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Nerve growth factor induces vascular endothelial growth factor expression in granulosa cells via a trkA receptor/mitogen-activated protein kinase-extracellularly regulated kinase 2-dependent pathway.

ABSTRACT: Acquisition of ovulatory competence by antral follicles requires development of an adequate vascular supply. Although it is well established that ovarian angiogenesis is cyclically regulated by vascular endothelial growth factor (VEGF), the factors controlling VEGF production by ovarian follicles remain largely unknown. Nerve growth factor (NGF) may be one of these factors, because NGF promotes angiogenesis and synthesis of angiogenic factors in other tissues and is produced by human granulosa cells (hGCs).The aim of the study was to determine whether NGF influences the production of VEGF by hGCs and to identify a potential signaling pathway underlying this effect.We conducted a prospective experimental study.hGCs were obtained from 41 women participating in the in vitro fertilization program of our institution.Changes in VEGF mRNA after exposure to NGF were evaluated in cultured hGCs by PCR and real-time PCR. The effect of NGF on VEGF secretion was determined by ELISA. The involvement of trkA, the high affinity NGF receptor, was examined by inhibiting the receptor's tyrosine kinase activity with K252a. The contribution of an ERK1/ERK2-mediated signaling pathway was identified by detecting NGF-dependent phosphorylation of these proteins and by blocking their activity with the inhibitor U0126.NGF promotes VEGF production in cultured hGCs. Blockade of trkA receptor tyrosine kinase activity blocks this effect. NGF induces MAPK-ERK2 phosphorylation, and blockade of this signaling pathway prevents the NGF-induced increase in VEGF production.NGF promotes ovarian angiogenesis by enhancing the synthesis and secretion of VEGF from hGCs via a trkA- and ERK2-dependent mechanism.

SUBMITTER: Julio-Pieper M

PROVIDER: S-EPMC2730870 | biostudies-literature | 2009 Aug

REPOSITORIES: biostudies-literature

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Nerve growth factor induces vascular endothelial growth factor expression in granulosa cells via a trkA receptor/mitogen-activated protein kinase-extracellularly regulated kinase 2-dependent pathway.

Julio-Pieper Marcela M Lozada Patricia P Tapia Veronica V Vega Margarita M Miranda Cristián C Vantman David D Ojeda Sergio R SR Romero Carmen C

The Journal of clinical endocrinology and metabolism 20090519 8

<h4>Context</h4>Acquisition of ovulatory competence by antral follicles requires development of an adequate vascular supply. Although it is well established that ovarian angiogenesis is cyclically regulated by vascular endothelial growth factor (VEGF), the factors controlling VEGF production by ovarian follicles remain largely unknown. Nerve growth factor (NGF) may be one of these factors, because NGF promotes angiogenesis and synthesis of angiogenic factors in other tissues and is produced by h ...[more]

PMID: 19454577

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Project description:INTRODUCTION:We have previously demonstrated that transforming growth factor-? (TGF-?) in the presence of elevated levels of Smad3, its primary signaling protein, stimulates rat vascular smooth muscle cell (VSMC) proliferation and intimal hyperplasia. The mechanism is partly through the nuclear exportation of phosphorylated cyclin-dependent kinase inhibitor p27. The objective of this study is to clarify the downstream pathways through which Smad3 produces its proliferative effect. Specifically, we evaluated the role of extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase (MAPK) in TGF-?-induced VSMC proliferation. METHODS:Cultured rat aortic VSMCs were incubated with TGF-? at varying concentrations and times, and phosphorylated ERK was measured by Western blotting. Smad3 was enhanced in VSMCs using an adenovirus expressing Smad3 or inhibited with small interfering RNA (siRNA). For in vivo experiments, male Sprague-Dawley rats underwent carotid balloon injury, followed by intraluminal infection with an adenovirus expressing Smad3. Arteries were harvested at 3 days and subjected to immunohistochemistry for Smad3, phospho-ERK MAPK, and proliferating cell nuclear antigen. RESULTS:In cultured VSMCs, TGF-? induced activation and phosphorylation of ERK MAPK in a time-dependent and concentration-dependent manner. Overexpression of the signaling protein Smad3 enhanced TGF-?-induced activation of ERK MAPK, whereas inhibition of Smad3 with a siRNA blocked ERK MAPK phosphorylation in response to TGF-?. These data suggest that Smad3 acts as a signaling intermediate between TGF-? and ERK MAPK. Inhibition of ERK MAPK activation with PD98059 completely blocked the ability of TGF-?/Smad3 to stimulate VSMC proliferation, demonstrating the importance of ERK MAPK in this pathway. Immunoprecipitation of phospho-ERK MAPK and blotting with Smad3 revealed a physical association, suggesting that activation of ERK MAPK by Smad3 requires a direct interaction. In an in vivo rat carotid injury model, overexpression of Smad3 resulted in an increase in phosphorylated ERK MAPK as well as increased VSMC proliferation as measured by proliferating cell nuclear antigen. CONCLUSIONS:Our findings demonstrate a mechanism through which TGF-? stimulates VSMC proliferation. Although TGF-? has been traditionally identified as an inhibitor of proliferation, our data suggest that TGF-? enhances VSMC proliferation through a Smad3/ERK MAPK signaling pathway. These findings at least partly explain the mechanism by which TGF-? enhances intimal hyperplasia. Knowledge of this pathway provides potential novel targets that may be used to prevent restenosis.

Dataset Information

Nerve growth factor induces vascular endothelial growth factor expression in granulosa cells via a trkA receptor/mitogen-activated protein kinase-extracellularly regulated kinase 2-dependent pathway.

Publications

Nerve growth factor induces vascular endothelial growth factor expression in granulosa cells via a trkA receptor/mitogen-activated protein kinase-extracellularly regulated kinase 2-dependent pathway.

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