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PubMed2000

Acute regulation of the bovine gene for the steroidogenic acute regulatory protein in ovarian theca and adrenocortical cells.

Upregulation of the steroidogenic acute regulatory protein (StAR) is implicated in the rapid synthesis and secretion of steroidogenic cells to produce steroids in response to stimulation by trophic hormones of the gonadal and stress axes. In the present study, we have assessed the kinetics of both StAR gene transcription and protein biosynthesis in primary cell cultures of bovine adrenocortical and ovarian theca cells, under conditions of acute stimulation by corticotrophin (ACTH) and luteinizing hormone (LH), respectively. In both cell systems, detectable upregulation of StAR gene transcription occurred within 1-2 h, reaching maxima at 4 h (theca cells) or 6 h (adrenocortical cells). mRNA levels returned rapidly to baseline, by 12 h or 24 h, respectively. Specific StAR protein levels were assessed by western blotting using a novel antibody raised against a bovine StAR peptide, and showed a similar fast upregulation, albeit delayed by 1-2 h compared with the mRNA. The response of the cultured theca cells was more acute than that of the adrenocortical cells, possibly reflecting the propensity of the LH receptor to desensitize rapidly, unlike the ACTH receptor. The primary bovine theca cell cultures were also used for fully homologous transfection studies using various deletion promoter-reporter constructs of the bovine StAR gene. Kinetic analysis of the results indicated that the acute transcriptional response resides within the proximal (-315 bp) promoter region, which includes two putative responsive elements for the steroidogenic factor-1. More distal promoter regions may be involved in modulating the specificity of expression by combining enhancer and inhibitory functions.

Adrenal Cortex↗
PubMed1988

Sclerosing stromal tumor of the ovary associated with endometrial adenocarcinoma: a case report.

The first case of ovarian sclerosing stromal tumor associated with endometrial adenocarcinoma in a 44-year-old woman is presented. By ultrastructural study, typical steroid-secreting type theca cells and various transitional tumor cells from fibroblast-like cells to theca-like cells were observed in the ovarian tumor. The clinical history of the patient and the ultrastructural study of this tumor suggested hormonal activity, but no apparent evidence of active hormone secretion was detected by plasma analysis for steroid hormones.

Adenocarcinoma↗
PubMed2003

A novel PEPP homeobox gene, TOX, is highly glutamic acid rich and specifically expressed in murine testis and ovary.

The homeobox gene superfamily has been highly conserved throughout evolution. These genes act as transcription factors during several important developmental processes. To explore the functional roles of homeobox genes in spermatogenesis, we performed a degenerate oligonucleotide polymerase chain reaction (PCR) screening of a testis cDNA library and isolated a novel mouse homeobox gene. This gene, which we named Tox, encodes a homeodomain protein distantly related to members of the Paired/Pax (Prd/Pax) family. A phylogenetic analysis revealed Tox to be a member of the recently defined PEPP subfamily of Paired-like homeobox genes. Tox was mapped to chromosome X, with its homeodomain organized into three exons. A special feature of Tox is that the encoded protein sequence contains two poly-glutamic acid (poly E) stretches, which make Tox highly acidic. Tox transcripts were detected predominately in the testis and ovary of mice. Tox expression in testes was initiated soon after birth, mainly in Sertoli cells and spermatogonia; however, in adult mice, Tox expression shifts to the spermatids and spermatozoa. Tox expression in ovaries was detected in somatic cells of follicles, early on in theca cells, and in both granulosa and theca cells at the later stages of follicular development. Based on these results, Tox may play an important role during gametogenesis.

Amino Acid Sequence↗
PubMed1992

Cellular and microvascular changes of the ovarian follicle during folliculogenesis: a scanning electron microscopic study.

