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PubMed2002

Characterization of integrin expression in the mouse ovary.

Integrin alpha:beta heterodimers mediate cell contacts to the extracellular matrix and initiate intracellular signaling cascades in response to a variety of factors. Integrins interact with many determinants of cellular phenotypes and play roles in controlling the development, structural integrity, and function of every type of tissue. Despite their importance, little is known about the regulation of integrin subunits in the mammalian ovary and how they function in folliculogenesis. To determine their relevance to ovarian physiology, we have studied the expression of integrin subunit mRNAs by Northern blot analysis and in situ hybridization in ovaries of wild-type, growth differentiation factor 9 (Gdf 9) knockout, FSHbeta (Fshb) knockout, and inhibin alpha (Inha) knockout mice. Integrin alpha6 mRNA is expressed in oocytes and granulosa cells of single-layer follicles and in oocytes and theca cells of multilayer follicles. Integrin alpha6 is highly expressed in Gdf 9 knockout ovaries, which are enriched in oocytes and primary (single layer) follicles because of a block at this stage of follicular development. Integrin alpha(v) mRNA is most highly expressed in the granulosa cells of multilayer growing follicles, and therefore only low levels of expression are detectable in the Gdf 9 knockout ovaries. Integrin beta1 mRNA exhibits a broad expression pattern in ovaries, including oocytes, granulosa cells, theca cells, and corpora lutea. Integrin beta3 mRNA is expressed in theca and interstitial cells and is upregulated in corpora lutea. It is nearly undetectable in ovaries of Fshb knockout mice, which develop preantral follicles but have no luteal cells. Integrin beta5 mRNA is predominantly expressed in granulosa cells of multilayer follicles. It is expressed at high levels in the Fshb knockout mice and in a compartmentalized manner in the granulosa cell/Sertoli cell tumors that develop in the Inha knockout mice. Specific integrins are associated with ovarian cellular phenotypes in mice, which raises intriguing possibilities as to integrin functions in oocyte competence, follicular development, luteinization, and granulosa cell proliferation.

Animals↗
PubMed2004

Bone morphogenetic proteins (BMP) -4, -6, and -7 potently suppress basal and luteinizing hormone-induced androgen production by bovine theca interna cells in primary culture: could ovarian hyperandrogenic dysfunction be caused by a defect in thecal BMP signaling?

We reported recently that bovine theca interna cells in primary culture express several type-I and type-II receptors for bone morphogenetic proteins (BMPs). The same cells express at least two potential ligands for these receptors (BMP-4 and -7), whereas bovine granulosa cells and oocytes express BMP-6. Therefore, BMPs of intrafollicular origin may exert autocrine/paracrine actions to modulate theca cell function. Here we report that BMP-4, -6, and -7 potently suppress both basal (P < 0.0001; respective IC(50) values, 0.78, 0.30, and 1.50 ng/ml) and LH-induced (P < 0.0001; respective IC(50) values, 5.00, 0.55, and 4.55 ng/ml) androgen production by bovine theca cells while having only a moderate effect on progesterone production and cell number. Semiquantitative RT-PCR showed that all three BMPs markedly reduced steady-state levels of mRNA for P450c17. Levels of mRNA encoding steroidogenic acute regulatory protein, P450scc, and 3beta-hydroxy- steroid dehydrogenase were also reduced but to a much lesser extent. Immunocytochemistry confirmed a marked reduction in cellular content of P450c17 protein after BMP treatment (P < 0.001). Exposure to BMPs led to cellular accumulation of phosphorylated Smad1, but not Smad2, confirming that the receptors signal via a Smad1 pathway. The specificity of the BMP response was further explored by coincubating cells with BMPs and several potential BMP antagonists, chordin, gremlin, and follistatin. Gremlin and chordin were found to be effective antagonists of BMP-4 and -7, respectively, and the observation that both antagonists enhanced (P < 0.01) androgen production in the absence of exogenous BMP suggests an autocrine/paracrine role for theca-derived BMP-4 and -7 in modulating androgen production. Collectively, these data indicate that an intrafollicular BMP signaling pathway contributes to the negative regulation of thecal androgen production and that ovarian hyperandrogenic dysfunction could be a result of a defective autoregulatory pathway involving thecal BMP signaling.

