College of Veterinary Medicine, Yangzhou University, Yangzhou, 225009, Jiangsu, China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, 225009, Jiangsu, China.
College of Veterinary Medicine, Yangzhou University, Yangzhou, 225009, Jiangsu, China.
Mol Cell Endocrinol. 2020 Jul 1;511:110842. doi: 10.1016/j.mce.2020.110842. Epub 2020 May 4.
ER oxidoreduclin 1α (ERO1α), an oxidase that exists in the ER, participates in protein folding and secretion and inhibiting apoptosis, and regulates tumor progression, which is a novel factor of poor cancer prognosis. However, the other physiological functions of ERO1α remain undiscovered. Although our preliminary results of this study indicated that ERO1α revealed the robust expression in ovary, especially in granulosa cells, the role of ERO1α in follicular development is not well known. Therefore, the aims of the present study were to explore the role of ERO1α and the possible mechanisms in regulating cell apoptosis and steroidogenesis in ovarian granulosa cells. ERO1α was mainly localized in granulosa cells and oocytes in the adult ovary by immunohistochemistry. Western blot analysis showed that the expression of ERO1α was highest at oestrous stage during the estrous cycle. The effect of ERO1α on cell apoptosis and steroidogenesis was detected by transduction of ERO1α overexpression and knockdown lentiviruses into primary cultured granulosa cells. Flow cytometry analysis showed that ERO1α decreased granulosa cells apoptosis. Western bolt and RT-qPCR analysis found that ERO1α increased the ratio of BCL-2/BAX, and decreased BAD and Caspase-3 expression. ELISA analysis showed that ERO1α enhanced estrogen (E2) secretion. Western bolt and RT-qPCR analysis found that ERO1α increased StAR, CYP11A1, 3β-HSD, CYP17A1, and CYP19A1 expression, and decreased CYP1B1 expression. Furthermore, Western bolt analysis found that ERO1αincreased PDI and PRDX 4 expression, and activated the PI3K/AKT/mTOR signaling pathway through increasing the phosphorylation of AKT and P70 S6 kinase. In summary, these results suggested that ERO1α might play an anti-apoptotic role and regulate steroidogenesis in granulosa cells, at least partly, via activation of the PI3K/AKT/mTOR signaling pathway.
内质网氧化还原酶 1α（ERO1α）是一种存在于内质网中的氧化酶，参与蛋白质折叠和分泌以及抑制细胞凋亡，并调节肿瘤进展，是癌症预后不良的新因素。然而，ERO1α 的其他生理功能尚未被发现。尽管我们本研究的初步结果表明 ERO1α在卵巢中表现出强烈的表达，特别是在颗粒细胞中，但 ERO1α 在卵泡发育中的作用尚不清楚。因此，本研究的目的是探讨 ERO1α在调节卵巢颗粒细胞细胞凋亡和类固醇生成中的作用及其可能的机制。免疫组织化学染色显示 ERO1α 主要定位于成年卵巢中的颗粒细胞和卵母细胞。Western blot 分析显示，在发情周期中，发情期 ERO1α 的表达最高。通过转导 ERO1α 过表达和敲低慢病毒进入原代培养的颗粒细胞，检测 ERO1α 对细胞凋亡和类固醇生成的影响。流式细胞术分析显示 ERO1α 降低了颗粒细胞凋亡。Western blot 和 RT-qPCR 分析发现 ERO1α 增加了 BCL-2/BAX 的比值，并降低了 BAD 和 Caspase-3 的表达。ELISA 分析显示 ERO1α 增强了雌激素（E2）的分泌。Western blot 和 RT-qPCR 分析发现 ERO1α 增加了 StAR、CYP11A1、3β-HSD、CYP17A1 和 CYP19A1 的表达，并降低了 CYP1B1 的表达。此外，Western blot 分析发现 ERO1α 增加了 PDI 和 PRDX4 的表达，并通过增加 AKT 和 P70 S6 激酶的磷酸化激活了 PI3K/AKT/mTOR 信号通路。总之，这些结果表明，ERO1α 可能通过激活 PI3K/AKT/mTOR 信号通路，在颗粒细胞中发挥抗细胞凋亡作用并调节类固醇生成。
