Benten W P, Lieberherr M, Giese G, Wrehlke C, Stamm O, Sekeris C E, Mossmann H, Wunderlich F
Division of Molecular Parasitology and Centre of Biological-Medical Research, Heinrich Heine University, 40225 Duesseldorf, Germany.
FASEB J. 1999 Jan;13(1):123-33. doi: 10.1096/fasebj.13.1.123.
T cells are considered to be unresponsive to testosterone due to the absence of androgen receptors (AR). Here, we demonstrate the testosterone responsiveness of murine splenic T cells in vitro as well as the presence of unconventional cell surface receptors for testosterone and classical intracellular AR. Binding sites for testosterone on the surface of both CD4(+) and CD8(+) subsets of T cells are directly revealed with the impeded ligand testosterone-BSA-FITC by confocal laser scanning microscopy (CLSM) and flow cytometry, respectively. Binding of the plasma membrane impermeable testosterone-BSA conjugate induces a rapid rise (<5 s) in [Ca2+]i of Fura-2-loaded T cells. This rise reflects influx of extracellular Ca2+ through non-voltage-gated and Ni2+-blockable Ca2+ channels of the plasma membrane. The testosterone-BSA-induced Ca2+ import is not affected by cyproterone, a blocker of the AR. In addition, AR are not detectable on the surface of intact T cells when using anti-AR antibodies directed against the amino and carboxy terminus of the AR, although T cells contain AR, as revealed by reverse transcription-polymerase chain reactions and Western blotting. AR can be visualized with the anti-AR antibodies in the cytoplasm of permeabilized T cells by using CLSM, though AR are not detectable in cytosol fractions when using the charcoal binding assay with 3H-R1881 as ligand. Cytoplasmic AR do not translocate to the nucleus of T cells in the presence of testosterone, in contrast to cytoplasmic AR in human cancer LNCaP cells. These findings suggest that the classical AR present in splenic T cells are not active in the genomic pathway. By contrast, the cell surface receptors for testosterone are in a functionally active state, enabling T cells a nongenomic response to testosterone.
由于缺乏雄激素受体（AR），T细胞被认为对睾酮无反应。在此，我们证明了小鼠脾T细胞在体外对睾酮有反应，并且存在睾酮的非常规细胞表面受体和经典的细胞内AR。通过共聚焦激光扫描显微镜（CLSM）和流式细胞术，分别直接揭示了T细胞的CD4(+)和CD8(+)亚群表面上睾酮的结合位点，其中阻碍性配体睾酮 - BSA - FITC用于标记。质膜不可渗透的睾酮 - BSA偶联物的结合会导致负载Fura - 2的T细胞的[Ca2+]i迅速升高（<5秒）。这种升高反映了细胞外Ca2+通过质膜的非电压门控和Ni2+可阻断的Ca2+通道流入。睾酮 - BSA诱导的Ca2+内流不受AR阻滞剂环丙孕酮的影响。此外，当使用针对AR氨基和羧基末端的抗AR抗体时，在完整T细胞表面未检测到AR，尽管通过逆转录 - 聚合酶链反应和蛋白质印迹法显示T细胞含有AR。通过CLSM可以在透化T细胞的细胞质中用抗AR抗体观察到AR，尽管在使用以3H - R1881作为配体的活性炭结合试验时，在细胞溶质组分中未检测到AR。与人类癌症LNCaP细胞中的细胞质AR不同，在睾酮存在的情况下，细胞质AR不会转移到T细胞的细胞核中。这些发现表明，脾T细胞中存在的经典AR在基因组途径中不活跃。相比之下，睾酮的细胞表面受体处于功能活跃状态，使T细胞能够对睾酮产生非基因组反应。
