Effect Of Gaoganjiang On Airway Inflammation In Bronchial Asthma And Its Mechanism

Part1The effect of galangin on airway inflammation in a murine model of asthmaObjective:To investigate the influence of galangin on airway inflammation and airway hyperresponsiveness in a murine model of asthma.Methods:36female BALB/c mice were randomly divided equally into6groups: control, OVA, OVA+dimethyl sulfoxide (DMSO), OVA+GA15(galangin15mg/kg), OVA+GA5(galangin5mg/kg), and OVA+dexamethasone (DXM,3mg/kg). The mice were sensitized on days0and14by intraperitoneal injection of20μg OVA emulsified in2mg aluminum hydroxide gel in a total volume of200μL. On days22,23, and24after initial sensitization, the mice were challenged for30min with an aerosol of1%OVA in0.9%saline. Galangin (15and5mg/kg) or vehicle (20μl1DMSO) was given by intraperitoneal injection from day21to day24. For a drug control,3mg/kg dexamethasone was administered in the same manner. Airway responsiveness to acetylcholine chloride was measured with animal lung function analysis systems24hours after the last OVA challenge. The lung sections were stained with either hematoxylin&eosin to assess the inflammatory cell infiltrates, periodic acid schiff (PAS) to quantify airway global cells. Interleukin (IL)-4、IL-5and IL-13levels in bronchoalveolar lavage fluid (BALF), and OVA-specific immunoglobulin E (IgE) levels in serum were measured by enzyme-linked immunosorbent assay (ELISA). The expression of iNOS and VCAM-1in lungs was evaluated by immunohistochemistry and Western blot. The expression of IκBα and phosphorylated p65in the whole cell extractions of the lung tissue, the expression of p65in the cytoplasmic and nuclear extracts of the lung tissue was detected by Western blot.Results:The airway resistance in the OVA group was obviously increased in a dose-dependent manner by administration of ACh, whereas only a slight increase could be detected in the control group. There were no significant differences in baseline airway resistance among the six groups (P>0.05). The LR generated by the administration of Ach at doses from30-270μg/kg increased dramatically in the OVA group compared with the control mice (Figure2). The OVA group had significantly greater airway resistance than the control group (P<0.05). Treatment with galangin and DXM resulted in a sharp decrease in airway resistance compared with the OVA group (P<0.05). The number of total leukocytes, eosinophils, neutrophils, and lymphocytes were markedly elevated in the OVA group (P<0.05). Treatment with galangin before OVA aerosol challenge dose-dependently prevented this increase (P>0.05). IL-4, IL-5, and IL-13in the BALF and OVA-specific IgE in serum were notably increased in the OVA group compared with the control group (P<0.05). Administration of galangin dose dependently reduced the levels of T-helper type2(Th2) cytokines in BALF and ova-specific IgE in serum compared with those in OVA group mice (P<0.05). OVA-challenged mice developed marked goblet cell hyperplasia and mucus hypersecretion in the lumen of the bronchioles (P<0.05). In contrast, OVA+GA group mice and OVA+DXM group mice showed a reduction in the number of PAS-stained goblet cells (P<0.05). OVA sensitization increased iNOS and VCAM-1expression in the lung compared with saline-challenged mice (P<0.05). iNOS and VCAM-1were dramatically decreased in OVA+GA group and OVA+DXM group mice (P<0.05). OVA challenge induced degradation of IxBa (P<0.05) and phosphorylation of p65subunit (P<0.05), raised the level of p65subunit in the nuclear extracts of the lung tissue (P<0.05), and reduced the level of p65in the cytoplasmic extracts (P<0.05). Galangin dose-dependently reduced degradation of IκBα (P<0.05), phosphorylation of p65(P<0.05), and nuclear translocation of p65(P<0.05). Conclusion:Galangin alleviate the airway inflammation and AHR in a murine model of asthma via negative regulation of NF-κB. Part2Galangin down-regulates TNF-a induced MCP-1, eotaxin, CXCL10and VCAM-1expression via negative regulation of NF-kB in human airway smooth muscle cellsObjective:To investigate whether galangin can down-regulate TNF-α induced monocyte chemotactic protein1(MCP-1), eotaxin, C-X-C motif chemokine ligand-10(CXCL10) and vascular cell adhesion molecule-1(VCAM-1) expression on human airway smooth muscle cells and its mechanism involved.Methods:Human primary airway smooth muscle cells were cultured in vitro. The cytotoxicity of galangin was measured by Cell Counting Kit-8(CCK-8) assay. Real-time PCR was used to detect the effect of galangin on TNF-α induced MCP-1, eotaxin, CXCL10and VCAM-1mRNA expression. Western blot was used to detect the effect of galangin on the activity of NF-κB.Results:Cell viability was not significantly altered by galangin up to20μM (P>0.05), cytotoxicity was observed at50μM and100μuM galangin (P<0.05). MCP-1, Eotaxin, CXCL10, and VCAM-1mRNA expression in normal human airway smooth muscle cells can be induced by10ng/mL TNF-α (P<0.05), and which can be blocked by galangin (P<0.05) as well as inhibitor of IκB kinase β,TPCA-1. Galangin can dose dependently decrease p65nuclear translocation induced by TNF-α (P<0.05).Conclusions:These results may support that galangin exerts its anti-inflammatory activity by suppressing TNF-α-induced expression of MCP-1, eotaxin, CXCL10and VCAM-1via blockage of NF-κB in human airway smooth muscle cells.