Reproductive Medicine Center, The First Affiliated Hospital of Sun Yat-sen University, Guangdong, Guangzhou, China.
Guangdong Provincial Key Laboratory of Reproductive Medicine, First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.
Reproduction. 2020 Dec;160(6):955-967. doi: 10.1530/REP-20-0340.
Luteinization is the event of corpus luteum formation, a way of follicle cells transformation and a process of steroidogenesis alteration. As the core clock gene, Bmal1 was involved in the regulation of ovulation process and luteal function afterwards. Till now, the underlying roles of luteinization played by Bmal1 remain unknown. To explore the unique role of Bmal1 in luteal steroidogenesis and its underlying pathway, we investigated the luteal hormone synthesis profile in Bmal1 knockout female mice. We found that luteal hormone synthesis was notably impaired, and phosphorylation of PI3K/NfκB pathway was significantly activated. Then, the results were verified in in vitro cultured cells, including isolated Bmal1 interference granulosa cells (GCs) and theca cells (TCs), respectively. Hormones levels of supernatant culture media and mRNA expressions of steroidogenesis-associated genes (star, Hsd3β2, cyp19a1 in GCs, Lhcgr, star, Hsd3β2, cyp17a1 in TCs) were mutually decreased, while the phosphorylation of PI3K/NfκB was promoted during in vitro luteinization. After PI3K specific-inhibitor LY294002 intervention, mRNA expressions of Lhcgr and Hsd3β2 were partially rescued in Bmal1 interference TCs, together with significantly increased androstenedione and T synthesis. Further exploration in TCs demonstrated BMAL1 interacted directly but negatively with NfκB p65 (RelA), a subunit which was supposed as a mediator in Bmal1-governed PI3K signaling regulation. Taken together, we verified the novel role of Bmal1 in luteal steroidogenesis, achieving by negative interplay with RelA-mediated PI3K/NfκB pathway.
黄体化是黄体形成的事件，是卵泡细胞转化的一种方式，也是类固醇生成改变的过程。作为核心时钟基因，Bmal1 参与了排卵过程的调节和随后的黄体功能。到目前为止，Bmal1 在黄体类固醇生成中的作用尚不清楚。为了探索 Bmal1 在黄体类固醇生成中的独特作用及其潜在途径，我们研究了 Bmal1 敲除雌性小鼠的黄体激素合成谱。我们发现黄体激素合成明显受损，PI3K/NfκB 途径的磷酸化显著激活。然后，在体外培养的细胞中分别验证了这一结果，包括分离的 Bmal1 干扰颗粒细胞（GCs）和间质细胞（TCs）。上清液培养介质中的激素水平和类固醇生成相关基因（GCs 中的 star、Hsd3β2、cyp19a1，TCs 中的 Lhcgr、star、Hsd3β2、cyp17a1）的 mRNA 表达相互降低，而体外黄体化过程中 PI3K/NfκB 的磷酸化增强。在 PI3K 特异性抑制剂 LY294002 干预后，Bmal1 干扰 TCs 中的 Lhcgr 和 Hsd3β2 的 mRNA 表达部分得到挽救，同时雄烯二酮和 T 的合成显著增加。在 TCs 中的进一步探索表明，BMAL1 直接但负性地与 NfκB p65（RelA）相互作用，后者被认为是 Bmal1 调控的 PI3K 信号转导的调节因子。综上所述，我们验证了 Bmal1 在黄体类固醇生成中的新作用，通过与 RelA 介导的 PI3K/NfκB 途径的负性相互作用实现。
