| 1 Objective : To observe the therapeutic effect of Yifei Jianpi Recipe on chronic obstructive pulmonary disease(COPD)model rats,and based on EGFR/MAPK/MUC5 AC pathway to explore the mechanism of Yifei Jianpi Recipe from the perspective of airway mucus hypersecretion.2 Methods : The experiment selected 96 male SD rats,which were divided into 6groups according to the random table(normal group,model group,Yifei Jianpi high,medium and low dose group,ambroxol group).Each group had 16 rats.The rat model of COPD lung and spleen deficiency was established by cigarette smoke exposure combined with intratracheal instillation of lipopolysaccharide(LPS)combined with senna leaf gavage and diarrhea.The modeling period was 49 days.After the model was completed,each group was given the corresponding drug for 6 weeks.The normal group and the model group were given normal saline.After the administration,the lung and bronchial tissue samples were taken.The bronchus and lung tissues of the right lower lobe were taken.HE staining was used to observe the pathological changes of the right lower lobe bronchus and lung.The right middle lobe bronchus was taken.AB-PAS staining was used to observe the bronchial goblet cells and mucus secretion.The expression of EGFR,MUC5 AC protein and ERK1/2,JNK,p38 total protein and phosphorylated protein in bronchopulmonary tissues of rats in each group were detected by Western blot.The expressions of EGFR,ERK,JNK,p38 and MUC5 AC mRNA in bronchopulmonary tissues of rats in each group were detected by RT-qPCR.3 Results :(1)HE staining results: the bronchial lumen of the normal group was unobstructed,no thickening of the bronchial wall,no inflammatory cell infiltration under the mucosa,no detachment of the cilia,normal alveolar morphology andstructural integrity;Infiltration of inflammatory cells,epithelial cells are largely shed,cilia fluctuate and fall off obviously,tracheal wall thickening is obvious,lumen is narrow,alveolar wall becomes thin or even ruptured,and a large number of fused into bullae;Yifei Jianpi Recipe group and ammonia bromide In the cord group,the local tracheal lumen was slightly deformed,the wall was slightly thickened,a small amount of epithelial cells were necrotic,some cilia were lodging,and the shedding was not obvious.Some alveolar fusion showed a small amount of inflammatory cell infiltration compared with the model group.There were different degrees of improvement,and the improvement of Yifei Jianpi Recipe high dose group and ambroxol group was more obvious.(2)AB-PAS staining results: Only a small amount of goblet cells were observed in the bronchus of the normal group;a large number of goblet cell hyperplasia was observed in the bronchus of the model group,accompanied by more mucus secretion;the middle and low dose groups of Yifei Jianpi were visible.The distribution of goblet cells was reduced compared with the model group;the number of goblet cells was significantly reduced in the high dose group and the ambroxol group.(3)Western blot analysis of EGFR,MUC5 AC protein and ERK1/2,JNK,p38 protein and phosphorylated protein expression in bronchopulmonary tissues of rats in each groupCompared with the normal group,the expression of EGFR,MUC5 AC protein and ERK1/2,JNK,p38 total protein and phosphorylated protein in the bronchopulmonary tissues of the model group were significantly increased(P<0.01).Compared with the model group,the results of the high-dose group of Yifei Jianpi Recipe showed that the protein expression decreased significantly(P<0.01);the expression of EGFR,P-p38 and p38 protein in Yizhong Jianpi Recipe group were obvious.The expressions of ERK1/2,JNK,P-ERK1/2,P-JNK and MUC5 AC were decreased(P<0.05).The expression of EGFR and P-p38 protein in low dose group of Yifei Jianpi Recipe The amount of p38 protein decreased significantly(P<0.01),and the expression of ERK1/2,JNK,P-ERK1/2,P-JNK and MUC5 AC was notstatistically different from the model group.The significance of learning(P>0.05);the expression of EGFR,P-p38 and p38 protein in bronchopulmonary tissues of rats in ambroxol group were significantly decreased(P<0.01),ERK1/2,JNK,P-ERK1/2,P-JNK and MUC5 AC protein expression levels were all decreased(P<0.05).The detection results of EGFR,P-p38,p38,ERK1/2,JNK,P-ERK1/2,P-JNK and MUC5 AC in the high dose group of Yifei Jianpi Recipe showed the lowest results.The four groups were compared with each other: the expression of EGFR and P-p38 protein in the high dose group of Yifei Jianpi Recipe was significantly different from that in the middle and low dose groups(P<0.01),and the difference was compared with the ambroxol group.Statistically significant(P<0.05);the expression levels of p38,ERK1/2,JNK,P-ERK1/2,P-JNK and MUC5 AC in high dose group of Yifei Jianpi Recipe were compared with low dose group of Yifei Jianpi Recipe The difference was statistically significant(P<0.05),and there was no significant difference between the middle dose group and the ambroxol group(P>0.05).(4)RT-qPCR was used to detect the expression of EGFR,ERK,JNK,p38 and MUC5 AC mRNA in bronchopulmonary tissues of rats in each group.Compared with the normal group,the expressions of EGFR,ERK,JNK,p38 and MUC5 AC mRNA in the bronchopulmonary tissues of the model group were significantly increased(P<0.01).Compared with the model group,the expressions of EGFR,JNK and MUC5 AC mRNA in bronchopulmonary tissues of Yifei Jianpi Recipe high-dose group were significantly decreased(P<0.01),and the expression levels of ERK and p38 MAPK mRNA were decreased(P<0.05).The expression of MUC5 AC mRNA in the middle dose group of Yifei Jianpi Recipe was significantly decreased(P<0.01),and the expression of MUC5 AC mRNA in the low dose group of Yifei Jianpi Recipe decreased(P<0.05).The middle and low dose groups of Yifei Jianpi Recipe The expressions of EGFR,JNK,ERK and p38 MAPK mRNA were not significantly different from those in the model group(P>0.05).The expressions of EGFR,JNK,ERK,p38 MAPK and MUC5 AC mRNA in ambroxol group were significantlydecreased(P<0.01).There were significant differences in the expressions of EGFR,JNK,ERK,p38 MAPK and MUC5 AC mRNA between the four groups(P<0.01).The expressions of EGFR,JNK,ERK and p38 MAPK mRNA in high dose group were not significantly different from those in middle dose group and ambroxol group(P>0.05).The expression of MUC5 AC mRNA and middle dose group in high dose group of Yifei Jianpi Recipe The difference was statistically significant(P<0.05),and there was no significant difference compared with ambroxol group(P>0.05).4 Conclusion:(1)Through the experiment to observe the general condition of each group of rats and the interpretation of HE staining and AB-PAS staining results,it has been successfully verified that cigarette smoke exposure combined with intratracheal instillation of lipopolysaccharide(LPS)combined with senna leaf gavage and diarrhea to establish a COPD model of lung and spleen deficiency is scientific and reproducible.(2)Yifei Jianpi Recipe can alleviate the pathological damage of bronchopulmonary tissue and relieve airway remodeling in rats,and the improvement of Yifei Jianpi Recipe high dose group is more obvious.(3)Based on the results of this experimental study,it can be inferred that Yifei Jianpi Recipe can inhibit the activation of various links in the EGFR/MAPK pathway,regulate the expression of MUC5 AC,inhibit the high secretion of airway mucus,and thus reduce airway inflammation. |
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