Cooperative Effects of FOXL2 with the Members of TGF-β Superfamily on FSH Receptor mRNA Expression and Granulosa Cell Proliferation from Hen Prehierarchical Follicles

FSH, acting through multiple signaling pathways, regulates the proliferation and growth of granulosa cells, which are critical for ovulation. The present study investigated whether AMP-activated protein kinase (AMPK), which controls the energy balance of the cell, plays a role in FSH-mediated increase in granulosa cell proliferation. Cells isolated from immature rat ovaries were grown in serum-free, phenol red free DMEM-F12 and were treated with FSH (50 ng/ml) for 0, 5, and 15 min. Western blot analysis showed a significant reduction in AMPK activation as observed by a reduction of phosphorylation at thr 172 in response to FSH treatment at all time points tested. FSH also reduced AMPK phosphorylation in a dose-dependent manner with maximum inhibition at 100 ng/ml. The chemical activator of AMPK (5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside, 0.5 mm) increased the cell cycle inhibitor p27 kip expression significantly, whereas the AMPK inhibitor (compound C, 20 μm) and FSH reduced p27kip expression significantly compared with control. FSH treatment resulted in an increase in the phosphorylation of AMPK at ser 485/491 and a reduction in thr 172 phosphorylation. Inhibition of Akt phosphorylation using Akt inhibitor VIII reversed the inhibitory effect of FSH on thr 172 phosphorylation of AMPK, whereas ERK inhibitor U0126 had no effect. These results show that FSH, through an Akt-dependent pathway, phosphorylates AMPK at ser 481/495 and inhibits its activation by reducing thr 172 phosphorylation. AMPK activation by 5-amino-imidazole-4-carboxamide-1-β-d-ribofuranoside treatment resulted in a reduction of cell cycle regulatory protein cyclin D2 mRNA expression, whereas FSH increased the expression by 2-fold. These results suggest that FSH promotes granulosa cell proliferation by increasing cyclin D2 mRNA expression and by reducing p27 kip expression by inhibiting AMPK activation through an Akt-dependent pathway. FSH stimulates granulosa cell proliferation by reducing cell cycle inhibitor p27 kip through AMP kinase inhibition.

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Effect of follitropin on adenylate cyclase activity and on FSH receptor mRNA expression during the ovarian cycle ofPodarcis sicula

Italian Journal of Zoology ◽

10.1080/11250000409356617 ◽

2004 ◽

Vol 71 (sup2) ◽

pp. 105-108

Author(s):

Lucia Borrelli ◽

Roberta de Stasio ◽

Elio Parisi ◽

Silvana Filosa

Keyword(s):

Adenylate Cyclase ◽

Mrna Expression ◽

Ovarian Cycle ◽

Cyclase Activity ◽

Adenylate Cyclase Activity ◽

Fsh Receptor ◽

Receptor Mrna

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Relationship between adenylate cyclase sensitivity to follitropin and FSH receptor mRNA expression in the ovary of the lizardPodarcis sicula

Molecular Reproduction and Development ◽

10.1002/mrd.10090 ◽

2002 ◽

Vol 62 (2) ◽

pp. 210-215 ◽

Cited By ~ 1

Author(s):

Lucia Borrelli ◽

Roberta De Stasio ◽

Elio Parisi ◽

Silvana Filosa

Keyword(s):

Adenylate Cyclase ◽

Mrna Expression ◽

Fsh Receptor ◽

Receptor Mrna

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Dihydrotestosterone Inhibits Insulin-Stimulated Cyclin D2 Messenger Ribonucleic Acid Expression in Rat Ovarian Granulosa Cells by Reducing the Phosphorylation of Insulin Receptor Substrate-1

Endocrinology ◽

10.1210/en.2005-0818 ◽

2006 ◽

Vol 147 (1) ◽

pp. 464-471 ◽

Cited By ~ 10

Author(s):

Pradeep P. Kayampilly ◽

K. M. J. Menon

Keyword(s):

