Tang Jian-feng, Guan Shu-hong, Wang Zhi-gang
Department of Respiratory Medicine, Third Affiliated Hospital, Soochow University, Changzhou 213000, China.
Zhonghua Yi Xue Za Zhi. 2012 Oct 30;92(40):2851-4.
To explore the roles of interleukin (IL)-10 differentiated peripheral blood monocyte-derived dendritic cell (DC-10) of allergic asthma patients in T-lymphocytes proliferation in vitro.
From January to June 2011, 10 subjects with dust mite allergic asthma treated at Third Affiliated Hospital of Soochow University were enrolled. Their peripheral blood monocytes were isolated by Ficoll-Hypaque solution density gradient centrifugation and adherent method. And the adherent monocytes were routinely cultured with granulocyte-macrophage colony-stimulating factor (GM-CSF)+interleukin-4 (IL-4)+tumor necrosis factor-alpha (TNF-α), stimulated with/without interleukin-10 (IL-10), pulsed with dust mite allergen and finally harvested. The cell surface molecules including CD80, CD83, CD86, human leukocyte antigen (HLA)-DR and immunoglobulin-like transcript 2 (ILT2) were detected by immunofluorescent labeling and flow cytometry. And cellular functions were estimated by detecting the capacities of DC uptake antigens with fluorescein isothiocyanate (FITC)-dextran capturing assay. The IL-10 differentiation DC (DC-10) were cultured with autologous peripheral T cells (DC-10 group), either alone (DC-TNF group) or together (combined group) with autologous immunostimulatory DC (DC-TNF). And the impact of this treatment on T-cell responses was assessed for each donor by 3-(4, 5)-dimethylthiahiazo (-z-y1)-3, 5-di-phenytetrazoliumromide (MTT) assay. The production of interferon-γ (IFN-γ), IL-4, interleukin-5 (IL-5) and interleukin-13 (IL-13) were measured with the quantification of enzyme linked immunosorbent assay (ELISA) kits.
In DC-10, the levels of some mature DC's markers (CD80, CD83, CD86 & HLA-DR) decreased, ILT2 increased and there were the higher capacities of up-taking FITC-dextran particle (72.32%±2.93% vs 54.41%±2.95%, P<0.01). Compared with the DC-TNF group (1.74±0.15), the T cell proliferation of the DC-10 group (1.06±0.18) and that of the combined group (1.34±0.16) were significantly inhibited (P<0.01). The secretion levels of IFN-γ, IL-4, IL-5 and IL-10 were (2998±141), (157±17), (2608±254) and (55±11) ng/L in the DC-10 group versus (3223±203), (149±19), (2465±183) and (88±10) ng/L respectively in the combined group. They were all significantly reduced versus (3639±209), (173±16), (2771±183) and (127±11) ng/L in the DC-TNF group (all P<0.01).
The IL-10-treated human DC may express a tolerogenic phenotype and induce the allergen tolerance of T cells. And the induction of DC-10 represents a promising target for therapeutic intervention of effectively managing the clinical outcomes of allergic asthma.
探讨变应性哮喘患者白细胞介素（IL）-10 分化的外周血单核细胞来源树突状细胞（DC-10）在体外对 T 淋巴细胞增殖的作用。
选取 2011 年 1 月至 6 月在苏州大学附属第三医院就诊的 10 例尘螨变应性哮喘患者。采用 Ficoll-Hypaque 溶液密度梯度离心法和贴壁法分离其外周血单核细胞。将贴壁单核细胞用粒细胞-巨噬细胞集落刺激因子（GM-CSF）+白细胞介素-4（IL-4）+肿瘤坏死因子-α（TNF-α）进行常规培养，分别在有/无白细胞介素-10（IL-10）刺激下，用尘螨变应原脉冲处理，最后收获细胞。通过免疫荧光标记和流式细胞术检测细胞表面分子，包括 CD80、CD83、CD86、人类白细胞抗原（HLA）-DR 和免疫球蛋白样转录物 2（ILT2）。通过异硫氰酸荧光素（FITC）-葡聚糖摄取试验检测 DC 摄取抗原的能力来评估细胞功能。将 IL-10 分化的 DC（DC-10）与自体外周 T 细胞共培养（DC-10 组），或单独（DC-TNF 组）或与自体免疫刺激性 DC（DC-TNF）一起（联合组）培养。通过 3-（4,5）-二甲基噻唑（-z-y1）-3,5-二苯基四氮唑溴盐（MTT）试验评估这种处理对每个供体 T 细胞反应的影响。用酶联免疫吸附测定（ELISA）试剂盒定量检测干扰素-γ（IFN-γ）、IL-4、白细胞介素-5（IL-5）和白细胞介素-13（IL-13）的产生。
在 DC-10 中，一些成熟 DC 标志物（CD80、CD83、CD86 和 HLA-DR）水平降低，ILT2 增加，且摄取 FITC-葡聚糖颗粒的能力更高（72.32%±2.93%对 54.41%±2.95%，P<0.01）。与 DC-TNF 组（1.74±0.15）相比，DC-10 组（1.06±0.18）和联合组（1.34±0.16）的 T 细胞增殖均受到显著抑制（P<0.01）。DC-10 组中 IFN-γ、IL-4、IL-5 和 IL-10 的分泌水平分别为（2998±141）、（157±17）、（2608±254）和（55±11）ng/L，联合组分别为（3223±203）、（149±19）、（2465±183）和（88±10）ng/L。与 DC-TNF 组（3639±209）、（173±16）、（2771±183）和（127±11）ng/L 相比，它们均显著降低（均 P<0.01）。
经 IL-10 处理的人 DC 可能表达一种致耐受性表型并诱导 T 细胞对变应原的耐受性。DC-10 的诱导代表了有效管理变应性哮喘临床结局的治疗干预的一个有前景的靶点。
