The Effects Of PKB/AKt Pathway On Chaipu Decoction Down Regulating The Expression Of TPD52L1 In Asthma Tic Rats

Objective:To investigate the effects of protein kinase B/Akt(PKB/AKt)pathway on Chaipu decoction down regulating the expression of tumor protein D53(TPD52L1)in asthmatic rats.Methods:20 healthy male SD rats,each weighing about(240 ± 10)g,were randomly divided into 4 groups,normal control group,asthma group,chaipu decoction group and chaipu decoction +PKB/AKt pathway inhibitor Wortmannin hereinafter referred to as the chai inhibition group,each group had 6 rats.First of all,asthma group,chaipu decoction group and chai inhibition group were given ovalbumin(OVA)sensitized to excite asthmatic model respectively.Chaipu decoction group was given chaipu decoction(1.5 g/kg)by intragastric administration half an hour before each atomization.Chai inhibition group was added Wortmannin(0.7 mg/kg)based on Chaipu decoction group.Normal control group was given the same amount of physiological saline to sensitization,isopyknic distilled water by intragastric administration in the same conditions.Rats were sacrificed after last inhalation inducing asthma by using OVA within 24 h.HE staining method was used to detect the pathological changes of lung tissue in each group;determinating the expression level of TPD52L1 mRNA and Bcl-2 mRNA by fluorescence quantitative PCR method;immunohistochemical determination was applied to mensurate the expression level of Bcl-2 and TPD52L1 protein;detecting the apoptosis of rats lung tissue in each group by TUNEL.Results: 1.The changes of histopathologicalIn the normal control group,alveolus and airway wall layer had good continuity,the cells were arranged in order,airway wall was no thickening,there was no inflammatory cell,no goblet cell hyperplasia and a little mucus secretion in the lumen.The average thickness of airway in asthmatic group increased significantly,bronchial wall layers were observed in a large number of inflammatory cells,submucosal edema,hyperplastic mucus gland and goblet cells leaded to excessive secretion of mucus in the lumen.Chaipu decoction group and chai inhibition groups were compared with asthma group,the smooth muscle was thicker,the degree of lumen diameter was larger,the level of inflammatory cell infiltration was alleviative and the number of mucus gland was reduced.2.The expression of TPD52L1 and Bcl-2 mRNA by fluorescent quantitative PCR methodThe contents of TPD52L1 and Bcl-2 mRNA in lung tissues of rats in asthma group were significantly increased compared with normal control group(P < 0.01);the chai inhibition group and chaipu decoction group compared with asthma group respectively,the TPD52L1 and Bcl-2 mRNA expression in that two groups were significantly reduced,the difference was statistically significant(P < 0.01);The expression amount of the two genes in the chai inhibition group was lower than chaipu decoction group(P< 0.05);while there was no significant difference between chai inhibition group and the normal group(P > 0.05).3.The expression of TPD52L1 and Bcl-2 protein by immunohistochemical methodThe TPD52L1 and Bcl-2 protein in lung tissue were high expressed in asthma group,which were significantly increased compared with the normal group(P < 0.01);after using chaipu decoction and wortmannin,chaipu decoction group and chai inhibition group were decreased compared with asthma group,the difference was statistically significant(P < 0.01);all of the protein expression in chai inhibition group were less than chaipu decoction group(P < 0.05),while the chai inhibition group had no significant difference compared with the normal group(P > 0.05).4.The apoptosis of lung tissue in rats by TUNELAirway smooth muscle was thickening in lung tissue of asthma group,a large number of inflammatory cells were infiltrated in submucosal,cell apoptosis rate of lung tissue was significantly lower compared with the normal group(P < 0.01);compared with the asthma group,chaipu decoction group and chai inhibition group airway wall was thinner,rate of apoptosis in lung tissue were increased,the apoptotic cells were mainly epithelial cells and eosinophils,there was significant difference(P < 0.01);lung tissue apoptosis rate was increased in chai inhibition group contrast with chaipu decoction group(P < 0.05),The apoptosis of airway epithelial cells in chai inhibition group was more than that in chaipu decoction group,the difference was significant(P < 0.05);when compared with the normal group,the apoptosis rate of lung tissue was also increased in chai inhibition group,the difference was statistically significant(P < 0.01).Conclusion: 1.Chaipu decoction can reduce the content of TPD52L1 and Bcl-2 in asthma rats lung tissue,induce the apoptosis of airway epithelial cells in lung tissue in lung tissue;2.The mechanism of Chaipu decoction down regulating the expression Bcl-2 and TPD52L1 is related to the inhibition of PKB/AKt pathway.