In order to study the three-dimensional topographic arrangement of the oocyte and zona pellucida, follicular cells and follicle microvasculature, this study applied alkali maceration methods for tissue exposure, the ruthenium red-detergent method for the extracellular matrix visualization, and the vascular corrosion cast technique to rabbits, guinea-pigs and mice ovaries at different stages of follicular development. Macerated samples showed a gradual differentiation of the oocyte surface. This, in primordial follicles, appeared rather smooth, but, with the follicular development, displayed a gradual increase of blebs and microvilli. The latter widely covered the surface of oocytes contained in large or mature follicles. The outer surface of the zona pellucida showed numerous fenestrations, whereas the inner one was smooth. The ruthenium red-detergent method permitted a well detailed view of the filamentous texture of the zona pellucida. The three-dimensional distribution of the contacts between oocyte and neighbouring follicular cells was clearly evaluated in macerated samples. Follicular cells of primary follicles were characterized by their short cytoplasmic processes reaching the oocyte surface. In secondary follicles, these processes issued secondary processes. In larger follicles, the secondary processes of the corona cells were much longer and thinner, and took a tortuous course to reach the oocyte surface, which ran among the numerous oocyte microvilli. This microvillous arrangement greatly increases contact between the oocyte and corona cells, and suggests a coordinated reciprocal control of the activities of both cell types. These data also showed that the spongy and filamentous nature of the zona pellucida is closely dependent upon the temporal differentiation and enormous increase in number of follicular cell projections and their ramifications. Maceration revealed the theca cells surface. In smaller follicles these appeared as fusiform cells which resembled fibroblasts. In larger or mature follicles, many theca cells differentiated to possess morphological features of steroidogenic cells. In addition, these cells delimited a series of intercellular communicating lacunae, continuous with wide pericapillary spaces. The gradual differentiation of the follicle towards a structure having an endocrinal role was further emphasized in vascular corrosion casts. A simple microvascular net made of thin capillaries supplying primary follicles was seen to transform into an elaborate sinusoidal network made of thick permeable capillaries, supplying mature follicles.

Animals↗
PubMed1987

Gonadotropin regulation of tissue-type and urokinase-type plasminogen activators in rat granulosa and theca-interstitial cells during the periovulatory period.

Plasminogen activators (PAs) are believed to be involved in ovulation. Because both tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA) are secreted by cultured rat granulosa cells, we have examined the activities of these proteins in ovarian homogenates as well as granulosa and theca-interstitial (TI) cells during gonadotropin-induced ovulation. Immature rats were injected with 20 IU pregnant mare serum gonadotropin (PMSG) to initiate follicle development, followed by treatment with 10 IU hCG 48 h later to induce ovulation. Ovarian proteins were separated by SDS-PAGE and PA activity determined by fibrin overlay. The activity of tPA, but not uPA, was stimulated following PMSG treatment in ovarian homogenates. Subsequent hCG injection further increased the tPA activity in a time-dependent manner, reaching a maximum (12 h after hCG treatment) immediately prior to ovulation and declined thereafter. Similar preovulatory increases in tPA activity were detected in isolated granulosa cells. Although both tPA and uPA activities were increased in TI cells after PMSG administration, no further increases were detected after hCG treatment. To estimate enzyme secretion, ovarian cells obtained at various preovulatory periods were incubated for 24 h in vitro. The ability of granulosa cells to secrete tPA, but not uPA, increased following in vivo PMSG and hCG treatment in a time-dependent manner, reaching a maximum immediately prior to ovulation. During the preovulatory period, an abrupt increase in tPA secretion by TI cells was also detected. Using immunohistochemical staining for tPA, it was found that ovarian sections from preovulatory rats at 12 h after hCG injection stained positively in granulosa, theca interna, and interstitial gland cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2000

Gonadotropin- and cytokine-regulated expression of the chemokine interleukin 8 in the human preovulatory follicle of the menstrual cycle.

