雄激素是由肾上腺和生殖细胞产生，借助特定的酶的作用。我们研究了蛋白激酶B (Akt)在小鼠莱迪格肿瘤细胞系mLTC1中调节Δ4类类固醇激素酶活性的作用。细胞以人绒黄体激素（LH）和绒毛膜促性腺激素（hCG）的3倍50％有效浓度处理0-24小时，在存在和缺乏特定的Akt抑制剂3CAI的情况下。通过双荧光共振能量转移（BRET）和Western blotting进行细胞信号分析，同时通过实时PCR调查关键靶基因的表达。通过免疫测定法测定孕酮、17α-羟基（OH）孕酮和睾酮的合成。同时还进行了关于细胞存活性和半胱天冬酶3活化的控制实验。我们发现这两种激素都激活了cAMP和下游效应物，如细胞外信号调控的激酶1/2（Erk1/2）和cAMP反应元件结合蛋白（Creb），以及Akt，以及Stard1，Hsd3b1，Cyp17a1和Hsd17b3基因的转录，促进了Δ4类类固醇途径。有趣的是，Akt的阻断有选择地降低Cyp17a1的表达水平，抑制其17,20-裂解酶活性，但不抑制17-羟化酶活性。这种效果与Cyp17a1对17α-OH-孕酮的亲和力低于对孕酮的亲和力一致。结果，细胞接受3CAI处理后，在16-24小时内17α-OH-孕酮累积，24小时后睾酮水平下降。综上所述，在小鼠莱迪细胞系mLTC1中，我们发现17,20-裂酶活性和睾酮合成显著依赖Akt。我们的结果表明不同的细胞内信号通路有选择地调节Cyp17a1的双重活性。版权所有©2023 Elsevier Inc.发表。

Androgens are produced by adrenal and gonadal cells thanks to the action of specific enzymes. We investigated the role of protein kinase B (Akt) in the modulation of Δ4 steroidogenic enzymes' activity, in the mouse Leydig tumor cell line mLTC1. Cells were treated for 0-24 h with the 3 × 50% effective concentration of human luteinizing hormone (LH) and choriogonadotropin (hCG), in the presence and in the absence of the specific Akt inhibitor 3CAI. Cell signaling analysis was performed by bioluminescence resonance energy transfer (BRET) and Western blotting, while the expression of key target genes was investigated by real-time PCR. The synthesis of progesterone, 17α-hydroxy (OH)-progesterone and testosterone was measured by immunoassay. Control experiments for cell viability and caspase 3 activation were performed as well. We found that both hormones activated cAMP and downstream effectors, such as extracellularly-regulated kinase 1/2 (Erk1/2) and cAMP response element-binding protein (Creb), as well as Akt, and the transcription of Stard1, Hsd3b1, Cyp17a1 and Hsd17b3 genes, boosting the Δ4 steroidogenic pathway. Interestingly, Akt blockade decreased selectively Cyp17a1 expression levels, inhibiting its 17,20-lyase, but not the 17-hydroxylase activity. This effect is consistent with lower Cyp17a1 affinity to 17α-OH-progesterone than progesterone. As a result, cell treatment with 3CAI resulted in 17α-OH-progesterone accumulation at 16-24 h and decreased testosterone levels after 24 h. In conclusion, in the mouse Leydig cell line mLTC1, we found substantial Akt dependence of the 17,20-lyase activity and testosterone synthesis. Our results indicate that different intracellular pathways modulate selectively the dual activity of Cyp17a1.Copyright © 2023. Published by Elsevier Inc.