基于miRNA测序技术探讨黄芪-丹参药对干预自发性高血压大鼠肾损害的机制研究

目的基于微小RNA (microRNA,miRNA)高通量测序技术,探讨黄芪-丹参药对通过调控miRNA改善高血压肾损害的作用机制。方法以9只WKY大鼠作为对照组,将自发性高血压大鼠随机分为模型组和黄芪-丹参药对(3.4 g/kg)组,每组9只;黄芪-丹参药对组给予黄芪-丹参药对进行干预,观察各组大鼠血压及肾组织病理变化,并对各组大鼠肾组织进行miRNA测序。结果经黄芪-丹参药对干预4周后,与模型组比较,大鼠血压显著降低(P0.01),大鼠肾小球分叶不明显,系膜区增生减轻。与对照组相比,模型组共筛选出115个差异表达miRNA,其中68个差异表达miRNA上调、47个差异表达miRNA下调;与模型组相比,黄芪-丹参药对组筛选得到91个差异表达miRNA,其中67个差异表达miRNA上调、24个差异表达miRNA下调。差异表达miRNA的qRT-PCR验证结果显示,各组大鼠肾组织miRNA-142-5p、miRNA-3585-5p、miRNA-219a-5p、miRNA-122-5p和miRNA-125b-1-3p表达与miRNA测序结果趋势一致。差异表达miRNA的靶基因预测及功能富集结果显示,基因本体(gene ontology,GO)功能主要富集于阴离子跨膜转运、突触前、蛋白质丝氨酸/苏氨酸激酶活性等方面;京都基因与基因组百科全书(Kyoto encyclopedia of genes and genomes,KEGG)通路主要富集于哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)、丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)、自噬、单磷酸腺苷活化蛋白激酶(adenosine monophosphate-activated protein kinase,AMPK)信号通路等途径。结论miR-219a-5p、miR-3585-5p和miR-142-5p可能是黄芪-丹参药对延缓高血压肾损害进程的直接靶点。

Mechanism of Astragali Radix and Salviae Miltiorrhizae Radix et Rhizoma herbal pair on renal damage of spontaneously hypertensive rats based on miRNA sequencing technology

Objective To explore the mechanism of Huangqi (Astragali Radix) and Danshen (Salviae Miltiorrhizae Radix et Rhizoma) herbal pair on renal damage of spontaneously hypertensive rats by regulating miRNA based on miRNA high-throughput sequencing. Methods Nine WKY rats were used as control group, spontaneously hypertensive rats were randomly divided into Astragali Radix and Salviae Miltiorrhizae Radix et Rhizoma herbal pair (3.4 g/kg) group and model group, with nine mice in each group. Astragali Radix and Salviae Miltiorrhizae Radix et Rhizoma herbal pair were given to intervene, blood pressure was measured and pathological changes of renal tissue were observed. miRNA sequencing experiments were performed to determine the differential expression profiles of miRNAs. Results After 4 weeks of intervention with Astragali Radix and Salviae Miltiorrhizae Radix et Rhizoma herbal pair, compared with model group, blood pressure of rats was significantly reduced (P<0.01), glomerular lobules of rats were not obvious, and mesangial hyperplasia was reduced. Compared with control group, 115 differentially expressed miRNAs in model group were screened, of which 68 differentially expressed miRNAs were up-regulated and 47 differentially expressed miRNAs were down-regulated; Compared with model group, 91 differentially expressed miRNAs in Astragali Radix and Salviae Miltiorrhizae Radix et Rhizoma herbal pair group were screened, of which 67 differentially expressed miRNAs were up-regulated and 24 differentially expressed miRNAs were down-regulated. The results of qRT-PCR verification of differentially expressed miRNAs showed that expressions of miRNA-142-5p, miRNA-3585-5p, miRNA-219a-5p, miRNA-122-5p and miRNA-125b-1-3p were consistent with the trend of miRNA sequencing results. The target gene prediction and function enrichment results of differentially expressed miRNAs showed that the function of gene ontology (GO) was mainly enriched in anion transmembrane transport, presynaptic, protein serine/threonine kinase activity, etc., Kyoto encyclopedia of genes and genomes (KEGG) pathway was mainly enriched in mammalian target of rapamycin (mTOR), mitogen-activated protein kinase (MAPK), autophagy, adenosine monophosphate-activated protein kinase (AMPK) signaling pathway and other pathways. Conclusion miR-219a-5p, miR-3585-5p and miR-142-5p may be the direct targets of Astragali Radix and Salviae Miltiorrhizae Radix et Rhizoma herbal pair to delay the progression of renal damage in hypertension.