ABSTRACT
ObjectiveTo observe the regulatory effects of Yiqi Wenyang Huwei decoction （YWHD） on autophagy and phosphatidylinositol 3-kinase （PI3K）/protein kinase B（Akt）/mammalian target of rapamycin （mTOR） signaling pathway in asthmatic rats and bronchial epithelial cells （16HBE） and further reveal the mechanism of YWHD in treating bronchial asthma （BA）. MethodForty-eight rats were randomly assigned into normal group， model group， dexamethasone group， and low-， medium-， and high-dose YWHD groups， with 8 rats in each group. The rat model of BA was established by intraperitoneal injection with ovalbumin （OVA） + aluminum hydroxide suspension and atomizing inhalation with OVA for 2 weeks. The normal group was administrated with an equal dose of normal saline. The bronchial maximum airway resistance （Max Rrs） induced by methacholine chloride （Mch） was determined by an animal lung function evaluation system. Enzyme-linked immunosorbent assay （ELISA） was employed to measure the levels of interleukin （IL）-4， IL-13， IL-6， IL-33， IL-25， tumor necrosis factor-α （TNF-α）， and immunoglobulin E （IgE） in the bronchial alveolar lavage fluid. Hematoxylin-eosin （HE） and Masson staining were used for observation of the pathological changes of bronchi in the lung tissue. The immunofluorescence assay was employed to measure the levels of the autophagy-associated proteins LC3B and Beclin1. The IL-13-induced autophagy of 16HBE cells exposed to the YWHD-containing serum was observed， and the autophagy level was traced by mRFP-GFP-LC3 adenovirus infection. The protein levels of LC3Ⅱ/Ⅰ， p-PI3K， p-Akt and p-mTOR were determined by Western blot. ResultCompared with the normal group， the model group showed increased Max Rrs （P<0.01） and elevated levels of IL-4， IL-13， IL-6， IL-33， IL-25， TNF-α， and IgE in the bronchial alveolar lavage fluid （P<0.05，P<0.01）. The modeling caused focal infiltration of inflammatory cells and lymphocytes around bronchus and blood vessels， epithelial goblet cell metaplasia， and increased subepithelial collagen deposition. Furthermore， it up-regulated the protein levels of LC3B and Beclin1 （P<0.01）， promoted the autophagy flux of GFP to mRFP in 16HBE cells induced by IL-13， down-regulated the protein levels of p-PI3K， p-Akt， and p-mTOR， and increased the LC3Ⅱ/Ⅰ ratio （P<0.01）. Compared with the model group， medium- and high-dose YWHD decreased Max Rrs （P<0.01）， lowered the levels of IL-4， IL-13， IL-6， IL-33， IL-25， TNF-α， and IgE in the bronchial alveolar lavage fluid （P<0.05， P<0.01）， and reduced lymphocyte and granulocyte infiltration in bronchi of the lung tissue， epithelial goblet cell metaplasia， and subepithelial collagen deposition. Moreover， they down-regulated the protein levels of LC3B and Beclin1 （P<0.05， P<0.01）， decreased the autophagy flux of GFP to mRFP， up-regulated the protein levels of p-PI3K， p-Ak， and p-mTOR， and decreased the LC3Ⅱ/Ⅰ ratio （P<0.05， P<0.01）. ConclusionYWHD ameliorates airway hyperresponsiveness and airway inflammation and inhibits the autophagy of airway epithelial cells in the lung tissue of BA rats by activating the PI3K/Akt/mTOR signaling pathway.
ABSTRACT
Objective: To observe the effects of Small Qinglong Decoction medicine-containing serum on ASMC proliferation action induced by ET-1.Methods: There were six groups in the experiment: normal group(10% normal control serum),model group(ET-1 added 10% normal control serum),Small Qinglong Decoction high dose group(ET-1 added 10% Small Qinglong Decoction high dose serum),Small Qinglong Decoction middle dose group(ET-1 added 10% Small Qinglong Decoction middle dose serum),Small Qinglong Decoction low dose group(ET-1 added 10% Small Qinglong Decoction low dose serum) and Dexamethasone group(ET-1 added 10% Dexamethasone serum),eight slots every group.ASMC proliferation status of 24h,48h and 72h were detected with MTT chromometry.Results: Compared with model group,ASMC proliferation in Small Qinglong Decoction low dose group medicine-containing serum each stage and middle dose group24h and 72h all had significant difference(P