Role of granulosa and theca cells on in vitro progesterone production in preovulatory follicles of the Japanese quail.

IGFs are known to be key regulators of ovarian follicular growth in eutherian mammals, but little is known regarding their role in marsupials. To better understand the potential role of IGFs in the regulation of follicular growth in marsupials, expression of mRNAs encoding IGF1, IGF2, IGF1R, IGF-binding protein 2 (IGFBP2), IGFBP4 and IGFBP5 was localized by in situ hybridization in developing ovarian follicles of the brushtail possum. In addition, the effects of IGF1 and IGF2 on granulosa cell function were tested in vitro. Both granulosa and theca cells synthesize IGF mRNAs, with the theca expressing IGF1 mRNA and granulosa cell expressing IGF2 mRNA. Oocytes and granulosa cells express IGF1R. Granulosa and theca cells expressed IGFBP mRNAs, although the pattern of expression differed between the BPs. IGFBP5 mRNA was differentially expressed as the follicles developed with granulosa cells of antral follicles no longer expressing IGFBP5 mRNA, suggesting an increased IGF bioavailability in the antral follicle. The IGFBP protease, PAPPA mRNA, was also expressed in granulosa cells of growing follicles. Both IGF1 and IGF2 stimulated thymidine incorporation but had no effect on progesterone production. Thus, IGF may be an important regulator of ovarian follicular development in marsupials as has been shown in eutherian mammals.

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Prostaglandin receptors and role of G protein-activated pathways on corpora lutea of pseudopregnant rabbit in vitro

Journal of Endocrinology ◽

10.1677/joe.0.1680141 ◽

2001 ◽

Vol 168 (1) ◽

pp. 141-151 ◽

Cited By ~ 31

Author(s):

C Boiti ◽

D Zampini ◽

M Zerani ◽

G Guelfi ◽

A Gobbetti

Keyword(s):

Protein Kinase ◽

G Protein ◽

Corpora Lutea ◽

Progesterone Production ◽

Kinase C ◽

Ligand Interactions ◽

Pka Pathway ◽

Oxide Synthase

Studies were conducted to characterize receptors for prostaglandin (PG) F(2alpha) (PGF(2alpha)) and PGE(2), and the signalling pathways regulating total nitric oxide synthase activity and progesterone production in rabbit corpora lutea (CL) of different luteal stages. CL were obtained at days 4, 9 and 13 of pseudopregnancy and cultured in vitro for 2 h with PGF(2alpha) or PGE(2) and with activators and inhibitors of G protein (Gp), phospholipase C (PLC), protein kinase C (PKC), adenylate cyclase (AC) and protein kinase A (PKA). High affinity PGF(2alpha) receptor (K(d)=1.9+/-0.6 nM mean+/-s.e.m. ) concentrations increased (P< or =0.01) four- to five-fold from early to mid- and late-luteal phases (50.6+/-8.5, 188.3+/-36.1 and 231.4+/-38.8 fmol/mg protein respectively). By contrast, PGE(2) receptor (K(d)=1.6+/-0.5 nM) concentrations decreased (P< or =0.01) from day 4 to day 9 and 13 (27.5+/-7.7, 12.4+/-2.4 and 16.5+/-3.0 fmol/mg protein respectively). The Gp-dependent AC/PKA pathway was triggered only on day 4 CL, mimicking the PGE(2) treatment and increasing progesterone production. In both day 9 and day 13 CL, the Gp-activated PLC/PKC pathway evoked a luteolytic effect similar to that induced by PGF(2alpha). The time-dependent selective resistance to PGF(2alpha) and PGE(2) by rabbit CL is mediated by factors other than a lack of luteal receptor-ligand interactions.

