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PubMed2000

Expression of mRNA encoding IGF-I, IGF-II and type 1 IGF receptor in bovine ovarian follicles.

IGFs regulate gonadotrophin-stimulated proliferation and differentiation of granulosa and theca cells in vitro. However, the detailed pattern of mRNA expression of IGFs in bovine follicles remains controversial. The objectives of this study were therefore to describe the temporal and spatial pattern of expression of mRNA encoding IGF-I, IGF-II and the type 1 IGF receptor in bovine follicles in vivo. The expression of mRNA encoding IGF-II was detected in theca tissue from around the time of antrum formation up to and during the development of dominance. No IGF-II mRNA expression was detected in granulosa cells. In the majority of follicles we were unable to detect mRNA encoding IGF-I in either granulosa or theca tissue from follicles at any stage of development. Occasionally low amounts of mRNA encoding IGF-I were detected in the theca externa and connective tissue surrounding some follicles. Type 1 IGF receptor mRNA was detected in both granulosa and theca cells of preantral and antral follicles. Expression was greater in granulosa tissue compared with theca tissue. We also measured IGF-I and -II mRNA in total RNA isolated from cultured granulosa and theca cells using reverse transcriptase PCR. In contrast to the in vivo results, IGF-II mRNA was detected in both granulosa and theca tissue. IGF-I mRNA was detected in theca tissue and in very low amounts in granulosa cells. Using a specific IGF-I RIA we were unable to detect IGF-I immunoreactivity in granulosa conditioned cell culture media. Using immunohistochemistry we detected IGF-I immunoreactivity in some blood vessels within the ovarian stroma. We conclude from these results that IGF-II is the principal intrafollicular IGF ligand regulating the growth of bovine antral follicles. In preantral follicles the expression of mRNA encoding type 1 IGF receptor but absence of endogenous IGF-I or -II mRNA expression, highlights a probable endocrine mechanism for the IGF regulation of preantral follicle growth.

Animals↗
PubMedDate not supplied

Histochemical changes in acid and alkaline phosphatase activities in the growing follicles and corpora lutea of the rat ovary.

Changes in acid and alkaline phosphatase activities are studied histochemically in different components of the growing follicles and corpora lutea of different generations. Theca cells of the growing follicles showed strong acid phosphatase activity as compared to that observed in the oocyte and granulosa cells. Alkaline phosphatase activity is localized only in the theca interna cells of growing follicles. However, after ovulation with the formation of corpora lutea, the granulosa lutein cells also showed this activity. Changes in the histochemical distribution of the acid and alkaline phosphatases in the follicles and corpora lutea are discussed in relation to folliculogenesis and corpus luteum and regression.

Acid Phosphatase↗
PubMed1996

Changes in lysosomal enzyme activities in pre-ovulatory follicles and endometrium of PMSG superovulated rats.

Large superovulatory doses of gonadotrophins result in reduced fertility in laboratory and large domestic animals and it has been postulated that some of the superovulated oocytes are derived from abnormal follicles which would not ovulate under normal physiological stimuli. Follicular growth, follicular maturation and atresia, ovulation and the nidation of the fertilized oocyte require intense tissue remodelation which can be accomplished only through the action of hydrolytic enzymes. We have studied the activities and sub-cellular distribution of three lysosomal enzymes (acid phosphatase, N-acetyl-beta-D-glucosaminidase and beta-glucuronidase) in the follicular fluid, granulosa and theca cells of preovulatory follicles and in the endometrial tissue of immature Wistar rats injected with 4 (control) or 40 (superovulated) IU of pregnant mare serum gonadotrophin (PMSG). Enzyme activities were from four to ten times higher in theca than in granulosa cells. This difference was particularly important in the case of beta-glucuronidase. Large preovulatory follicles tended to have higher activities of lysosomal enzymes in the free fraction of all the compartments studied. This difference was remarkable in theca cells where free enzymes would be required to help ovulation. Forty IU of PMSG induced higher activities of acid phosphatase in theca and granulosa cells than 4 IU, but in endometrial tissue this latter dose of PMSG was more efficient to induce higher activities of this enzyme. The endometrial bound fraction of N-acetyl-beta-D-glucosaminidase was almost three times higher than the free activity. This behavior was also observed in endometrial beta-glucuronidase but only in the control rats. The results observed in follicular fluid were less homogeneous. The activities of glucosaminidase and acid phosphatase were two to three times higher in rats overstimulated with 40 IU of PMSG than in the control rats, whereas the activities of beta-glucuronidase were lower in the superovulated rats. Our results suggest that alterations in the process of tissue remodeling required for ovulation of mature, normal oocytes and for nidation of the fertilized ovum may be important factors to explain pregnancy failure in the PMSG superovulated female.

