先天性巨结肠患儿结肠组织中miR-145-5p异常表达与GFRα1的关系

来源 :中华新生儿科杂志 | 被引量 : 0次 | 上传用户:zhouqidhi
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目的:探讨miR-145-5p对胶质细胞源性神经营养因子受体(glial cell-derived neurotrophic factor receptor,GFR)α1表达的调控及其与先天性巨结肠(Hirschsprung's disease,HSCR)的关系。方法:选择2016年3月至2018年6月湖南省儿童医院新生儿外科收治的HSCR患儿巨结肠根治术后痉挛段结肠组织为HSCR组,同期新生儿坏死性小肠结肠炎(necrotizing enterocolitis,NEC)患儿手术切除坏死肠段两端正常结肠组织为NEC组作为对照,进行前瞻性研究。采用实时定量聚合酶链反应检测两组结肠组织内miR-145-5p及GFRα1表达水平,荧光素酶活性实验检测GFRα1与miR-145-5p的关系;采用实时定量聚合酶链反应及蛋白免疫印迹检测SH-SY5Y细胞转染miR-145-5p模拟物后miR-145-5p及GFRα1表达水平,CCK8法检测转染后SH-SY5Y细胞活力。结果:HSCR组24例,NEC组18例。HSCR组miR-145-5p mRNA水平明显高于NEC组[(5.62±2.04)比(1.11±0.48)],GFRα1 mRNA水平明显低于NEC组[(0.39±0.18)比(1.00±0.37)],差异有统计学意义(n P<0.01),且miR-145-5p表达水平与GFRα1表达水平成负相关(n r=-0.676,n P<0.01)。荧光素酶活性实验证实GFRα1为miR-145-5p的靶基因。SH-SY5Y细胞转染miR-145-5p模拟物后GFRα1的表达水平明显降低(n P<0.01),SH-SY5Y细胞活力下降。n 结论:GFRα1是miR-145-5p的靶基因,HSCR患儿病变结肠组织中GFRα1低表达可能与miR-145-5p高表达有关。“,”Objective:To study the regulation of miR-145-5p on glial cell-derived neurotrophic factor receptor(GFR)α1 expression in Hirschsprung's disease (HSCR).Method:From March 2016 to June 2018, colon tissue resected from HSCR patients who received radical operation in the neonatal surgery department of our hospital were reviewed as the HSCR group. During the same period, normal colon tissue at both ends of the necrotic intestinal segment from patients with necrotizing enterocolitis (NEC) were used as control (NEC group). Real-time PCR was used to detect the expression levels of miR-145-5p and GFRα1 in colon tissues of both groups. The modulation effects of miR-145-5p on GFRα1 was detected using luciferase activity assay. Real-time PCR and Western blotting were used to detect the expression levels of miR-145-5p and GFRα1 in miR-145-5p-mimic-transfected SH-SY5Y cells. CCK8 assay kit was used to examine the cell viability of miR-145-5p-mimic-transfected SH-SY5Y cells.Result:A total of 24 cases were in the HSCR group and 18 cases in the NEC group. Compared with the NEC group, the miR-145-5p expression level in HSCR group was significantly higher (5.62±2.04 vs. 1.11±0.48) and the mRNA level of GFRα1 was significantly lower (0.39±0.18 vs. 1.00±0.37) (n P<0.01). The expression level of miR-145-5p was negatively correlated with the mRNA level of GFRα1 (n r=-0.676,n P<0.01). Luciferase activity assay confirmed that GFRα1 was the target gene of miR-145-5p. After transfected with miR-145-5p mimic, the expression level of GFRα1 in SH-SY5Y cells was significantly decreased (n P<0.01) and the cell viability was also decreased.n Conclusion:GFRα1 is the target gene of miR-145-5p.Insufficient expression of GFRα1 in diseased colon tissue of HSCR patients may be related to high level of miR-145-5p.
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