miRNA表达谱在肾纤维化中的差异表达及抗纤灵的干预研究
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摘要
目的筛选观察在阿霉素肾病大鼠模型特异性miRNA表达谱,观察抗纤灵方干预的作用。方法将SD雄性大鼠随机分为6组,每组8只。其中假手术组仅于右侧肾切除,灌于生理盐水,余下40只SD大鼠均采取右侧肾切除加尾静脉注射阿霉素制备大鼠硬化模型。将造模成功后的大鼠分成模型组、科素亚组、抗纤灵低,中,高5组,分别给予相对应的药物共8周,观察各组大鼠肾脏组织TGF-β、Smad3 mRNA水平改变。同时对各组实验肾脏标本,运用miRNA高通量测序,检测miRNA表达谱,分析差异表达谱,观察抗纤灵方干预的作用。结果抗纤灵方对阿霉素肾病模型大鼠能降低TGF-β、Smad3 mRNA水平,差异有统计学意义(P <0.05)。随着抗纤灵剂量增加,其抗纤维化效应更明显,差异有统计学意义(P <0.05)。通过筛选miRNA表达谱,差异表达分析中发现假手术组与模型组表达谱差别最大,假手术组与抗纤灵高剂量治疗组差别最小。从差异表达分析中,观察到了药物处理剂量越大,样本的表达谱越与假手术组相近。结论抗纤灵方能改善肾纤维化,并且随着剂量增大抗纤维化的效果越好,推测其机制可能通过调节miRNA谱的差异表达,降低TGF-β、Smad3 mRNA水平,来改善肾纤维化。
Objective To observe the mechanism of Kangxianling Decoction and screen the specific miRNA expression profile on renal fibrosis of adriamycin-induced nephropathic rats. Methods SD male rats were randomly divided into 6 groups with 8 rats in each group. The sham operation group was treated with normalsaline only after right nephrectomy. The other 40 SD rats were treated with right nephrectomy and intravenous injection of adriamycin. The operation group was randomly divided into the model group, the Kangxianling Decoction low, medium and high dose group and the Losartan Potassium group. Corresponding drugs administered to rats in all groups for 8 weeks. After the treatment, the changes of TGF-β and Smad3 mRNA levels in kidney tissues of model rats were observed. For each group of experimental kidney specimens, miRNA high-throughput sequencing was used to detect miRNA expression profiles, analyze differential expression profiles, and observe the effects of anti-fibrosis intervention. Results Compared with the model group, Kangxianling Decoction could significantly lower the expression of TGF-β and Smad3(P < 0.05). By increasing of the dose of Kangxianling Decoction, the anti-fibrosis effect was more obvious, and the difference was significant(P < 0.05). By screening miRNA expression profiles, the analysis showed that the difference between the sham operation group and the model group was the largest, and the difference between the sham operation group and the Kangxianling Decoction high-dose treatment group was the smallest. So it was observed that the larger the drug treatment dose, the more the expression profile of the sample was similar to the sham operation group. Conclusion Kangxianling Decoction can improve renal fibrosis, and the better the anti-fibrosis effect with increasing dose. It is speculated that it may regulate differential expression of miRNA profiles and decreasing TGF-β and Smad3 m RNA levels to protect the kidney.
Objective To observe the mechanism of Kangxianling Decoction and screen the specific miRNA expression profile on renal fibrosis of adriamycin-induced nephropathic rats. Methods SD male rats were randomly divided into 6 groups with 8 rats in each group. The sham operation group was treated with normalsaline only after right nephrectomy. The other 40 SD rats were treated with right nephrectomy and intravenous injection of adriamycin. The operation group was randomly divided into the model group, the Kangxianling Decoction low, medium and high dose group and the Losartan Potassium group. Corresponding drugs administered to rats in all groups for 8 weeks. After the treatment, the changes of TGF-β and Smad3 mRNA levels in kidney tissues of model rats were observed. For each group of experimental kidney specimens, miRNA high-throughput sequencing was used to detect miRNA expression profiles, analyze differential expression profiles, and observe the effects of anti-fibrosis intervention. Results Compared with the model group, Kangxianling Decoction could significantly lower the expression of TGF-β and Smad3(P < 0.05). By increasing of the dose of Kangxianling Decoction, the anti-fibrosis effect was more obvious, and the difference was significant(P < 0.05). By screening miRNA expression profiles, the analysis showed that the difference between the sham operation group and the model group was the largest, and the difference between the sham operation group and the Kangxianling Decoction high-dose treatment group was the smallest. So it was observed that the larger the drug treatment dose, the more the expression profile of the sample was similar to the sham operation group. Conclusion Kangxianling Decoction can improve renal fibrosis, and the better the anti-fibrosis effect with increasing dose. It is speculated that it may regulate differential expression of miRNA profiles and decreasing TGF-β and Smad3 m RNA levels to protect the kidney.
引文
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[8]钟利平,麻志恒,余柯娜,等.抗纤灵方对5/6肾切除小鼠肾组织纤维化作用机制研究[J].中国实验方剂学杂志,2016,22(2):118-121.
[9]张长明,周家俊,何立群,等.抗纤灵方对CKD 3~4期患者疗效及机制的临床多中心随机对照研究[J].辽宁中医杂志,2013,40(1):122-124.
[2]吴旭,林凡,黄朝兴,等.大鼠腹膜纤维化相关miRNA的初步筛选[J].中国中西医结合肾病杂志,2013,14(3):203-206.
[3]CHUNG A C,LAN H Y.MicroRNAs in renal fibrosis[J].Frontiers in Physiology,2015(6):50.
[4]MENG X M,NIKOLICPATERSON D J,LAN H Y.TGF-β:the master regulator of fibrosis[J].Nature Reviews Nephrology,2016,12(6):325-338.
[5]WANG B,JHA J C,HAGIWARA S,et al.Transforming growth factor-β1-mediated renal fibrosis is dependent on the regulation of transforming growth factor receptor 1 expression by let-7b[J].Kidney Int,2014,85(2):352-361.
[6]LOPEZHERNANDEZ,F J,LOPEZNOVOA,J M.Role of TGF-beta in chronic kidney disease:an integration of tubular,glomerular and vascular effects[J].Cell Tissue Res,2012(347):141-154.
[7]DA C,LIU Y,ZHAN Y,et al.Nobiletin inhibits epithelialmesenchymal transition of human non-small cell lung cancer cells by antagonizing the TGF-β1/Smad3 signaling pathway[J].Oncol Rep,2016,35(5):2767-2774.
[8]钟利平,麻志恒,余柯娜,等.抗纤灵方对5/6肾切除小鼠肾组织纤维化作用机制研究[J].中国实验方剂学杂志,2016,22(2):118-121.
[9]张长明,周家俊,何立群,等.抗纤灵方对CKD 3~4期患者疗效及机制的临床多中心随机对照研究[J].辽宁中医杂志,2013,40(1):122-124.