摘要
建立大鼠急性酒精性肝损伤(acute alcoholic liver injury,ALI)模型,研究金银花Lonicerae Japonicae Flos水提取物对ALI大鼠丝裂原活化蛋白激酶(MAPK)信号通路的影响,探讨其保护ALI的作用机制。首先,利用网络药理学方法预测金银花预防ALI的靶点,构建成分-靶点-通路关系网络,筛选金银花活性成分作用于MAPK信号通路的关键靶点。SD大鼠随机分为空白组、模型组、阳性药组以及金银花高、中、低剂量组。各给药组预防性灌胃给予相应药物,每天灌胃1次,连续7 d,空白组和模型组每天灌胃给予等体积的蒸馏水。除空白组外,大鼠灌胃56度白酒,持续灌胃3 d后建立ALI模型。检测各组大鼠血清谷草转氨酶(AST)、谷丙转氨酶(ALT)、白细胞介素-6(IL6)、肿瘤坏死因子α(TNF-α)含量及肝组织中丙二醛(MDA)、谷胱甘肽(GSH)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)水平,并利用荧光定量PCR法研究金银花对ALI大鼠肝脏组织MAPK通路关键靶点的影响。TCMSP数据库筛选得到28种金银花活性成分,与肝损伤相关的靶点317个,KEGG富集分析得到226条相关信号通路,主要富集在肝病信号通路、炎症、免疫、细胞凋亡等相关的通路。对"MAPK信号通路-靶点-活性成分"网络进行可视化分析,得到金银花关键活性成分如绿原酸、金丝桃苷、常春藤皂苷元等作用于MAPK信号通路的25个关键靶点。体内实验结果表明,金银花高、中、低剂量组大鼠血清AST、ALT和肝脏组织MDA水平均显著降低(P<0.01或P<0.05),金银花低、高剂量组大鼠血清IL6含量均显著降低(P<0.01或P<0.05),金银花中、高剂量组肝组织SOD和GSH含量显著升高(P<0.01或P<0.05)。荧光定量PCR结果表明,金银花高、中、低剂量组均可显著降低MAP2K4和MAPK3基因的表达水平(P<0.01)。网络药理学和实验验证结果表明,金银花中活性成分可能通过抑制MAPK信号通路关键靶点的表达以降低炎症因子水平,增加抗氧化酶的活性而抑制氧化应激反应,从而改善和预防酒精对肝脏造成的损伤。
This study aims to explore underlying mechanism of Lonicerae Japonicae Flos(LJF)in protecting rats against acute alcoholic liver injury(ALI)based on mitogen-activated protein kinase(MAPK)pathway.First,the targets of LJF in preventing ALI were predicted by network pharmacology and the component-target-pathway network was constructed,so that the key targets of LJF components acting on MAPK pathway were screened.Second,male SD rats were randomized into the control(KB)group,model(MX)group,positive(YX)group,and LJF high-(GJ),medium-(ZJ),and low-(DJ)dose groups.Each administration group was given(ig)corresponding drugs for 7 days and KB group and MX group received(ig)equal volume of distilled water every day.Except for KB group,rats were given Chinese spirit(56%,3 days)for ALI modeling.The levels of aspartate transaminase(AST),alanine transaminase(ALT),interleukin-6(IL6)and tumor necrosis factor-α(TNF-α)in serum and malondialdehyde(MDA),glutathione(GSH),superoxide dismutase(SOD)and glutathione peroxidase(GSH-Px)in liver tissue of rats in each group were detected.Furthermore,we employed quantitative real-time PCR(qRT-PCR)to probe the effects of LJF on the key targets of MAPK pathway in ALI rats.A total of 28 active components of LJF were screened from TCMSP database,and 317 intersected with ALI-related targets.According to Kyoto encyclopedia of genes and genomes(KEGG)pathway enrichment analysis,the 317 targets involved 226 pathways,which were mainly liver disease,inflammation,immunity,apoptosis and other related pathways.According to the MAPK pathway-target-active component network,the key active components of LJF,such as chlorogenic acid,hederagenol,and hyperoside,acted on 25 key targets of MAPK pathway.The results of in vivo experiments showed decreased levels of AST,ALT,and MDA in DJ,ZJ,and GJ groups(P<0.01 or P<0.05),reduced levels of IL6 in DJ and GJ groups(P<0.01 or P<0.05),and improved levels of SOD and GSH in ZJ and GJ groups(P<0.01 or P<0.05).The results of qRT-PCR demonstrated that the expression levels of mitogen-activated protein kinase kinase 4(MAPK2 K4)and mitogen-activated protein kinase 3(MAPK3)were decreased in DJ,ZJ,and GJ groups(P<0.01).The network pharmacology and experimental verification showed that the active components in LJF can reduce the inflammatory factor level and enhance the activities of SOD and GSH-Px by inhibiting the expression of key targets of MAPK pathway,thus alleviating and preventing liver damage caused by alcohol.
作者
刘畅
丁晶鑫
周英
尹志刚
罗洪涛
孔维涛
LIU Chang;DING Jing-xin;ZHOU Ying;YIN Zhi-gang;LUO Hong-tao;KONG Wei-tao(School of Pharmacy/Research Center for Application and Development of Medicine and Food Dual-use Resources,Guizhou University of Traditional Chinese Medicine,Guiyang 550025,China;Guizhou Engineering Center for Innovative Traditional Chinese Medicine and Ethnic Medicine,Guiyang 550025,China;College of Life Science,Guizhou University,Guiyang 550025,China)
出处
《中国中药杂志》
CAS
CSCD
北大核心
2021年第17期4531-4540,共10页
China Journal of Chinese Materia Medica
基金
贵州省科学技术基金项目(黔科合基础-ZK[2021]一般515)
贵州省科技攻关计划项目(黔科合[2016]支撑2909)
贵州中医药大学本科教学工程建设项目(贵中医教学工程合字(2019)54号)。
关键词
金银花
网络药理学
MAPK信号通路
酒精性肝损伤
Lonicerae Japonicae Flos
network pharmacology
MAPK pathway
alcoholic liver injury
