Department of Immunology, Institute of Biomedical Sciences, University of São Paulo, Brazil.
Department of Immunology, Institute of Biomedical Sciences, University of São Paulo, Brazil
J Leukoc Biol. 2014 May;95(5):755-762. doi: 10.1189/jlb.0413239. Epub 2014 Jan 16.
CD83 is a marker of mDCs directly related to their lymphostimulatory ability. Some data suggest that it has a central role in the immune system regulation, but how this function is performed remains to be determined. This work aimed to analyze the influence of CD83, present in mDCs, in the modulation of calcium signaling in T lymphocytes. Mo were differentiated into iDCs and activated with TNF-α. iDCs were treated, 4 h before activation, with siRNA, to reduce CD83 expression. Purified allogeneic T lymphocytes were labeled with the calcium indicator Fluo-4-AM, and calcium mobilization in the presence of mDCs was analyzed. CD83 knockdown mDCs induced lower calcium signal amplitude in T lymphocytes (29.0±10.0) compared with siRNA-treated mDCs (45.5±5.3). In another set of experiments, surface mDC CD83 was blocked with a specific mAb, and again, decreased calcium signaling in T lymphocytes was detected by flow cytometry and microscopy (fluorescence and confocal). In the presence of antibody, the percentage of responding T cells was reduced from 58.14% to 34.29%. As expected, anti-CD83 antibodies also reduced the proliferation of T lymphocytes (as assessed by CFSE dilution). Finally, in the absence of extracellular calcium, CD83 antibodies abrogated T cell signaling induced by allogeneic mDCs, suggesting that the presence of CD83 in mDC membranes enhances T lymphocyte proliferation by boosting calcium release from intracellular stores in these cells.
CD83 是与 mDC 淋刺激能力直接相关的标志物。一些数据表明，它在免疫系统调节中具有核心作用，但这种功能如何发挥作用仍有待确定。本工作旨在分析 mDC 中存在的 CD83 对 T 淋巴细胞钙信号转导的调节作用。单核细胞分化为未成熟树突状细胞（iDC），并通过 TNF-α 激活。在激活前 4 小时，用 siRNA 处理 iDC，以降低 CD83 的表达。用钙指示剂 Fluo-4-AM 标记纯化的同种异体 T 淋巴细胞，并分析 mDC 存在时钙动员情况。与 siRNA 处理的 mDC 相比，CD83 敲低 mDC 诱导的 T 淋巴细胞钙信号幅度较低（29.0±10.0）。在另一组实验中，用特异性 mAb 阻断表面 mDC CD83，通过流式细胞术和显微镜（荧光和共聚焦）检测到 T 淋巴细胞钙信号再次降低。在抗体存在的情况下，响应 T 细胞的百分比从 58.14%降至 34.29%。正如预期的那样，抗 CD83 抗体也降低了 T 淋巴细胞的增殖（如 CFSE 稀释所评估的）。最后，在没有细胞外钙的情况下，抗 CD83 抗体阻断了同种异体 mDC 诱导的 T 细胞信号转导，这表明 mDC 膜中 CD83 的存在通过增强这些细胞内钙库的钙释放来增强 T 淋巴细胞的增殖。