Animals↗
PubMed2001

Different action of ovine GH on porcine theca and granulosa cells proliferation and insulin-like growth factors I- and II-stimulated estradiol production.

Growth hormone (GH) and insulin-like growth factors (IGFs) are recognized as regulators of ovarian function. This study was designed to compare the effect of GH and IGFs added alone or together on porcine theca interna and granulosa cells proliferation and steroidogenesis. Moreover, the effect of GH on IGF-I secretion was examined. Cells were isolated from medium size follicles and cultured in vitro for 48 h in serum free medium. Estradiol and IGF-I medium concentrations were determined by radioimmunoassays. Proliferation was evaluated by alamar blue assay and by radiolabelled thymidine incorporation. GH increased IGF secretion by granulosa cells while decreased its secretion by theca cells. Proliferation of both cell types was stimulated by IGF-I and IGF-II (30 ng/ml) and modestly inhibited by GH (100 ng/ml). Insulin-like growth factor II increased, in a statistically significant manner, estradiol secretion by both cell types, while IGF-I stimulated estradiol secretion to a greater extent by granulosa then by theca cells. The synergistic action of GH and IGFs on estradiol secretion was stimulatory in theca cells and inhibitory in granulosa cells. These data demonstrate that despite its direct action on estradiol secretion by granulosa and theca cells, GH also modulated estradiol secretion induced by IGFs. Differences in the estradiol production in response to GH alone and the effect of the synergistic action of GH and IGFs suggest that different cellular mechanisms for these hormones are triggered in each cell type.

Animals↗
PubMed1977

[Light and electronmicroscopic examinations on granulosa and theca of preovulatory and freshly ruptured follicles of human ovaries (author's transl)].

In a first paper, the structural and functional relationship of granulosa and theca of follicles during early development stages was reported. On the special question of whether and from which moment these two tissue formations are involved in the steroidbiosynthesis, our electronmicroscopic examinations have given a further insight to this. Neither the granulosa nor the theca folliculi of primordial, primary and secondary follicles show definite morphologic submicroscopic criteria of the steriodbiosynthesis. In the resting tertiary follicle, the electronmicroscopy defines the theca cells as steroidbiosynthetic active cells, whereas the granulosa cells of this stages of follicle demonstrate the morphologic characteristics of protein-synthetic active cells. Our, under physiological conditions, systematically conducted light and electronmicroscopic studies of preovulatory and freshly ruptured follicles showed remarkable structural changes in the granulosa, as well as in the theca folliculi. For the follicle granulosa cells of the preovulatory follicle, the studies could demonstrate a structural transformation process of proteinsynthetic active to steroidbiosynthetic active cells. This transformation process can be seen in the nucleus. Especially recognizable is the continuous change from rough to smooth endoplasmatic reticulum in the cytoplasm of most of the follicle granulosa cells, which apart from that, often displayed whorle-like formations. Furthermore, we noticed striking changes of the paraplasmatic structures, especially of the fat, present in various stages in the cytoplasm of the follicle granulosa cells. We also noticed large mitochondria, which showed a lot of vesicular cristae. Numerous existing Golgi-fields contained many little homogenous fat-like droplets, which were surrounded by a thin osmiophilic membrane. This recognizable transformation process of the proteinsynthetic active granulosa cells to the steroid cells is quite completed in freshly ruptured follicles, according to our examinations. Although the theca cells have already been defined submicroscopically in the resting tertiary follicle as steroidbiosynthetic active cells, we see in this cellgroup in the preovulatory, as well as in the freshly ruptured follicle a remarkable conspicuous size-increase of the mitochondria. Apart from that, there are wide areas which are solely occupied by smooth endoplasmatic reticulum, whereas other structures, for example fat, are scarely seen. As our examinations have shown, there is a close relationship between the transformation process in the granulosa and the theca of preovulatory follicles and the theca of preovulatory follicles that, the increasing concentration of progesteron in serum, which begins prior to the ovulation can be regarded as a product of the follicle granulosa cells being transformed to steroid cells.

