目的 运用网络药理学和细胞生物学验证探讨黄芪治疗糖尿病肾病的分子作用机制。方法 通过中药系统药理学数据库(TCMSP)筛选出黄芪有效成分;利用Drugbank数据库、GeneCards和OMIM数据库筛选黄芪有效成分和糖尿病肾病的相关靶点;应用String数据库绘制PPI网络;通过Cytoscape软件绘制黄芪-成分-靶点-疾病网络;借助DAVID数据库对靶点进行GO和KEGG通路分析;利用KEGG数据库构建黄芪治疗糖尿病肾病的通路图;细胞生物学验证分析黄芪中的有效成分对糖尿病肾病的保护作用。结果 筛选得到黄芪12个有效成分,涉及56个作用靶点,主要涉及氧化应激、炎症和细胞凋亡等生物过程,主要调节有丝分裂原激活的蛋白激酶(MAPK)、缺氧诱导因子(HIF)、ErbB、p53和核因子-κB(NF-κB)等信号通路,其作用机制包含抗氧化、抗炎和抑制细胞凋亡等。细胞生物学验证分析表明槲皮素和山柰酚能够降低高糖引起的细胞凋亡,降低活性氧(reactive oxygen species,ROS) 水平(P＜0.05)、提高过氧化氢酶(catalase,CAT)和超氧化物歧化酶(superoxide dismutase,SOD)活性(P＜0.05)。槲皮素能够显著降低高糖引起的HK-2细胞磷酸化P38 MAPK和JNK蛋白表达水平(P＜0.01)。结论 本研究通过网络药理学层面探讨了黄芪治疗糖尿病肾病的作用机制,发现可能是通过多成分、多靶点、多通路等途径发挥其治疗作用,黄芪中的活性成分槲皮素和山柰酚可能是其治疗糖尿病肾病的物质基础。体外水平验证了黄芪中的主要活性成分槲皮素和山柰酚能够降低高糖引起的细胞损伤,降低氧化应激反应,提高抗氧化性,为黄芪治疗糖尿病肾病提供新思路。
Abstract
OBJECTIVE To explore the molecular mechanism of Astragali Radix in the treatment of diabetic nephropathy (DN) by the network pharmacology and cell biology verification. METHODS The main active ingredients of Astragali Radix were obtained through TCMSP. Drugbank, Gene Cards and OMIM were used to predict and screen the targets of Astragali Radix and DN. The PPI network was constructed using the String database. The GO and KEGG pathways involved in the targets were analyzed by DAVID. The ingredients-targets-diseases network was constructed by Cytoscape software. Cell biology verification analyzes the protective effect of active ingredients in Astragali Radix on DN. RESULTS The results showed that 12 active components and 56 targets of Astragali Radix were involved. The network results showed that the process of oxidative stress, inflammation, apoptosis were mainly involved, which played a role in the treatment of DN by adjusting the MAPK, HIF, ErbB, p53, NF-κB and other signal pathways. Its mechanism of action involved antioxidation, anti-inflammatory and anti-apoptosis. Cell biology verification analysis showed that quercetin and kaempferol could reduce cell apoptosis caused by high glucose, reduce ROS level (P＜0.05), and improve CAT and SOD capacity (P＜0.05). Quercetin could significantly reduce the protein expression levels of phosphorylated P38 MAPK and JNK in HK-2 cells caused by high glucose (P＜0.01). CONCLUSION In the current study, the mechanism of Astragali Radix in the treatment of DN by network pharmacology was explored. RESULTS showed that Astragali Radix could act on multi-ingredient, multi-target and multi-pathway played a potential therapeutic role on DN. The prediction results of network pharmacology suggest that the quercetin and kaempferol in Astragali Radix might be the material basis for treatment of DN. The analysis proved the quercetin and kaempferol in Astragali Radix can reduce cell damage caused by high glucose, reduce oxidative stress, and improve antioxidant capacity by in vitro experiment, providing a new idea for Astragali Radix in the treatment of DN.
关键词
黄芪 /
糖尿病肾病 /
网络药理学 /
MAPK信号通路
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Key words
Astragali Radix /
diabetic nephropathy /
network pharmacology /
MAPK signal pathway
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中图分类号：
R966
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脚注
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基金
国家自然科学基金项目资助(81673656);浙江省自然科学基金项目资助(LGF20H300003);浙江省医药卫生厅项目资助(2019RC085);绍兴市科技局医卫类项目资助(2018C30115)
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