Cell Proliferation ◽

Insulin Receptor ◽

Mrna Expression ◽

Granulosa Cell ◽

Insulin Signaling ◽

Pi3 Kinase ◽

Insulin Receptor Substrate ◽

Cyclin D2 ◽

Insulin Receptor Substrate 1 ◽

Erk Phosphorylation

The effect of 5α-dihydrotestosterone (DHT) on insulin-stimulated granulosa cell proliferation was examined using cyclin D2 mRNA as a marker. Granulosa cells from 3-d estradiol-treated immature rats showed a concentration-dependent increase in cyclin D2 mRNA expression in response to insulin. Exposure to DHT reduced the insulin-stimulated cyclin D2 mRNA expression. Inhibition of the two insulin-signaling pathways, ERK and phosphatidylinositol 3 kinase (PI3 kinase), by using specific inhibitors, also reduced this insulin-stimulated response. These results suggest that both ERK and PI3 kinase signaling are involved in insulin stimulated granulosa cell proliferation. DHT exposure resulted in reduced insulin-stimulated ERK phosphorylation. DHT treatment also reduced the insulin mediated insulin receptor substrate-1 and Raf-1 phosphorylation, the upstream molecules of ERK in insulin signaling pathway. Additionally, inhibition of insulin stimulated PI3 kinase activation reduced ERK phosphorylation. The present study therefore shows that the inhibitory effect of DHT on insulin-stimulated granulosa cell proliferation occurs early in the signaling pathway at the level of insulin receptor substrate-1 phosphorylation, leading to reduced ERK phosphorylation and subsequent inhibition of cyclin D2 mRNA expression.

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Reduction of bFGF-induced smooth muscle cell proliferation and endothelin receptor mRNA expression by mevastatin and atorvastatin

Biochemical Pharmacology ◽

10.1016/s0006-2952(02)01189-9 ◽

2002 ◽

Vol 64 (3) ◽

pp. 497-505 ◽

Cited By ~ 21

Author(s):

Cang-Bao Xu ◽

Emelie Stenman ◽

Lars Edvinsson

Keyword(s):

Cell Proliferation ◽

Smooth Muscle ◽

Smooth Muscle Cell ◽

Mrna Expression ◽

Muscle Cell ◽

Smooth Muscle Cell Proliferation ◽

Endothelin Receptor ◽

Receptor Mrna

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Vascular endothelial growth factor-A isoform and (co)receptor expression are differentially regulated by 17β-oestradiol in the ovariectomised mouse uterus

Reproduction ◽

10.1530/rep-10-0047 ◽

2010 ◽

Vol 140 (2) ◽

pp. 331-341 ◽

Cited By ~ 12

Author(s):

Lisa M Walter ◽

Peter A W Rogers ◽

Jane E Girling

Keyword(s):

Vascular Endothelial Growth Factor ◽

Cell Proliferation ◽

Mrna Expression ◽

Receptor Expression ◽

Endothelial Cell Proliferation ◽

Vascular Endothelial ◽

Uterine Tissue ◽

Time Points ◽

Receptor Mrna ◽

Endothelial Growth Factor

The angiogenic effects of 17β-oestradiol (E2) in the mouse endometrium are mediated by vascular endothelial growth factor-A (VEGFA). We analysed the temporal and spatial changes in VEGFA isoform and (co)receptor expression in ovariectomised mouse uteri following E2 treatment. VEGFA isoform and receptor mRNA were quantified in whole uterine tissue collected 2, 6, 12 and 24 h after E2 or vehicle treatment. Laser capture microdissection was used to investigate mRNA expression in epithelial, stromal and myometrial tissues separately. Endothelial cell proliferation, VEGFA and VEGF receptor-2 (VEGFR2) protein were visualised using immunohistochemistry. Endometrial endothelial cell proliferation was only observed 24 h after E2 treatment. In whole uterine tissue, total Vegfa, Vegfa164 and Vegfa120 mRNA expression increased 2 h post E2 treatment, and then decreased by 24 h. Vegfa188 expression was lower in E2-treated animals at all time points relative to control animals. Vegfr2 and neuropilin-1 (Nrp1) mRNA expression did not change following E2 treatment; Nrp2 expression decreased by 24 h. When uterine compartments were considered separately at 24 h post E2 or vehicle, stromal Vegfa120, Vegfa188 and Vegfr2 mRNA expression and myometrial Vegfa120 and Vegfa188 mRNA expression were reduced in E2-treated mice relative to controls, whereas epithelial Vegfa188 mRNA expression increased. The highest VEGFA immunoexpression was observed in luminal epithelium; expression increased at 24 h relative to other time points. No changes were noted in VEGFR2 immunoexpression among treatment groups. We have provided the first evidence that VEGFA isoform and receptor mRNA expression are differentially regulated by E2 in different uterine cell compartments.

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