Effect Of ZhiXiaoPingChuanFang On Eosinophil Apoptosis And Expression Of Controlling Gene In Asthmatic Rats

1 Objective:To investigate the effects of ZhiXiaoPingChuanFang on asthmatic symptom scores,pulmonary overflow amount,tension of tracheal strips in vitro,the count of leukocyte and eosinophil in whole blood and bronchial alveolar lavage fluid(BALF),the content of IgE,IL-5 and IL-13 in serum,Histomorphology of bronchopulmonary tissue,airway remodeling,expression of Fas,Bcl-2mRNA and eosinophil apoptosis in lung tissue.The effective mechanism of ZhiXiaoPingChuanFang on immune response,airway inflammation, airway hyperresponsiveness and airway remodeling of asthma were explored.2 Methods:The rat asthma model was established by sensitization and challenge with ovalbumin(OVA).Sixty SD rats were evenly assigned to six groups at random:normal control group,asthma model group, dexamethasone treated group,low dosage group,middle dosage group and high dosage group of ZhiXiaoPingChuanFang.After the rat models were formed successfully,the decoction of ZhiXiaoPingChuanFang was used for treatment.The asthmatic symptom scores,pulmonary overflow amount,tension of tracheal strips,the count of leukocyte and eosinophil in whole blood and BALF were determined.The content of IgE,IL-5 and IL-13 in serum were detected by enzyme linked immuoabsorbent assay(ELISh).The histomorphology of the bronchopulmonary tissue was observed by the light microscop. Parameters of airway remodeling were determined by medical image analysis system.Apoptosis was detected by TUNEL,the expressions of FasmRNA and Bcl-2 mRNA in the lung tissue were detected in situ hybridization.3 Results:3.1 The general condition and symptoms of rats at the time of provocation:After inhalation of OVA,model rats came forth symptom of hypersensitiveness such as restlessness and scratching fur on their face;and were in tachypnea or dyspnea(visible abdominal breathing) with slight cyanosis.Some of model rats sounded sonorous wheeze during expiratory phase.After repeated provocation, fur of model rats lost luster,depressed and responsed slowly, the weight growth of model group was slow obviously after repeatedly provocation.There was significant difference on the weight between normal group and model group(P＜0.05).Compared with model group, after treated with medicine,the symptoms of each treatment group alleriated obviously.3.2 Asthmatic symptom scores:Compared with model group,the asthmatic symptom scores of each treatment group decreased significantly(P＜0.01 or P＜0.05).3.3 Determination of pulmonary overflow amount and tension of tracheal strips in vitro:Compared with model group,the pulmonary overflow amount and tracheal strips tension of each treatment group decreased significantly(P＜0.01 or P＜0.05). 3.4 Count of leukocyte and eosinophil in whole blood and BALF: Compared with normal group,the count of leukocyte and eosinophil in whole blood and BALF in model group increased significantly(P＜0.01).Compared with model group,the count of leukocyte and eosinophil in whole blood and BALF of each treatment group decreased significantly(P＜0.01 or P＜0.05).Compared with normal group,there was no significant difference on count of leukocyte and eosinophil in whole blood and BALF among dexamethasone group, high dosage group and normal group(P＞0.05).3.5 The content of IgE,IL-5 and IL-13 in serum:Compared with normal group,the content of IgE,IL-5 and IL-13 in model group increased significantly(P＜0.01).Compared with model group,the content of IgE,IL-5 and IL-13 in each treatment group decreased significantly(P＜0.01 or P＜0.05).Compared with normal group, there was no significant difference on IgE in dexamethasone group and high dosage group(P＞0.05),and there was no significant difference on IL-5 and IL-13 between normal group and each treatment group(P＞0.05).3.5 Histomorphology of bronchopulmonary tissue:There were obvious infiltration of inflammatory cells,severe luminal stenosis and thickening of alveolar septa in model group,significant hyperplasias of basement membrane and smooth muscle layer of central and peripheral airways were observed in the bronchopulmonary tissue too.The inflammation in the bronchopulmonary was significantly improved in dexamethasone group,high dosage group and middle dosage group.3.7 Parameters of airway remodeling:Compared with normal group, the thickness of bronchial wall(WAt/Pbm),thickness of bronchial smooth muscle(WAm/Pbm) and the number of airway smooth muscle cells(N/Pbm) increased significantly in model group(P＜0.01). Compared with model group,the parameters of WAt/Pbm,WAm/Pbm and N/Pbm decreased significantly in dexamethasone group,high dosage group and middle dosage group(P＜0.01 or P＜0.05),the parameters of WAt/Pbm and N/Pbm decreased significantly in low dosage group(P＜0.01 or P＜0.05),there was no significant difference on parameters of WAm/Pbm between low dosage group and model group(P＞0.05).3.8 Expression of Fas,Bcl-2mRNA and eosinophil apoptosis in lung tissue:Compared with normal group,the count of eosinophil increased and eosinophil apoptosis rates decreased,the expression of FasmRNA significantly decreased and the expression of Bcl-2mRNA significantly increased in model group(P＜0.01).Compared with model group,eosinophil apoptosis rates and the expression of FasmRNA increased significantly,while the expression Bcl-2mRNA decreased significantly in each treatment group(P＜0.01 or P＜0.05).The mechanism of ZhiXiaoPingChuanFang on the expressions of Fas,Bcl-2mRNA and eosinophil apoptosis was in a dose-dependent manner,A positively significant correlation was found between the FasmRNA expression and eosinophil apoptosis in each group(P＜0.01),and a very significant negative correlation was found between the Bcl-2mRNA expression and eosinophil apoptosis(P＜0.01).4 Conclusion:Sensitization and challenge with OVA could replicate asthma model in rats.ZhiXiaoPingChuanFang could relax the airway smooth muscle directly,relieve the airway spasm,could reduce the count of leukocyte and eosinophil in whole blood and BALF,decrease the content of IgE,IL-5 and IL-13 in serum,promote eosinophil apoptosis,FasmRNA expression,and inhibit Bcl-2mRNA expression. Consequently,by means of reducing immune response and airway inflammation,inhibiting airway hyperresponsiveness and airway remodeling,ZhiXiaoPingChuanFang might play a important role in relieving asthma.