An integrated RNA-Seq and network study reveals that valproate inhibited progesterone production in human granulosa cells

In addition to somatic cell-derived growth factors, oocyte-derived growth differentiation factor (GDF)9 and bone morphogenetic protein (BMP)15 play essential roles in female fertility. However, few studies have investigated their effects on human ovarian steroidogenesis, and fewer still have examined their differential effects or underlying molecular determinants. In the present study, we used immortalized human granulosa cells (SVOG) and human granulosa cell tumor cells (KGN) to compare the effects of GDF9 and BMP15 on steroidogenic enzyme expression and investigate potential mechanisms of action. In SVOG cells, neither GDF9 nor BMP15 affects the mRNA levels of P450 side-chain cleavage enzyme or 3β-hydroxysteroid dehydrogenase. However, treatment with BMP15, but not GDF9, significantly decreases steroidogenic acute regulatory protein (StAR) mRNA and protein levels as well as progesterone production. These suppressive effects, along with the induction of Sma and Mad-related protein (SMAD)1/5/8 phosphorylation, are attenuated by cotreatment with 2 different BMP type I receptor inhibitors (dorsomorphin and DMH-1). Furthermore, depletion of activin receptor-like kinase (ALK)3 using small interfering RNA reverses the effects of BMP15 on SMAD1/5/8 phosphorylation and StAR expression. Similarly, knockdown of ALK3 abolishes BMP15-induced SMAD1/5/8 phosphorylation in KGN cells. These results provide evidence that oocyte-derived BMP15 down-regulates StAR expression and decreases progesterone production in human granulosa cells, likely via ALK3-mediated SMAD1/5/8 signaling. Our findings suggest that oocyte may play a critical role in the regulation of progesterone to prevent premature luteinization during the late stage of follicle development.

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Regulatory role of angiotensin II on progesterone production by cultured human granulosa cells. Expression of angiotensin II type-2 receptor

MHR Basic science of reproductive medicine ◽

10.1093/molehr/3.8.663 ◽

1997 ◽

Vol 3 (8) ◽

pp. 663-668 ◽

Cited By ~ 18

Author(s):

M. Johnson

Keyword(s):

Angiotensin Ii ◽

Granulosa Cells ◽

Regulatory Role ◽

Human Granulosa Cells ◽

Progesterone Production

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High- and low-density lipoproteins stimulate progesterone production in cultured human granulosa cells

Human Reproduction ◽

10.1093/oxfordjournals.humrep.a136537 ◽

1987 ◽

Vol 2 (4) ◽

pp. 291-295 ◽

Cited By ~ 11

Author(s):

L. Enk ◽

N. Crona ◽

T. Hillensjö

Keyword(s):

Granulosa Cells ◽

Low Density Lipoproteins ◽

Low Density ◽

Human Granulosa Cells ◽

Progesterone Production

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Stimulation of Apoptosis in Human Granulosa Cells from in Vitro Fertilization Patients and Its Prevention by Dexamethasone: Involvement of Cell Contact and Bcl-2 Expression

The Journal of Clinical Endocrinology & Metabolism ◽

10.1210/jcem.87.7.8676 ◽

2002 ◽

Vol 87 (7) ◽

pp. 3441-3451 ◽

Cited By ~ 40

Author(s):

Ravid Sasson ◽

Abraham Amsterdam

Keyword(s):

In Vitro Fertilization ◽

Actin Cytoskeleton ◽

Gap Junctions ◽

Granulosa Cells ◽

Connexin 43 ◽

Pronounced Increase ◽

Vitro Fertilization ◽

Human Granulosa Cells ◽

Progesterone Production

Human granulosa cells obtained from in vitro fertilization patients are highly luteinized, but can still be stimulated by LH/cAMP for production of progesterone. This stimulation involved enhancement of apoptosis. Incubation of the cells with dexamethasone (Dex) reduced the apoptotic incidence compared with nontreated cells and completely abolished the increase in apoptosis stimulated by LH or forskolin, concomitantly with a pronounced increase in progesterone production. Organization of the actin cytoskeleton was dramatically reduced after LH/forskolin stimulation. In contrast, Dex prevented disorganization of the actin filament networks. LH and forskolin also decreased the organization of gap junctions, which could be prevented by Dex. However, the intracellular level of connexin 43 was elevated in the presence of LH, forskolin, and Dex. Endogenous levels of the survival gene protein Bcl-2 were significantly elevated in all cultures treated with Dex compared with either nonstimulated cultures or cultures stimulated with LH and forskolin. Our data suggest that LH/cAMP can stimulate steroidogenesis even during the initial stage of apoptosis of human granulosa cells, whereas Dex, which blocks apoptosis, could further elevate progesterone production. Moreover, the integrity of gap junctions and the actin cytoskeleton as well as elevated levels of Bcl-2 may play an important role in the suppression of apoptosis of human granulosa cells.