Interleukin 8 (IL-8) is a chemotactic cytokine involved in the recruitment and activation of neutrophils as well as in cell proliferation and angiogenesis. Because these events are essential components of folliculogenesis, ovulation, and subsequent repair of the ruptured follicle, the presence and regulation of IL-8 in the human follicle of the menstrual cycle was investigated. The concentrations of IL-8 were higher in follicular fluids from dominant follicles of late follicular/ovulatory phase compared with those of midfollicular phase. IL-8 was detected in the media from cultured granulosa and theca cells, with 10-fold higher levels in the theca cell cultures. Exposure to FSH and LH increased the IL-8 secretion from granulosa cells, but no effect was seen in theca cell cultures. Estradiol and progesterone did not affect IL-8 secretion from any cell type. The cytokines IL-1alpha and IL-1beta, but not tumor necrosis factor alpha, enhanced IL-8 secretion from both cell types. IL-8 levels in cultures of granulosa-lutein cells from hyperstimulated in vitro fertilization cycles were not affected by either gonadotropins or steroids. These data provide evidence that ovarian IL-8 is gonadotropin and cytokine induced and may be involved in the hormonally regulated stages of follicular development and ovulation.

Adult↗
PubMed2006

Distribution of extracellular matrix proteins type I collagen, type IV collagen, fibronectin, and laminin in mouse folliculogenesis.

The extracellular matrix (ECM) plays a prominent role in ovarian function by participating in processes such as cell migration, proliferation, growth, and development. Although some of these signaling processes have been characterized in the mouse, the relative quantity and distribution of ECM proteins within developing follicles of the ovary have not been characterized. This study uses immunohistochemistry and real-time PCR to characterize the ECM components type I collagen, type IV collagen, fibronectin, and laminin in the mouse ovary according to follicle stage and cellular compartment. Collagen I was present throughout the ovary, with higher concentrations in the ovarian surface epithelium and follicular compartments. Collagen IV was abundant in the theca cell compartment with low-level expression in the stroma and granulosa cells. The distribution of collagen was consistent throughout follicle maturation. Fibronectin staining in the stroma and theca cell compartment increased throughout follicle development, while staining in the granulosa cell compartment decreased. Heavy staining was also observed in the follicular fluid of antral follicles. Laminin was localized primarily to the theca cell compartment, with a defined ring at the exterior of the follicular granulosa cells marking the basement membrane. Low levels of laminin were also apparent in the stroma and granulosa cell compartment. Taken together, the ECM content of the mouse ovary changes during follicular development and reveals a distinct spatial and temporal pattern. This understanding of ECM composition and distribution can be used in the basic studies of ECM function during follicle development, and could aid in the development of in vitro systems for follicle growth.

Animals↗
PubMed1991

A novel mechanism for the induction of aromatase in ovarian cells in vitro: role of transforming growth factor alpha-induced protein tyrosine kinase.

In the developing follicle, follicle-stimulating hormone (FSH) and luteinizing hormone (LH) are the primary stimulators of steroidogenesis in granulosa and theca cells. The steroidogenic actions of both these gonadotropins are largely if not exclusively mediated through cAMP. Previous studies have shown that EGF and TGF alpha do not affect basal estrogen production but attenuate FSH-stimulated estrogen production in vitro in rat granulosa cells. Here we present evidence that TGF alpha stimulates basal estrogen production in prepubertal porcine ovarian granulosa and theca cells in culture under defined conditions. In granulosa cells, TGF alpha is more potent than FSH in stimulating estrogen production. LH does not stimulate estrogen production in prepubertal porcine theca cells but rather attenuates that stimulated by TGF alpha. Treatment of follicular cells with genistein (inhibitor of protein tyrosine kinase) attenuates TGF alpha-induced estrogen biosynthesis suggesting that the action of TGF alpha is mediated through protein tyrosine kinase. These studies provide evidence for an alternative cAMP-independent and TGF alpha-induced tyrosine kinase-dependent mechanism for the induction of ovarian aromatase.

Animals↗
PubMed1979

Follicular growth in the rat: a reevaluation of the roles of FSH and LH.

The growth of preovulatory ovarian follicles involves hormonally induced proliferation and differentiation of theca cells and granulosa cells resulting ultimately in an increased ability of follicles to produce estradiol and to respond to the pituitary gonadotropins. The increased ability of follicles to produce estradiol appears to depend on an increased ability of theca cells to produce androgen as well as an increased ability of granulosa cells to aromatize androgens to estradiol. Estradiol in turn, appears to be required for FSH or FSH and LH to stimulate the appearance of functional receptors for LH in granulosa cells. Thus, the intrafollicular hormone estradiol enhances the response of granulosa cells to the gonadotropin. Therefore production of estradiol appears to determine which follicles will gain the mechanisms, including LH receptor, necessary for ovulation and luteinization. Since LH can act to increase its own receptor in the presence of estradiol and low amounts of FSH, it is possible that LH plays a predominant role in the final stages of preovulatory follicular growth both to promote estradiol production stimulation of theca cell androgen production as well as by facilitating an increase in its receptor by acting directly on granulosa cells.

