POD-1/Tcf21 overexpression reduces endogenous SF-1 and StAR expression in rat adrenal cells

During gonad and adrenal development, the POD-1/capsulin/TCF21transcription factor negatively regulates SF-1/NR5A1expression, with higher SF-1 levels being associated with increased adrenal cell proliferation and tumorigenesis. In adrenocortical tumor cells, POD-1 binds to the SF-1 E-box promoter region, decreasing SF-1 expression. However, the modulation of SF-1 expression by POD-1 has not previously been described in normal adrenal cells. Here, we analyzed the basal expression of Pod-1 and Sf-1 in primary cultures of glomerulosa (G) and fasciculata/reticularis (F/R) cells isolated from male Sprague-Dawley rats, and investigated whether POD-1 overexpression modulates the expression of endogenous Sf-1 and its target genes in these cells. POD-1 overexpression, following the transfection of pCMVMycPod-1, significantly decreased the endogenous levels of Sf-1 mRNA and protein in F/R cells, but not in G cells, and also decreased the expression of the SF-1 target StAR in F/R cells. In G cells overexpressing POD-1, no modulation of the expression of SF-1 targets, StAR and CYP11B2, was observed. Our data showing that G and F/R cells respond differently to ectopic POD-1 expression emphasize the functional differences between the outer and inner zones of the adrenal cortex, and support the hypothesis that SF-1 is regulated by POD-1/Tcf21 in normal adrenocortical cells lacking the alterations in cellular physiology found in tumor cells.在性腺与肾上腺发育过程中，POD-1/包膜素/TCF21转录因子（POD-1/capsulin/TCF21）负调控SF-1/核受体亚家族5组A成员1（SF-1/NR5A1）的表达；SF-1水平升高与肾上腺细胞增殖亢进及肿瘤发生风险增加密切相关。在肾上腺皮质肿瘤细胞中，POD-1可结合SF-1的E-box启动子区域，下调SF-1的表达。然而，此前尚未见关于POD-1在正常肾上腺细胞中调控SF-1表达的相关报道。本研究以雄性斯普拉-道来大鼠（Sprague-Dawley rats）分离获得的肾上腺球状带（G）与束状带/网状带（F/R）细胞原代培养物为样本，分析了Pod-1与Sf-1的基础表达水平，并探究了POD-1过表达对该类细胞中内源性Sf-1及其靶基因表达的调控作用。通过转染pCMVMycPod-1质粒实现POD-1过表达后，可显著降低束状带/网状带细胞中内源性Sf-1的mRNA与蛋白水平，而对球状带细胞无此调控效果；同时，该操作还下调了SF-1靶基因类固醇急性调节蛋白（StAR）在束状带/网状带细胞中的表达。在过表达POD-1的球状带细胞中，未检测到SF-1靶基因类固醇急性调节蛋白（StAR）与细胞色素P450家族11亚家族B成员2（CYP11B2）的表达发生变化。本研究结果显示，球状带与束状带/网状带细胞对异位POD-1表达的响应存在显著差异，这进一步凸显了肾上腺皮质外层与内层区域的功能异质性，并支持了“在未出现肿瘤细胞所特有的细胞生理改变的正常肾上腺皮质细胞中，SF-1可受POD-1/Tcf21调控”这一假说。