Androgens↗
PubMed1990

Site at which ovarian nerve extracts inhibit thecal androgen production.

We have recently reported that extracts of the superior ovarian nerve (SON) of rats inhibit porcine theca cell androstenedione production. Theca cells obtained from prepubertal gilts were cultured under serum-free conditions for 48 h. The inhibitory effect of SON extracts occurred within 6 h of treatment, was irreversible and independent of the dose of luteinizing hormone (LH) employed. The SON extracts' actions were exerted at a site(s) distal to the generation of adenosine 3',5'-monophosphate (cyclic AMP), since they did not affect extracellular cyclic AMP accumulation, while causing a significant inhibition of androstenedione production. The SON extract decreased 17 alpha-hydroxyprogesterone, androstenedione, testosterone, estradiol and estrone production while increasing progesterone and pregnenolone production, suggesting that the SON extract causes an inhibition of the 17 alpha-hydroxylase:C17-20 lyase complex. These results indicate that a factor(s) in the SON may play an important role in the regulation of follicular development, since thecal androgens are substrates for granulosa cell estrogen biosynthesis and are also involved in follicular atresia.

17-alpha-Hydroxyprogesterone↗
PubMed1994

Insulin-like growth factor-I regulation of luteinizing hormone (LH) receptor messenger ribonucleic acid expression and LH-stimulated signal transduction in rat ovarian theca-interstitial cells.

Currently available evidence supports the hypothesis that insulin-like growth factor-I (IGF-I) secreted by small preantral follicles may be involved in stimulating the initial differentiation of the theca interna and, in particular, expression of the LH receptor in pre-theca cells. To test this hypothesis, we examined the effects of IGF-I on LH receptor mRNA expression in theca-interstitial cells (TIC) isolated from the ovaries of hypophysectomized immature rats by percoll gradient centrifugation. TIC (3.5 x 10(4) viable cells/well) were cultured up to 6 days with and without LH (0-10 ng/ml) and IGF-I (0-100 ng/ml). Androsterone in the medium was measured by RIA, and LH receptor mRNA was measured by specific reverse transcriptase-polymerase chain reaction assay. LH receptor mRNA was low in control (untreated) TIC. IGF-I stimulated a dose-related increase (2-fold) in LH receptor mRNA at 2 days (ED50 = 9.0 +/- 1.9 ng/ml) that remained constant at 4 days and then declined to basal levels at 6 days. LH stimulated a dose-related (ED50 = 17.6 +/- 1.0 pg/ml) increase in LH receptor mRNA that reached a maximum of 4-fold at 2 days. At 4 days, LH down-regulated LH receptor mRNA below basal levels, and it had no effect at 6 days. Addition of IGF-I (30 ng/ml) to LH-treated TIC abolished the stimulatory effect of LH throughout the culture period. LH receptor mRNA was highly sensitive to LH since the ED50 was approximately 2.5-fold lower than for stimulation of androsterone production (39.8 +/- 3.8 pg/ml). To understand the molecular mechanism of the synergistic stimulation of androgen production by IGF-I and LH, the effects of IGF-I on the cAMP/protein kinase A (PKA) signaling pathway were examined. When freshly isolated TIC were challenged with IGF-I alone (30 ng/ml), there was no effect on cAMP production or PKA activity, but IGF-I augmented LH stimulation of cAMP production slightly at high concentrations of LH and blocked stimulation of PKA activity by a saturating concentration of LH (3 ng/ml), suggesting that IGF-I increased LH down-regulation of PKA. We next examined the effects of IGF-I on LH receptor number. When TIC were placed into culture, LH/hCG binding sites decreased to approximately 35% of the initial number at 24 h and 25% at 2 days. This decrease was accompanied by a similar loss of cholera toxin- and hCG-stimulated cAMP production.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1995

Epidermal growth factor and its receptor gene expression and peptide localization in porcine ovarian follicles.

