Qingjin Huatan Decoction And Baicalin Inhibit LPS-induced Acute Lung Injury Through The TLR4/MyD88/NF-κB/NLRP3 Pathway

This paper is divided into two parts:review and experimental research.ReviewReview oneThis review summarizes the domestic and foreign references on the treatment of acute lung injury with Chinese herbal compound and Chinese herbal medicine monomer..Review twoThis article reviews the etiology and clinical manifestations of acute lung injury,and focuses on the pathogenesis of acute lung injury.Experimental StudyThe experimental study is divided into two parts.The first part mainly studies the intervention effect of Qingjin Huatan Decoction on acute lung injury,which is divided into four experiments;the second part mainly studies the intervention effect of baicalin on acute lung injury,which is divided into five Experiments.Part 1:Intervention of Qingjin Huatan Decoction on Acute Lung InjuryExperiment 1Objective:To evaluate the effect of Qingjin Huatan Decoction on the inflammatory response in rats with acute lung injury.Methods:An acute lung injury model was established by airway instillation of LPS solution,and low-dose Qingjin Huatan Decoction,high-dose Qingjin Huatan Decoction and clarithromycin were given by gavage.Detect lung wet/dry ratio,BALF protein concentration,inflammatory factor secretion and lung tissue pathological changes.Results:Airway instillation of LPS significantly increased the wet/dry ratio of rat lung weight and the protein concentration of rat BALF,increased the permeability of alveolar capillary membranes,promoted the release of inflammatory factors CXCL-1,IL-6 and MPO,induced the aggregation and infiltration of neutrophils,and caused alveolar wall thickening and bronchial epithelial cell edema.Qingjin Huatan Decoction significantly reduced lung weight wet/dry ratio and protein concentration of rat BALF,reduced alveolar capillary membrane permeability in ALI rats,reduced protein infiltration from capillaries and interstitium to alveolar,reduce the penetration of protein from capillaries and interstitial to alveoli,reduced neutrophil aggregation,and inhibit the secretion of inflammatory factors CXCL-1,IL-6 and MPO.Compared with the model group,the difference was statistically significant(P<0.05).Experiment 2Objective:To evaluate the effect of Qingjin Huatan Decoction on the inflammatory response in rats with acute lung injury.Methods:An acute lung injury model was established by airway instillation of LPS solution,and low-dose Qingjin Huatan Decoction,high-dose Qingjin Huatan Decoction and clarithromycin were given by gavage.WB method was used to detect changes in protein expression,and IHC was used to detect changes in lung tissue protein expression.Results:Airway instillation of LPS significantly promoted the increase of TLR4,MYD88,NLRP3 protein expression and NF-κB phosphorylation.Qingjin Huatan Decoction significantly reduced the expression of TLR4,MYD88,NLRP3 protein,and inhibited the phosphorylation of NF-κB.Compared with the model group,the difference was statistically significant(P<0.05).Experiment 3Objective:Explore the effect of Qingjin Huatan Decoction containing serum freeze-dried powder on the expression of inflammatory factors in BEAS-2B cell injury.Method:Lyophilize medicated serum into lyophilized powder.The bronchial epithelial cells were treated with blank medicated serum freeze-dried powder,Qingjin Huatan Tang medicated serum freeze-dried powder,and clarithromycin medicated serum freeze-dried powder.The CCK8 method was used to detect the cytotoxicity of each drug,and detect the effect of each drug in inhibiting the secretion of inflammatory factors.Results:Qingjin Huatan Decoction medicated serum freeze-dried powder and clarithromycin medicated serum freeze-dried powder caused obvious cell edema,and black particulate matter can be seen in the cells.And it does not significantly inhibit the secretion of inflammatory factors IL-8,IL-6,TNF-α,MPO.Compared with the model group,the difference was statistically significant(P<0.05).Experiment 4Objective:To analyze the main chemical components and blood components of the traditional Chinese medicine compound Qingjin Huatan Decoction.Methods:Rats were intragastrically administered with pure water,Qingjin Huatan Decoction,and clarithromycin 7 days later,blood was collected and centrifuged to prepare medicated serum.Using HPLC-MS detect and analyze the main chemical components of each drug and its drug-containing serum.Results:Qingjin Huatan Decoction contains 9 chemical components,namely baicalin,pseudoephedrine,chlorogenic acid,caffeic acid,rutin,forsythin,rhein,ammonium glycyrrhizinate,oleanolic acid,and three of them are blood components,namely baicalin,pseudoephedrine,and ammonium glycyrrhizinate.Part 2:Intervention effect of baicalin on acute lung injuryExperiment 5Objective:Explore the effect of baicalin on the expression of inflammatory factors in LPS-induced BEAS-2B cell injury.Methods:Intervention of bronchial epithelial cells with baicalin standards and clarithromycin standards.The CCK8 method was used to detect the cytotoxicity of each drug and the effect of each drug on inhibiting the secretion of inflammatory factors.Results:When the concentration of baicalin standard is 