Anti-asthmatic Effect Of Xuebijing And Potential Role Of Heme Oxygenase-1in An Ovalbumin-induced Murine Allergic Rhinitis Model

Objective:Allergic rhinitis is nasal mucosa of allergic inflammationwhich is susceptible individuals exposed to the allergen, immunoglobulinE-mediated, main symptoms is paroxysmal sneezing, runny nose and nasalcongestion. The final performance is Abundantly expressed Th cells for highexpression of Th2cytokines, eosinophils, and mast cell infiltration, whileinitiate the inflammatory reaction. An important feature of the process ofinflammation is inflammatory cell adhesion,through the vascular endothelium,chemokines infiltration into sites of inflammation. The past50years,the trendof the incidence of allergic rhinitis were increased year by year, foreignreports is in over the past decade, allergic rhinitis prevalence increasedsignificantly.Therefore the refractory disease has become a global healthproblem. The disease involved in many complex factors, the exact mechanismis still unclear, the existing treatment unsatisfactory.FVB mice is the research study to establish the allergic rhinitis mousemodel, submucosal capillary endothelial cells of the nasal in mice is the entrypoint research, expression of HO-1is observed Xuebijing(XBJ) treatmentbefore and after, and to explore therapeutic effect in mice OVA-inducedallergic rhinitis which XBJ treated and its impact on HO-1, provides a newway of thinking for the treatment of allergic rhinitis.Methods: SPF grade mice40(FVB Department) were randomly dividedinto normal control group, the OVA model group, OVA+Dexamethasoneintervention group and OVA+XBJ intervention group, n=10.0VA modelgroup:ovalbumin (OVA)15mg powder was dissolved in saline75ml. TheOVA1.5ml is added hydrogen alumina1.5ml, finally made of10%OVAsuspension. Mice were sensitized with intraperitoneal injection of the theOVA suspension250μl of (OVA25μg) in experimental section l to9days. 10~17days after initial sensitization. OVA+Dexamethasone interventiongroup, OVA+XBJ intervention group model is established: The mice are withintraperitoneal injection of the the OVA suspension250μl of (OVA25μg) inexperimental section l to9days, after the completion of the foundationsensitization, dexamethasone (10mg/kg/d) or XBJ (0.41ml/kg/d) is byintraperitoneal injection, the mice is excited after30min, Animal models arefor behavioral observation. Nose morphological change is observed throughHE staining. The precipitate which nasal lavage fluid was centrifuged countsinflammatory cell. OVA-specific IgE of blood and IL-4, IL-5, IL-13, IFN-γlevels in nasal lavage fluid are detected by Enzyme-linked immunosorbentassay(ELISA). Immunohistochemical staining probe HO-1expression of thenasal mucosa. Immunomagnetic bead technology separate nasal submucosalcapillary endothelial cells of normal mice, subculture, cultured in DMEMmedium contain20%fetal calf serum,capillary endothelial cells are dividedinto the normal control group, the inflammatory model group, XBJintervention group, TNF+Dexamethasone intervention group, TNF+XBJintervention group, Znpp+TNF+XBJ intervention group. Western blot,Real-time PCR detect of expression of HO-1, ICAM-1of capillary endothelialcells. Confocal laser microscopy and flow cytometry detect expression ofROS. All information use SPSS13.0statistical analysis using analysis ofvariance and related analytical processing data, α=0.05as a test, P <0.05considered statistically significant.Results:The OVA model group of10animals were modeling success, the typicalsymptoms of allergic rhinitis is obvious; The symptoms ofOVA+Dexamethasone intervention group, OVA+XBJ intervention groupcompared to the control group have not significant changes.1Hematoxlin-eosin is used to observe morphology change: the normalcontrol group had no significant inflammation in the nasal mucosa; the OVAmodel group show different degree edema, disorder of epithelial cell structure,cup shape cell hyperplasia, infiltration of submucosal eosinophilic acid granulocyte (eosinophils, Eos), lymphocytes cells. The symptoms ofinfiltrating inflammatory cells in the nasal mucosa, and mainly located in OVA+Dexamethasone intervention group, OVA+XBJ intervention groupcompared to normal control group significantly reduced. ELISA detect thelevels of OVA-specific IgE in serum:The levels of the OVA-specific IgE, inOVA model group compared to the normal control group was significantlyhigher, but OVA+Dexamethasone intervention group, OVA+XBJ interventiongroup compared to the OVA model group was significantly lower, thedifference was statistically significant (P<0.05). Inflammatory cell countwhile Nasal lavage fluid pelleted by centrifugation: the OVA model groupeosinophils, neutrophils compared with normal control group wassignificantly higher, but OVA+Dexamethasone intervention group,OVA+XBJ intervention group compared to the OVA model group wassignificantly