Effects of metabolic hormones and growth factors on forskolin and dibutyryl adenosine 3′,5′-cyclic monophosphate induced steroidogenic responses by porcine granulosa cells in vitro

1995 ◽  
Vol 75 (1) ◽  
pp. 85-91 ◽  
Author(s):  
Y. Xu ◽  
R. N. Kirkwood ◽  
P. A. Thacker ◽  
K. Rajkumar
Keyword(s):  
Growth Factors ◽  
Granulosa Cells ◽  
Metabolic Hormones ◽  
Estradiol Secretion ◽  
Porcine Granulosa Cells ◽  
Igf I ◽  
The Face ◽  
Positive Effect

The present experiments were conducted to examine the influence of metabolic hormones and growth factors on progesterone and estradiol secretion by porcine granulosa cells in culture and to determine the possible site(s) of action of these agents on steroidogenesis. Porcine granulosa cells, obtained from medium-sized (4–6 mm) nonatretic follicles of prepubertal gilts slaughtered at a commercial abattoir, were cultured at a density of 2 × 106 viable cells well−1 in serum-free Eagle's Minumum Essential Medium for 72 h with 0, 10 or 100 ng mL−1 of GH, insulin or IGF-I or IGF-II. At 72 h, each treatment group was divided into three subgroups, one subgroup remained untreated (basal) while either 25 μM FK or 3 mM dbcAMP was added to the other two subgroups. Additionally, androstenedione (10−7 M) was provided to all cultures as a substrate for estradiol biosynthesis. Media were collected at 96 h for determination of progesterone and estradiol content. Both IGF-I and IGF-II increased (P < 0.01) basal progesterone and estradiol secretion and enhanced the FK- and dbcAMP-induced stimulation of progesterone and estradiol secretion (P < 0.01). Insulin, at 100 ng mL−1, had a modest but significant positive effect on both FK- and dbcAMP-stimulated steroidogenic responses. In contrast, GH inhibited basal, FK- and dbcAMP-induced progesterone and estradiol production. Based on the stimulatory effects of both IGFs above those seen with dbcAMP alone and the inhibitory effects of GH in the face of dbcAMP stimulation, we conclude that their effects are likely mediated by non-cAMP dependent pathways. Key words: Steroidogenesis, granulosa cells, growth hormone, IGF, insulin, pigs

Comparison of the facilitative roles of insulin and insulin-like growth factor I in the functional differentiation of granulosa cells: in vitro studies with the porcine model

1988 ◽  
Vol 117 (2) ◽  
pp. 230-240 ◽  
Author(s):  
Takeshi Maruo ◽  
Masato Hayashi ◽  
Hiroya Matsuo ◽  
Yasuo Ueda ◽  
Hajime Morikawa ◽  
...  
Keyword(s):  
Granulosa Cell ◽  
Granulosa Cells ◽  
Binding Sites ◽  
Physiological Concentration ◽  
Physiological Level ◽  
Estradiol Secretion ◽  
Porcine Granulosa Cells ◽  
Igf I ◽  
Hcg Receptor

Abstract. The facilitative effects of insulin and IGF-I were compared in vitro with regard to induction of differentiated functions of porcine granulosa cells. The monolayers were maintained under serum-free conditions in the absence or presence of porcine FSH (20 μg/l), with or without graded doses of insulin or IGF-I. Concurrent treatment with IGF-I and FSH produced morphological differentiation and augmented LH/hCG receptor binding together with an enhancement in progesterone and estradiol secretion relative to treatment with FSH alone. IGF-I alone was incapable of exhibiting these effects. Insulin synergized with FSH to facilitate the granulosa cell functions except estradiol secretion. Maximal effective dose of IGF-I was 100 μg/l which is within the physiological concentration in vivo, whereas that of insulin was 1.0 mg/l, which is 1000-fold higher than the physiological level. Although the maximal effective doses of IGF-I and insulin produced a comparable increment in progesterone secretion and LH/hCG receptor induction, combined treatment with IGF-I and insulin did not prove additive. [125I]IGF-I binding revealed that specific IGF-I receptors with two classes of binding sites are present on porcine granulosa cells. No distinct differences were detected between IGF-I receptors of granulosa cells from small, medium and large follicles. Insulin was approximately 100-fold less active than IGF-I in competing for [125I]IGF-I binding. These findings suggest that porcine granulosa cells possess specific IGF-I binding sites which may mediate the cytodifferentiative actions of insulin-like peptides. Since IGF-I is more potent than insulin in amplifying the actions of FSH and maximally exerts the cytodifferentiative effects at the physiological concentration, it is likely that IGF-I plays the more important role in granulosa cell differentiation in synergy with FSH.