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In vitro action of HMG-CoA reductase inhibitors on cholesterol synthesis, progesterone production and secretion in human granulosa cells

Atherosclerosis ◽

10.1016/0021-9150(94)94028-2 ◽

1994 ◽

Vol 109 (1-2) ◽

pp. 256

Author(s):

A.K. Van Vliet ◽

L.H. Cohen

Keyword(s):

Granulosa Cells ◽

Cholesterol Synthesis ◽

Hmg Coa Reductase ◽

Human Granulosa Cells ◽

Progesterone Production ◽

Reductase Inhibitors ◽

Hmg Coa Reductase Inhibitors ◽

Coa Reductase

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Growth Differentiation Factor-8 Decreases StAR Expression Through ALK5-Mediated Smad3 and ERK1/2 Signaling Pathways in Luteinized Human Granulosa Cells

Endocrinology ◽

10.1210/en.2015-1461 ◽

2015 ◽

Vol 156 (12) ◽

pp. 4684-4694 ◽

Cited By ~ 14

Author(s):

Lanlan Fang ◽

Hsun-Ming Chang ◽

Jung-Chien Cheng ◽

Yiping Yu ◽

Peter C. K. Leung ◽

...

Keyword(s):

Signaling Pathways ◽

Granulosa Cells ◽

Follicular Fluid ◽

Type I ◽

Mrna Levels ◽

Activin Receptor ◽

Human Granulosa Cells ◽

Progesterone Production ◽

Differentiation Factor ◽

Receptor Like Kinase

Growth differentiation factor-8 (GDF-8) has been recently shown to be expressed in human granulosa cells, and the mature form of GDF-8 protein can be detected in the follicular fluid. However, the biological function and significance of this growth factor in the human ovary remains to be determined. Here, we investigated the effects of GDF-8 on steroidogenic enzyme expression and the potential mechanisms of action in luteinized human granulosa cells. We demonstrated that treatment with GDF-8 did not affect the mRNA levels of P450 side-chain cleavage enzyme and 3β-hydroxysteroid dehydrogenase, whereas it significantly down-regulated steroidogenic acute regulatory protein (StAR) expression and decreased progesterone production. The suppressive effect of GDF-8 on StAR expression was abolished by the inhibition of the TGF-β type I receptor. In addition, treatment with GDF-8 activated both Smad2/3 and ERK1/2 signaling pathways. Furthermore, knockdown of activin receptor-like kinase 5 reversed the effects of GDF-8 on Smad2/3 phosphorylation and StAR expression. The inhibition of Smad3 or ERK1/2 signaling pathways attenuated the GDF-8-induced down-regulation of StAR and production of progesterone. Interestingly, the concentrations of GDF-8 were negatively correlated with those of progesterone in human follicular fluid. These results indicate a novel autocrine function of GDF-8 to down-regulate StAR expression and decrease progesterone production in luteinized human granulosa cells, most likely through activin receptor-like kinase 5-mediated Smad3 and ERK1/2 signaling pathways. Our findings suggest that granulosa cells might play a critical role in the regulation of progesterone production to prevent premature luteinization during the final stage of folliculogenesis.

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STEROIDOGENESIS BY CULTURED GRANULOSA CELLS ASPIRATED FROM HUMAN FOLLICLES USING PREGNENOLONE AND ANDROGENS AS PRECURSORS

Journal of Endocrinology ◽

10.1677/joe.0.0770171 ◽

1978 ◽

Vol 77 (2) ◽

pp. 171-183 ◽

Cited By ~ 19

Author(s):

R. E. FOWLER ◽

N. L. FOX ◽

R. G. EDWARDS ◽

D. E. WALTERS ◽

P. C. STEPTOE

Keyword(s):

Granulosa Cells ◽

Short Term ◽

Expected Time ◽

Human Granulosa Cells ◽

Progesterone Production ◽

Thecal Cells ◽

Short Term Test ◽

Endogenous Substrates ◽

Test Cultures

SUMMARY Human granulosa cells from Graafian follicles aspirated 3–4 h before the expected time of ovulation were incubated with various steroid substrates, including pregnenolone, androstenedione, testosterone and dehydroepiandrosterone (DHA). Steroid production after 3 and 10 h of incubation was determined by radioimmunoassay. Progesterone and 17α-hydroxyprogesterone were the major products of granulosa cells in control short-term cultures with endogenous substrates. The addition of pregnenolone increased the synthesis of progesterone and 17α-hydroxyprogesterone compared with the controls, although the response varied considerably between paired short-term cultures. Little or no oestradiol-17β was produced from endogenous precursors or short-term cultures to which pregnenolone had been added; one follicle, however, produced similar amounts of oestradiol-17β in the control cultures and after incubation with pregnenolone. When granulosa cells were cultured with various amounts of androstenedione, DHA or testosterone, large amounts of oestradiol-17β were produced, especially in short-term cultures in which larger amounts of substrate were added. Progesterone production continued and progesterone was synthesized more rapidly or in greater amounts in some short-term test cultures than in the controls. The results indicate that human granulosa cells are one source of oestradiol-17β during the preovulatory phase. The data support the two-cell theory for oestradiol synthesis, for granulosa cells do not appear to undertake steroid conversion via the 5-unsaturated pathway, but aromatize androgens known to be produced by thecal cells. It is also suggested that either androgens or oestradiol-17β stimulate progesterone production by granulosa cells, at least in vitro.

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