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Tokishakuyakusan Stimulates Progesterone and Estradiol-17 β Production by Rat Granulosa Cells and Progesterone, Testosterone and Estradiol-17 β by the Residual Portion of the Follicle in Vitro

The American Journal of Chinese Medicine ◽

10.1142/s0192415x91000223 ◽

1991 ◽

Vol 19 (02) ◽

pp. 155-161 ◽

Cited By ~ 4

Author(s):

Satoshi Usuki

Keyword(s):

Granulosa Cells ◽

Cell Level ◽

Testosterone Production ◽

Progesterone Production ◽

Preovulatory Follicles ◽

Immature Rats ◽

Serum Gonadotropin ◽

Tissue Compartments ◽

Stimulation Of

To examine the possible effects of Tokishakuyakusan (TS) on steroidogenesis by preovulatory follicles at the cell level, the expressed granulosa cells and remaining portion of follicles from pregnant mare's serum gonadotropin (PMS)-treated immature rats were incubated in vitro with increasing concentrations of TS for 3 h. TS significantly stimulated progesterone and estradiol-17 b production, with a predominant stimulation of progesterone, by the expressed granulosa cells, while testosterone production was not stimulated. In the remaining portion of the follicle, TS also significantly stimulated progesterone, testosterone and estradiol-17 b production. Similar to the effect produced by granulosa cells, the stimulatory effect of TS was stronger on progesterone than on testosterone and estradiol-17 b production. These results suggest that TS has a potent, direct stimulatory effect on steroidogenesis, especially progesterone production, by constituent tissue compartments of rat preovulatory follicles in vitro.

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Regulation of interleukin-11 expression in ovulatory follicles of the rat ovary

Reproduction Fertility and Development ◽

10.1071/rd16460 ◽

2017 ◽

Vol 29 (12) ◽

pp. 2437 ◽

Cited By ~ 4

Author(s):

You-Jee Jang ◽

Jae-Il Park ◽

Seong-Eun Jeong ◽

You-Mi Seo ◽

Phuong T. M. Dam ◽

...

Keyword(s):

Protein Kinase ◽

Regulatory Protein ◽

In Situ Hybridisation ◽

Mitogen Activated Protein Kinase ◽

Mrna Levels ◽

Α Subunit ◽

Theca Cells ◽

Progesterone Production ◽

Preovulatory Follicles ◽

Mitogen Activated Protein

The aim of the present study was to examine the regulation of interleukin (IL)-11 expression, as well as the role of IL-11, during ovulation in gonadotropin-primed immature rats. Injection of equine chorionic gonadotropin (eCG), followed by human CG (hCG) to induce superovulation stimulated expression of the Il11 gene in theca cells within 6 h, as revealed by northern blot and in situ hybridisation analyses. Real-time reverse transcription–polymerase chain reaction analysis showed that the IL-11 receptor, α subunit gene was expressed in granulosa and theca cells and that injection of hCG had no effect on its expression. IL-11 protein expression was stimulated in theca cells by hCG. LH-stimulated increases in Il11 mRNA levels in cultured preovulatory follicles were inhibited by protein kinase A and mitogen-activated protein kinase kinase inhibitors. Toll-like receptor (TLR) 2 and TLR4 were detected in preovulatory follicles, and the TLR4 ligand lipopolysaccharide, but not the TLR2 ligand Pam3Cys, increased Il11 mRNA levels in theca cells, but not in granulosa cells. Treatment of preovulatory follicles with IL-11 stimulated progesterone production and steroidogenic acute regulatory protein (Star) gene expression. Together, these results indicate that IL-11 in theca cells is stimulated by mitogen-activated protein kinase signalling and TLR4 activation, and increases progesterone production during ovulation.

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Spatiotemporal expression of aquaporin 9 is critical for the antral growth of mouse ovarian follicles†

Biology of Reproduction ◽

10.1093/biolre/ioaa108 ◽

2020 ◽

Vol 103 (4) ◽

pp. 828-839

Author(s):

Sungeun Lee ◽

Hee-Gyoo Kang ◽

Chongsuk Ryou ◽

Yong-Pil Cheon

Keyword(s):