Effects of growth factors and hormones on basal and FSH-stimulated inhibin production by porcine granulosa cells in vitro

1991 ◽  
Vol 3 (2) ◽  
pp. 201 ◽  
Author(s):  
U Michel ◽  
S Ludemann ◽  
H Jarry ◽  
W Wuttke
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Granulosa Cells ◽  
Type I ◽  
Tgf Beta ◽  
G Cells ◽  
Porcine Granulosa Cells ◽  
Igf I ◽  
Factor Type

The effect of several growth factors, protein and steroid hormones on follicle stimulating hormone (FSH)-stimulated and basal inhibin secretion by mature porcine granulosa cells (g-cells) in culture was examined in order to elucidate the putative role of growth factors and hormones in the regulation of inhibin secretion by porcine g-cells in vitro. Cells were incubated with the respective hormones over a timespan of 0-144 h and immunoreactive inhibin was measured with a radioimmunoassay against porcine inhibin. Epidermal growth factor (EGF) and human transforming growth factor type beta (TGF-beta) decreased basal and gonadotrophin-stimulated inhibin and progesterone in a dose-dependent manner. In the absence of insulin, insulin-like growth factor type I (IGF-I) caused a 4-fold enhancement of basal inhibin secretion, but inhibin secretion was elevated only to 20% above control in the presence of 500 nM insulin. Porcine platelet-derived growth factor (PDGF) had no significant effect on basal or FSH-induced inhibin secretion by g-cells. In addition, neither gonadotrophin-releasing hormone (GnRH) nor prolactin (PRL), arginine vasopressin (AVP) and oxytocin affected basal or FSH-stimulated inhibin release by porcine g-cells. Oestradiol caused a slight but significant (P less than 0.01) rise of basal inhibin production (158% of control) in the last 2 days of culture (96-144 h) and the effect of androstenedione on basal (158% of control) and FSH-stimulated (140% of control) inhibin release (P less than 0.01) was also only visible on Days 4-6 of culture. In contrast to androstenedione and oestradiol, progesterone did not show any effect during 6 days of culture in a dose range of 10(-5) to 10(-9) M. Like steroids, prostaglandin E2 (PGE2) had a stimulatory effect on basal inhibin production (250% of control) by porcine g-cells, visible on Days 3-6 of culture, but an inhibitory effect on FSH-stimulated release (less than 40% of control). Over all the experiments with different hormones and growth factors, tested in varying doses and over a time span of 0-144 h, there was a strong correlation between progesterone and inhibin secretion by g-cells (0-48 h = 0.78; 48-96 h = 0.92; 96-144 h = 0.92). These results suggest that EGF, TGF-beta, IGF-I, oestradiol and androstendione as well as PGE2 have para- and/or autocrine modulatory effects on basal and FSH-stimulated inhibin secretion by mature porcine g-cells in vitro and further demonstrate that the secretion of the proteohormone inhibin and the steroid progesterone are closely related.

scholarly journals Granulosa cells of the cumulus oophorus are different from mural granulosa cells in their response to gonadotrophins and IGF-I

