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Objective To investigate the effect of budesonide ( BUD) inhalation on the proliferation and apoptosis of airway smooth muscle cells ( ASMCs) in asthmatic rats and its molecular biological mechanism. Methods Totally 40 SD rats were randomly divided into control group, asthma model group, low (0. 25 mg/kg) and high (2 mg/kg) BUD group. The rat asthma model was induced by ovalbumin (VOA) combined with aluminium hydroxide Gel sensitization stimulation. After sensitization, the intervention group inhaled different doses of BUD before stimulation. The related parameters of lung tissue and airway were measured and calculated by medical image analysis system, immunofluorescence was used to detect the expression of type I collagen ( Col I ) and Col III in rat airway smooth muscle ( ASM ), and the protein expressions of Bcl-2, Bax, Caspase-3, phosphorylated ERK 1 and 2(p-ERK 1 / 2), p-p38 MAPK were detected by Western blotting. The proliferation activity of ASMCs was detected by MTT method, and the apoptosis rate of ASMCs was detected by flow cytometry. Results Compared with the control group, airway remodeling occurred in the asthmatic model group, and the airway wall thickness ( WAt/Pbm ), inner wall thickness ( WAi/Pbm ) and smooth muscle thickness ( WAm/Pbm ) increased, compared with the model group, the airway remodeling was inhibited in BUD intervention group, and the tracheal WAt/Pbm, WAi/Pbm and WAm/Pbm decreased in bud treatment group. BUD could decrease the proliferation activity of ASMCs, increase the apoptosis rate of ASMCs, inhibit the expression of Col I and Col III, deregulate the expression of Bcl-2, upregulate the expression of Bax and Caspase-3 ( all P<0. 05), and inhibit the activity of ERK 1/2 and p38 MAPK signal pathway. Conclusion BUD can inhibit the proliferation and the promote apoptosis of ASMCs in asthmatic rats, which may be related to the inhibition of ERK 1/2 and p38 MAPK signal pathways.
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Objective To explore the mechanism of azithromycin (AZM) for inhibiting the proliferation of rat airway smooth muscle cells (ASMCs).Methods Thirty Sprague-Dawley (SD) rats were divided into the control group,asthma model group and AZM group.The rat model of asthma was established by ovalbumin (OVA) sensitization and stimulation in vitro.The airway related parameters of rat lung tissue were determined by using the medical image analysis system.Primary passage ASMCs were isolated and cultured using the tissue-sticking method,and the vascular endothelial growth factor (VEGF) overexpression vector or tumor necrosis factor receptor-associated factor 6 (TRAF6) overexpression vector was transfected into ASMCs in the AZM group.The protein levels of VEGF,NF-κB p65 and TRAF6 were detected by Western blotting,and the proliferation of ASMCs was evaluated by CCK-8 kit.Results AZM significantly inhibited the increase of thickness of total airway wall,thickness of inner airway wall and thickness of airway smooth muscle layer in asthma rats (P<0.05),also significantly inhibited the proliferation of ASMCs in the asthma model group (P<0.05).AZM significantly inhibited the protein expression of VEGF and NF-κB p65 induced by asthma (P<0.05),and the overexpression of VEGF significantly reduced the inhibiting effects of AZM on proliferation of ASMCs (P<0.05).AZM significantly inhibited the high expression of TRAF6 induced by asthma (P<0.05),and the overexpression of TRAF6 significantly reduced the inhibiting effects of AZM on expression of VEGF and NF-κB p65 as well as proliferation of ASMCs (P<0.05).Conclusion AZM can suppress the proliferation of ASMCs,its partial mechanism may be realized through inhibiting TRAF6/NF-κB/VEGF signaling pathway.
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Hyperplasia of smooth muscle cells is a major aspect of airway remodeling in asthma .In recent years.It is found that the epigenetic regulation plays an important role on the proliferation of airway smooth muscle cells and the secretion of inflammatory cytokines , including DNA methyltransferase inhibitors to inhibit the phenotype trans-formation;histone acetylation related with hypertrophy .In addition , the microRNA is potentially related with the regulation of a variety of physiological functions of airway smooth muscle cells of asthma model , including inhibition of proliferation and release of inflammatory factors .Hope that epigenetics can become a new target for the treatment of asthma .