Endoplasmic Reticulum↗
PubMed2003

Abundant expression of sodium-potassium-activated adenosinetriphosphatase alpha 1 subunit in corpus luteum of porcine ovary.

Follicular development is accompanied by the accumulation of follicular fluid. During corpus luteum formation, follicular fluid is diminished and antrum is replaced by lutein cells. These dynamic changes in fluid distribution suggest the existence of control mechanism of fluid transport and membrane permeability. One of the major factors regulating membrane permeability is the sodium-potassium-activated adenosinetriphosphatase (Na(+)-K(+)-ATPase). To elucidate the possible involvement of Na(+)-K(+)-ATPase in follicular growth and luteinization, immunohistochemical localization of Na(+)-K(+)-ATPase alpha1 subunit and enzyme activity in porcine ovary were investigated. In primordial follicles, Na(+)-K(+)-ATPase alpha1 subunit immunostaining was localized only in the oocyte and the surrounding stromal cells. In preantral follicles, immunostaining for Na(+)-K(+)-ATPase alpha1 subunit became apparent in granulosa and theca cells. As the follicle matured, the staining intensity in the oocyte, theca, and granulosa cells increased, which corresponded with the enzyme activity. Na(+)-K(+)-ATPase alpha1 subunit immunostaining became most abundant in granulosa and theca lutein cells in corpus luteum, and decreased in the regressing corpus luteum. Enzyme activity in corpus luteum was significantly higher than that in the follicles. This is the first study indicating that Na(+)-K(+)-ATPase alpha1 subunit expression is augmented in granulosa cells by follicular growth and most abundant in lutein cells in the corpus luteum, suggesting its possible involvement in corpus luteum formation.

Animals↗
PubMed1998

Two steroidogenic pathways present in the chicken ovary: theca layer prefers delta 5 pathway and granulosa layer prefers delta 4 pathway.

It has been suggested that there is a shift in steroidogenic pathways of the theca layer from delta 5 to delta 4 during follicular maturation in the chicken ovary. However, the preferred steroid metabolic pathway in the theca and granulosa layers of small and preovulatory follicles has not been identified. Therefore, the goal of our study was to identify the preferred steroidogenic pathways used by these follicles. Previously, we found that small follicles that had not entered the follicular hierarchy did not use cholesterol but did use pregnenolone (P5) to produce testosterone and estradiol. For this reason, P5 was used as the substrate for steroid production in the present study. The theca layer of small follicles (2-4 mm in diameter), and the theca and granulosa layers of the fifth largest (F5), third largest (F3), and largest (F1) preovulatory follicles were isolated from the chicken ovary 22 hr before ovulation. The granulosa layer of small follicles was excluded from the study because it is steroidogenically incompetent. Tissues were incubated with [3H]-P5 for 1 hr at 37 degrees C. Production of steroid metabolites was determined on the basis of comparison with known steroid standards using two-dimensional thin layer chromatography (2D-TLC). Conversion of [3H]-P5 to [3H]-DHEA was greater than that of [3H]-P4 in the theca layer of small and preovulatory follicles. Even though the amount of [3H]-P4 was less than that of [3H]-DHEA, theca cells from F3 and F1 follicles did produce [3H]-P4. In contrast to the theca layer, conversion of [3H]-P4 was greater than that of [3H]-DHEA in the granulosa layer of preovulatory follicles. Conversion of [3H]-P5 to [3H]-DHEA by the granulosa layer gradually decreased as the follicle matured and approached ovulation. We conclude that both delta 5 and delta 4 pathways are functional in theca and granulosa cells. However, in contrast to earlier speculation, the theca layer preferentially metabolizes steroids via the delta 5 pathway regardless of the maturational stage of the follicle, whereas the granulosa layer of preovulatory follicles preferentially metabolizes steroids via the delta 4 pathway.

Animals↗
PubMed1993

Expression of epidermal growth factor and its receptor in the human ovary during follicular growth and regression.