Chorionic Gonadotropin ◽

Follicle Cell ◽

Cell Types ◽

Physiological Status ◽

Dependent Manner ◽

Specific Expression ◽

Theca Cells ◽

Spatiotemporal Expression ◽

Preovulatory Follicles

Abstract Although a few aquaporins (AQPs) expressed in granulosa cells have been postulated to mediate fluid passage into the antrum, the specific expression of AQPs in different follicle cell types and stages and their roles have not been evaluated extensively. The spatiotemporal expression of aquaporin (Aqp) 7, 8, and 9 and the functional roles of Aqp9 in antral growth and ovulation were examined using a superovulation model and 3-dimensional follicle culture. Aqp9 was expressed at a high level in the rapid growth phase (24–48 h post equine chorionic gonadotropin (eCG) for superovulation induction) compared to Aqp7 (after human chorionic gonadotropin (hCG)) and Aqp8 (8–24 h post eCG and 24 h post hCG). A dramatic increase in the expression and localization of Aqp9 mRNA in theca cells was observed, as evaluated using quantitative reverse transcription-polymerase (RT-PCR) coupled with laser capture microdissection and immunohistochemistry. AQP9 was located primarily on the theca cells of the tertiary and preovulatory follicles but not on the ovulated follicles. In phloretin-treated mice, the diameter of the preovulatory follicles and the number of ovulated oocytes decreased. Consistent with these findings, knocking down Aqp9 expression with an Aqp9 siRNA inhibited follicle growth (0.28:1 = siRNA:control) and decreased the number of ovulated follicles (0.36:1 = siRNA:control) during in vitro growth and ovulation induction. Based on these results, the expression of AQPs is under the control of the physiological status, and AQP9 expression in theca during folliculogenesis is required for antral growth and ovulation in a tissue-specific and stage-dependent manner.

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Modulating effect of growth hormone on the functional state of cultured cells from hen preovulatory follicles

Genetika i razvedenie zhivotnyh ◽

10.31043/2410-2733-2021-4-108-113 ◽

2022 ◽

pp. 108-113

Author(s):

A. Smekalova ◽

O. Mityashova ◽

O. Aleinikova ◽

E. Montvila ◽

I. Lebedeva

Keyword(s):

Granulosa Cells ◽

Final Stage ◽

Proliferative Activity ◽

Laying Hens ◽

Theca Cells ◽

Apoptotic Protein ◽

Thecal Cells ◽

Preovulatory Follicles ◽

Regulation Of Growth

Somatotropic hormone (STH) is an important positive modulator of ovarian function in mammals. Local production of STH and the expression of the corresponding specific receptors were also detected in hen ovarian follicles, which indicates the participation of this hormone in the endocrine/paracrine control of folliculogenesis in birds. Nevertheless, the role of STH in the regulation of growth of avian follicles at the final stage of maturation is still not clear.Objective: To study in vitro the effect of STH on the proliferative activity and apoptotic changes of granulosa and theca cells from preovulatory follicles of domestic hens.Materials and methods. Young laying hens aged 34-35 weeks with a long clutch were used in the experiments. Granulosa and theca cells were isolated from the largest yellow follicle in the hierarchy (F1). The cells were cultured in a medium containing 10% fetal bovine serum until a monolayer was formed, and then for 24 h in the medium without serum in the absence (control) or in the presence of STH at various concentrations (1-100 ng/ml). The proliferative activity and apoptotic changes in the cells were assessed by immunocytochemical assay, based on the expression level of proliferating cell nuclear antigen PCNA and pro-apoptotic protein Bax, respectively.Results. The proportion of PCNA-positive granulosa cells increased 1.3-1.8 times (P<0.01-0.05) as compared to control with increasing the content of STH in the medium to 10-100 ng/ml. Furthermore, within this concentration range, the studied hormone reduced 1.2-1.6 times (P<0.05) the relative number of granulosa cells with the positive reaction to Bax. The sensitivity of theca cells to the growth-stimulating effect of STH was lower than that of granulosa cells. Such the effect of STH led to an increase in the proportion of PCNA-positive thecal cells by 1.2-1.3 times (P<0.05) and was detected only at concentrations of 25 and 100 ng/ml. Meanwhile, STH (25-100 ng/ml) increased 1.3 times (P<0.05) the level of Bax expression in theca cells.Conclusions. The results of the present study indicate the stimulating effect of STH in vitro on the proliferative activity of granulosa and theca cells from the most mature hen preovulatory follicle. In addition, STH is able to reduce the expression of the pro-apoptotic protein Bax in granulosa cells and increase this expression in thecal cells. Thus, the data obtained indicate the possible participation of STH in the regulation of growth and development of follicles at the final stage of maturation during the period of maximum egg-laying intensity in laying hens.

