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首页> 外文期刊>Journal of Neuroimmunology: Official Bulletin of the Research Committee on Neuroimmunology of the World Federation of Neurology >Identification of dopamine plasma membrane and vesicular transporters in human peripheral blood lymphocytes.
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Identification of dopamine plasma membrane and vesicular transporters in human peripheral blood lymphocytes.

机译：人外周血淋巴细胞中多巴胺质膜和水泡转运蛋白的鉴定。

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Plasma membrane dopamine transporter (DAT), vesicular monoamine transporters (VMAT) type-1 and -2 and the expression of the dopaminergic markers dopamine and tyrosine hydroxylase were assessed in membranes and/or in cytospin centrifuged human peripheral blood lymphocytes. The radiolabeled DAT ligand [3H]GBR12935 was bound to peripheral lymphocytes in a manner consistent with the specific binding to a dopamine uptake system, with a dissociation constant similar to that found in striatum, but with a lower density of binding sites. On the other hand, no specific binding occurred in cerebellum used as a test tissue not expressing DAT. Western blot analysis using antibodies raised against amino or carboxy terminus of DAT or against VMAT-1 or VMAT-2 revealed labeling of single bands of approximately 76, 55 or 68 KDa, respectively, displaying similar migration characteristics in lymphocytes and test tissues used for comparison. Immunofluorescence revealed that anti-dopamine, anti-tyrosine hydroxylase, anti-DAT, anti-VMAT-1 and anti-VMAT-2 antibodies labeled the total population of cytospin-centrifuged lymphocytes mounted on microscope slides. Confocal laser microscopy demonstrated that dopamine and VMAT-2 immunoreactivity was developed mainly in cytoplasmic punctiform areas likely corresponding to vesicles and to a lower extent was associated to plasma membrane. Tyrosine hydroxylase immunoreactivity was diffused to cytoplasm and to plasma membrane of lymphocytes, whereas DAT and VMAT-1 immunoreactivity were located almost exclusively in lymphocyte plasma membrane and cytoplasm, respectively. Lymphocyte DAT characterized in this study has probably functional relevance as [3H]dopamine was taken up by intact lymphocytes and uptake was inhibited specifically by compounds known to affect dopamine transport. These findings indicate that human peripheral blood lymphocytes possess DAT plasma membrane and VMAT-1 and VMAT-2 transporters. Increasing evidence indicates that dopamine transporter changes may be related to neuronal injury. In view of this assessment of lymphocyte DAT and VMAT transporters can be considered for identifying pathologies characterized by impaired dopaminergic neurotransmission.
机译：评估了膜和/或细胞旋转离心的人外周血淋巴细胞中的质膜多巴胺转运蛋白（DAT），囊泡单胺转运蛋白（VMAT）1和-2以及多巴胺能标记多巴胺和酪氨酸羟化酶的表达。放射性标记的DAT配体[3H] GBR12935以与对多巴胺摄取系统的特异性结合一致的方式与外周淋巴细胞结合，其解离常数类似于纹状体中发现的解离常数，但结合位点的密度较低。另一方面，在用作不表达DAT的测试组织的小脑中没有发生特异性结合。使用针对DAT的氨基或羧基末端或针对VMAT-1或VMAT-2的抗体进行的蛋白质印迹分析表明，分别标记了约76、55或68 KDa的单条带，在淋巴细胞和用于比较的测试组织中显示出相似的迁移特性。免疫荧光显示，抗多巴胺，抗酪氨酸羟化酶，抗DAT，抗VMAT-1和抗VMAT-2抗体标记了载于离心显微镜载玻片上的细胞离心离心淋巴细胞的总数。共聚焦激光显微镜检查表明，多巴胺和VMAT-2免疫反应性主要在可能与囊泡相对应的细胞质点状区域发展，而在较低程度上与质膜有关。酪氨酸羟化酶的免疫反应性扩散到淋巴细胞的细胞质和质膜，而DAT和VMAT-1免疫反应性几乎分别位于淋巴细胞的质膜和细胞质中。这项研究中表征的淋巴细胞DAT可能与功能相关，因为[3H]多巴胺被完整的淋巴细胞吸收，而吸收被已知影响多巴胺转运的化合物特异性抑制。这些发现表明人外周血淋巴细胞具有DAT质膜和VMAT-1和VMAT-2转运蛋白。越来越多的证据表明，多巴胺转运蛋白的变化可能与神经元损伤有关。考虑到这种评估，可以考虑将淋巴细胞DAT和VMAT转运蛋白用于鉴定以多巴胺能神经传递受损为特征的病理。

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