Immunohistochemical studies were performed using specific antibodies to epidermal growth factor (EGF) and EGF receptor to determine their presence and cellular localization in the human ovary during follicular growth and regression. There was no immunostaining for EGF or EGF receptor in primordial follicles. In the preantral follicle stage, immunostaining for EGF and EGF receptor was observed only in the oocyte. The staining intensity of the oocyte increased as the oocyte reached the preovulatory stage. In the antral follicle stage, immunostaining for EGF and EGF receptor became apparent in the granulosa and theca interna cell layers, without appreciable staining in the surrounding stromal cells. The immunostaining for EGF and EGF receptor in the granulosa cells and theca interna cells persisted in preovulatory follicles and corpus luteum, and intensified in the midluteal phase. The stromal cells surrounding the corpus luteum were negative for EGF and EGF receptor staining. In the regressing corpus luteum, immunostaining for EGF and EGF receptor was present in the peripheral lutein cells adjacent to the central core of scar tissue, but absent in the scar tissue of the central core. Corpus albicans showed no staining for EGF and EGF receptor. By contrast, the stromal cells surrounding the corpus albicans in the cortex region demonstrated intense staining for EGF and EGF receptor, while the stromal cells surrounding the corpus albicans in the medullary region were negative for immunostaining. In the case of atretic follicles, the theca interna cells showed intense staining for EGF and EGF receptor, but immunostaining in the scattered granulosa cells was negligible. This is the first study to demonstrate a remarkable change in the expression of EGF and EGF receptor in the oocyte, granulosa cells, thecal cells, and surrounding stromal cells over the course of follicular growth and regression. The results obtained support EGF participation in oocyte maturation and in follicular growth and atresia. The intense immunostaining for EGF and EGF receptor observed in the theca interna cells in atretic follicles and the stromal cells surrounding corpus albicans in the cortex region raises the possibility of EGF involvement in transformation of thecal cells into stromal cells. Furthermore, the cell type-specific simultaneous expression of EGF and EGF receptor in follicular and stromal compartments in the various stages of follicular development suggests that an autocrine mode of EGF action may exist to regulate follicular growth and regression in the human ovary.

Adult↗
PubMed2004

Cell proliferation in the atretic follicles of buffalo and cattle ovary.

The present study was carried out to describe the proliferative activity of granulosa and theca cells in healthy antral and atretic follicles of Philippine buffaloes (BU) and Holstein-Friesian (HF) cows. Paraffin sections of ovary were immunostained with mouse monoclonal antibody to proliferating cell nuclear antigen (PCNA). Then the follicles were classified into healthy and various stages of atretic follicles. The granulosa layer of healthy follicles had a significantly higher frequency of PCNA-positive cells than the early and advanced atretic follicles in both breeds. In the theca interna, significantly reduced populations of the PCNA-positive cells were found in both breeds as atresia progressed. Moreover, HF had significantly higher PCNA-positive cells in the theca interna of healthy, early atretic and advanced atretic follicles than BU. A reduction of PCNA-positive cells during atresia was also noted in the theca externa in both animals although differences were not significant. The results of the present work suggest that the proliferative activity of granulosa and theca cells decreases in association with follicular atresia in the BU similar to HF. Furthermore, a significantly deficient cell proliferative activity of theca interna was found in BU compared with HF.

Animals↗
PubMed2006

The identification of novel ovarian proteases through the use of genomic and bioinformatic methodologies.

Proteolytic activities are essential for follicular growth, ovulation, as well as for luteal formation and regression. Using suppression subtractive hybridization (SSH), a novel mouse ovary-selective gene (termed protease serine 35, Prss35) was identified. Analysis of the mouse genome database using the Prss35 sequence led to the identification of a homologous protease (protease serine 23, Prss23). PRSS35 possesses general features that are characteristic of serine (Ser) proteases, but is unique in that the canonical Ser that defines this enzyme family is replaced by a threonine (Thr). In contrast, PRSS23 possesses the standard catalytic Ser typical for this family of proteases. As determined by real-time polymerase chain reaction (PCR), the Prss35 mRNA levels increased around the time of ovulation and remained elevated in the developing corpus luteum. Steroid ablation/replacement studies demonstrated progesterone-dependent regulation of Prss35 gene expression prior to follicle rupture. Prss35 gene expression was localized to the theca cells of pre-antral follicles, the theca and granulosa cells of pre-ovulatory and ovulatory follicles, as well as to the developing corpus luteum. In contrast, Prss23 mRNA levels decreased transiently after ovulation induction and again in the postovulatory period. Prss23 gene expression was noted primarily in the granulosa cells of the secondary/early antral follicles. PRSS35 and PRSS23 orthologs in the rat, human, rhesus macaque, chimpanzee, cattle, dog, and chicken were identified and found to be highly homologous to one another (75-99% homology). Collectively, these results suggest that the PRSS35 and PRSS23 genes have been conserved as critical ovarian proteases throughout the course of vertebrate evolution.