2001 ◽  
Vol 170 (3) ◽  
pp. 565-573 ◽  
Author(s):  
F Khamsi ◽  
S Roberge
Keyword(s):  
Granulosa Cells ◽  
Cumulus Cells ◽  
Saline Control ◽  
Cell Replication ◽  
Progesterone Secretion ◽  
Igf I ◽  
Negative Effect ◽  
Positive Effect

There are two types of granulosa cells: those which surround the oocyte are cumulus cells (CC) and those which surround the antrum are mural granulosa cells (MGC). These cells are under the influence of several hormones and growth factors, the most important of which are gonadotrophins and IGF-I. In this article, we report novel observations on the differences between these two types of granulosa cells and their interaction with gonadotrophins and IGF-I. We were able to conduct physiological studies on the role of IGF-I by using an analogue of IGF-I which does not bind to IGF-I-binding proteins (LR3-IGF-I). Immature rats received saline, equine chorionic gonadotrophin (eCG), LR3-IGF-I or eCG plus LR3-IGF-I by infusion using a pump from 24-29 days of age. The rats were killed and the ovaries removed. Surface follicles were punctured and MGC and oocyte cumulus complexes were removed. These were cultured in saline (control) and in three different doses of FSH. Cell replication was assessed by 3H-thymidine incorporation and differentiation was evaluated by the measurement of progesterone secretion. It was noted that CC replicated ten times more than MGC. Similarly, progesterone secretion by CC was six times more than by MGC. In vivo exposure to gonadotrophins (eCG) positively influenced in vitro treatment with FSH in both cell types. This phenomenon was observed in both cell replication and progesterone secretion. The IGF-I analogue had a positive effect on cell replication of MGC but a negative effect on the cell replication of CC. With respect to progesterone secretion, the IGF-I analogue had a negative effect on CC but a positive effect on MGC. In conclusion, CC behaved differently from MGC in response to gonadotrophins and the IGF-I analogue. IGF-I and FSH acted additively, synergistically or antagonistically in different circumstances.

Effects of gonadotropins and insulin-like growth factors-I and -II on in vitro steroid production by bovine granulosa cells

1998 ◽  
Vol 78 (4) ◽  
pp. 587-597 ◽  
Author(s):  
M. Y. Yang ◽  
R. Rajamahendran
Keyword(s):  
Growth Factors ◽  
Granulosa Cells ◽  
Culture System ◽  
Physiological Role ◽  
Serum Free ◽  
Igf I ◽  
The Media ◽  
Estradiol 17Β ◽  
Insulin Like Growth Factors

The objectives of this study were: 1) to develop a bovine granulosa cell (GC) culture system; and 2) to use this system to evaluate the effects of gonadotropins (FSH and LH) and insulin-like growth factors-I and -II (IGF-I and IGF-II) on steroidogenesis of bovine GC derived from small, medium, and large antral follicles (diameters ≤4, 5–8 and >8 mm, respectively). Granulosa cells were cultured (concentration, 5 × 105 cells per well) in serum-free medium for 48 h with variable doses of hormones and growth factors. Concentrations of progesterone (P4) and estradiol-17β (E2) in the media were determined by radioimmunoassay. Basal E2 production by GC from follicles of all sizes decreased with time of culture (P < 0.01) while basal P4 production increased (P < 0.01). Basal E2 and P4 production increased with increasing size of follicles (P < 0.01). Only very low concentrations of FSH stimulated E2 production from medium and large follicles. Follicle-stimulating hormone stimulated P4 production by GC of follicles of all sizes (P < 0.05). Luteinizing hormone inhibited E2 production by GC in medium and large follicles (P < 0.05), suggesting that LH is responsible for the rise in plasma E2 through effects on both theca cells and GC. A dose of 100 ng mL−1 of IGF-I increased E2 production by GC from medium and large follicles (P < 0.05). Progesterone production by GC from all categories of follicles was also stimulated by IGF-I (P < 0.05). Estradiol-17β production by GC from large follicles decreased in response to IGF-II (P < 0.05). The physiological role of IGF-II on steroidogenesis in the bovine ovary remains to be elucidated. In summary, these results demonstrate the development of a serum-free culture system for bovine GC, and that FSH, LH, IGF-I and IGF-II have different effects on steroidogenesis by bovine GC from different size follicles. Key words: Granulosa cells, gonadotropins, Insulin-like growth factors, progesterone, estradiol-17β, cows

scholarly journals IGF I autoregulation in porcine granulosa cells in vitro.