10 μg/ml and below,it has no obvious effect on the cell survival rate.The cells grow well,and the cells are flat and polygonal.Baicalin has a certain inhibitory effect on the expression of inflammatory factors IL-8,IL-6,TNF-α and MPO.Experiment 6Objective:Explore the effect of baicalin on the inflammatory response of BEAS-2B cell injury induced by LPS.Methods:The baicalin standard and clarithromycin standard were used to intervene the LPS-induced acute injury model of epithelial cells.Detectd the content of inflammatory factors in the cell culture supernatant,and observed cell morphology.The Transwell chamber was used to detect the effect of epithelial cells on chemotaxis of neutrophils.Results:LPS intervention in BEAS-2B cells significantly promoted the secretion of cytokines IL-8,IL-6,TNF-α,IFN-y,IL-1 β and GM-CSF,stimulated bronchial epithelial cells to chemoattract neutrophils,and induced epithelial cell damage.Baicalin significantly inhibited the secretion of inflammatory factors such as IL-8,IL-6,TNF-α,IFN-γ,IL-1β and GM-CSF,and reduced the migration number of neutrophils.Experiment 7Objective:Explore the effect of baicalin on the TLR4 signaling pathway of LPS-induced BEAS-2B cell injury.Methods:Baicalin standards and clarithromycin standards were used to intervene the LPS-induced acute injury model of epithelial cells.To detect TLR4,MYD88,TRIF,NF-κB,NLRP3 protein content and mRNA expression in cells.Results:LPS intervention in BEAS-2B cells activated the TLR4 signaling pathway,significantly up-regulate the expression of TLR4,MyD88,NLRP3 protein and the phosphorylation of NF-κB.Baicalin significantly down-regulated the protein content and mRNA expression of TLR4,MyD88,p-NF-κB and NLRP3.Compared with the model group,the difference was statistically significant(P<0.05).Experiment 8Objective:Explore the effects of baicalin on the inflammatory response in rats with acute lung injury.Methods:LPS solution was instilled into the airway to establish acute lung injury rat model,and low-dose baicalin,high-dose baicalin and clarithromycin were administered by gavage.Detected lung wet/dry ratio,BALF protein concentration,inflammatory factor secretion,and lung tissue pathological changes.Results:LPS intervention up-regulated the secretion of inflammatory factors CXCL-1,IL-6,IL-1β,TNF-α and MPO in BALF and serum,and increased the permeability of alveolar capillary membranes.Baicalin intervention significantly reduced the wet/dry ratio of rat lung tissue weight and the protein concentration of BALF,inhibited the infiltration of a large number of neutrophils into the alveolar interstitium and alveolar space,reduced bronchial epithelial cell edema,inhibited epithelial cell mucus secretion,and inhibited the secretion of inflammatory factors CXCL-1,IL-6,IL-1β,TNF-α and MPO in BALF and serum.Compared with the model group,the difference was statistically significant(P<0.05).Experiment 9Objective:Explore the effects of baicalin on TLR4 and MAPK signaling pathways in rats with acute lung injury.Methods:LPS solution was instilled into the airway to establish acute lung injury rat model,and low-dose baicalin,high-dose baicalin and clarithromycin were administered by gavage.WB method was used to detect changes in protein expression,and IHC was used to detect changes in lung tissue protein expression.Results:LPS intervention promotes the activation of TLR4 signaling pathway and MAPK signaling pathway.Baicalin significantly inhibited the expression of TLR4,MYD88,P-NF-κB,NLRP3,P-ERK and P-p38 protein.Compared with the model group,the difference was statistically significant(P<0.05).Conclusion1.Airway instillation of 5 mg/kg LPS solution can induce increased alveolar capillary permeability,increased secretion of inflammatory factors,and pathological changes in lung tissue in rats.It conforms to the pathological changes of acute lung injury and can be used for experimental research of acute lung injury.2.Qingjin Huatan Decoction can inhibit the activation of the TLR4/MYD88/NF-κB/NLRP3 signaling pathway,inhibit the secretion of inflammatory factors and the infiltration of inflammatory cells,reduce the inflammatory response induced by LPS,and alleviate the acute lung injury model rats Increased alveolar capillary permeability and acute lung injury in rats.3.Baicalin,the main chemical component of Qingjin Huatan Decoction,can significantly inhibit LPS-induced damage to BEAS-2B cells and the secretion of inflammatory factors.4.Baicalin can inhibit the secretion of inflammatory factors and the infiltration of inflammatory cells in LPS-induced acute lung injury model rats,reduce the inflammatory response induced by LPS,and relieve the increase in alveolar capillary permeability in acute lung injury model rats.Pulmonary edema and acute lung injury in rats.5.Baicalin can inhibit the activation and transcription of the TLR4/MYD88/NF-κB/NLRP3 signaling pathway,reduce the secretion of inflammatory factors and chemokines,and inhibit the aggregation of inflammatory cells and the outbreak of inflammatory storms.6.Baicalin can inhibit the activation of MAPK signaling pathway,reduce the secretion of inflammatory factors and chemokines,inhibit the aggregation of inflammatory cells and the expansion of inflammatory response.