| Objective:This study aims to explore the possible mechanism of Pingchuan Formula in the treatment of asthma by observing its clinical efficacy in the treatment of childhood asthma and its influence on expression of NLRP3 and downstream factors in blood.At the same time,this study explores the effects of Pingchuan Formula on NLRP3 inflammasomes and its related mechanism in airway inflammation and airway remodeling.By comparing the "eliminating stasis" method and "resolving phlegm"method in TCM treatment of asthma,this study explores various adaptations of the academic experience of Xu’s pediatrics to provide reference for future clinical practice.Methods:The prospective cohort study involves 240 children with chronic persistent asthma equally assigned to the exposed group and the non-exposed group.The exposed group was treated with inhaled Pingchuan Formula+Western medicine(ICS+LABA),and the non-exposed group was treated with inhaled Western over a period of 3 months.Changes in the standard of asthma control,C-ACT score changes,TCM syndrome score and lung function(FVC%,FEV1%,FEV1/FVC%,PEF%,FEF25%,FEF50%,FEF75%)were compared between the exposed group and the non-exposed group.RT-PCR was used to detect the expression of NLRP3 mRNA in blood before and after treatment,and IL-1β and IL-18 content was detected using ELISA.Experiment 1:60 balb/c mice were randomly divided into Sham group,MOD group,DEX group,PCF group,QYF group and HTF group,and sensitized with 10%OVA+aluminum-magnesium adjuvant to establish an acute airway inflammation model.Then,the mice were nebulized with 5%OVA and sacrificed 7 days later.General behavior of mice and airway histopathological changes were observed,and expression of NLRP3 and caspase-1 was determined using immunohistochemistry.Expression levels of NLRP3,caspase-1 protein,ASC protein and mRNA in lung tissues,as well as IL-1βand IL-18 levels in BALF were also quantified.Experiment 2:60 balb/c mice were randomly divided into Sham group,MOD group,DEX group,PCF group,QYF group,HTF group,and sensitized with 0.5%OVA+aluminum-magnesium adjuvant to establish an asthma model with airway remodeling.2.5%OVA was nebulized 3 times a week with daily intragastric administration of the respective drugs.The mice were sacrificed 6 weeks later.Airway inflammation and collagen fiber infiltration were visualized after histopathological staining,and expression of NLRP3 and caspase-1 in lung tissue was determined using immunohistochemistry.Expression levels of NLRP3,ASC,caspase-1 protein and mRNA in lung tissue,JNK,p-JNK,MMP-9,T-NC,E-cadherin,N-cadherin,and IL-1β,IL-18 and TGF-β in BALF were also quantified.Results:After treatment,improved asthma control,C-ACT score and lung function indexes,as well as reduced expression of IL-1β,IL-18 and NLRP3 mRNA in blood(P<0.05)are observed in both groups.Improvements in asthma control and TCM syndrome score with Pingchuan Formula+western medicine was better than treatment with western medicine alone(P<0.05.There was no significant difference in C-ACT score changes between both groups after treatment(P>0.05).The FEF75 index of the exposed group was higher than that of the non-exposed group,and the expression levels of NLRP3 mRNA,IL-1β,and IL-18 in blood were lower than those of the non-exposed group(P<0.05).Experiment 1:Compared with the MOD group,Pingchuan Formula and decomposed formula can improve wheezing behavior of mice in acute asthma and reduce inflammation in airways and lung tissue visualized through HE staining.Positive expression of NLRP3 and caspase-1(P<0.05),reduced NLRP3,ASC,caspase-1 protein and gene expression(P<0.05),and reduced BALF levels IL-1β and IL-18 in BALF(P<0.05)are also observed.There was no statistical difference between the phlegm resolving formula and blood stasis resolving formula(P>0.05)Experiment 2:Compared with the MOD group,Pingchuan Formula and decomposed formula can reduce inflammatory cells and collagen fiber infiltration in airway and lung tissue histopathology,reduce NLRP3,ASC,and caspase-1 protein and mRNA expression(P<0.05),reduce expression levels of NLRP3 and caspase-1 determined using immunohi stochemi stry(P<0.05),and reduce BALF levels of IL-1β,IL-18 and TGF-β(P<0.05).It also works through the NLRP3/IL-1β/JNK axis to reduce the expression of p-JNK,as well as relevant airway remodeling related proteins MMP-9,N-cadherin,and T-NC(P<0.05),and increase the expression of E-cadherin(P<0.05).The phlegm resolving formula has a statistical tendency to reduce NLRP3、caspase-1、p-JNK protein expression than the blood stasis resolving formula(P>0.05).Conclusion:Pingchuan formula and western medicine both effectively control asthma attacks and improve the TCM syndrome scores of children.Both treatments promote recovery of small airway function and reduce expression of NLRP3mRNA,IL-1β and IL-18 in blood.The combined treatment of Pingchuan formula and western medicine is more effective than western medicine alone.Animal experiment results show that Pingchuan formula can relieve airway inflammation by inhibiting the activation of NLRP3 inflammasomes and reducing the secretion of downstream inflammatory factors.It is also shown to reduce airway collagen fiber infiltration through the NLRP3/IL-1β/JNK axis,and regulate expression of downstream factors involved in airway remodeling.There is no significant difference between both TCM treatment methods in an acute asthma model;but the phlegm removing method shows greater efficacy than the blood stasis removing method in a chronic asthma model with airway remodeling. |
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