Zhou Ge, Hu Rong-Kui, Xia Gui-Cheng, Yan Shi-Hai, Ren Qing-Ling, Zhao Juan, Wang Fei-Hong, Huang Cheng-Cai, Yao Qi, Tan Yong, Zhao Ning-Wei
Department of Reproductive Medicine, Affiliated Hospital of Nanjing University of Chinese Medicine, Jiangsu Province Hospital of Chinese Medicine, Nanjing, China.
Laboratory of Pharmacology, Affiliated Hospital of Nanjing University of Chinese Medicine, Jiangsu Province Hospital of Chinese Medicine, Nanjing, China.
Aging (Albany NY). 2019 May 15;11(10):3094-3116. doi: 10.18632/aging.101964.
Many infertile women suffered from poor ovarian response, and increased reactive oxygen species with age might mediate the poor ovarian response to FSH. In this study, we collected follicular fluids and isolated granulosa cells from female patients. Increased levels of peroxynitrite, tyrosine nitrations of FSH receptor (FSHR) and apoptosis were obviously detectable with decreased FSHR protein expressions in granulosa cells of the poor ovarian responders. In KGN (a human ovarian granulosa cell line) cells, exogenous peroxynitrite could sequester FSHR in the cytoplasm, and these dislocated FSHR might suffer from proteasome-mediated degradations. Here, we identified four peroxynitrite-mediated nitrated tyrosine residues of FSHR. Site-directed mutagenesis of FSHR revealed that Y626 was pivotal for intracellular trafficking of FSHR to the cell surface. Akt-induced inactivation of FoxO3a was required for the repression of FSH on granulosa cell apoptosis. However, peroxynitrite impaired FSH-induced Akt-FoxO3a signaling, while FSHR-Y626A mutant took similar effects. In addition, FoxO3a knockdown indeed impaired FSH-mediated cell survival, while FoxO3a-S253A mutant reversed that significantly.
许多不孕女性存在卵巢反应不良的情况，并且随着年龄增长活性氧增加可能介导了卵巢对促卵泡激素（FSH）反应不良。在本研究中，我们收集了女性患者的卵泡液并分离出颗粒细胞。在卵巢反应不良者的颗粒细胞中，明显可检测到过氧亚硝酸盐水平升高、FSH受体（FSHR）酪氨酸硝化增加以及细胞凋亡增加，同时FSHR蛋白表达降低。在KGN（一种人卵巢颗粒细胞系）细胞中，外源性过氧亚硝酸盐可将FSHR隔离在细胞质中，这些错位的FSHR可能会遭受蛋白酶体介导的降解。在此，我们鉴定了FSHR的四个过氧亚硝酸盐介导的硝化酪氨酸残基。FSHR的定点诱变显示Y626对于FSHR向细胞表面的细胞内转运至关重要。FSH对颗粒细胞凋亡的抑制需要Akt诱导的FoxO3a失活。然而，过氧亚硝酸盐损害了FSH诱导的Akt - FoxO3a信号传导，而FSHR - Y626A突变体产生了类似的效果。此外，敲低FoxO3a确实损害了FSH介导的细胞存活，而FoxO3a - S253A突变体显著逆转了这种情况。
