Effect And Mechanism Of Bufei Huoxue Capsule In The Intervention Of Lung Injury In Rats With Chronic Obstructive Pulmonary Diseas

The content of the thesis is divided into two parts:literature review and animal experiment research.The first part is an overview of the pathological manifestations,main pathogenesis,and signal pathways of chronic obstructive pulmonary disease.Summary Two summarizes the western medicine treatment of chronic obstructive pulmonary disease,non-pharmaceutical treatment,the study of the mechanism of action of western medicine,the prevention and treatment of chronic obstructive pulmonary by ancient physicians,the prevention and treatment of chronic obstructive pulmonary by modern physicians,and the research on the mechanism of traditional Chinese medicine treatment.Summary Three:Summarize the efficacy and pharmacological effects of Bufeihuoxue Capsule,and review its clinical application and basic research status.The second part,animal experiment research.Objective:1.To clarify the intervention effect of Bufei Huoxue Capsule on chronic inflammation,mucus hypersecretion and airway remodeling in COPD rats;2.To explore the intervention of Bufei Huoxue Capsule on chronic inflammation,mucus hypersecretion and airway of COPD rats The specific mechanism of remodeling.Methods:40 SPF male Wistar rats were randomly divided into normal control group,model group,nourishing lung and blood circulation group,and theophylline group.The chronic obstructive pulmonary rat model was established by continuous cigarette smoke exposure combined with airway instillation of LPS.The normal control group did not smoke or drip,and was fed normally;the model group,the lung and blood circulation group,and the theophylline group were smoked for 28 consecutive days.LPS was instilled in the first,11,and 21 days of the experiment,and the drug was not smoked on the day of dripping.smoke.From the 21st day of the experiment,the normal control group,model group,Bufei Huoxue group,and theophylline group were given normal saline,normal saline,Bufei Huoxue capsule(0.42g/kg),and theophylline sustained-release tablets(0.02g/kg).On the 61st day of the experiment,samples were collected,and peripheral blood,alveolar lavage fluid and lung tissue were collected.HE staining was used to observe the pathological changes of the bronchi and alveoli of each group of rats,AB-PAS staining was used to observe the airway epithelial goblet cell metaplasia and mucus secretion of each group of rats,Masson staining was used to observe the collagen deposition in the rat lung tissue,and ELISA was used to detect The contents of IL-1β,IL-6,IL-13,TNF-α,TGF-β1,NE and EGF in rat serum and alveolar lavage fluid,immunohistochemical staining to observe the expression of Muc5ac and TGF-β1 in lung tissue,Western Blot method to detect TLR4,TLR2,MyD88,ICAM-1,MCP-1,MPO,NE,CD206,JNK,P-JNK,ERK1/2,P-ERK1/2,P38 MAPK in the lung tissue of each group of rats,P-P38 MAPK,NF-κB P65,Muc5ac,EGFR,P-EGFR,PI3K,P-PI3K,AKT,P-AKT,TGF-β1,Smad2/3,Smad4,Smad7,α1-AT,MMP-9,TIMP-1 protein expression,real-time fluorescent quantitative PCR to detect TLR4,MyD88,ICAM-1,MPO,NE,MAPK1,MAPK3,MAPK9,MAPK14,NFκB,Muc5ac,PI3K,AKT,TGF-β1,Smad2,Smad3,Smad4,Smad7,αl-AT,MMP-9,TIMP-1 mRNA expression.Result:The results of HE staining showed that compared with the normal control group,the airway epithelium of the model group was significantly thickened,cilia adhered and fell,submucosal inflammatory cells infiltrated,the alveolar structure was obviously destroyed,the alveolar collapsed and merged,the alveolar septum thickened,and the lungs Infiltration of inflammatory cells and red blood cells.Compared with the model group,the airway epithelium of the Bufeihuoxue group and the theophylline group is relatively complete,the columnar epithelium is arranged more regularly,the ciliary adhesion,lodging,and shedding are improved,the alveolar structure is relatively intact,the alveolar interval becomes smaller,and the alveolar fusion decreases.The infiltration of inflammatory cells and red blood cells is reduced.The results of AB-PAS staining showed that compared with the normal control group,a large number of blue-stained goblet cells were seen in the airway epithelium of the model group;compared with the model group,the airway epithelial goblet cells of the Bufeihuoxue