| objectives:Through detecting the change of CD4+CD25+regulatory T cells and the expression of TLR4 on CD4+CD25+regulatory T cells in mouse with asthma,To observe The effect of glucocorticoids on CD4+CD25+ regulatory T cells and TLR4 expression on CD4+CD25+regulatory T cells in a murine model of asthma; Assessment of early Mycobacterium phlei intervention on CD4+CD25+regulatory T cells and the expression of TLR4 in asthmatic mice investigate the effects of Mycobacterium phlei in immune regulation, to find new ways to better prevention and treatment of asthma and provide experimental Basis. Methods:Forty female BALB/c mice were divided into four groups according to random digits table. They are control group (NS), the asthma group (OVA), asthma Dexamethasone treatment group (OVA/GC) and asthma Mycobacterium phlei F.U.36 treatment group (OVA/Utilin). the mice of OVA group, OVA/GC group and OVA/Utilin group undergone ovalbumin(OVA) and aluminum hydrate(Al(OH)3) sensitization by i.p. on day 0 and day 14,and Ondays 21～25, they were challenged by atomization with 2%OVA for 30 minutes a day, OVA/GC group were injected intraperitoneally with dexamethasone (1mg/Kg) before 30 minutes of every challenge;OVA/Utilin group were injected intraperitoneally with mycobacterium phlei F.U.36 injection 0.57ug total of three times every other day, before 2 weeks of the first sensitization;NS group undergone physiologic saline by i.p. and atomization as compared group.After 24 hours of the last challenge, all mice were killed. Left lung tissue of mice for pathology, Airway inflammation were observed with HE staining method.Spleen cells were grinded on cell sieve, isolated mononuclear cells from it with mouse lymphocyte isolation liquid, and we obtained mononuclear cells suspension, The percentage of CD4+CD25+regulatory T cells in CD4+T cells and the mean fluorescence intensity of TLR4 on CD4+CD25+regulatory T cells was detected by flow cytometry. Results:1) successfully established murine asthma model, The mice of OVA group demonstrated similar symptoms of human asthma when exposed to aerosolized OVA, such as breathing deeply and fast, standing still, bowing the back, urinary and fecal incontinence, and so on;the mice of control group didn't show these symptoms of acute asthma. Pathology showed:NS group is normal, Airway and perivascular are visible multifocal mainly eosinophils increase in inflammatory cell infiltrates in OVA group,the inflammation in OVA/GC and OVA/Utilin group is lessened.2) The flow cytometry results showed that:(1) the percentage of CD4+CD25+regulatory T cells in CD4+T lymphocyte subsets in OVA group was significantly lower than those in NS group, OVA/ GC group and OVA/Utilin group significantly increased compared with OVA group, the difference was statistically significant (P<0.01), the percentage of CD4+CD25+regulatory T cell in OVA/GC group and OVA Utilin group was slightly higher than that in NS group(P>0.05), there was no significant difference between OVA/Utilin group and OVA/GC group (P>0.05);(2) the mean fluorescence intensity of TLR4 on CD4+CD25+ regulatory T cells:OVA group compared with NS group were decreased, but the difference was not statistically significant (P>0.05); OVA/GC group and OVA/Utilin group were significantly higher than the OVA(P<0.01); OVA/ GC group compared with OVA/Utilin group was slightly lower, but the difference was not statistically significant (P>0.05); (3) the percentage of CD4+CD25+regulatory T cells in splenic mononuclear cells and the mean fluorescence intensity of TLR4 on CD4+CD25+regulatory T cells had no correlation(P>0.05). Conclusion:1) CD4+CD25+regulatory T cells involved in immune homeostasis of asthma, reducing their number and dysfunction can lead to and aggravate the symptoms of asthma. OVA can not change TLR4 expression, the TLR4 on CD4+CD25+regulatory T cell has no effect in OVA-induced asthma.2) glucocorticoids can promote CD4+CD25+regulatory T cell numbers increase and TLR4 expression in asthma mice may be one of the immunological mechanisms in the treatment of asthma.3) Early Mycobacterium phlei intervention treatment, through promotion of TLR4 expression on CD4 CD25+cells and CD4+CD25+regulatory T cells increase in the number,correct the body's immune disorders play its therapeutic effects, there may be new ways for future prevention and treatment of asthma.4) Asthma is a complex immune network in vivo, glucocorticoids and Mycobacterium phlei on regulation of CD4+CD25+regulatory T cells and the expression of TLR4 is affected by many factors. |
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