Role of mitogen-activated protein kinase in prostaglandin F2α action in human granulosa-luteal cells
Role of mitogen-activated protein kinase in prostaglandin F2α action in human granulosa-luteal cells
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DOI：
10.1210/jc.86.1.375
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发表时间：
2001-01-01
影响因子：
5.8
通讯作者：
Leung, PCK
中科院分区：
文献类型：
--
作者：
Tai, CJ;Kang, SK;Leung, PCK
在卵巢中，已经证明PGF（2 α）激活磷脂酶C（PLC）/二酰甘油/蛋白激酶C途径。然而，很少有人知道下游信号事件，介导随后的细胞反应，如类固醇。本研究旨在检测PGF（2 α）对人颗粒黄体细胞（hGLC）中丝裂原活化蛋白激酶（MAPK）信号通路激活的影响及其生理作用。从接受体外受精-胚胎移植的妇女中获得的人GLC用浓度递增的PGF（2 α）处理对于时程实验，用1 μ mol/L PGF（2 α）处理hGLC 1、5、10或20分钟。使用单克隆抗体检测细胞外信号调节激酶1和2（分别为p42（mapk）和p44（mapk））的磷酸化形式，证明PGF（2 α）以剂量和时间依赖性方式激活hGLC中的MAPK。用新霉素（10 mmol/L; PLC抑制剂）、双吲哚马来酰亚胺I（5 μ mol/L; PKC抑制剂）或PD 98059（50 μ mol/L; MEK抑制剂和MAPK激酶抑制剂）处理细胞显著减弱PGF（2 α）诱导的MAPK活化。相比之下，MAPK活化不受百日咳毒素（200 ng/mL; G（i）抑制剂）预处理的显著影响。为了确定MAPK在类固醇生成中的作用，在存在或不存在PD 98059的情况下，用PGF（2 α）（1 μ mol/L）、hCG（1 IU/mL）或PGF（2 α）加hCG处理hGLC。放免法检测培养液中孕酮水平。用PGF（2 α）处理hGLC显著抑制hCG诱导的孕酮产生。MEK抑制剂PD 98059的存在逆转了PGF（2 α）对hCG诱导的孕酮产生的抑制作用。据我们所知，这是第一次证明PGF（2 α）诱导的人卵巢MAPK信号通路的激活。这些结果表明，PGF（2 α）激活MAPK后PLC和PKC激活通过百日咳毒素不敏感的G蛋白在hGLC。此外，我们证明了PGF（2 α）诱导的MAPK激活与孕酮产生的调节有关。这些结果支持MAPK信号通路参与介导人卵巢中PGF（2 α）作用的观点。
In the ovary it has been demonstrated that PGF(2 alpha) activates the phospholipase C (PLC)/diacylglycerol/protein kinase C pathway. However, little is known about the downstream signaling events that mediate subsequent cellular responses such as steroidogenesis. The present study was designed to examine the effect of PGF(2 alpha) on activation of the mitogen-activated protein kinase (MAPK) signaling pathway and its physiological role in human granulosa-luteal cells (hGLCs). Human GLCs, obtained from women undergoing in vitro fertilization-embryo transfer, were treated with increasing concentrations of PGF(2 alpha) (10 nmol/L to 10 mu mol/L) for 5 min. For time-course experiments, hGLCs were treated with 1 mu mol/L PGF(2 alpha) for 1, 5, 10, or 20 min. Western blot analysis, using a monoclonal antibody that detected the phosphorylated forms of extracellular signal-regulated kinases 1 and 2 (p42(mapk) and p44(mapk), respectively), demonstrated that PGF(2 alpha) activated MAPK in hGLCs in a dose- and time-dependent manner. Treatment of the cells with neomycin (10 mmol/L; a PLC inhibitor), bisindolylmaleimide I (5 mu mol/L; a PKC inhibitor), or PD98059 (50 mu mol/L; a MEK inhibitor and a MAPK kinase inhibitor) significantly attenuated the PGF(2 alpha)-induced activation of MAPK. In contrast, MAPK activation was not significantly affected by pertussis toxin (200 ng/mL; a G(i) inhibitor) pretreatment. To determine the role of MAPK in steroidogenesis, hGLCs were treated with PGF(2 alpha) (1 mu mol/L), hCG (1 IU/mL), or PGF(2 alpha) plus hCG in the presence or absence of PD98059. Progesterone levels in the culture medium were examined by RIA. Treatment of hGLCs with PGF(2 alpha) significantly inhibited hCG-induced progesterone production. The presence of the MEK inhibitor, PD98059, reversed the inhibitory effect of PGF(2 alpha) on hCG-induced progesterone production. To our knowledge, it is the first demonstration of PGF(2 alpha)-induced activation of the MAPK signaling pathway in the human ovary. These results indicated that PGF(2 alpha) activated MAPK subsequent to PLC and PKC activation through pertussis toxin-insensitive G protein in hGLCs. Further, we demonstrated that PGF(2 alpha)-induced MAPK activation is associated with modulation of progesterone production. These results support the idea that the MAPK signaling pathway is involved in mediating PGF(2 alpha) actions in the human ovary.