group and theophylline group were significantly reduced.Masson staining results showed that compared with the normal control group,a large number of blue collagen fibers were seen in the airway epithelium and submucosa of the model group,and the staining was significantly deepened,and part of the red smooth muscle was destroyed by inflammatory cells in the submucosa.Quantitative analysis showed collagen The proportion of fibers in the total airway area increased significantly.Compared with the model group,the blue collagen fibers between the airway epithelium and the submucosa of the Bufeihuoxue group and the theophylline group were significantly reduced,and the red smooth muscle was thin and uniform without breaking;the proportion of collagen fibers in the total airway area was significantly reduced.There was no significant difference between the two groups.Immunohistochemical staining showed that compared with the normal control group,the positive expression of Muc5ac and TGF-β1 in the model group was significantly increased;compared with the model group,the positive expression of Muc5ac and TGF-β1 in the Bufeihuoxue group and theophylline group was significantly reduced.The ELISA test results showed that compared with the normal control group,the levels of IL-1β,IL-6,NE,TGF-β1,and EGF in the alveolar lavage fluid of the model group increased significantly,and the serum levels of IL-13 and TNF-α The level has risen significantly.Compared with the model group,the levels of IL-1β,IL-6,NE,TGF-β1 IL-13,TNF-α,and EGF in the Bufeihuoxue group and theophylline group were significantly reduced;compared with the theophylline group,The level of TGF-β1 in the alveolar lavage fluid of rats in the Bufeihuoxue group was lower,and there was no significant difference in other indicators.Western Blot results showed that compared with the normal control group,the model group TLR4,MyD88,ICAM-1,MPO,NE,CD206,P-EGFR/EGFR,P-ERK1/2/ERK1/2,P-P38/P38The expressions of MAPK,NF-κB,Muc5ac,P-EGFR/EGFR,P-PI3K/PI3K,P-AKT/AKT,TGF-β1,Smad2/3,Smad4,NE,αl-AT,and MMP-9 were significantly increased,Smad7 protein expression was significantly reduced,and there was no significant difference in TLR2,MCP-1,P-JNK/JNK,TIMP-1.Compared with the model group,the TLR4,MyD88,ICAM-1,NE,CD206,P-EGFR/EGFR,P-ERK1/2/ERK1/2,P-P38/P38-MAPK,theophylline group and the theophylline group were compared with the model group.The expression of NF-κB,Muc5ac,P-EGFR/EGFR,P-PI3K/PI3K,P-AKT/AKT,TGF-β1,Smad2/3,Smad4,NE,α1-AT,MMP-9 was significantly reduced,and the expression of Smad7 protein Significantly increased;the expression of MPO decreased in the nourishing lung and blood circulation group,and there was no significant difference in the expression of TLR2,MCP-1,P-JNK/JNK,and TIMP-1;the expression of TLR2,MPO,MCP-1,and TIMP-1 in the theophylline group decreased Trend,but not statistically significant,the expression of P-JNK/JNK decreased.Compared with theophylline group,the P-P38/P38 expression was higher in the Bufeihuoxue group,and there was no significant difference in other indicators.Real-time fluorescent quantitative PCR results showed that compared with the normal control group,the model group TLR4,MyD88,IL-1β,IL-6,MPO,ICAM-1,NE,MAPK1,MAPK3,MAPK14,NF-κB,Muc5ac,PI3K,AKT,TGF-β1,Smad2,Smad4,MMP-9,TIMP-1,NE,al-AT mRNA expression increased significantly,Smad7 mRNA expression decreased significantly,and Smad3,MAPK9 mRNA expression had no significant difference.Compared with the model group,TLR4,MyD88,IL-1β,IL-6,MPO,ICAM-1,NE,MAPK1,MAPK3,MAPK14,NF-κB,Muc5ac,PI3K,AKT,and theophylline group were compared with the model group.TGF-β1,Smad2,Smad4,MMP-9,TIMP-1,NE,and αl-AT mRNA expression were significantly reduced,Smad7 mRNA expression was significantly increased,Smad3,MAPK9 mRNA expression was not significantly different,and there was no significant difference between the two groups.Conclusion:1.Bufei Huoxue Capsule can reduce the chronic inflammatory response of COPD by inhibiting TLR4-MyD88-NF-κB,P38MAPK,ERK1/2 signal transduction pathway;2.Bufei Huoxue Capsule can inhibit EGFR-PI3K-AKT signal Transduction pathway,downregulate the expression of Muc5ac,and interfere with the hypersecretion of COPD airway mucus;3.Bufei Huoxue Capsules can improve the airway remodeling of COPD by inhibiting the TGF-β1/Smad signal transduction pathway.