancer patients with cancer and paracancerous tissues were detected. The expression of miR-155 and miR-21 in cancer tissues was significantly higher than that in paracancerous tissues, and the expression of SOCS6 was significantly lower than that in paracancerous tissues. MiR-155 and miR-21 were inversely related to SOCS6. With the progress of tumor stage, the expression of miR-155 and miR-21 increased gradually, while the expression of SOCS6 decreased gradually. (2) promoting or inhibiting the expression of miR-155,miR-21 and miR-155/miR-21 in vitro could inhibit or promote the expression of SOCS6 mRNA and protein, but by promoting or inhibiting miR-155/miR- 21:00, the inhibitory or promoting effect of miR-155/miR- on SOCS6 was better than that of miR-155 or miR-21 on SOCS6. When the wild type miR-155,miR- was overexpressed at 21:00, the fluorescence activity of SOCS6-3'-UTR decreased significantly, but the fluorescence activity of overexpression mutant miR-155,miR-21,SOCS6-3'-UTR did not change. It was confirmed that miR-155,miR-21 decreased the expression of miR-155,miR-21 by directly targeting the 3'-UTR of SOCS6. (3) inhibition of miR-155,miR-21 and miR-155/miR-21 could promote the apoptosis of A549 and H1299 cells in vitro. At the same time, it also inhibited the proliferation ratio (cell number, activity, clone number) of A549 and H1299 cells, but the synergistic inhibitory effect of miR-155/miR-21 was better than that of miR-155 or miR-21 alone in promoting the apoptosis of A549 and H1299 cells and inhibiting the proliferation ratio of A549 and H1299 cells. (4) in vivo experiment, The tumor size of miR-155/21 inhibitor group was significantly lower than that of miR-155 or miR-21 inhibitor group. The expression of SOCS6 in miR-155/21 inhibitor group was significantly higher than that in single miR-155 or miR-21 inhibitor group. Conclusion: (1) in vitro and in vivo experiments, miR-155 and miR-21 are the key factors to promote the occurrence and development of lung cancer; (2) miR-155 and miR-21 promote the occurrence and development of lung cancer through direct targeting SOCS6; (3) the inhibitory effect of synergistic inhibition of miR-155/miR-21 on lung cancer is better than that of single miR-155 or miR-21 on lung cancer.
【学位授予单位】：中国人民解放军医学院
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R734.2

【相似文献】