miR-203 靶向调节 PEG3/NF-κB 信号通路对糖尿病肾病大鼠肾纤维化的 影响.

nscription factor-κB (NF-κB) signaling pathway, and its effect on renal fibrosis in diabetic nephropathy (DN) rats. Methods: A rat DN model was established, and the renal tubular epithelial cells (NRK-52E) were cultured with high sugar to simulate an in vitro DN model, qRT-PCR was used to detect the expression level of miR-203, Western blot was used to detect the expressions of PEG3 and NF-κB proteins. Dual luciferase reporter gene technology was used to detect the targeting relationship between miR-203 and PEG3. The miR-203 mimic (miR-203 mimic) and negative control (mimic NC), PEG3 high expression recombinant vector (pcDNA-PEG3) and negative control (pcDNA-NC) were constructed, and injected and transfected into DN rats and DN model cells respectively to set the control group, model group, miR-203 mimic group, mimic NC group, miR-203 mimic+pcDNA-PEG3 group, and miR-203 mimic+pcDNA-NC group. The 24 h urine protein, blood sugar, serum creatinine (SCr) and urea nitrogen (BUN) in rats were detected, electron microscopy and Masson staining were used to detect renal tissue lesions and fibrosis changes; ELISA was used to detect the contents of IL-6, IL-1β, Collagen Ⅰand TGF-β1 in cells； Western blot was used to detect the protein expressions of IL-6, IL-1β, Collagen Ⅰ，α-SMA and TGF-β1 in kidney tissue. Results: The expressions of miR-203 in kidney tissue and renal tubular epithelial cells of DN rats were decreased (P<0.05), the expression of PEG3/NF-κB was increased in kidney tissue and renal tubular epithelial cells of DN rats (P<0.05). There was a targeted binding site between miR-203 and PEG3. Up-regulating the expression of miR-203 could inhibit the activation of PEG3/NF-κB in DN rats and cell models, reduce the expression of inflammation-fibrosis related factors, and alleviate the pathological symptoms of DN such as renal tissue structural damage and decreased renal function in rats (P<0.05). pcDNA-PEG3 could partially reverse the above effects of miR-203(P<0.05). Conclusion: miR-203 can target to negatively regulate the PEG3/NF-κB pathway and up-regulate the expression of miR-203, which can target to inhibit the activation of the PEG3/NF-κB pathway and play the role of anti-inflammatory-fibrosis in the kidney of DN rats. [ABSTRACT FROM AUTHOR]

目的:探究微小RNA-203(miR-203)对父本表达基因3(PEG3)/核转录因子-κB(NF-κB)信号通路的靶向调控作用及其对糖尿病肾病(DN)大鼠肾纤维化的影响.方法:建立大鼠DN模型,高糖培养大鼠肾小管上皮细胞(NRK-52E)模拟体外DN模型,qRT-PCR检测miR-203表达水平,Western blot检测PEG3、NF-κB蛋白表达.双荧光素酶报告基因技术检测miR-203与PEG3的靶向关系.构建miR-203模拟物(miR-203 mimic)及阴性对照(mimic NC),PEG3高表达重组载体(pcDNA-PEG3)及阴性对照(pcDNA-NC),分别注射及转染至DN大鼠及DN模型细胞中,设置为:对照组、模型组、miR-203 mimic组、mimic NC组、miR-203 mimic+pcDNA-PEG3组、miR-203 mimic+pcDNA-NC组.检测大鼠24 h尿蛋白、血糖、血清肌酸酐(SCr)和尿素氮(BUN),电镜及Masson染色检测肾组织病变及纤维化变化;ELISA检测细胞中IL-6、IL-1β、Ⅰ型胶原(Collagen Ⅰ)、TGF-β1含量;Western blot检测肾组织中IL-6、IL-1β、Collagen Ⅰ、α-SMA、TGF-β1蛋白表达.结果:miR-203在DN大鼠肾组织及肾小管上皮细胞中表达降低(P＜0.05),PEG3/NF-κB在DN大鼠肾组织及肾小管上皮细胞中表达升高(P＜0.05).miR-203与PEG3之间有靶向结合位点.上调miR-203表达,可抑制DN大鼠及细胞模型PEG3/NF-κB激活,降低炎症-纤维化相关因子表达,缓解大鼠肾组织结构损伤、肾功能下降等DN病理症状(P＜0.05).pcDNA-PEG3可部分逆转miR-203的上述作用(P＜0.05).结论:miR-203可靶向负调控PEG3/NF-κB通路,上调miR-203表达,可靶向抑制PEG3/NF-κB通路活化,发挥抗DN肾脏炎症-纤维化病变. [ABSTRACT FROM AUTHOR]