Androgens↗
PubMed1994

Detection of manganese superoxide dismutase mRNA in the theca interna cells of rat ovary during the ovulatory process by in situ hybridization.

To determine the possible involvement of reactive oxygen species in ovulation, dynamic aspects of superoxide dismutase (SOD) isozyme were studied in the ovaries of rats by in situ hybridization histochemistry. Previously, mRNA levels of ovarian manganese superoxide dismutase (Mn-SOD) were reported markedly to increase whilst enzymic activity of Mn-SOD decreased during the ovulatory process after treating immature rats with 10 and 5 Units, respectively, of pregnant mare serum gonadotrophin (PMSG) and human chorionic gonadotrophin (HCG). Levels of Cu/Zn-SOD activity and Cu/Zn-SOD mRNA were reported to remain unchanged throughout ovulation. This increase in the Mn-SOD mRNA level was shown in the present study by in situ hybridization to be localized to the theca interna cells throughout the PMSG/HCG-induced ovulatory process. The observations suggest that the turnover rate of Mn-SOD but not Cu/Zn-SOD increases specifically in the mitochondria of these cells. SOD has been postulated to play important roles in steroidogenesis. The relationship is discussed between mitochondrial functions in steroid-secreting cells and superoxide radicals and related metabolite(s).

Animals↗
PubMed1998

Cyclic and maturation-dependent regulation of follicle-stimulating hormone receptor and luteinizing hormone receptor messenger ribonucleic acid expression in the porcine ovary.

We sought to characterize the ovarian expression of mRNAs for the receptors for FSH (FSHr) and LH (LHr) at the different stages of the estrous cycle of the pig and to localize the receptor mRNAs to individual cell types in follicles and corpora lutea as a function of their developmental status. Northern blot analyses indicated that multiple FSHr and LHr mRNA transcripts occur on Days 0, 4, 7, 12, and 16 of the estrous cycle. In situ hybridization analyses revealed considerable cyclic variation in expression among small, medium, and large follicles. In small follicles of both immature prepubertal ovaries and mature adult ovaries at all days of the cycle studied, FSHr mRNA expression was strongly positive and was spatially restricted to granulosa cells. FSHr mRNA expression was strongly positive in granulosa cells of medium follicles but had declined in granulosa cells of mature follicles on Day 0. LHr expression in small follicles, in contrast, was spatially restricted to theca cells and was estrous cycle-dependent. LHr expression in theca cells was weakly positive in small follicles on Days 12, 0, and 4, was strongly positive in small follicles on Day 7 and in medium follicles on Days 12 and 16, and had declined in large mature follicles on Day 0. LHr expression in granulosa cells was negative in small follicles on Days 4 and 7, weakly positive in medium follicles on Days 12 and 16, and positive in large follicles on Day 0. In degenerating follicles, LHr mRNA was expressed weakly in the theca cells, and FSHr mRNA expression in granulosa cells was absent. In the corpus luteum, FSHr mRNA was not expressed, but LHr mRNA was expressed in some cells on Days 4 and 7, was expressed maximally on Day 12, and had declined partially by Day 16. These studies indicate that there is maturation-dependent regulation of FSHr and LHr expression in the adult porcine ovary. FSHr mRNA content decreases in parallel with FSHr protein content as follicles increase in size.

Animals↗
PubMed2005

Oocyte-somatic cell-endocrine interactions in pigs.

