目的 研究miR-199b-5p在新肾病1号方抗肾纤维化中的作用。方法 36只大鼠随机分为假手术组、模型组及新肾病1号方低、中、高剂量（9.69、19.38、38.76 g/kg）组和氯沙坦（50 mg/kg）组，每组6只。除假手术组外，其余各组采用左侧单侧输尿管梗阻方法建立大鼠肾纤维化模型，给予药物干预14 d后，检测各组大鼠肾功能；苏木素-伊红（HE）和Masson染色观察肾脏组织病理变化；对假手术组、模型组和新肾病1号方高剂量组大鼠肾脏组织进行miRNA测序，筛选差异表达miRNAs，qRT-PCR验证，并进行靶基因的基因本体（gene ontology，GO）功能及京都基因与基因组百科全书（Kyoto encyclopedia of genes and genomes，KEGG）通路富集分析，生物信息学分析miRNAs与上皮间质转化（epithelial-mesenchymal transition，EMT）、肾纤维化的关联性。免疫荧光法和Western blotting检测肾组织EMT相关蛋白表达情况。结果 与模型组比较，新肾病1号方组大鼠血清肌酐（serum creatinine，SCr）和血清尿素氮（blood urea nitrogen，BUN）水平均明显降低（P＜0.05、0.01），肾组织病理变化有所改善。miRNA测序分析发现，miR-199b-5p在模型组表达上调，而在新肾病1号方高剂量组表达下调，qRT-PCR验证了其mRNA相对表达量与测序结果基本一致；miR-199b-5p的靶基因中与EMT、肾纤维化相关功能的90个。与模型组比较，新肾病1号方组肾组织中EMT相关蛋白α-平滑肌肌动蛋白（α-smooth muscle actin，α-SMA）和波形蛋白（Vimentin）表达下调（P＜0.05、0.001），E-钙黏蛋白（E-cadherin）和紧密连接蛋白-1（Zonula occludens-1，ZO-1）表达上调（P＜0.05）。结论 新肾病1号方可能通过下调miR-199b-5p表达，抑制EMT，从而发挥抗肾纤维化的作用。

Objective To investigate the role of miR-199b-5p in the treatment of anti-renal fibrosis of Improved-Nephropathy Formula 1 (新肾病1号方, N1F). Methods A total of 36 rats were randomly divided into sham group, model group, N1F low-, medium- and high-dose (9.69, 19.38, 38.76 g/kg) groups and losartan (50 mg/kg) group, with six rats in each group. Except the sham group, renal fibrosis model was established by left unilateral ureteral obstruction method in the other groups. After 14 d of drug intervention, renal function was detected in each group of rats. The pathological changes of renal tissue were observed by hematoxylin-eosin (HE) and Masson staining. Renal tissues of rats in sham group, model group and N1F high-dose group were sequenced by miRNA, screened for differentially expressed miRNAs and verified by qRT-PCR, gene ontology (GO) function and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis of target genes were performed. The correlation between miRNAs and epithelial-mesenchymal transition (EMT) and renal fibrosis was analyzed by bioinformatics. The expressions of EMT-related proteins in renal tissue were detected by immunofluorescence and Western blotting. Results Compared with model group, serum creatinine (SCr) and blood urea nitrogen (BUN) levels were significantly decreased (P < 0.05, 0.01), the pathological changes of renal tissue were improved in N1F groups. miRNA sequencing analysis showed that the expression of miR-199b-5p was up-regulated in model group, but down-regulated in N1F high-dose group. qRT-PCR verified that the relative mRNA expression level was basically consistent with the sequencing results. A total of 90 target genes in miR-199b-5p were related to EMT and renal fibrosis. Compared with model group, the expressions of EMT-related proteins α-smooth muscle actin (α-SMA) and Vimentin in renal tissue were down-regulated (P < 0.05, 0.001), but the expressions of E-cadherin and Zonula occludens-1 (ZO-1) were up-regulated in N1F groups (P < 0.05). Conclusion N1F may inhibit EMT by down-regulating the expression of miR-199b-5p, and play an anti-renal fibrosis role.

浙江省科学技术厅研发攻关计划立项项目（2022C03160）；全国名老中医药专家学术传承工作室项目（国中医药办人教函[2021]270号）