Effet Of Transcription Factor T-bet/GATA-3 On Th1/Th2 In Asthmatic Mice And Intervention Of Inactivated Mycobacterium Phlei

Objective To investigate the effect of inhalation of inactivated Mycobacterium phlei on the expression of T-bet/GATA-3 in asthmatic mice,to understand the immunoregulatory effect of Mycobacterium phlei.Methods Twenty-four male Balb/c mices were randomly divided into 3group?8 per group?:control group,asthma group and treatment group.Asthma group and treatment group on day 0,7 and 14 days for 25?g OVA and 1 mg aluminum hydroxide of 0.2 ml mixed suspension of PBS in the mice by intraperitoneal injection.From 21^th and 27^th,the mice inhale with 2%atomizing OVA PBS suspension,30 min a day.Treatment group was inactivated by Mycobacterium phlei from 28^th to 32^nd,1 time a day.Control group were intraperitoneally injected and inhaled with control saline instead of OVA,asthma group with control saline instead of inhaling inactivated Mycobacterium phlei.Observe the behavior and activity changes of the mice in the process of inhaling.The last inhalation the mice were performed invasive airway resistance test within 12 hours and were executed within 24h.The lung tissues and bronchoalveolar lavage fluid?BALF?were harvested.Lung inflammation changes was observed by HE staining and Periodic acid Schiff?PAS?staining.Then consult literature methods pathological semi-quantitative score line to estimate the severity of airway inflammation in mice.The levels of IFN-?and interleukin?IL?-4 in BALF were measured by ELISA.The expressions of T-bet,GATA-3,STAT1,STAT6 mRNA in lung tissue were measured by real time-PCR.The percentage of Th1 and Th2 cells in CD4⁺T cells in lung mononuclear cells was measured by flow cytometry.Results In asthma group,sRaw was significantly higher than the control group?p<0.05?after Mch challenge at 12.5mg/ml,25mg/ml,and 50mg/ml,but also higher than the treatment group?p<0.05?.The treatment group sRaw was significantly higher than the control group after Mch challenge at 25mg/ml and and 50mg/ml?p<0.05?.Compared with control group,the asthma group had significantly decreased IFN-?,STAT1 mRNA,T-betmRNA and protein levels and percentage of Th1 cells?p<0.05?and significantly increased IL-4,STAT6 mRNA levels and protein and percentage of Th2 cells?p<0.05?.Compared with the asthma model group,treatment group had significantly increased IFN-?,STAT1mRNA,T-bet mRNA and protein levels and percentage of Th1 cells?p<0.05?and significantly decreased IL-4,STAT6 mRNA levels and protein levels and percentage of Th2 cells?p<0.05?.Correlation analysis showed that the levels of T-bet mRNA in lung tissue was positively correlated with the percentage of Th1cells?r=0.7,p<0.01?,the levels of GATA-3 mRNA was positively correlated with the percentage of Th2 cells?r=0.76,p<0.01?.Conclusions Inhalation of inactivated Mycobacterium phlei could alleviate airway inflammation and reducing airway responsiveness in asthmatic mice.It can inhibit the secretion of Th2 cytokines by increasing T-bet mRNA expression,while inhibiting GATA-3 mRNA expression at transcriptional level.Objective To investigate the effect of inactivated Mycobacterium phlei on the expression of ??T cells in asthmatic mice,to understand the immunoregulatory effect of Mycobacterium phlei.Methods Twenty-four male Balb/c mices were randomly divided into 3 group?8 per group?: control group,asthma group and treatment group.Asthma group and treatment group on day 0,7 and 14 days for 25?g OVA and 1 mg aluminum hydroxide of 0.2 ml mixed suspension of PBS in the mice by intraperitoneal injection.From 21 th and 27 th,the mice inhale with 2% atomizing OVA PBS suspension,30 min a day.Treatment group was inactivated by Mycobacterium phlei from 28 th to 32 nd,1 time a day.Control group were intraperitoneally injected and inhaled with normal saline instead of OVA,asthma group with normal saline instead of inhaling inactivated.Observe the behavior and activity changes of the mice in the process of inhaling.The last inhalation the mice were executed within 24 h.The lung tissues and bronchoalveolar lavage fluid?BALF?were harvested.Lung inflammation changes was observed by HE staining and Periodic acid Schiff?PAS?staining.The levels of IFN-? and interleukin?IL?-4 in BALF were measured by ELISA.The percentage of IFN-? and IL-4 in ??T cells in lung mononuclear cells was measured by flow cytometry.??T cells were isolation from spleens of asthma mice by magnetic beads,divided into A,B,C group.After 48 h,the expression of IFN-? and IL-4 expression in ??T cells were detemined by Flow Cytometry.Results In vivo,Mycobacterium phlei treatment alleviated lung inflammation,attenuated mucus production.Compared with control group,the asthma group had significantly decreased IFN-?,and percentage of IFN-?+??T cells?p<0.05?and significantly increased IL-4,and percentage of IL-4+??T cells?p<0.05?.Compared with the asthma group,treatment group had significantly increased IFN-? levels and percentage of IFN-?+??T cells?p<0.05?and significantly decreased IL-4 levels and percentage of IL-4+??T cells?p<0.05?.In vitro,the expression of IFN-? reduced and IL-4 increased in group B,compared with group A?p<0.05?;the expression of IFN-? increased and IL-4 reduced in group C,compared with group B?p<0.05?.Conclusions The expression of IL-4 increased and IFN-? decreased in sensitization mice ??T cells,show that ??T cells also exists ??T1/??T2 imbalance.Inactivated Mycobacterium phlei can promote IFN-? and inhibition of IL-4 expression by ??T cells,regulating ??T1/??T2 imbalance.