Developmental regulation of intracellular and surface androgen receptors in T cells
Introduction
Steroid hormones play a critical role in growth, differentation and physiology of cells and, hence also, in development and physiology of multicellular tissues and organisms [1]. Testosterone, for example, is known to exert characteristic effects on T cells, thymus, and the immune system. Thus, testosterone is able to down-regulate autoimmune diseases [2], [3], [4], to increase susceptibility towards numerous infectious diseases [5], [6], [7], and to cause involution of the thymus largely due to elimination of thymic T cells [8], [9], [10], [11]. Currently, our understanding of actions of steroids including testosterone on cells is steadily proving to be much more refined than originally anticipated, since steroids exert their effects not only through the classical genomic signaling pathway, but also through nongenomic signaling pathways [12].
Cell responsiveness to testosterone has been considered until recently to be regulated exclusively by the genomic pathway. This is mediated through the intracellular androgen receptor (iAR), which belongs to the nuclear steroid receptor superfamily acting as ligand-inducible transcription factors [13], [14]. The iAR is a protein of approximately 110 kDa containing several domains for androgen binding, transactivation, DNA binding, nuclear localization and dimerization [15], [16]. The functional active iAR binds testosterone in the cytoplasm and the iAR translocates to the nucleus where it binds to androgen responsive element in target gene promotors causing ultimately activation or repression of transcription [13], [14]. During the past years, there is increasing information available that the iAR is also able to mediate nongenomic actions of testosterone through direct protein–protein interactions, for example, through direct interplay with mitogen-activated protein kinases [17].
A different mode of nongenomic testosterone action is mediated through membrane androgen receptors (mAR) on the surface of cells. Such mAR, first described in rat osteoblasts, belong to the class of membrane receptors coupled to phospholipase C via a pertussis toxin-sensitive G-protein [18]. The functional active mAR mediate rapid rises in [Ca2+]i due to influx of external Ca2+ and/or release of Ca2+ from intracellular Ca2+ stores. Such mAR were also found in splenic T cells [19], [20] as well as in macrophages [21].
The current evidence indicates variable expression patterns of iAR and mAR among different cell types. For instance, osteoblasts contain both functional active mAR [18] and iAR [22]. Macrophages of the cell line IC-21 with ligand-sequestrable mAR do not express any significant amounts of iAR [21]. By contrast, splenic T cells do not only express mAR but also iAR, but the latter appear to be functionally inactive in the genomic pathway [19], [20]. Here, we provide evidence for just a reverse situation in thymic T cells, the predecessors of splenic T cells. Thymic T cells possess functionally active iAR, but do not express any significant amounts of mAR. This indicates that the expression pattern of mAR and iAR can also vary within a given cell type during development.
Section snippets
Mice
Female mice of the inbred strain C57BL/10 were used in all experiments. They were obtained from the animal facilities of the Heinrich-Heine-University (Düsseldorf, Germany) and received the standard diet Nohrlin 10H10 (Nohrlin, Bad Salzuflen, Germany) and water ad libitum. The experiments were approved by the state authorities and followed the German law on animal protection.
Testosterone treatment
Mice, 9–12 weeks old, received subcutaneous injections of 100 μl sesame oil containing 0.9 mg testosterone
Testosterone sensitivity of thymocytes
Circulating testosterone levels in female mice of the inbred strain C57BL/10 at the age of 9–15 weeks vary between 0.15 and 0.36 ng/ml. These levels are raised to approximately 3.84±1.02 ng/ml after treatment with 0.9 mg testosterone twice per week for 3 weeks (Table 1). Incidentally, this level is similar to that of male mice of the same strain [25]. The testosterone treatment causes a dramatic involution of the thymus, which coincides with a decrease in the total number of T cells by about 90% (
Discussion
Testosterone responsiveness of T cells is subjected to more complex regulatory mechanisms than anticipated to date. This view is supported by the present study showing different expression patterns of mAR and iAR in thymic T cells in comparison with splenic T cells.
The testosterone sensitivity of thymic T cells in vivo manifests itself as a decrease in the total number of cells by about 90% after testosterone treatment of female C57BL/10 mice. In accordance, previous studies have revealed a
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