Autocrine/paracrine proliferative effect of ovarian GH and IGF-I in chicken granulosa cell cultures
Autocrine/paracrine proliferative effect of ovarian GH and IGF-I in chicken granulosa cell cultures
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DOI：
10.1016/j.ygcen.2016.05.008
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发表时间：
2016-08-01
影响因子：
2.7
通讯作者：
Aramburo, Carlos
中科院分区：
文献类型：
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作者：
Marisela Ahumada-Solorzano, S.;Martinez-Moreno, Carlos G.;Aramburo, Carlos
众所周知，在母鸡卵巢卵泡发育过程中，生长激素（GH）及其受体（GHR）在颗粒细胞（GC）和鞘细胞中表达，这表明GH参与雌性生殖系统中的自分泌/旁分泌作用。在这项工作中，我们表明，在 GC 培养物中用特定的 cGH siRNA 敲低局部卵巢 GH 显着降低了 cGH mRNA 表达和 GH 向培养基的分泌，并且还降低了它们的增殖率。因此，我们分析了卵巢 GH 和重组鸡 GH (rcGH) 对原代培养物中分级前 GC 增殖的影响。将 GC 与 rcGH 或主要含有 15-kDa GH 同工型的条件培养基一起孵育，结果显示，根据 3-(4,5-二甲基噻唑-2-y1)-2,5-二苯基四唑溴化物 (MTT) 测定、增殖细胞核抗原 (PCNA) 定量和 (H-3)-胸苷掺入 (H-3-T) 测定，两者的增殖均显着增加以剂量反应方式。本地产生的 GH 和 rcGH 也在 GC 培养物中诱导 Erk1/2 磷酸化。此外，GH 增加了 IGF-I 的合成及其向 GC 培养孵化介质中的释放。这些结果表明GH可能通过局部IGF-I诱导GC增殖，因为IGF-I免疫中和完全消除了GH诱导的增殖作用。这些数据表明，GH 和 IGF-I 可能在分级前阶段母鸡卵巢卵泡发育过程中发挥自分泌/旁分泌调节剂的作用。 (C) 2016 Elsevier Inc. 保留所有权利。
It is known that growth hormone (GH) and its receptor (GHR) are expressed in granulosa cells (GC) and thecal cells during the follicular development in the hen ovary, which suggests GH is involved in autocrine/paracrine actions in the female reproductive system. In this work, we show that the knockdown of local ovarian GH with a specific cGH siRNA in GC cultures significantly decreased both cGH mRNA expression and GH secretion to the media, and also reduced their proliferative rate. Thus, we analyzed the effect of ovarian GH and recombinant chicken GH (rcGH) on the proliferation of pre-hierarchical GCs in primary cultures. Incubation of GCs with either rcGH or conditioned media, containing predominantly a 15-kDa GH isoform, showed that both significantly increased proliferation as determined by the 3-(4,5-dimethylthiazol-2-y1)-2,5-diphenyltetrazolium bromide (MTT) assay, proliferating cell nuclear antigen (PCNA) quantification and (H-3)-thymidine incorporation (H-3-T) assays in a dose response fashion. Both, locally produced GH and rcGH also induced the phosphorylation of Erk1/2 in GC cultures. Furthermore, GH increased IGF-I synthesis and its release into the GC culture incubation media. These results suggest that GH may act through local IGF-I to induce GC proliferation, since IGF-I immunoneutralization completely abolished the GH-induced proliferative effect. These data suggest that GH and IGF-I may play a role as autocrine/paracrine regulators during the follicular development in the hen ovary at the pre-hierarchical stage. (C) 2016 Elsevier Inc. All rights reserved.