Oocyte-somatic cell communication is bi-directional and essential for both oocyte and follicular granulosa and theca cell function and development. We have shown that the oocyte secretes factors that stimulate porcine granulosa cell proliferation in serum-free culture, and suppress progesterone production, thereby preventing premature luteinisation. Possible candidates for mediating some of these effects are the bone morphogenetic proteins (BMPs) that belong to the transforming growth factor beta family. They are emerging as a family of proteins critical for fertility and ovulation rate in several mammals, and they are expressed in various cell types in the ovary. We have evidence for a functional BMP system in the porcine ovary and BMP receptors are present in the egg nests in the fetal ovary and in the granulosa cells, oocytes and occasional theca cells throughout subsequent development. In addition to paracrine interactions in the ovary, the porcine oocyte and its developmental potential can also be influenced by nutritional manipulation in vivo. We have demonstrated that feeding a high plane of nutrition to gilts for 19 days prior to ovulation increased oocyte quality compared to control animals fed a maintenance diet, as determined by oocyte maturation in vitro. This was associated with a number of changes in circulating reproductive and metabolic hormones and also in the follicular fluid in which the oocyte is nurtured. Further studies showed a similar increase in prenatal survival on Day 30 of gestation, demonstrating a direct link between oocyte quality/maturation and embryo survival. Collectively, these studies emphasise the importance of the interactions that occur between the oocyte and somatic cells and also with endocrine hormones for ovarian development, and ultimately for the production of oocytes with optimal developmental potential.

Animal Nutritional Physiological Phenomena↗
PubMed1984

Prostaglandin production by dispersed granulosa and theca interna cells from porcine preovulatory follicles.

Prepubertal gilts were treated with 750 IU pregnant mares' serum gonadotropin (PMSG) and 72 h later with 500 IU human chorionic gonadotropin (hCG) to induce follicular growth and ovulation. Dispersed granulosa (GC) and theca interna (TIC) cells were prepared by microdissection and enzymatic digestion from follicles obtained 36, 72 and 108 h after PMSG treatment and incubated for up to 6 h in a chemically defined medium in the presence or absence of arachidonic acid, follicle-stimulating hormone (FSH), luteinizing hormone (LH) and indomethacin. Production of prostaglandin E2 (PGE) and prostaglandin F2 alpha (PGF) was measured by radioimmunoassay. Both GC and TIC had the capacity to produce prostaglandins, with production by each cell type increasing markedly with follicular maturation. PGE was the major prostaglandin produced by both cellular compartments. Only PGE production by GC was consistently enhanced by addition of arachidonic acid to the incubation medium. Neither cell type was responsive to FSH and LH in vitro. Indomethacin inhibited the production of PGE and PGF by both cell types. These results provide convincing evidence for an intrafollicular source of prostaglandins and indicate that both cellular compartments contribute significantly to the increased production of prostaglandins associated with follicular rupture.

Animals↗
PubMed1988

An immunocytochemical study of the progesterone receptor in rabbit ovary.

Distribution of progesterone receptor (PR) was studied immunohistochemically in rabbit ovary and uterus using a monoclonal anti-receptor antibody. At the subcellular level, PR was located in cell nuclei of prepubertal rabbits, which were either non-treated, primed with estrogen or made pseudopregnant. At the tissue level, germinal epithelium, the external theca cell layer and granulosa cell layer were PR-positive. The internal theca cell layer had no PR immunoreactivity. The corpora lutea of pseudopregnant rabbits contained small amounts of PR in some of the animals. During pseudopregnancy, the distribution and staining intensity of PR-positive cells in the ovary was essentially similar to that in non-treated rabbits except for weak PR immunoreactivity in the internal theca cell layer. This differed significantly from the uterus, in which pseudopregnancy caused a marked decrease in PR immunoreactivity. This implies that receptor downregulation by endogenous progesterone is not the same in different organs and cell types. Immunohistochemical techniques give valuable information as to the steroid hormone target cell types and their distribution which are not available by conventional steroid-binding assays.

Animals↗
PubMed2004

Chronic somatostatin treatment affects pituitary gonadotrophs, ovaries and onset of puberty in rats.