1993 ◽  
Vol 33 (1) ◽  
pp. 83-83
Author(s):  
F. Hatey ◽  
V. Dupouy ◽  
I. Langlois ◽  
A. Bonnet ◽  
P. Mulsant ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Porcine Granulosa Cells ◽  
Igf I

scholarly journals Amphetamine-Decreased Progesterone and Estradiol Release in Rat Granulosa Cells: The Regulatory Role of cAMP- and Ca2+-Mediated Signaling Pathways

Biomedicines ◽  
2021 ◽  
Vol 9 (5) ◽  
pp. 493
Author(s):  
 Chung-Yu Chen ◽  
Chien-Rung Chen ◽  
Chiao-Nan Chen ◽  
Paulus S. Wang ◽  
Toby Mündel ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Prostaglandin F2α ◽  
Inhibitory Effect ◽  
Inhibitory Effects ◽  
Steroidogenic Enzyme ◽  
Estradiol Secretion ◽  
Basal Release ◽  
Ca2 Channels

The purpose of this study is to evaluate the amphetamine effects on progesterone and estradiol production in rat granulosa cells and the underlying cellular regulatory mechanisms. Freshly dispersed rat granulosa cells were cultured with various test drugs in the presence of amphetamine, and the estradiol/progesterone production and the cytosolic cAMP level were measured. Additionally, the cytosolic-free Ca2+ concentrations ([Ca2+]i) were measured to examine the role of Ca2+ influx in the presence of amphetamine. Amphetamine in vitro inhibited both basal and porcine follicle-stimulating hormone-stimulated estradiol/progesterone release, and amphetamine significantly decreased steroidogenic enzyme activities. Adding 8-Bromo-cAMP did not recover the inhibitory effects of amphetamine on progesterone and estradiol release. H89 significantly decreased progesterone and estradiol basal release but failed to enhance a further amphetamine inhibitory effect. Amphetamine was capable of further suppressing the release of estradiol release under the presence of nifedipine. Pretreatment with the amphetamine for 2 h decreased the basal [Ca2+]i and prostaglandin F2α-stimulated increase of [Ca2+]i. Amphetamine inhibits progesterone and estradiol secretion in rat granulosa cells through a mechanism involving decreased PKA-downstream steroidogenic enzyme activity and L-type Ca2+ channels. Our current findings show that it is necessary to study the possibility of amphetamine perturbing reproduction in females.

RGD-mediated adhesion of porcine granulosa cells modulates their differentiation response to FSH in vitro

1995 ◽  
Vol 83 (2-3) ◽  
pp. 169-177 ◽  
Author(s):  
Béatrice Goxe ◽  
Jacques E. Flechon ◽  
Solange Delasalle ◽  
Roland Salesse
Keyword(s):  
Granulosa Cells ◽  
Porcine Granulosa Cells

COMPARISON OF THE STIMULATORY EFFECTS OF OVINE, PORCINE AND HUMAN FOLLICLE-STIMULATING HORMONE AND OF OVINE AND HUMAN LUTEINIZING HORMONE ON THE ACCUMULATION OF CYCLIC AMP BY PORCINE GRANULOSA CELLS

1979 ◽  
Vol 80 (1) ◽  
pp. 9-20 ◽  
Author(s):  
ADA M. LINDSEY ◽  
CORNELIA P. CHANNING
Keyword(s):  
Cyclic Amp ◽  
Granulosa Cells ◽  
Incubation Medium ◽  
Follicle Stimulating Hormone ◽  
Similar Response ◽  
Intracellular Accumulation ◽  
Porcine Granulosa Cells ◽  
Tenfold Increase ◽  
Incubation Periods