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Objective Investigate the effect of substance P and receptor antagonist on the current of NCX in ASMC.Methods Primary rat ASMC cultures were established.Immunofluorescence was used to identify ASMCs.Patch clamp exam was used to assess NCX currents in ASMC before and after intervented by Ach,substance P,NK-1R receptor antagonist and nimodiping,Voltage-current curves were drawn according to variation of voltage and current.PA indicates current amplitude,PF indicates cell area,and PA/PF indicates current density.Results As the voltage increased,the current density increased.When voltage up to-40mV the reverse current appeard.When the voltage is 60mV the current density in the Ach intervention group was highest,but lowest in the nimodipine intervention group;the current density in the substance P intervention group is more than in control group but lower than in Ach interevention group.The current density in the substance P receptor antagonist intervention group higher than nimodipine interevention goup,but lower than the other group (P ＜0.05).Conclusion In asthma airways,Ca2 + overload is related to the appearance of reverse-mode NCX.NK-1R antagonist plays a role in decreasing the Ca2 + concentration,which can relieve the current of reverse-mode NCX,and may benefit alleviating airway inflammation and responsiveness.As a result,NK-1R antagonist may be an attractive target for therapeutic approaches to asthma.
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Objective To investigate the effect of neuropeptide substance P on airway smooth muscle cell contraction amplitude.Methods According to random method, 10 Wistar rats were divided into normal group and asthmatic group.By inhaled OVA to make asthmatic rat model;primary culture ASMC;confocal microscopy were used to observe the morphological changes and measure the length before and after different intervention.The percent of contraction length come from different group ASMC were used for statistical analysis.Results The ASMCs volume in acetylcholine intervented group and substance P intervented group decreased significantly,cell diameters shorten, cytoplasm reduction and cell arranged densely.The ASMCs volume in substance P receptor antagonist intervented group and nimodipine intervented group are about the same size as the ones in normal control group, were spindle-shaped, abundant cytoplasm and arrangement regularly.The contraction length percent of Ach intervened group is the biggest(19.60 ± 3.47) ％, contraction length percent of nimodiping intervented group is the shortest(3.25 ± 1.14)％ ,the contraction length percent in substance Precepter antagonist intervented group is bigger than the one in control group (3.54 ± 1.26) ％, but less than the one in Ach intervented group, asthmatic (14.36 ± 2.37) ％ and substance P intervened group (17.79 ± 3.19) ％.Conclusion Substance P can increase the amplitude of airway smooth muscle cell contraction, but the effect less than Ach;substance P receptor antagonists can inhibit smooth muscle cell contractility, but the effect less than nimodipine.Substance P participates in acute attack of asthma, increases airway reactivity by increasing airway smooth muscle contraction intensity.
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Asthma is a extensive airway limited disease caused by allergens or other factors and emerging under bronchial hyper-responsiveness,which is a common chronic respiratory disease.Airway inflammation and airway remodeling is the main feature of bronchial asthma,and airway remodeling is the main pathological basis causing the irreversible airflow obstruction and pulmonary function injury of asthma.The main pathological change of airway remodeling is airway smooth muscle layer thickening induced by airway smooth muscle cell (ASMC) proliferation,and the mast and increasing number of ASMC under the action of various cytokines is the main cause of smooth muscle layer thickening.The function of airway smooth muscle cells in the airway remodeling is described.
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Asthma is an important respiratory disease that endangers human health, while its complex mechanisms of pathobiology have not been fully understood. Recently, environmental factors are increasingly recognized to play important roles in the pathogenesis of asthma. In particular, the effect of PM2.5 (particulate matter with diameter smaller than 2.5 μm) on the structure and function of pulmonary airways at cell level has become a research hotspot and frontier, and led to many important findings. In this article, the main pathological features, i.e. airway inflammation and hyperresponsiveness were discussed, and recent progress and important findings in pathological effects of PM2.5 on the airway and its mechanism were reviewed, including PM2.5 transport and deposition in the airway, PM2.5 and airway inflammation and damage, PM2.5 and airway remodeling, PM2.5 and airway hyperresponsiveness, PM2.5 and airway smooth muscle cell mechanics via either indirect regulation or direct interaction. The analysis on the role of PM2.5 in airway biomechanics in relation to asthma pathobiology will provide a valuable reference for studying effects of PM2.5 on the respiratory system.