The effects of chronic somatostatin (SRIH-14) treatment on the pituitary gonadotrophs (FSH and LH cells) and ovaries of female Wistar rats were examined. Females were given 20 microg/100 g b.w. twice per day from the immature (23rd day) till the adult period of life (71st day). The onset of puberty was determined by daily examination for vaginal opening. The peroxidase-antiperoxidase immunocytochemical procedure was used to study the gonadotrophs. Changes in the number per unit area (mm2), cell volume and volume densities of LH- and FSH-immunoreactive cells were evaluated by morphometry and stereology. Ovaries were analysed by simple point counting of follicles and corpora lutea (CL). Follicles were divided by size according to the classification of Gaytán and Osman. The mitotic indexes of granulosa and theca cells in the follicles were estimated at all stages of folliculogenesis. The number, volume and the volume density of FSH- and LH-immunoreactive cells decreased after chronic SRIH-14 treatment, particularly the latter. In the ovary, SRIH-14 treatment decreased the number of healthy follicles at all phases of folliculogenesis, lowered the mitotic indexes of granulosa and theca cells but increased the number of atretic follicles. Healthy CL were fewer in number, while regressive CL were increased. Vaginal opening occurred at a later age in treated females. It can be concluded that chronic SRIH-14 treatment markedly inhibited LH cells and to a lesser extent FSH cells. In the ovary SRIH-14 inhibited folliculogenesis, enhanced atretic processes and lowered proliferative activity of granulosa and theca cells. It also delayed puberty onset.

Animals↗
PubMed1997

Immunohistochemical localization of basic fibroblast growth factor (bFGF) during folliculogenesis in the human ovary.

Basic fibroblast growth factor (bFGF) has been suggested to be one of the intraovarian regulators of ovarian folliculogenesis, but its localization in the human ovary remains to be determined. We examined the immunohistochemical reactivity for bFGF in the course of follicular development and corpora lutea formation in the human ovary. Pregranulosa cells in the primordial follicle were negative, but at the preantral stage both granulosa and theca cells showed weakly positive immunostaining for bFGF. In the antral follicles, both granulosa and theca interna cells showed stronger staining for bFGF with the increase in follicular diameter. In atretic follicles at various stages, granulosa cells were negative or weakly positive for bFGF, whereas luteinized theca cells showed strong immunoreactivity. In the corpora lutea during the early luteal phase, granulosa lutein cells were strongly positive for bFGF but in the late luteal phase became immunonegative. On the other hand, bFGF staining in theca lutein cells was strong throughout the course of corpora lutea formation and regression. These findings suggest that bFGF is localized not only in granulosa cells but also in theca cells, and in the latter, bFGF immunoreactivity is associated with luteinization.

Adult↗
PubMed1976

Luteoma of pregnancy: ultrastructural features.

The ultrastructural configuration of the cells in one case of pregnancy luteoma was studied by conventional electron microscopy. The fine structure of these cells conforms closely to that of steroid hormone producing cells in other human organs, such as the adrenal cortex, testicular interstitium, and corpus luteum, particularly in terms of the presence of abundant smooth endoplasmic reticulum, dispersed Golgi apparatus, and tubular cristae in the mitochondria. Similarities were noted between the pregnancy luteoma cells and, as described by other authors, the luteinized granulosa and theca cells of the human corpus luteum and the cells of spontaneous mouse luteoma. There was a more pronounced resemblance to the granulosa cells of the corpus lutemum than to the luteinized theca cells or the mouse luteoma cells but with considerable overlap among all of them. Distinctive features, not previously described, were noted in the pregnancy luteoma cells in the form of deep cytoplasmic invaginations of the cell membrane closed by tight junctions, and peculiar pleated sinuous membranous arrays within the cytoplasm. The significance of these structures is not known. In addition mesenchymal cells of ambiguous differentiation were noted in the stroma of the tumor, suggesting that the stimulus to luteinized cell proliferation may transcend the specific ovarian mesenchymal cells and may also be exerted on the supporting stroma. On the basis of previous reports concerning the biochemical and biosynthetic patterns of sex steroid hormone production by these tumors, it is concluded that although the latter recapitulate the endocrine functions of the normal ovarian stroma rather than those of the corpus luteum, the cytoarchitecture of these tumors covers a much broader spectrum of differentiation, which may come to resemble closely although not exclusively that of luteinized granulosa cells.

Adult↗