The present study was undertaken to determine the expression of genes for epidermal growth factor (EGF) and its receptor (EGF-R) in various components of medium-sized porcine ovarian follicles by reverse transcription-polymerase chain reaction (RT-PCR), and to localize their peptides during folliculogenesis by immunocytochemistry. A strong band for EGF mRNA transcript was detected in the oocyte, whereas the signal in cumulus, granulosa, and theca cells was very weak but detectable. In contrast, a very strong EGF-R mRNA signal was observed in cumulus, granulosa, and theca cells, whereas the signal in the oocyte was very weak. EGF peptide was localized in the oocyte, cumulus, and granulosa cells of all stages of follicle. In the oocyte, the intensity of immunostaining was more pronounced in primordial and primary follicles, compared to atrial follicles. In large antral follicles, immunostaining was pronounced in granulosa cells, whereas theca cells showed little or no detectable staining for EGF. EGF staining was also observed in the cumulus and granulosa cells of follicles undergoing atresia. EGF-R immunostaining was observed in the oocytes of primordial and primary follicles, and in cumulus, granulosa, and theca cells of all stages of follicle, including atretic follicles. In large antral follicles, the intensity of immunostaining was more pronounced in theca cells than in granulosa cells, and the oocyte showed little or no detectable staining. No immunostaining was observed when the primary antibody was replaced with preimmune serum (EGF), or preabsorbed with the control peptide (EGF-R), confirming the specificity of the staining procedures.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1996

Steroid metabolism in theca externa cells from preovulatory follicles of domestic hen (Gallus domesticus).

The purpose of this study was to evaluate the ability of theca externa cells from preovulatory follicles of the domestic hen to metabolize tritiated steroid precursors. Theca externa cells were isolated from ovarian preovulatory follicles at three different developmental stages; F1 (35 mm), F3 (26 mm), and F5 (13 mm). Tritiated pregnenolone (P5), progesterone (P4), dehydroepiandrosterone (DHEA), androstenedione (A4), and testosterone (T) were employed as precursors and their metabolic products were evaluated after separation by thin-layer chromatography. The major metabolite of P5 by theca externa cells was 5-pregnen-3 beta, 20 beta-diol (F5, 47%; F3, 39%; and F1, 24%), but minimal quantities of P4 were detected. Progesterone metabolism yielded mainly 4-pregnen-20 beta-ol-3-one (F5, 52%; F3, 34%; and F1, 49%). When DHEA was used as precursor, A4 was produced in higher amounts (F5, 29%; F3, 23%; and F1, 11%) than estrone (E1) (F5, 1.5%; F3, 0.9%; and F1, 0.4%). Androstenedione was mainly transformed into E1 (F5, 11.9%; F3, 12.2%; and F1, 0.2%) but lower quantities of T and 17 beta-estradiol (E2) were found. Testosterone was actively transformed into A4 (F5, 50%; F3, 50%; and F1, 30%), but a low transformation to E2 (F5, 1.9%; F3, 1.7%; and F1, 1.4%) and E1 (F5, 2%; F3, 1%; and F1, 0.5%) was found. These results show that theca externa cells from preovulatory follicles of hen have enzymatic activities of 20 beta-reductase (from P5 and P4), 3 beta-hydroxysteroid dehydrogenase/5-4 isomerase (from P5 and DHEA), 17 beta-hydroxysteroid dehydrogenase (from A4 and T), and aromatase (from A4 and T). Furthermore, the enzyme activities decrease with follicular maturation, except for 20 beta-reductase which is constant. These data support the concept that theca externa cells have the ability to synthesize different steroids than reported in theca externa cells. In addition, since theca externa cells did not show the capacity to produce androgens but these steroids were aromatized to estrogens by these cells, it was suggested that the interaction between theca interna cells and theca externa cells occurs in vivo, thus supporting the multicellular theory for estrogen production.

Androstenedione↗
PubMed1995

Expression of vascular permeability factor/vascular endothelial growth factor by human granulosa and theca lutein cells. Role in corpus luteum development.

Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) is a cytokine that is overexpressed in many tumors, in healing wounds, and in rheumatoid arthritis. VPF/VEGF is thought to induce angiogenesis and accompanying connective tissue stroma in two ways: 1), by increasing microvascular permeability, thereby modifying the extracellular matrix and 2), as an endothelial cell mitogen. VPF/VEGF has been reported in animal corpora lutea and we investigated the possibility that it might be present in human ovaries and have a role in corpus luteum formation. We here report that VPF/VEGF mRNA and protein are expressed by human ovarian granulosa and theca cells late in follicle development and, subsequent to ovulation, by granulosa and theca lutein cells. Therefore, VPF/VEGF is ideally positioned to provoke the increased permeability of thecal blood vessels that occurs shortly before ovulation. VPF/VEGF likely also contributes to the angiogenesis and connective tissue stroma generation that accompany corpus luteum/corpus albicans formation. Finally, VPF/VEGF was overexpressed in the hyperthecotic ovarian stroma of Stein-Leventhal syndrome in which it may also have a pathophysiological role.

