Explore The Liver Protection Of Pleurotus Placentodes Based On Network Pharmacology And LC-MS/MS Technology

Liver injury is a common disease in daily life.Long-term use of tiopronin and bidentate and other commonly used liver injury drugs will produce a series of adverse reactions,and bring great pain and economic burden to patients.Pleurotus placentodes（Pleurotus placentodes）is a fungus that can be used as both medicine and food.Our research group found that the polysaccharide of Pleurotus placentodes has good anti-inflammatory activity.The preliminary screening of Pleurotus placentodes alcohol extract（PP-a）showed that it could significantly reduce the contents of GOT and GPT in the serum of mice（P<0.01）.Therefore,the in-depth study of the hepatoprotective activity of PP-a also provides a material basis for the development and utilization of Pleurotus placentodes.Based on the previous literature investigation and experimental studies,it has been shown that Pleuropleura oospore has potential liver protection activity.Therefore,this study first used non-targeted metabolomics technology to identify the chemical components of PP-a,then predicted the potential active ingredients,signaling pathways,and mechanisms for treating liver injury through network pharmacology technology,and verified the liver protection efficacy and target of PP-a through RT-qPCR technology in combination with animal experiments.Firstly,the chemical constituents of PP-a were analyzed by LC-MS/MS technique,and 107 chemical constituents of PP-a were obtained.Secondly,effective chemical components were screened out through TCMSP and Swiss ADME databases,and the potential hepatoprotective targets and mechanisms of PP-a were studied through network pharmacology technology.Eight potential targets were identified by PPI topological attribute analysis,which was as follows:PTGS2,PPARA,EP300,NR3C1,RARA,RXRA,HDAC1,RARG,and the results of KEGG enrichment analysis,and GO enrichment analysis suggested that the hepatoprotective effect of PPA might be related to the arachidonic acid metabolic pathway and PPAR signaling pathway.Secondly,in this study,a mouse model of acute liver injury was established by intraperitoneally injecting 0.2%CCl₄ olive oil solution,and RT-qPCR was used to verify the potential liver-protective targets of PP-a.The KM mice were randomly divided into 6 groups:blank control group,model group,PP-a 3dose groups（high-dose group 1000 mg/kg,medium-dose group 500 mg/kg,low-dose group 250 mg/kg）and silymarin positive control group 140 mg/kg.The liver index of mice was measured,the changes in liver pathological tissue sections were analyzed,and the effect of PP-a on target organs was evaluated.Serum was collected and the changes in liver function and immune cytokine content in mice were measured.The anti-oxidative stress ability of mice was measured by liver tissue.The results showed that compared with the model group,PP-a could reduce the liver index（P<0.001）,and improve liver swelling and hypertrophy;In serum,the contents of IL-6,IL-1β,and TNF-αin the high-dose PP-a group were significantly decreased（P<0.001）;The activities of AKP,GOT,and GPT in serum were significantly decreased（P<0.001）;In the liver,SOD and GSH-Px activities increased（P<0.05）,while MDA content decreased（P<0.001）,increased the ability of mice to resist oxidative stress.RT-qPCR was used to detect the m RNA expression levels of 8 targets,and the mRNA expression levels of PTGS2,NR3C1,and PPARA showed significant changes.Compared with the blank group,PTGS2 and NR3C1 in the model group were significantly increased（P<0.001）,PPARA m RNA expression decreased（P<0.01）,compared with the model group,and PP-a high-dose group was significantly reversed（P<0.01）.The potential chemical components,targets,and signaling pathways of PP-a’s hepatoprotective effect were identified through network pharmacological prediction.Combined with animal models,it was found that PP-a can effectively improve liver function,reduce inflammatory factors,increase antioxidant stress ability,and improve the pathological changes of liver tissue in mice with liver injury.Combined with the prediction results of network pharmacology,RT-qPCR was used to verify the target.PTGS2,NR3C1,and PPARA showed significant differences compared to the model group.It is speculated that L-Histidine,N-Acetyl-D-alloisoleucine,15-Oxo EDE,and Palmitic acid in the effective chemical components of PP-a may play a protective role in liver injury by regulating the mRNA expression levels of PTGS2,NR3C1,and PPARA.