Amino Acid Sequence↗
PubMed1982

Gynandroblastoma: its ultrastructure.

Gynandroblastoma is a rare, sex-cord stromal tumor of the ovary that shows morphologic evidence of female and male differentiation. Such a tumor produced masculinization in a 24-year-old woman, whose symptoms disappeared following removal of the tumor. By electron microscopy, the granulosa cell nests displayed Call-Exner (CE) bodies of the hyaline type composed of multiple layers of basal lamina resembling CE bodies of the normal graafian follicle. In contrast, CE bodies of a classic granulosa theca cell tumor were of the spongiform type, consisting of a space limited by a single basal lamina containing altered granulosa cells and cell processes. Both types of CE bodies are believed to arise following secretion by and/or degeneration of granulosa cells, the variation in morphology between the two resulting from differences in amounts of basal lamina deposited. The tubular components of the tumor resembled more closely the rete ovarii than did Sertoli cells, and its proposed that such structures be called by the alternate and less specific term "androblastoma." The identity of Leydig cells was established by demonstrated of intracytoplasmic Reinke crystals. Despite a difference in architectural pattern, there was a close ultrastructural resemblance between the different sex-cord components of the gynandroblastoma.

Adult↗
PubMed2004

The expression patterns of mRNA-encoding stem cell factor, internal stem cell factor and c-kit in the prepubertal and adult porcine ovary.

The receptor, c-kit, and its ligand, stem cell factor (SCF), are important regulators of ovarian follicle growth and development. The aim of this study was to identify the sites of expression of mRNA for c-kit and SCF in prepubertal and mature (pregnant and non-pregnant) animals. Ovaries were recovered from prepubertal animals, non-pregnant sows and five sows at approximately 3 months of gestation. Ovine SCF and c-kit DNA were cloned into plasmid vectors to produce RNA probes. Expression of mRNA encoding SCF and c-kit were detected via in situ hybridization. Both mRNA were detected throughout ovaries from all animals. This study provides evidence that the growth-factor complex is required throughout follicle development, and also for continued maintenance of the corpus luteum (CL) in the mature animal. SCF mRNA was localized to the granulosa cell layer and was also extensively expressed in endothelial tissue and throughout the CL. c-kit mRNA was detected in the theca layer, oocytes and also in CL. In conclusion, expression of SCF and c-kit mRNA in granulosa and theca cells, respectively, indicate an important interaction between somatic cells throughout follicle development and that in the mature animal, SCF and c-kit potentially have a role in maintaining progesterone secretion by the CL. The observations of continued expression of SCF and c-kit throughout development suggest that there may be differences in the role of this receptor-ligand complex between large mono- vs. poly ovulatory species, such as the pig.

Animals↗
PubMed2006

Induction of cytochromes P450, caspase-3 and DNA damage by PCB3 and its hydroxylated metabolites in porcine ovary.

