Dehydroepiandrosterone and its 7-hydroxylated metabolites do not interfere with the transactivation and cellular trafficking of the glucocorticoid receptor
详细信息 查看全文
- 作者：Muller ; Caroline ; Cluzeaud ; Fran??oise ; Pinon ; Gregory M. ; Rafestin-Oblin ; Marie-Edith ; et. al.
- 关键词：7????-Hydroxy-DHEA ; 7????-Hydroxy-DHEA ; Cortisol ; Dexamethasone ; COS-7 cells ; Transient transfection
- 刊名：Journal of Steroid Biochemistry and Molecular Biology
- 出版年：2004
- 出版时间：December, 2004
- 年：2004
- 卷：92
- 期：5
- 页码：469-476
- 全文大小：305 K
文摘
The human brain is a target tissue for glucocorticoids (GC). Dehydroepiandrosterone (DHEA) is a neurosteroid produced in the brain where it is transformed into 7α-hydroxy-DHEA and 7β-hydroxy-DHEA. The antiglucocorticoid effects of both 7-hydroxylated metabolites have been investigated with evidence in mice that neither form of DHEA interfered with the binding of GC to its glucocorticoid receptor (GR), but contributed to a decreased nuclear uptake of the activated GR. Our objective was to use COS-7 cell culture to research DHEA, 7α-hydroxy-DHEA and 7β-hydroxy-DHEA interferences with GR trafficking. These cells did not carry out the 7α-hydroxylation of DHEA and the oxidation of cortisol into cortisone. The cDNA of the human GR was inserted into pcDNA3 for a transient transfection of COS-7 cells. Human GR transactivation activity was measured from a luciferase-MMTV reporter gene. The transfected COS-7 cells were cultured using 10−12 to 10−5 M dexamethasone (DEX) or cortisol, which triggered the reporter expression. Treatment with 10−12 to 10−5 M DHEA, 7α-hydroxy-DHEA and 7β-hydroxy-DHEA caused no change in the GC-induced GR transactivation. A reconstruction of the process associated EGFP to the human GR cDNA. Confocal microscopic examination of COS-7 cells transiently expressing the fusion protein EGFP-GR showed nuclear fluorescence 60 min after incubation with 10−8 M DEX or cortisol. The addition of 10−5 M DHEA, 7α-hydroxy-DHEA or 7β-hydroxy-DHEA did not change its kinesis and intensity. These results contribute to the knowledge of DHEA, 7α-hydroxy-DHEA and 7β-hydroxy-DHEA, in relation to antiglucocorticoid activity. We conclude that direct interference with GR trafficking can be discounted in the case of these hormones, therefore proposing new possibilities of investigation.