清肺化痰汤联合地塞米松通过调控PI3K/AKT通路上调HDAC2治疗COPD的研究

Qingfei Huatan Decoction(清肺化痰汤)Combined with Dexamethasone in the Treatment of COPD by Regulating the PI3K/AKT Pathway and Up-regulating HDAC2

目的:探讨清肺化痰汤联合地塞米松通过调控PI3K/AKT信号通路上调组蛋白去乙酰化酶2(HDAC2)治疗慢性阻塞性肺疾病(COPD)。方法:将60只SD大鼠随机分为正常对照组、模型对照组、地塞米松0.5 mg/kg组、地塞米松+清肺化痰汤7.5、15、30 g/kg组,运用香烟烟雾(CS)吸入、气道脂多糖滴入建立COPD大鼠模型,造模28 d后给药,连续给药14 d,末次给药24 h后,以小动物肺功能仪测定大鼠呼吸功能;取肺组织,HE染色观察大鼠肺组织炎症程度及病理变化;Masson染色检测大鼠气道重塑情况;ELISA法检测肺组织匀浆中的肿瘤坏死因子(TNF-α)、白介素-1β(IL-1β)的含量;Real-time RT-qPCR法检测大鼠肺组织Hdac2 mRNA的表达;Western blot法检测大鼠肺组织HDAC2、p-PI3K、PI3K、p-AKT、AKT蛋白的表达。结果:与正常对照组相比,模型对照组COPD大鼠肺功能显著下降,大鼠气道周围肺组织结构大量破坏,肺泡隔结构消失,胶原沉积显著增多,TNF-α与IL-1β的含量显著升高,肺组织中Hdac2 mRNA及HDAC2蛋白表达显著下调(P<0.01),p-PI3K及p-AKT蛋白表达明显上调(P<0.05);与模型对照组相比,各给药组的肺功能显著提高(P<0.01),肺组织形态明显改善,肺泡壁变薄,胶原沉积明显降低,TNF-α与IL-1β的含量明显降低,肺组织中Hdac2 mRNA及HDAC2蛋白表达明显上调,p-PI3K及p-AKT蛋白表达明显下调(P<0.05或P<0.01);与地塞米松0.5 mg/kg组相比,地塞米松+清肺化痰汤15 g/kg组肺功能显著提高,肺组织形态明显改善,肺泡壁变薄,胶原沉积率显著降低,TNF-α与IL-1β的含量显著降低(P<0.01),肺组织中Hdac2 mRNA及HDAC2蛋白表达明显上调(P<0.05或P<0.01),p-PI3K蛋白表达明显下调(P<0.05)。结论:清肺化痰汤联合地塞米松治疗COPD优于仅使用地塞米松,可能与调控PI3K/AKT信号通路,上调HDAC2的表达有关。

Objective:To explore the therapeutic effect of Qingfei Huatan Decoction(清肺化痰汤)combined with dexamethasone on chronic obstructive pulmonary disease(COPD)by regulating the PI3K/AKT signaling pathway and up-regulating the expression of histone deacetylase 2(HDAC2).Methods:Sixty SD rats were randomly divided into a normal control group,a model control group,a dexamethasone(0.5 mg/kg)group,and dexamethasone+Qingfei Huatan Decoction(7.5,15,and 30 g/kg)groups.A COPD model was established using cigarette smoke(CS)inhalation and airway lipopolysaccharide injection,and after 28 days of modeling,the rats were treated for 14 consecutive days.24 hours after the last administration,the rats'respiratory function was measured using a small animal lung function meter.Lung tissues were then obtained,and the degree of inflammation and pathological changes in the lung tissues were observed by HE staining.Masson staining was used to detect airway remodeling,while ELISA was used to detect the content of tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)in the lung tissue homogenate.Real-time RT-qPCR was used to detect the expression of Hdac2 mRNA in the rat lung tissues,while Western blot was used to detect the expression of HDAC2,p-PI3K,PI3K,p-AKT,and AKT proteins in the lung tissues.Results:Compared with the normal control group,the model control group had weakened lung function(P<0.O1),with extensive destruction of the lung tissue structure,disappearance of alveolar septal structure,increased collagen deposition(P<0.O1),and elevated levels of TNF-αand IL-1β(P<0.01),and the expression of Hdac2 mRNA and HDAC2 protein in the lung tissues was down-regulated(P<0.01),while the expression of p-PI3K and p-AKT proteins was up-regulated(P<0.05).Compared with the model control group,the treatment groups showed improved lung function(P<0.O1),significant improvement in lung tissue morphology,thinning of alveolar walls,decreased collagen deposition(P<0.05 or P<0.01),and decreased levels of TNF-αand IL-1β(P<0.05 or P<0.01),and the expression of Hdac2 mRNA and HDAC2 protein in the lung tissues was up-regulated(P<0.05 or P<0.01),while the expression of p-PI3K and p-AKT proteins was down-regulated(P<0.05 or P<0.01).Compared with the dexamethasone(0.5 mg/kg)group,the dexamethasone+Qingfei Huatan Decoction(15 g/kg)group showed significantly improved lung function(P<0.01),significant improvement in lung tissue morphology,thinning of alveolar walls,significant decrease in collagen deposition(P<0.01),significant decrease in levels of TNF-αand IL-1β(P<0.01),and up-regulation of Hdac2 mRNA and HDAC2 protein expression in the lung tissues(P<0.05 or P<0.01),while the expression of p-PI3K protein was down-regulated(P<0.05).Conclusion:The combination of Qingfei Huatan Decoction and dexamethasone is superior to dexamethasone alone in the treatment of COPD,which may be related to the upregulation of HDAC2 expression in the PI3K/AKT signaling pathway.