Polychlorinated biphenyl (PCBs) levels of tens and hundreds of pg/ml for individual congeners are measured in human follicular fluid. PCB3 (4-chlorobiphenyl), caused a significant increase in estradiol secretion in porcine granulose-theca cell co-cultures and its two metabolites, 4-OH-PCB3 and 3,4-diOH-PCB3, were even more potent than PCB3 itself [Ptak, A., Ludewig, G., Lehmler, H.J., Wojtowicz, A.K., Robertson, L.W., Gregoraszczuk, E.L. 2005. Comparison of the actions of 4-chlorobiphenyl and its hydroxylated metabolites on estradiol secretion by ovarian follicles in primary cells in culture. Reprod. Toxicol. 20, 57-64]. The question is whether these follicle cells are potentially able to metabolize PCB3 to hydroxylated and genotoxic or cytotoxic intermediates. We report here that granulose-theca co-cultures express xenobiotic-metabolizing cytochrome P450 activities, with CYP1A1>CYP2B>>CYP1A2. A significant increase in CYP1A1 and 2B, but not CYP1A2, activity was seen in cells that were exposed to 6 ng/ml PCB3 or 20 nM 17-beta-estradiol. An increase in caspase-3 activity, indicative for apoptosis, was only observed in PCB3-exposed cells after 24 h exposure. Genotoxicity, determined with the Comet assay, was initially reduced after 24 h exposure to PCB3 and both metabolites compared to untreated controls, followed by a significant transient increase in Comets at the 4 and 24 h time point with PCB3 and 4-OH-PCB3. 3,4-diOH-PCB3 induced a significant increase only after 72 h of recovery. We hypothesize that these biphasic damage kinetics may be due to cross-links caused by adduct formation. These results show for the first time that granulose-theca cells in co-culture express CYP1A1, 2B and 1A2 activities and that PCBs at concentrations that are reached in the environment induce genotoxicity in granulosa cells.

Animals↗
PubMed1996

Transient periovulatory expression of pituitary adenylate cyclase activating peptide in rat ovarian cells.

Pituitary adenylate cyclase activating polypeptide (PACAP), a potent stimulator of cAMP formation, has a widespread distribution in neuronal elements of the central nervous system and a number of peripheral organs. Recently, PACAP has also been shown to be stage specifically expressed in the spermatogenic cells from the rat testis. This prompted us to examine the cell-specific expression of PACAP during the estrous cycle in the rat ovary by in situ hybridization with a digoxiginin UTP-labeled RNA probe and by immunohistochemistry using a specific monoclonal antibody. Ovaries from PMS gonadotropin (PMSG)/human CG (hCG) treated immature animals were examined 0, 12, 24, 48, 54, 57, 58.5, 60, 61.5, 63, 66, 72, and 96 h after PMSG treatment. Ovaries from adult cyclic rats were examined at 1000 h on each day of the estrous cycle and at 0400 h on estrus morning. PACAP positive cells were observed in the preovulatory and ovulatory period, i.e. 6-18 h following hCG injection in the PMSG induced cycle and at 0400 h and 1000 h on estrus morning in adult cyclic animals. Both PACAP messenger RNA (mRNA) and PACAP immunoreactivity were observed in the majority of granulosa and cumulus cells from large preovulatory follicles, in the majority of the cells comprising the interstitial glandular tissue and in solitary theca cells of growing and mature follicles. In PMSG/hCG treated animals, positive signals were also observed in solitary theca and interstitial cells 12 h after PMSG injection. PACAP immunoreactive nerve fibers were observed in all ovaries examined, located in the interstitial glandular tissue or in the loose connective tissue. Northern blotting of ovarian tissue confirmed the transient preovulatory and ovulatory expression of PACAP. Three transcripts were observed. The most predominant band had a size of approximately 1.2 kilobase pairs, corresponding to the transcript observed in the testis. Two bands of 2.4 and 3.0 kilobase pairs, corresponding to the transcripts from neural tissue, were also observed. The complex distribution of PACAP positive cells and nerve fibers suggests that the peptide could be a local regulator of a number of the events that take place in the periovulatory period as well as during early folliculogenesis in the ovary.

Animals↗
PubMed1988

Steroidogenic relationships of gonadotrophin hormones in the ovary of the hen (Gallus domesticus).