lower, the difference was statistically significance (P<0.05).Those Prove that the XBJ have a therapeutic effect to OVA-induced allergicrhinitis mice.2ELISA detect the levels of IL-4, IL-5, IL-13, INF-γ in nasal lavagefluid: The levels of the IL-4, IL-5, IL-13in OVA model group compared tothe normal control group was significantly higher, but OVA+Dexamethasoneintervention group, OVA+XBJ intervention group compared to the OVAmodel group was significantly lower, the difference was statisticallysignificant (P<0.05), and the INF-γ level in the OVA model group comparedto the control group was significantly lower, but OVA+Dexamethasoneintervention group, OVA+XBJ intervention group compared to the OVAmodel group was significantly higher, the difference was statisticallysignificant (P<0.05).Those explain that XBJ play a therapeutic role throughregulating Th1/Th2imbalance, to achieve the treatment of OVA-inducedallergic rhinitis mice.3Confocal laser microscopy and flow cytometry detect ROS: ROS in theTNF+Dexamethasone intervention group, the TNF+XBJ intervention groupdecreased significantly compared with the inflammation model group in capillary endothelial cells, the difference was statistically significant (P<0.05).SOD,CAT,GSH detection: SOD,CAT,GSH in the TNF+XBJ interventiongroup compared to the inflammation model group increased significantly, andthe difference was statistically significant (P<0.05). Those illustrate that XBJplay a therapeutic role through reversing the oxidant/antioxidant imbalance,toachieve the treatment of OVA-induced allergic rhinitis mice.4Western blot analysis showed that: after the XBJ intervention incapillary endothelial cells, compared with the control group, ICAM-1expression was decreased; compared to the inflammation model group, theTNF+XBJ interfere with the capillary endothelial cells, ICAM-1alsoexpression decreased.Monocyte adhesion: The TNF+Dexamethasoneintervention group, TNF+XBJ intervention group compared to theinflammation model group, monocyte adhesion to vascular endothelial cellssignificantly reduced, and the difference was statistically significant (P<0.05).Those illustrate that XBJ play a therapeutic role through regulating Th1/Th2imbalances, the oxidant/antioxidant imbalance, inhibiting submucosalcapillary cell activation to achieve the treatment of OVA-induced allergicrhinitis mice.5Immunohistochemistry results showed that: HO-1in the nasal mucosaof OVA model group mice is mainly located in the cytoplasm of epithelialcells and glandular epithelial cells, rare in nuclei, the cytoplasm of stromalvascular endothelial cells and infiltrating inflammatory cells also have apositive coloring expression, which is brown particles. OVA model group,OVA+Dexamethasone intervention group, OVA+XBJ intervention groupcompared with the normal control group, HO-1expression was significantlyhigher, and the difference was statistically significant (P<0.05). Increasing thenasal mucosa of HO-1expression, and thus XBJ play a therapeutic role. HOvitality detection: XBJ intervention group, TNF+XBJ intervention groupcompared to the inflammation of the model group was significantly increased,and the difference was statistically significant (P<0.05). Western blot resultsShow: The capillary endothelial cells were intervened by XBJ, HO-1 expression presents dose-dependent and time-dependent manner; After TNF-αintervene capillary endothelial cells, HO-1expression first high, then low(4hhighest expression); TNF+XBJ to intervene in the capillary endothelial cells,HO-1was longly high expression. Compared to the inflammation modelgroup, the TNF+XBJ interfere with the capillary endothelial cells, ICAM-1expression decreased; Znpp is a blocker of HO-1, after Znpp blockingHO-1expression in the capillary endothelial cells, ICAM-1expression wasremain significantly increased. SOD,CAT,GSH in the Znpp+TNF+XBJintervention group compared to the TNF+XBJ intervention group decreasedsignificantly. Those declare that XBJ play a therapeutic role throughimproving HO-1expression in the nasal submucosal capillary endothelialcells, its expression in a dose-dependent and time-dependent manner,and thusthrough regulating Th1/Th2imbalances,the oxidant/antioxidant imbalance,to achieve the treatment of the OVA-induced allergic rhinitis mice.Conclusion:1XBJ have a therapeutic effect in the OVA-induced allergic rhinitismice.2XBJ play a therapeutic role through through r through regulatingTh1/Th2imbalances the oxidant/antioxidant imbalance, inhibiting submucosalcapillary cell activation to achieve the treatment of the OVA-induced allergicmice.3XBJ play a therapeutic role through inducting HO-1expression in thenasal submucosal capillary endothelial cells, its expression in adose-dependent and time-dependent manner,and thus through regulatingTh1/Th2imbalances the oxidant/antioxidant imbalance, to achieve thetreatment of the OVA-induced allergic rhinitis mice.