抗敏止嗽颗粒对哮喘小鼠PI3K/Akt通路和Th1/Th2的调控作用研究

Regulation of Anti-Allergic Anti-tussive Granules on PI3K/Akt pathway and Th1/Th2 in asthmatic mice

目的 观察抗敏止嗽颗粒对哮喘小鼠磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)通路和Th1/Th2的影响,探究其作用机制。方法 将48只BALB/c小鼠随机分为空白组、模型组、抗敏止嗽颗粒低剂量组、抗敏止嗽颗粒中剂量组、抗敏止嗽颗粒高剂量组和地塞米松组,每组8只。除空白组外,其余组小鼠均采用卵白蛋白腹腔注射联合雾化吸入方法制备哮喘模型。于实验第15~21天雾化激发前,抗敏止嗽颗粒低、中、高剂量组分别给予0.35 g/kg、0.7 g/kg、1.4 g/kg抗敏止嗽颗粒灌胃,地塞米松组给予地塞米松0.075 mg/kg灌胃,空白组和模型组给予蒸馏水灌胃,均1次/d。实验末次雾化激发24 h后,ELISA法检测血清中白细胞介素-4(IL-4)、干扰素-γ(IFN-γ)水平并计算IFN-γ/IL-4比值,HE和Masson染色观察小鼠肺组织病理形态,Western blot技术检测肺组织中PI3K、p-PI3K、Akt和p-Akt蛋白表达情况,荧光定量PCR技术检测肺组织中PI3K、Akt mRNA表达情况。结果 与空白组比较,模型组血清IL-4水平明显升高(P<0.05),血清IFN-γ水平和IFN-γ/IL-4均明显降低(P均<0.05),肺组织可见大量炎症细胞浸润和胶原沉积,肺组织中p-PI3K、p-Akt蛋白相对表达量和PI3K、Akt mRNA相对表达量均明显升高(P均<0.05);与模型组比较,抗敏止嗽颗粒各组和地塞米松组血清IL-4水平均明显降低(P均<0.05),血清IFN-γ水平和IFN-γ/IL-4均明显升高(P均<0.05),肺组织中炎症细胞浸润和胶原沉积减少,肺组织中p-PI3K、p-Akt蛋白相对表达量和PI3K、Akt mRNA相对表达量均明显降低(P均<0.05)。结论 抗敏止嗽颗粒可以有效调节哮喘导致的Th1/Th2比例失衡,机制可能与抑制PI3K/Akt信号通路相关。

Objective It is to observe the effects of Anti-Allergic Anti-tussive Granules on phosphatidylinositol-3-kinase/protein kinase B(PI3K/Akt)pathway and Th1/Th2 in asthmatic mice,and to investigate its mechanism of action.Methods Forty-eight BALB/c mice were randomly divided into blank group,model group,low-dose,medium-dose,high-dose groups of Anti-Allergic Anti-tussive Granules and dexamethasone group,with 8 mice in each group.Except for the blank group,the mice of all other groups were given ovalbumin(OVA)by intraperitoneal injection combined with nebulization inhalation to establish asthma models.Before nebulization stimulation on the 15th to 21st day of the experiment,the low,medium,and high dose groups of Anti-Allergic Anti-tussive Granules were respectively given Anti-Allergic Anti-tussive Granules 0.35 g/kg,0.7 g/kg,and 1.4 g/kg by gavage,the dexamethasone group was given dexamethasone 0.075 mg/kg by gavage,and the blank group and model group were given distilled water by gavage,all once a day.After 24 hours of the last atomization stimulation in the experiment,the levels of interleukin-4(IL-4)and interferon-γ(IFN-γ)in serum were determined by ELISA,and IFN-γ/IL-4 ratio was calculated,the pathological morphology of lung tissue in the mice were observed by HE and Masson staining,the expression of PI3K,p-PI3K,Akt,and p-Akt proteins in lung tissue were detected by Western blot technology,and the expression of PI3K,Akt mRNA in lung tissue was detected by fluorescence quantitative PCR technology.Results Compared with the blank group,the level of serum IL-4 was significantly increased(P<0.05),and the levels of serum IFN-γand IFN-γ/IL-4 were significantly reduced in the model group(both P<0.05),with a large number of inflammatory cell infiltration and collagen deposition in the lung tissue,the relative expression of p-PI3K and p-Akt proteins and the relative expression of PI3K and Akt mRNA in the lung tissue were significantly increased(all P<0.05);Compared with the model group,the levels of serum IL-4 were significantly decreased(all P<0.05),and the levels of serum IFN-γand IFN-γ/IL-4 were significantly increased in each dose group of Anti-Allergic Anti-tussive Granules and dexamethasone group(all P<0.05),the inflammatory cell infiltration and collagen deposition in the lung tissue were reduced,the relative expression of p-PI3K and p-Akt proteins and the relative expression of PI3K and Akt mRNA in the lung tissue were significantly decreased(all P<0.05).Conclusion Anti-Allergic Anti-tussive Granules can effectively regulate the Th1/Th2 imbalance caused by asthma,and the mechanism may be related to the inhibition of PI3K/Akt signaling pathway.