特异性免疫治疗对屋尘螨致敏哮喘小鼠NKT细胞的影响

Effect of Specific Immunotherapy on the Expression of NKT Cells in House Dust Mite Extracts-Sensitized Murine Model

目的:研究特异性免疫治疗(SIT)对哮喘小鼠自然杀伤T(NKT)细胞的影响。方法:24只BALB/C小鼠随机分为对照组(A组)、哮喘模型组(B组)、哮喘免疫治疗(SIT)组(C组),各8只。通过屋尘螨提取液(HDM)诱导建立哮喘小鼠模型并进行SIT治疗。检测各组小鼠的气道反应性、支气管肺泡灌洗液(BAlF)细胞计数及分类、ELISA检测IL-4、IFN-γ以及应用流式细胞仪检测NKT细胞数目,通过RT-PCR方法检测T-bet和GATA-3mRNA表达水平;HE染色观察小鼠肺组织的改变。结果:与B组相比,C组气道反应性明显下降(P<0.01);BALF中细胞总数及嗜酸性粒细胞(EOS)数显著减少(P<0.01);血清IL-4分泌显著降低(P<0.01),IFN-γ显著升高(P<0.01);NKT细胞数及其成熟型比例明显升高(P<0.05);T-betmRNA表达水平明显升高(P<0.01),且与NKT细胞数及其成熟型比例呈正相关性,GATA-3mRNA表达水平明显降低(P<0.05),且与NKT细胞数及其成熟型比例呈负相关性。B组肺部管腔周围炎性细胞聚集,组织上皮损伤,组织水肿,而C组肺部变应性炎症明显减轻。C组其他各项指标接近A组。结论:哮喘的发生可能与NKT细胞失调相关,通过改变NKT细胞数目及其成熟型比例来调节GATA-3/T-bet的表达可能是SIT治疗哮喘的作用机制之一。

Objective： To investigate the effects of specific immunotherapy on the NKT cells in house dust mite（HDM） extracts-sensitized murine mode.Methods： Twenty-four BALB/C mice were randomly divided into three groups（with 8 in each group）：control group（group A）,asthma group（group B）,SIT group（group C）.The model of asthma were sensitized and challenged by HDM extracts while SIT group were subcutaneously immunized by HDM extracts.Twenty-four hours after the challenge,airway hyperresponsiveness of mice,total cellular score and cell classification in bronchoalveolar lavage fluid（BALF）,the serum level of IL-4 were examined.IFN-γ was detected by ELISA,the number of NKT cells was determined by flow-cytometry,the expression of T-bet and GATA-3 mRNA was detected by RT-PCR,lung inflammation was detected by HE stain.Results： Compared with that in the group B,airway hyperresponsiveness in group C decreased（P0.01）.Specific immunotherapy significantly inhibit inflammation in mouse lung tissue pathological changes;Total cells and eosinophils（EOS） in BALF reduced significantly（P 0.01）;Serum IL-4 was significantly lower（P 0.01）;IFN-γ secretion was significantly higher（P 0.01）;NKT cells number and the proportion of mature cells increased significantly（P 0.05）;The level of T-bet mRNA expression increased significantly（P 0.01）,and it was positively correlated with the number of NKT cells and its proportion of mature;The level of GATA-3mRNA expression were significantly lower（P 0.05）,and it was negatively correlated with the number of NKT cells and its proportion of mature.The interstitial space surrounding the airway lumen was characterized by a densely mixed cellular infiltrate,epithelium tissue damage and tissue edema in group B,while lung inflammation of group C reduced considerably.Each test value of group C was substantially similar with that of group A.Conclusion： The occurrence of asthma may be associated with NKT cell dysfunction.By changing the number of NKT cells and its proportion of mature to regulate the GATA-3 / T-bet expression may be one of the mechanisms by which SIT was in the treatment of asthma.