Adolescent↗
PubMed1999

Expression of the steroidogenic acute regulatory protein and luteinizing hormone receptor and their regulation by tumor necrosis factor alpha in rat corpora lutea.

Expression of both mRNA and protein of the steroidogenic acute regulatory protein (StAR), in correlation with progesterone (P) production and LH receptor (LHR) mRNA expression, was studied in the corpora lutea (CL) of gonadotropin-induced-pseudopregnant and pregnant rats at various stages of CL development. Immature female rats, 21-22 days old, were injected s.c. with 20 IU eCG to stimulate follicle growth and then with 20 IU hCG 48 h later to induce ovulation. The ovaries were removed at various stages of CL development; either CL were isolated and snap frozen for total RNA analysis, or whole ovaries were fixed in Bouin's fluid for paraffin sectioning. The results of in situ hybridization, immunohistochemistry, and Northern blotting showed that the increase in StAR mRNA and protein expression was well correlated with the increase in serum P concentration. StAR expression was restricted to the luteal cells or theca cells in antral follicles. Both StAR mRNA and protein in the CL of pseudopregnant rats increased steadily on Day 1 and Day 4, reached highest levels on Day 4, and then dropped sharply on Day 8 when luteolysis takes place. LHR mRNA content was high on Day 1 but dropped significantly on Day 2. LHR mRNA increased to high levels on Day 4 and 8 and then declined on Day 12. StAR mRNA and protein levels in the CL of pregnant rats were high during early luteal development (Day 2, 4), increased even further on Day 9, and decreased on Day 13 when luteolysis takes place. It is therefore suggested that the expression of StAR coincides well with the capacity of P production in the CL and that StAR expression can be used as a functional "marker" of CL development. To study the possible effect of cytokines on StAR expression, pseudopregnant rats on Day 5 were injected s.c. with 10 IU hCG plus 20 microg prolactin (PRL), with or without 500 IU tumor necrosis factor alpha (TNFalpha) 30 min later. TNFalpha significantly inhibited hCG/PRL-induced StAR and LHR mRNA expression at 1 and 3 h post-TNFalpha. It is suggested that the luteolytic effect of TNFalpha may be mediated by its direct inhibition on StAR expression or by an indirect decrease in LHR expression.

Animals↗
PubMed2001

Hepatocyte growth factor and stem cell factor involvement in paracrine interplays of theca and granulosa cells in the human ovary.

OBJECTIVE: To examine gene expression of hepatocyte growth factor (HGF), the receptor for HGF, c-met, and the receptor for stem cell factor (SCF), c-kit, in the human ovary and to investigate the effects of HGF and SCF on the proliferation and function of granulosa and theca cells. DESIGN: Prospective study. SETTING: University hospital. PATIENT(S): Six premenopausal women. INTERVENTION(S): Follicular fluid and granulosa cells were collected during IVF cycles. Ovarian tissues were obtained from women who underwent surgery. MAIN OUTCOME MEASURE(S): Gene expression of HGF, c-met, and c-kit in the human ovary was determined. RESULT(S): Reverse-transcription polymerase chain reaction showed the presence of HGF and c-kit mRNA in the theca and stroma cells of the ovary, whereas c-met mRNA was observed in the granulosa, theca, and stroma cells. HGF increased the expression of SCF gene in granulosa cells, and SCF reciprocally increased the expression of HGF gene in theca cells. SCF stimulated the proliferation of theca cells. HGF stimulated progesterone production in granulosa cells. CONCLUSION(S): A positive feedback loop between theca cells and granulosa cells was identified that is mediated by HGF and SCF. HGF and SCF modulate the interplay between theca and granulosa cells by promoting cell proliferation and steroid hormone production.

Adult↗
PubMed2004

Ovarian granulosa cell lines.

