基于网络药理学和体外实验探讨金银花治疗IgA肾病的作用机制

Mechanism of honeysuckle for IgA nephropathy based on network pharmacology and in vitro experimental verification

目的运用网络药理学预测金银花治疗IgA肾病(IgAN)潜在作用机制,进一步通过体外实验进行验证。方法通过TCMSP平台获取金银花的主要活性成分,运用GeneCards、OMIM数据库检索IgAN相关靶点,并映射到金银花的潜在作用靶点中,将IgAN和金银花靶点取交集,通过构建药物-成分-疾病-靶点网络图,进行KEGG通路富集分析。最后通过重组人IgA刺激共培养人肾小球血管内皮细胞(HRGECs)和人肾小球系膜细胞(HMCs)构建IgAN细胞模型,并给予不同浓度木犀草素进行干预,验证网络药理学结果。结果网络药理学分析结果发现PI3K-AKT信号通路可能是金银花治疗IgAN的潜在作用靶点。网络图拓扑学分析显示,活性成分木犀草素与PI3K-AKT信号通路关系最大。体外实验发现,木犀草素能抑制HMCs增殖,减少IL-6分泌,从而增强HRGECs间黏附分子的表达,改善细胞间的紧密连接,减少HRGECs单层通透性。对HMCs内PI3K-AKT信号通路相关蛋白检测发现,木犀草素可以抑制细胞内PI3K和AKT的磷酸化水平。结论木犀草素通过抑制HMCs内PI3K-AKT信号通路,抑制HMCs增殖和IL-6分泌,进而改善肾小球滤过屏障发挥治疗IgAN作用。

Objective To predict the mechanism of honeysuckle for IgA nephropathy(IgAN)by network pharmacology,and to verify the results by in vitro experiments.Methods The main active components of honeysuckle were obtained through TCMSP platform.The related targets of IgAN were retrieved by GeneCards and OMIM database,and mapped to the potential targets of honeysuckle.The targets of IgAN and honeysuckle were overlapped,and the KEGG pathway enrichment analysis was performed by constructing drug-component-disease-target network diagram.Finally,an cell model of IgAN was constructed by co-culturing human glomerular endothelial cells(HRGECs)and human mesangial cells(HMCs)stimulated with recombinant human IgA.Different concentrations of luteolin were given to verify the network pharmacology results.Results Network pharmacology analysis showed that PI3K-AKT signaling pathway might be a potential target of honeysuckle for IgAN.The topological analysis of network diagram showed that the active component luteolin was most related to PI3K-AKT signaling pathway.In vitro experiments showed that luteolin inhibited the proliferation of HMCs and reduced the secretion of inflammatory factor IL-6,thus enhancing the expression of adhesion molecules in HRGECs,improving the junction between cells and reducing the permeability of HRGECs monolayers.The expression of PI3K-AKT signaling pathway-related proteins in HMCs was detected,and luteolin inhibited the phosphorylation of PI3K and AKT in cells.Conclusion Luteolin inhibits the proliferation of HMCs and IL-6 secretion by inhibiting PI3K-AKT signaling pathway to improve glomerular filtration barriers in IgAN.