Wang Gang, Guo Feng, Wang Hefeng, Liu Wei, Zhang Lijuan, Cui Mingyu, Wu Xiangyu
Department of Pediatric Surgery, Shandong Province Hospital Affiliated to Shandong University , Jinan, China .
DNA Cell Biol. 2017 Nov;36(11):930-937. doi: 10.1089/dna.2017.3821. Epub 2017 Nov 1.
Recent studies have suggested the critical roles of miRNAs for disease progression. miRNA-483-5p (miR-483-5p) was previously found to have a relationship with tumor cell behavior, but its biological function in Hirschsprung's disease (HSCR) remains undefined. Thus, we explored the role of miR-483-5p in the pathogenesis of HSCR. Histological changes of colonic tissues were evaluated by hematoxylin and eosin (HE) staining. Quantitative real-time PCR and western blotting were used to determine relative expression levels of miRNA, mRNA, and proteins in 20 HSCR patients and 20 normal colon tissues. In this study, we found that miR-483-5p expression in HSCR tissues was significantly increased and their downregulation promoted cell proliferation, cell cycle progression and invasion and inhibited cell apoptosis in human 293T and SH-SY5Y cell lines by the CCK-8, flow cytometry, and Transwell assay. GNDF family receptor alpha 4 (GFRA4) was confirmed as a downstream target of miR-483-5p by dual-luciferase reporter gene assay and inversely correlated with miR-483-5p expression in cell lines. Taken together, miR-483-5p may play a crucial role in the pathogenesis of HSCR by targeting GFRA4.
最近的研究表明，微小RNA（miRNA）在疾病进展中起着关键作用。先前发现miRNA-483-5p（miR-483-5p）与肿瘤细胞行为有关，但其在先天性巨结肠症（HSCR）中的生物学功能仍不明确。因此，我们探讨了miR-483-5p在HSCR发病机制中的作用。通过苏木精和伊红（HE）染色评估结肠组织的组织学变化。采用定量实时聚合酶链反应（qRT-PCR）和蛋白质免疫印迹法检测20例HSCR患者和20例正常结肠组织中miRNA、mRNA和蛋白质的相对表达水平。在本研究中，我们发现HSCR组织中miR-483-5p表达显著增加，通过细胞计数试剂盒-8（CCK-8）、流式细胞术和Transwell实验发现，其下调可促进人293T和SH-SY5Y细胞系的细胞增殖、细胞周期进程和侵袭，并抑制细胞凋亡。通过双荧光素酶报告基因实验证实胶质细胞源性神经营养因子家族受体α4（GFRA4）是miR-483-5p的下游靶点，且在细胞系中与miR-483-5p表达呈负相关。综上所述，miR-483-5p可能通过靶向GFRA4在HSCR发病机制中起关键作用。
