Role of MiR-215 in Hirschsprung's Disease Pathogenesis by Targeting SIGLEC-8
Role of MiR-215 in Hirschsprung's Disease Pathogenesis by Targeting SIGLEC-8
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通过靶向 SIGLEC-8，miR-215 在先天性巨结肠发病机制中的作用。
DOI：
10.1159/000453214
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发表时间：
2016-01-01
影响因子：
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通讯作者：
Tang, Weibing
中科院分区：
文献类型：
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作者：
Lei, Hao;Li, Hongxing;Tang, Weibing
背景/目标：先天性巨结肠症（Hirschsprung's disease，HSCR）是一种罕见的先天性肠道运动障碍，以肠神经元缺失为特征。在本研究中，我们重点关注内含子miR-215及其宿主基因异亮氨酰-tRNA合成酶2（IARS 2）在HSCR发病机制中的作用。方法：采用真实的时间定量PCR和Western blot方法检测miRNA、mRNA和蛋白质水平。双荧光素酶报告基因测定证实了细胞系中特异性mRNA和miRNA的直接调控。使用Transwell测定、CCK 8测定和流式细胞术来测量人293 T和SH-SY 5 Y细胞的细胞功能。结果如下：HSCR患者结肠组织中miR-215的表达水平显著低于对照，这与宿主基因IARS 2的表达正相关，与预测的靶基因：唾液酸结合Ig样凝集素8（SIGLEC-8）负相关。miR-215的缺失抑制细胞迁移和增殖，这与IARS 2的减少一致。双荧光素酶报告基因测定证实miR-215通过直接结合SIGLEC-8的3 '-UTR导致SIGLEC-8的抑制。此外，敲低SIGLEC-8挽救了由miR-215减少引起的细胞迁移和增殖抑制的程度。结论：我们的研究结果表明，miR-215与宿主基因IARS 2协同作用，通过靶基因SIGLEC-8影响神经元迁移和增殖。我们推断IARS 2-miR-215-SIGLEC-8通路可能在HSCR的发病机制中起关键作用。(C)2016作者（s）由S. Karger AG，巴塞尔
Background/Aims: Hirschsprung's disease (HSCR), known as aganglionosis, is an infrequent congenital gut motility disorder characterized by absence of enteric neurons. In this study, we focus on the role of the intronic miR-215 and its host gene isoleucyl-tRNA synthetase 2 (IARS2) in the pathogenesis of HSCR. Methods: Quantitative real time PCR and Western blot were used to detect the miRNA, mRNAs, and proteins levels. The dual-luciferase reporter gene assay confirmed the direct regulation of the specific mRNA and miRNAs in cell lines. Transwell assays, CCK8 assay, and flow cytometry were used to measure cell function of the human 293T and SH-SY5Y cells. Results: Expression levels of miR-215 in HSCR patient colon tissues were outstandingly lower than controls, which was positively correlated with expression of the host gene IARS2 and negatively correlated with predicted target gene: sialic acid binding Ig-like lectin 8 (SIGLEC-8) The loss of miR-215 inhibited cell migration and proliferation, which was consistent with the reduction of IARS2. The dual-luciferase reporter gene assay confirmed that miR-215 resulted in the inhibition of SIGLEC-8 by directly binding to the 3'-UTR of SIGLEC-8. Moreover, knocking-down SIGLEC-8 rescued the extent of suppressed cell migration and proliferation that resulted from the diminishment of miR-215. Conclusions: Our findings indicate that miR-215 acts in concert with the host gene IARS2 to affect neuron migration and proliferation through the target gene SIGLEC-8. We infer that the IARS2-miR-215-SIGLEC-8 pathway may play a crucial role in the pathogenesis of HSCR. (C) 2016 The Author(s) Published by S. Karger AG, Basel