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Involvement of theca cells and steroids in the regulation of granulosa cell apoptosis in rabbit preovulatory follicles

Reproduction ◽

10.1530/rep.0.1250709 ◽

2003 ◽

pp. 709-716 ◽

Cited By ~ 5

Author(s):

G Maillet ◽

A Benhaim ◽

H Mittre ◽

C Feral

Keyword(s):

Granulosa Cell ◽

Granulosa Cells ◽

Cell Apoptosis ◽

Annexin V ◽

Rapid Loss ◽

Theca Cells ◽

Preovulatory Follicles ◽

Apoptotic Effect

Follicular atresia is characterized by a rapid loss of granulosa cells and, to a lesser extent, theca cells, via apoptosis. The aim of this study was to investigate the possible involvement of theca cell secretions in the regulation of apoptosis of rabbit granulosa cells. The annexin-V binding method based on externalization of phosphatidylserine to the outer layer of plasma membrane during apoptosis was used to detect apoptotic granulosa cells in flow cytometry. Regulation of apoptosis of granulosa cells was studied in three different culture systems: (i) isolated cultured granulosa cells, (ii) granulosa cells obtained from cultured preovulatory follicles and (iii) granulosa cells co-cultured with theca cells. The results of this study indicate that: (i) the rate of apoptosis of granulosa cells was significantly reduced when granulosa cells were co-cultured with theca cells or obtained from cultured preovulatory follicles in comparison with isolated cultured granulosa cells; (ii) FSH exerts its anti-apoptotic effect only on granulosa cells issued from cultured preovulatory follicles; (iii) ovarian steroids do not affect the percentage of isolated apoptotic granulosa cells; and (iv) the occurrence of an apoptotic process in rabbit theca cells could be upregulated in vitro by hCG and an analogue of the gonadotrophin second messenger cAMP. The results of this study indicate that in rabbits (i) steroids were ineffective in vitro in protecting isolated granulosa cells against apoptosis in comparison with observations in vivo in rats, and (ii) the presence of theca cells was efficient to reduce granulosa cell apoptosis but not sufficient to allow the anti-apoptotic effect of gonadotrophins observed in cultured follicles.

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Effects of inhibin-related peptides and oestradiol on androstenedione and progesterone secretion by bovine theca cells in vitro

Journal of Endocrinology ◽

10.1677/joe.0.1450491 ◽

1995 ◽

Vol 145 (3) ◽

pp. 491-500 ◽

Cited By ~ 58

Author(s):

J H M Wrathall ◽

P G Knight

Keyword(s):