The ovary is a complex endocrine gland responsible for production of sex steroids and is the source of fertilizable ova for reproduction. It also produces various growth factors, transcription factors and cytokines that assist in the complex signaling pathways of folliculogenesis. The ovary possesses two primary steroidogenic cell types. The theca cells (and to a lesser extent, the stroma) are responsible for androgen synthesis, and the granulosa cells are responsible for conversion of androgens to estrogens, as well as progesterone synthesis. These cells undergo a transformation in the luteal phase of the menstrual cycle, converting them from estrogen producing, to predominantly progesterone producing cells. Understanding the molecular mechanisms regulating these cells is essential in understanding the regulation of steroidogenesis and reproduction. Creation of appropriate in vitro cell model systems can provide important tools for the study of ovarian function. This has led to the development of ovarian steroidogenic cell lines in several laboratories. Developing theca cell lines has met with limited success. Conversely, numerous human and animal granulosa cell lines have been developed. This review will discuss the existing granulosa cell lines and their characteristics.

Cell Line↗
PubMed1985

[Functional interrelationship between granulosa and theca lutein cells in steroidogenesis in vitro by human corpus luteum].

To investigate the interrelation between steroidogenic function of granulosa and theca lutein cells of the human corpus luteum, twelve corpora lutea of menstruation and pregnancy were divided into inner and outer layers, each of which was histologically confirmed to be composed of pure granulosa lutein cells (GL) or GL plus theca lutein cells (TL), respectively. Slices of each layer were incubated with radioactive pregnenolone, 17-hydroxyprogesterone and androstenedione respectively, and the metabolites were analysed by reverse dilution technique with recrystallization to constant specific activity. Both slices converted pregnenolone-14C to progesterone, 17-hydroxyprogesterone, androstenedione, estrone and estradiol-17 beta, with progesterone being the major steroid formed. Formation of progesterone was significantly pronounced in the case of slices from the inner layer, while conversion into the other steroids was significantly higher in the case of slices from the outer layer. Formation of androstenedione from 17-hydroxyprogesterone-14C was also greater with the outer slices than the inner slices. However, conversion of androstenedione-14C to radioactive estrone and estradiol-17 beta was more efficient with the inner slices than the outer slices. These results suggest that the activity of aromatizing enzymes is higher in granulosa lutein cells, while that of 17 alpha-hydroxylase and C17-C20 lyase is higher in theca lutein cells. Therefore, cooperation of the two cell types would be necessary for the fulfillment of the function of estrogen biosynthesis by human corpus luteum.

Adult↗
PubMed2003

Effects of follicular cells and FSH on the resumption of meiosis in equine oocytes matured in vitro.

It has been suggested that preculturing immature oocytes in a manner that maintains them in meiotic arrest may improve cytoplasmic maturation and, thereby, the eventual developmental competence of oocytes matured in vitro. This study examined the ability of follicular cells to maintain meiotic arrest in equine oocytes. Cumulus-oocyte complexes (COCs) recovered from dead mares were cultured for 38 h in M199 either attached to, or together with, different follicle wall components, as follows: (1) attached to the follicle wall, (2) cocultured with separated follicle wall, (3) attached to membrana granulosa (COCG), (4) COCGs cocultured with sheets of theca cells, (5) COCGs cultured in theca-cell conditioned medium, and (6) control COCs without any follicle wall components. When oocytes were cultured attached to their follicle wall, 79% remained in the GV stage throughout the 38 h incubation. However, when oocytes were cocultured with separate pieces of follicle wall, meiosis resumed and a similar proportion of oocytes progressed to metaphase II (79%) as under control conditions (84%). Only 16% of oocytes cultured while still attached to the membrana granulosa (COCGs) maintained the GV stage, whereas when COCGs were cocultured with theca cells or in theca-cell conditioned medium, significantly more oocytes remained in the GV stage (64 and 52%, respectively), indicating that theca cells secrete a meiosis-inhibiting factor. The effect of FSH on the meiosis-inhibiting activity of follicular cells was investigated by culturing COCs attached to the follicle wall and COCGs in the presence or absence of theca cells in medium containing FSH. Addition of 0.05 iu recombinant human FSH ml(-1) to the culture medium did not affect nuclear maturation and failed to overcome the suppressive effect exerted by the follicle wall or by theca cells, despite the fact that mRNA for the FSH receptor was found using RT-PCR in both cumulus and granulosa cells. These results demonstrate that the maintenance of meiotic arrest in equine oocytes during culture can be promoted by theca cells, which appear to act via a secreted inhibitory factor that cannot be suppressed or counteracted by FSH.

