大黄牡丹汤调控MAPK信号通路防治重症急性胰腺炎相关肺损伤的作用机制OA

Objective To investigate the therapeutic mechanism of Dahuang Mudan Decoction(DHMDD)in severe acute pancreatitis-associated lung injury(SAP-ALI)by combining network pharmacology prediction with animal experiments.Methods(1)Network pharmacology was adopted to predict potential therapeutic targets of DHMDD against SAP-ALI.The"active ingredient-target"interaction network of DHMDD was constructed to screen key active ingredients.Protein-protein interaction(PPI)network analysis was performed on potential targets to identify core targets.Gene Ontology(GO)functional enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analyses were conducted on potential targets to screen critical signaling pathways.(2)Kunming mice were randomly divided into blank control group,model group and DHMDD group,with 10 mice in each group.The SAP mouse model was established by intraperitoneal injection of 20%L-arginine(3.5 g·kg-1)twice at an interval of 1 hour.Thirty minutes after L-arginine injection,mice in the DHMDD group received intragastric administration of DHMDD(22 g·kg-1·d-1)at a volume of 10 mL·kg-1,twice daily for consecutive 3 days.An automatic biochemical analyzer was used to detect serum amylase and lipase levels.Hematoxylin-eosin(HE)staining was applied to observe pathological changes in pancreatic and lung tissues.Immunohistochemistry was utilized to determine the expressions of IL-6,F4/80,Ly6G,MPO,p-ERK,p-P38 and p-JNK in lung tissues.Immunofluorescence co-staining was adopted to examine the co-expression of F4/80&p-ERK,F4/80&p-JNK,F4/80&p-P38 in lung tissues.Results(1)A total of 83 potential targets of DHMDD against SAP-ALI were screened.Quercetin,kaempferol,aloe-emodin and catechin were identified as key active components.TP53,AKT1,HIF-1α and IL-6 were core targets.The potential targets were mainly involved in key biological processes such as inflammatory response regulation,apoptosis and angiogenesis,as well as key signaling pathways including IL-17,MAPK and PI3K-AKT.(2)Compared with the blank control group,the model group exhibited disrupted pancreatic lobular structure,extensive acinar cell degeneration and necrosis,marked interstitial edema,and significant inflammatory cell infiltration,with significantly increased pancreatic pathological scores,pancreas-to-body weight ratio,and serum amylase and lipase levels(P＜0.01).Lung tissue showed typical acute injury features with significantly widened alveolar septa and diffuse inflammatory cell infiltration(predominantly neutrophils and macrophages),with occasional erythrocyte exudation and elevated lung pathological scores(P＜0.01).F4/80 and Ly6G levels were significantly increased in lung tissue(P＜0.01),mainly localized in alveolar septa;IL-6 and MPO levels were also significantly elevated(P＜0.01),mainly expressed in macrophages and neutrophils.Protein expression of p-ERK,p-JNK and p-P38 was significantly upregulated in lung tissue(P＜0.01)and co-localized with macrophages,with significantly increased co-expression levels of p-ERK/p-JNK/p-P38(red)and F4/80(green)(P＜0.01).Compared with the model group,the DHMDD group showed significantly alleviated pancreatic tissue injury,reduced acinar cell necrosis,inflammatory cell infiltration and interstitial edema,and partially restored pancreatic lobular structure,with significantly decreased pancreatic pathological scores,pancreas-to-body weight ratio,and serum amylase and lipase levels(P＜0.01).Lung injury was also markedly attenuated,with narrowed alveolar septa and reduced inflammatory cell infiltration,accompanied by significantly decreased lung pathological scores(P＜0.01).Levels of F4/80,Ly6G,IL-6 and MPO in lung tissue were all significantly reduced(P＜0.01).Protein expression of p-ERK,p-JNK and p-P38 was significantly downregulated(P＜0.01),and their co-expression levels with F4/80 were significantly decreased(P＜0.01).Conclusion DHMDD can ameliorate pathological damage of pancreatic and lung tissues in SAP-ALI mice,and inhibit macrophage infiltration,neutrophil aggregation and expression of inflammatory factors MPO and IL-6 in lung tissue.Its mechanism may involve the regulation of MAPK signaling pathway to suppress macrophage-mediated inflammatory response,thereby improving SAP-ALI.