The effects of ovine, porcine and human FSH, and ovine and human LH on the accumulation of cyclic AMP by porcine granulosa cells obtained from follicles at various stages of maturation were investigated. During incubation periods of 15 min, 10 μg ovine FSH pretreated with antiserum to LH or 10 μg human FSH resulted in an 11- to 18-fold, five-to ninefold, and less than a twofold increase in intracellular accumulation of cyclic AMP by granulosa cells from small (1–2 mm), medium (3–5 mm) and large (6–12 mm) follicles respectively. Similar patterns of response occurred with addition of porcine FSH. After incubation for 30 and 60 min with ovine, porcine or human FSH, significant accumulation of cyclic AMP in the incubation medium occurred with cells obtained from small and medium-sized follicles. After 60 min of incubation with FSH the accumulation of cyclic AMP in the incubation medium exceeded the intracellular cyclic AMP levels in granulosa cells from small and medium-sized follicles. During incubation periods of 15 min, 1·0 μg ovine LH resulted in less than a twofold, a fourfold and greater than a tenfold increase in intracellular accumulation of cyclic AMP by granulosa cells from small, medium and large follicles respectively. Addition of human LH brought about a similar response. Incubation periods of 30 and 60 min with 1·0 μg ovine or human LH resulted in significant accumulation of cyclic AMP in the incubation medium by granulosa cells from large follicles; cyclic AMP content in the incubation medium was greater after 60 min compared with 30 min of incubation. It was concluded that ovine FSH pretreated with an antiserum to LH had similar effects on cyclic AMP levels as did purified human and porcine FSH, and that the stimulatory effects of the less pure ovine FSH were probably not due to an impurity in the FSH preparation. Porcine granulosa cells obtained from small follicles should be suitable as an in-vitro FSH bioassay while granulosa cells obtained from large follicles should be suitable as an in-vitro LH bioassay.

scholarly journals GH/IGF e neoplasia: o que há de novo nesta associação

2005 ◽  
Vol 49 (5) ◽  
pp. 833-842 ◽  
Author(s):  
Angela M. Spinola e Castro ◽  
Gil Guerra-Júnior
Keyword(s):  
Growth Factors ◽  
Binding Proteins ◽  
De Novo ◽  
Igf Binding Proteins ◽  
Igf I ◽  
Ciclo Celular ◽  
Igf Binding ◽  
Insulin Like Growth Factors

Estudos in vitro e em animais sugerem que os membros do sistema insulin-like growth factors (IGFs), incluindo IGF-I, IGF-II, receptores de IGF-I e IGF-II (IGF-IR e IGF-IIR), e as IGF-binding proteins (IGFBPs) podem ter um importante envolvimento no desenvolvimento e na progressão de neoplasias. Mais especificamente, as IGFs promovem a progressão do ciclo celular e inibem a apoptose tanto por ação direta com outros fatores de crescimento como por ação indireta interagindo com outros sistemas moleculares intracelulares envolvidos na promoção e/ou progressão do câncer. Além disso, inúmeros estudos epidemiológicos têm sugerido que concentrações elevadas das IGFs, independente das alterações nas IGFBPs, podem estar associadas a um aumento no risco de desenvolver determinadas neoplasias. Esta revisão tem como objetivo apresentar o envolvimento do sistema IGF na regulação tumoral, os principais estudos epidemiológicos realizados e o risco de desenvolvimento de neoplasia em pacientes (com ou sem história pessoal de neoplasia prévia) que receberam hormônio de crescimento (rhGH). É importante salientar que o uso clínico de rhGH, nas indicações aprovadas internacionalmente, é seguro e não existem evidências, até o momento, da associação com o desenvolvimento de neoplasias.

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