Roche Jennifer, Ramé Christelle, Reverchon Maxime, Mellouk Namya, Cornuau Marion, Guerif Fabrice, Froment Pascal, Dupont Joëlle
Unité de Physiologie de la Reproduction et des Comportements, Institut National de la Recherche Agronomique, Nouzilly, France.
Service de Médecine et Biologie de la Reproduction, CHRU de Tours, Tours, France.
Biol Reprod. 2016 Nov;95(5):104. doi: 10.1095/biolreprod.116.141754. Epub 2016 Sep 28.
Apelin (APLN) is a recently discovered adipokine involved in the regulation of various metabolic functions. Its receptor, APLNR, is expressed in reproductive tissues, however, its role in human ovarian cells is unknown. In this study, we identified APLN and APLNR in human ovarian follicles and analyzed their expression in granulosa cells and follicular fluid obtained from obese and nonobese patients, with or without polycystic ovary syndrome (PCOS). We also investigated the effect of APLN on steroidogenesis in cultured human luteinized granulosa cells (hGCs) from nonobese patients without PCOS. Using RT-PCR and immunoblotting, we found that APLN and APLNR were expressed in hGCs and cumulus and theca cells. We confirmed these data immunohistochemically and observed that APLNR and APLN are present in human oocytes at different stages of follicular development. In patients with PCOS, we observed that follicular fluid APLN concentration and granulosa cell APLN and APLNR mRNA expression was higher than that observed in control patients. In cultured hGCs from nonobese patients without PCOS, insulin-like growth factor 1 (IGF1) increased APLNR expression, and recombinant human APLN (APLN-13 and APLN-17) increased both basal and IGF1-induced steroid secretion. These effects on steroid production were reversed when cultured in the presence of ML221, an APLNR antagonist, which was associated with an increased 3beta-hydrosteroid dehydrogenase (HSD3B) protein concentration. We showed that these effects were dependent on the activation of the AKT and MAPK3/1 pathways using pharmacological inhibitors. Our results show that APLN and APLNR are present in human ovarian cells and APLN increases IGF1-induced steroidogenesis in granulosa cells through an increase in HSD3B protein expression and activation of the MAPK3/1 and Akt pathways. Therefore, APLN and APLNR may play a role in human follicular development and the pathogenesis of PCOS.
阿片肽（APLN）是一种最近发现的脂肪因子，参与多种代谢功能的调节。其受体APLNR在生殖组织中表达，然而，其在人类卵巢细胞中的作用尚不清楚。在本研究中，我们在人类卵巢卵泡中鉴定出APLN和APLNR，并分析了它们在肥胖和非肥胖患者（伴或不伴多囊卵巢综合征（PCOS））的颗粒细胞和卵泡液中的表达。我们还研究了APLN对来自无PCOS的非肥胖患者的培养人黄素化颗粒细胞（hGCs）中类固醇生成的影响。使用逆转录聚合酶链反应（RT-PCR）和免疫印迹法，我们发现APLN和APLNR在hGCs、卵丘细胞和卵泡膜细胞中表达。我们通过免疫组织化学法证实了这些数据，并观察到APLNR和APLN存在于卵泡发育不同阶段的人类卵母细胞中。在PCOS患者中，我们观察到卵泡液中APLN浓度以及颗粒细胞中APLN和APLNR mRNA表达均高于对照患者。在来自无PCOS的非肥胖患者的培养hGCs中，胰岛素样生长因子1（IGF1）增加了APLNR表达，重组人APLN（APLN-13和APLN-17）增加了基础和IGF1诱导的类固醇分泌。当在APLNR拮抗剂ML221存在的情况下培养时，这些对类固醇产生的影响被逆转，这与3β-羟基类固醇脱氢酶（HSD3B）蛋白浓度增加有关。我们使用药理学抑制剂表明这些作用依赖于AKT和丝裂原活化蛋白激酶3/1（MAPK3/1）途径的激活。我们的结果表明，APLN和APLNR存在于人类卵巢细胞中，并且APLN通过增加HSD3B蛋白表达以及激活MAPK3/1和Akt途径来增加IGF1诱导的颗粒细胞类固醇生成。因此，APLN和APLNR可能在人类卵泡发育和PCOS发病机制中发挥作用。