Animals↗
PubMed1996

Expression of steroidogenic acute regulatory protein (StAR) in the human ovary.

Expression of steroidogenic acute regulatory protein (StAR) messenger ribonucleic acid (mRNA) in human ovary and cultured proliferating granulosa-lutein cells, theca interna cells, and luteinized granulosa cells was examined. The StAR transcripts were restricted in situ to theca of preovulatory follicles and luteinized granulosa and thecal cells of the corpus luteum. The cyclic nucleotide analog, 8-bromo-cAMP (8-Br-cAMP), increased StAR mRNA in all cell types studied by a process requiring on-going RNA and protein synthesis. Phorbol myristate acetate prevented the stimulatory effects of 8-Br-cAMP. In proliferating granulosa-lutein cells, 8-Br-cAMP increased StAR gene transcription and did not significantly affect StAR mRNA stability. Forskolin treatment was also found to increase the expression of a human StAR proximal promoter-luciferase fusion gene transfected into the proliferating granulosa-lutein cells. We conclude that 1) the StAR gene is expressed in the most steroidogenic compartments of the human ovary; 2) induction of StAR gene transcription by cAMP, produced in response to the LH surge, accounts for the appearance of StAR transcripts in luteinized granulosa cells; and 3) the effects of cAMP are antagonized by activators of protein kinase C.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1998

Insulin and insulin-like growth factor I stimulate the proliferation of human ovarian theca-interstitial cells.

OBJECTIVE: To determine whether insulin and insulin-like growth factor I (IGF-I) affect the proliferation of human theca-interstitial cells. DESIGN: In vitro assays. SETTING: University laboratory. PATIENT(S): Premenopausal women undergoing oophorectomy for benign conditions. INTERVENTION(S): Purified theca-interstitial cells were cultured in chemically defined media with or without insulin and IGF-I. MAIN OUTCOME MEASURE(S): The proliferation of cells was evaluated by determination of [3H] thymidine incorporation and cell counting. RESULT(S): Insulin and IGF-I stimulated DNA synthesis by theca-interstitial cells in a dose-dependent fashion. Insulin-like growth factor I had a greater potency than did insulin. The effects of both approached, but did not reach, the level of DNA synthesis observed in cultures exposed to 10% fetal bovine serum. Direct counting of theca-interstitial cells revealed that IGF-I significantly increased the total number of cells (36% above control), whereas insulin induced a modest and statistically nonsignificant increase in the cell number (14% above control). CONCLUSION(S): The present results support the hypothesis that insulin and IGF-I promote the mitotic activity of theca-interstitial cells. These effects may represent mechanisms that lead to hyperplasia of the thecal/stromal compartment in polycystic ovary syndrome.

Adult↗
PubMed1997

Effects of insulin and insulin-like growth factors on proliferation of rat ovarian theca-interstitial cells.

Hyperplasia of the theca-interstitial (T-I) compartment, such as observed in polycystic ovary syndrome, is associated with ovarian dysfunction. Yet the mechanisms regulating proliferation of T-I cells are virtually unknown. This study was an investigation of the effects of insulin and insulin-like growth factors (IGF-I and IGF-II) on proliferation of rat T-I cells. Purified T-I cells were incubated in chemically defined media. Insulin (1-100 nM) and both IGFs (0.3-30 nM) dose-dependently stimulated DNA synthesis as determined by radiolabeled thymidine incorporation assay. IGF-I was most potent with EC50 = 1.4 +/- 0.4 nM, while IGF-II had EC50 = 4.3 +/- 0.18 nM and insulin had EC50 = 8.4 +/- 3.9 nM. The maximal effects of all three treatments were comparable. A combination of IGF-I at 10 nM (a concentration producing a near-maximal effect) with insulin or IGF-II resulted in DNA synthesis comparable to that achieved by IGF-I alone. IGF-I mutants with decreased affinity to IGF-binding proteins (IGFBPs)-long R3-IGF-I and des(1-3)IGF-I-produced greater effects on DNA synthesis than did IGF-I. The effects of insulin and IGFs on cell proliferation were confirmed by counting the steroidogenically active cells (stained positive for 3 beta-hydroxysteroid dehydrogenase [3 beta-HSD]) and steroidogenically inactive cells (3 beta-HSD negative). The number of steroidogenically active T-I cells was increased by insulin (by 3.7-fold, p < 0.001), IGF-I (by 3.2-fold, p < 0.001), and IGF-II (by 2.1-fold, p < 0.001). The number of steroidogenically inactive cells was not significantly altered. These findings indicate that 1) insulin- and IGF-dependent synthesis of DNA by T-I cells is stimulated via a common pathway, probably via type I IGF receptors; 2) endogenous IGFBPs may modify the effects of IGF-I; and 3) the increased DNA synthesis is reflected by an increase in the number of steroidogenically active cells. Insulin and the IGFs may play a role in the regulation of proliferation and differentiation of T-I cells under physiological and pathological conditions. In particular, the present observations may explain thecal and stromal hyperplasia accompanying hyperinsulinemic conditions such as polycystic ovary syndrome or hyperthecosis.