Granulosa Cell ◽

Thecal Cell ◽

Theca Cells ◽

Progesterone Secretion ◽

Pretreatment Period ◽

Direct Contrast ◽

Primary Monolayer ◽

Dose Dependent

Abstract Primary monolayer cultures of bovine theca cells isolated from pooled ovarian follicles (3–10 mm diameter) were used to examine the effects of various granulosa cell-derived substances on basal and luteinizing hormone (LH)-induced androgen and progesterone secretion. After an overnight pretreatment period, cells were incubated with a range of treatments including LH, oestradiol-17β, inhibin, activin and follistatin. Media were collected after 48 h and assessment of androstenedione and progesterone secretion made by radioimmunoassay. Addition of LH (5–50 ng/ml) to the cells resulted in a dose-dependent stimulation of both androstenedione (2·5-to 3-fold rise; P<0·01) and progesterone (∼ 1·6-fold rise; P<0·001) production. Secretion of androstenedione was also raised (up to 5-fold; P<0·001) by addition of oestradiol-17β (0·3–300 ng/ml), whilst levels of the androgen in the presence of both LH (20 ng/ml) and oestradiol (300 ng/ml) were up to 12-fold higher (P<0·001) than control values. In contrast, oestradiol treatment inhibited by up to 50% both basal (P<0·001) and LH-stimulated (P<0·001) secretion of progesterone. Exposure of cells to purified bovine inhibin (5–125 ng/ml) consistently raised androstenedione secretion by up to 42% over basal levels (P<0·001). Inhibin also enhanced both LH-stimulated (∼20%; P<0·001) and oestradiol-stimulated (∼20%; P<0·05) secretion of androstenedione. In direct contrast, treatment of theca cells with human recombinant activin-A (1–50 ng/ml) inhibited both LH-stimulated (∼50%; P<0·001) and oestradiol-stimulated (∼30%; P<0·005) androstenedione secretion. Activin also reversed the positive effect of inhibin on basal (P<0·01), LH-stimulated (P<0·001) and oestradiol-stimulated (P<0·001) androstenedione secretion, though activin alone did not affect basal steroid output. Simultaneous addition of human recombinant follistatin reversed the inhibitory effects of activin on LH- and oestradiol-induced androstenedione secretion but did not modify the effects of inhibin. Follistatin alone did not alter either basal or LH-stimulated androstenedione output. Neither basal nor LH-stimulated secretion of progesterone were consistently affected by inhibin, activin or follistatin. As well as confirming the stimulatory effects of both LH and oestradiol on bovine thecal cell androgen production, these observations are indicative of opposing intrafollicular paracrine roles for granulosa cell-derived inhibin and activin in modulating thecal cell responses to gonadotrophins and steroids in the bovine ovary. Though inhibin and oestradiol had qualitatively similar effects in promoting thecal androgen secretion, the magnitude of the response to oestradiol was much greater. The results also support an intrafollicular role of follistatin as a binding protein capable of neutralizing the effect of activin, but not inhibin, on thecal androgen production. Journal of Endocrinology (1995) 145, 491–500

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Fibronectin production by chicken granulosa cells in vitro: effect of follicular development

Acta Endocrinologica ◽

10.1530/acta.0.1270466 ◽

1992 ◽

Vol 127 (5) ◽

pp. 466-470 ◽

Cited By ~ 11

Author(s):

Elikplimi K Asem ◽

Jacqueline A Carnegie ◽

Benjamin K Tsang

Keyword(s):

Granulosa Cells ◽

Follicular Development ◽

Enzyme Linked Immunosorbent Assay ◽

Dependent Manner ◽

Ovarian Granulosa Cells ◽

Preovulatory Follicles ◽

Vitro Effect ◽

Dose Dependent

In vitro studies were conducted to investigate the role of chicken ovarian granulosa cells in the production of fibronectin, a component of the basal lamina of ovarian follicles. Collagenase dispersed granulosa cells obtained from the first (F1; about 35 mm in diameter) and third (F3; 15–20 mm in diameter) largest preovulatory follicles, as well as from a pool of small yellow follicles (SF; 6–10 mm in diameter), were incubated in serum-free medium-199 for 24 to 96 h in the absence and presence of luteinizing hormone (LH) or forskolin. Fibronectin secreted in the medium was quantitated by enzyme linked immunosorbent assay. Basal fibronectin production (which increased with the duration of incubation) was significantly greater (p<0.001) in granulosa cells derived from mature follicle (F1) than in F3 or SF cells. Both LH and forskolin stimulated fibronectin production in SF and F3 cells in a dose-dependent manner; however, they were without effect in F1 cells. The magnitude of increase in fibronectin production elicited by LH or forskolin was greater in SF cells than in F3 cells. The cytoplasm of cultured granulosa cells taken at all stages of follicular development stained positively for fibronectin. These findings indicate that chicken granulosa cells produce fibronectin. This ability is acquired early in follicular development and the stimulatory effect of the gonadotropin (LH) diminished as the follicle approached ovulation.

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