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ObjectiveTo explore the effect of TLR4,PI3K and NF-κB on the migration of asthmatic airway smooth muscle cell(ASMCs) induced by airway epithelial cells.MethodsPrimary ASMCs were cultured by the method of cell digestion.Cell culture supernatant of RTE cells were collected by TNF-α stimulation of epithelial cells.Detected the IL-8 and RANTES levels in the supernatant.The effect of TLR4/PI3K-related signaling molecules on the migration of asthmatic ASMCs induced by epithelial cells were detected by Modified Boyden chemotaxis chamber with anti-TLR4 antibody,Wortmannin and PDTC drugs as a tool.ResultsThe levels of IL-8 and RANTES in the supernatant of TNF-αgroups were significantly increased,and that in the 20 ng/ml group was significantly higher than other groups ( P＜0.01 ).Compared with control group,the transmembrane migration of asthmatic ASMCs from other groups was significantly increased (P＜0.01 ).The transmembrane migration of asthmatic ASMCs from treated groups was significantly increased than asthma group (P＜0.01).The migration of asthmatic ASMCs from TNF-α+anti-TLR4 antibody group,TNF-α+Wortmannin group and TNF-α+Wortmannin+PDTC were significantly decreased than that of TNF-αgroup( P＜0.01 ).The migration of asthmatic ASMCs from TNF-α+Wortmannin+PDTC were significantly decreased than that of TNF-α+Wortmannin group (P＜0.05).ConclusionTLR4/PI3K-related signaling molecules involved in the transmembrane migration of asthmatic ASMCs induced by airway epithelial cells and may be one of the mechanisms of airway remodeling of asthma.
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Objective: To examine the serum level of IL-22 in asthmatic patients and the expression of IL-22R1 in human airway epithelial cells, human airway smooth muscle cells, and lung fibroblasts, so as to explore the target cells of IL-22. Methods: The serum levels of IL-22 and IL-17 in 36 asthmatic patients and 20 normal control subjects were measured by enzyme-linked immunosorbent assay (ELISA). And lung function of the asthmatic patients was assessed by Gaeger spirometry. According to the values of FEV1/FVC and FEV1%, the patients were divided into two groups, bronchodilation test positive group (19 patients) and bronchial provocation test positive group (17 patients). The expression of IL-22R1 mRNA in human airway epithelial cells, human airway smooth muscle cells, and lung fibroblasts was examined using real-time PCR, and IL-22R1 protein expression was detected by immunofluorescence staining and Western blotting analysis. Results: There was no significant difference in serum IL-22 and IL-17 levels between the asthmatic patients and normal controls. The serum IL-22 and IL-17 levels in asthmatic patients positive for bronchodilation test was significantly higher than those positive for bronchial provocation test(P<0.05). IL-22R1 mRNA and protein were detected in all the 3 types of cells. Conclusion: IL-22 may be involved in the pathogenesis of asthma, and human airway epithelial cells, human airway smooth muscle cells, and lung fibroblasts may all be the target cells of IL-22.
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Objective To investigate the expression and function of Fca receptor on human airway smooth muscle(ASM)cells.Methods RT-PCR and immunofluorescence technique were used to detect the expression of Fca receptor on human ASM cells.The effect of IgA on intracellular calcium concentration of human ASM cells was measured by using Fura-2/AM as a calcium indicator.Results RT-PCR and immunofluorescence staining confirmed the presence of Fca receptor on human ASM cells.Compared with control,secretory IgA(sIgA)induced a rise in intracellular calcium concentrations on human ASM cells after 90min incubation(P0.05).Conclusion Fca receptor was expressed on human ASM cells.SIgA increased the intracellular calcium concentration of human ASM cells.
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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
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Objective:To investigate the promoting effects of interleukin-13(IL-13)on the proliferation of airway smooth muscle cells(ASMC)in mice and its mechanism.Methods:Murine ASMC were isolated and subcultured.IL-13,IL-13plus AG1478,IL-13plus neutralizing antibody to TGF-?,IL-13plus neutralizing antibody to EGF,IL-13plus neutralizing antibody to HB-EGF,IL-13plus dexamethasone were added to the media,respectively.MTT and 3 H-TdR incorporation assays were used.Apical media were analyzed for the presence of soluble TGF-?using ELISA.Results:The D 595 value of MTT assay and the cpm value of 3 H-TdR incorporation assay of IL-13group were significantly higher than those of control group(P