3-Hydroxysteroid Dehydrogenases↗
PubMed1977

Evidence for interaction between granulosa cells and theca in early progesterone synthesis.

The temporal characteristics of steroidogenesis in vitro by hamster preovulatory follicles, were compared to granulosa cells and theca incubated separately. Gonadotropin-stimulated intact follicles or recombined granulosa cells and theca synthesized increased amounts of progesterone by 30-120 minutes of incubation. The granulosa cells and theca, when incubated separately, did not begin to accumulate progesterone until 4 to 6 hours. The relatively rapid rise in follicular progesterone synthesis after in vitro gonadotropin stimulation follows the same time course as the rapid rise in vivo of hamster and rat preovulatory progesterone after the gonadotropin surge. The sharp differences in the temporal characteristics of progesterone synthesis between follicles and separated follicular cell types suggest an interaction between granulosa cells and theca in at least one phase of progesterone synthesis.

Animals↗
PubMed1986

Hammond memorial lecture. New concepts of the control of corpus luteum function.

Five new concepts concerning the control of corpus luteum function in the cow have been developed in recent years. Prostacyclin (PGI-2) plays a luteotrophic role. Conversely, products of the lipoxygenase pathway of arachidonic acid metabolism, particularly 5 hydroxyeicosatetraenoic acid (5-HETE), play luteolytic roles. Luteal cells arise from two sources. The small luteal cells are all of theca cell origin; the large cells found early in the cycle (Days 2-6) are mainly of granulosa cell origin. However, a population of large cells found after Day 10 of the cycle are of theca cell origin. Oxytocin of luteal cell origin plays a role in development of the corpus luteum and possibly in its regression. The recently described Ca2+-polyphosphoinositol-protein kinase C second messenger system, as well as the LH-cAMP system, is involved in control of progesterone synthesis in the bovine corpus luteum. Progesterone synthesis in the small theca-derived luteal cells is primarily controlled by the cAMP system. However, elevated intracellular calcium diminishes cAMP-mediated progesterone synthesis in these cells. These findings modify our current concepts of the mechanisms of control of progesterone secretion by the corpus luteum and suggest several new lines of research.

Animals↗
PubMed1984

Establishment and characterization of an estrogen-producing human ovarian granulosa tumor cell line.

Cells designated HTOG and HTOT were established by long-term culture from a human ovarian granulosa cell and a theca cell tumor, respectively. The HTOG line grew well forming colonies and multilayered rapidly without contact inhibition; serial passages of HTOG were performed over 100 times successively within 25 months. HTOG were spindle cells, polygonal or spherical in shape, revealed neoplastic and pleomorphic features, and produced estrone (E1) and 17 beta-estradiol (E2). The chromosome number varied considerably and showed hyperploidy; the modal chromosome number was in the hypertriploid-tetraploid range. When HTOG cells were heterotransplanted into the subcutis of BALB/c nude mice, they produced a sarcomatous diffuse type of granulosa cell tumors. In contrast, HTOT cells grew slowly while forming monolayers and underwent five successive passages in about 100 days, but a theca cell tumor line could not be established. HTOT cells were fibroblastic in shape and also produced E1 and E2. The majority of the cells showed diploidy and karyologic normality.

Animals↗