摘要
基于网络药理学和体外试验,共同探讨人参皂苷Rg1和Re对感染LPS猪空肠上皮细胞IPEC-J2的保护作用.采用网络药理学获取并筛选Rg1、Re缓解肠道屏障损伤的交集靶点,并采用分子对接技术验证网络药理学的预测结果.本试验分组为Control、LPS、Rg1和Re组.观察不同浓度Rg1、Re对IPEC-J2细胞存活率、凋亡率、TEER值、FD4通透性和炎性因子的影响,同时采用荧光定量PCR检测不同浓度Rg1、Re对凋亡相关基因mRNA表达水平的影响.网络药理学结果显示,Rg1、Re预防肠道屏障损伤主要涉及PI3K-Akt、MAPK信号通路等过程.分子对接结果显示,Rg1与所有交集靶点结合能均小于0,而人参皂苷Re仅与SRC靶点的结合能小于0.体外试验显示不同浓度的Rg1、Re预处理均会不同程度地提高LPS处理的IPEC-J2的存活率、TEER值,减少细胞凋亡、FD4通透性降低、炎性因子TNF-α的分泌,表明Re、Rg1可有效降低LPS处理对IPEC-J2细胞的影响.Rg1显著上调 MAPK8、MAPK10、HRAS,显著下调 MAP2K1、PIK3CG、IL-2、SRC 的 mRNA 表达水平;Re 显 著上调 MAPK8、MAPK10、HRAS、PIK3R1、BCL2基因的mRNA表达水平.这些结果提示,人参皂苷Rg1、Re和含有Rg1、Re的人参皂苷产品在预防仔猪肠道屏障损伤方面值得进一步研究.
Abstract
Based on network pharmacology and in vitro assays,we conducted a collaborative investi-gation into the protective effects of ginsenosides Rg1 and Re on LPS-induced damage of porcine je-junal epithelial cells IPEC-J2.Network pharmacology was used to obtain and screen the intersec-ting targets of Rg1 and Re to alleviate intestinal barrier damage,and molecular docking technique was used to verify the predicted results of network pharmacology.The experiment included the Control group,LPS group,Rg1 group,and Re group.The effects of different concentrations of Rg1 and Re on cell survival rate,apoptosis rate,TEER value,FD4 permeability,and inflammatory fac-tors of IPEC-J2 were observed,and the effects of different concentrations of Rg1 and Re on the mRNA expression levels of apoptosis-related genes were also detected by fluorescence quantitative PCR.The results of network pharmacology showed that the prevention of intestinal barrier damage by Rg1,Re mainly involved the processes of PI3K-Akt and MAPK signaling pathways.The molec-ular docking results showed that the binding energy of Rg1 to all intersecting targets was less than 0,while that of ginsenoside Re to SRC targets only was less than 0.In vitro experiments showed that pretreatment with different concentrations of Rg1 and Re increased the survival rate and TEER value of LPS-treated IPEC-J2 to varying degrees,and reduced the apoptosis,the decrease of FD4 permeability,and the secretion of inflammatory factor TNF-α,suggesting that Re and Rg1 prevented the intestinal barrier from damage.It was shown that Re and Rg1 could effectively re-duce the effects of LPS treatment on IPEC-J2 cells.Rg1 significantly upregulated the mRNA ex-pression levels of MAPK8,MAPK10,HRAS,and significantly down-regulated the mRNA expres-sion levels of MAP2K1,PIK3CG,IL-2 and SRC;and Re significantly upregulated the mRNA ex-pression levels of MAPK8,MAPK10,HRAS,and PIK3R1,BCL2 gene mRNA expression levels.These results suggest that ginsenosides Rg1,Re and ginsenoside products containing Rg1 and Re deserve further investigation in preventing intestinal barrier damage in piglets.
基金项目
国家自然科学基金(32002325)
国家生猪技术创新中心先导科技资助项目(NCTIP-XD/B12)
重庆市自然科学基金(cstc2020jcyjmsxmX0418)
NETL
NSTL
万方数据