尘螨过敏性支气管哮喘患者肺泡巨噬细胞表面CD_(86)的表达及抗CD_(86)单抗对其炎性细胞因子生成的作用

CD_(86) expression on alveolar macrophages and effects of anti-CD_(86) monoclonal antibody on the production of inflammatory cytokines in dust-mite allergic asthma patients

目的 观察尘螨过敏性支气管哮喘 (简称哮喘 )患者肺泡巨噬细胞表面CD86的表达 ,探讨抗CD86单抗抑制哮喘炎症反应的机制。方法 从 10例尘螨过敏性哮喘患者支气管肺泡灌洗液(BALF)和外周血中分离出肺泡巨噬细胞 (AM)和CD+ 4 T淋巴细胞 ,AM经螨变应原的刺激活化后与CD+ 4 T淋巴细胞共培养。每例患者的培养细胞分为抗CD86单抗干预组和CD86表达组 ,每组内再分为实验组和对照组。抗CD86单抗干预组、实验组分别加抗CD86单抗至 0 1mg/L(低剂量 )和 1.0mg/L(高剂量 )两个剂量 ,对照组不加相应抗体。共培养 72h后分别留取培养上清液 ,用酶联免疫吸附测定(ELISA)法检测细胞因子γ干扰素 (INF γ)、白细胞介素 4 (IL 4 )、IL 5的产生量。CD86表达组、实验组加螨变应原 ,对照组不加变应原 ,共培养 2 4h后收集培养细胞 ,流式细胞仪 (FCM)检测AM表面CD86分子的表达水平。结果 AM在尘螨变应原的刺激活化下 ,实验组AM表面CD86分子平均荧光密度表达水平为 (5 0± 9) % ,对照组为 (2 3± 5 ) % ,两组比较差异有显著性 (P <0 0 1) ;实验低剂量组IL 4、IL 5的分泌量分别为 (135± 19)ng/L、(10 4± 2 1)ng/L ,实验高剂量组分别为 (90± 17)ng/L、(6 8± 14 )ng/L ,对照组IL 4、IL 5的分泌量分别为 (187± 2 4 )ng/L。

Objective To investigate the CD_ 86 molecule expression on alveolar macrophages(AMs) before and after AM activation with dust-mite allergen and effects of anti-CD_ 86 monoclonal antibody on the production of inflammatory cytokines produced by cultured CD +_4 T cells, and to explore the pathogenesis of anti-CD_ 86 monoclonal antibody on inhibiting asthmatic inflammation. Methods AMs and CD +_4 T cells were isolated from brochoalveolar lavage fluid (BALF) and from peripheral blood derived from 10 asthma patients allergic to house dust-mite dermatophagoides pteronyssinus. AMs were stimulated and activated by house dust-mite allergen and co-cultured with CD +_4 T cells. The co-cultured cells were divided into anti-CD_ 86 monoclonal antibody intervention group and CD_ 86 expression group with which the control groups were set up. The anti-CD_ 86 antibody was added to 0.1 mg/L and 1.0 mg/L in the experiment group and no antibody was added in the control group. The supernatants were harvested after 72 hour co-culture. The concentration of interferon-gamma(INF-γ), interleukin-4(IL-4) and IL-5 were determined by enzyme linked immunosorbent assay(ELISA). CD_ 86 expression group was divided into experiment and control groups,and house dust-mite allergen was added to the experiment group but no allergen was added in the control group. The cells of the two groups were harvested after 24 hour co-culture. The CD_ 86 molecule expression on the AMs was detected by flow cytometry. Results After activated by house dust-mite allergen,the average fluorescence density of CD_ 86 on AM in the experiment group [(50±9)%] was significantly higher than that in the control group [(23±5)%,P <0.01]. The production of IL-4 and IL-5 was (135±19)ng/L,and (104±21) ng/L in the low-dose group respectively;(90±17)ng/L and (68±14) ng/L in the high-dose group;(187±24)ng/L and (161±23)ng/L in the control group;the differences among groups were significant ( P <0.01). The production of IFN-γ was (193±39)ng/L in the low-dose group and (201±47)ng/L in the high-dose group respectively,and there was no significant difference between the two groups( P >0.05). Conclusion AMs may effectively present allergen to T cells in allergic asthma patients. Anti-CD_ 86 monoclonal antibody can inhibit the production of IL-4 and IL-5 by blocking the co-stimulatory signal of CD +_ 4 T cells. These data suggestes that anti-CD_ 86 monoclonal antibody may hold therapeutic potentials in asthma by inhibiting airway inflammation.