The effects of chicken luteinizing hormone (cLH: IRC-2 and PRC AE1-1), turkey LH (B221B and HS-5-18), bovine follicle-stimulating hormone (bFSH: HS-2-17), chicken FSH (cFSH: PRC DC3(2) and AGCQSQ113445C), and turkey FSH (B150A and HS-1-153) on steroid output were evaluated by in vitro incubation of various ovarian tissues with the gonadotrophins. Output of androstenedione and estradiol was determined by 3-hr incubations of individual whole small follicles, classified by size and color as follows: small white (SWF, less than 1 mm), large white (LWF, 2-3 mm), and small yellow follicles (SYF, 5-10 mm). The effects of gonadotrophin preparations were also evaluated in large preovulatory follicles (F1-F5). Androstenedione and estradiol output was measured in incubation media from 100,000 theca cells and progesterone content was determined in the incubation media of 100,000 granulosa cells. All incubations were conducted in 1 ml of Medium 199 at 37 degrees. Steroid output was quantitated by radioimmunoassay of incubation media. Potency estimates were derived by calculation of a peak stimulation index. The standard reference preparation was bLH (NIAMDD-LH-B4). Steroidogenesis was stimulated by three avian LH preparations. preparations. PRC AE1-1 was the most potent, with IRC-2 and B150A showing approximately 50% of the biological activity of PRC AE1-1 in most tissues. Turkey LH HS-5-18 was generally not potent. The presence of multiple isohormones of LH was implied, as various LH preparations exhibited different potency estimates in different tissues. The effects of FSH on steroidogenesis were not significant in most cases. Although the addition of cFSH AGCQSQ113445C failed to significantly increase output of estradiol from small follicles, potency estimates of this preparation were 0.15, 0.20, and 0.13 relative to NIAMDD-LH-B4 follicles was more highly stimulated by LH than by FSH, and thus it would seem that FSH does not play a significant role in steroidogenesis in the hen's ovary. The results of this study suggest that steroid biosynthesis in the hen's ovary may be regulated by multiple forms of LH.

Androstenedione↗
PubMed2005

Exogenous granulocyte-macrophage colony-stimulating factor promotes follicular development in the newborn rat in vivo.

BACKGROUND: Expression and selective cellular localization of granulocyte-macrophage colony-stimulating factor (GM-CSF) and its receptor in ovarian tissue imply an autocrine/paracrine role in ovarian function. Evidence indicating a functional role for GM-CSF in ovarian follicular cell function has been provided by studies with GM-CSF knockout (GM-/-) mice, which suggest that GM-CSF influences events associated with murine follicular maturation. METHODS: Immature female rats were treated with GM-CSF, FSH or saline for 5 or 10 days. Ovaries were collected for histologic examination and immunostaining determination of CYP17, a theca cell marker. In addition, ovarian section slides were evaluated by immunofluorescence for CD45, an ovarian leukocyte marker. To investigate the possible mechanism of GM-CSF action on follicular development, theca-interstitial cells (T-I) were separated and cultured. Cells were treated with increasing concentrations of GM-CSF, then evaluated for CYP17 mRNA and protein expression assays. RESULTS: After 10 days of treatment with GM-CSF, the number of small preantral and large preantral follicles was significantly increased compared with the control group (P < 0.05). Similarly, treatment with FSH increased the number of small preantral and large preantral follicles (P < 0.05). CD45 expression measured by immunofluorescence was not different in the three groups, indicating that the distribution of leukocytes was unchanged. In addition, CYP17 was increased in the T-I cells both in vivo and in vitro after GM-CSF treatment. CONCLUSION: The present results suggest that GM-CSF may play a significant role in follicular development.

Animals↗
PubMed1999

Characterization and regulation of type A endothelin receptor gene expression in bovine luteal cell types.

Our previous studies demonstrated that endothelin-1 (ET-1), a 21-amino acid vasoconstrictor peptide, has a paracrine regulatory role in bovine corpus luteum (CL). The peptide is produced within the gland where it inhibits progesterone production by acting via the selective type A endothelin (ETA) receptors. The present study was designed to characterize ETA receptor gene expression in different ovarian cell types and its hormonal regulation. ETA receptor messenger RNA (mRNA) levels were high in follicular cells as well as in CL during luteal regression. At this latter stage, high ETA receptor expression concurred with low prostaglandin F2alpha receptor mRNA. The ETA receptor gene was expressed by all three major cell populations of the bovine CL; i.e. small and large luteal cells, as well as in luteal endothelial cells. Among these various cell populations, the highest ETA receptor mRNA levels were found in endothelial cells. cAMP elevating agents, forskolin and LH, suppressed ETA receptor mRNA expression in luteinized theca cells (LTC). This inhibition was dose dependent and was evident already after 24 h of incubation. In luteinized granulosa cells (LGC), 10 and 100 ng/ml of insulin-like growth factor I and insulin (only at a concentration of 2000 ng/ml) markedly decreased ETA receptor mRNA levels. In both LGC and LTC there was an inverse relationship between ETA receptor gene expression and progesterone production; insulin (in LGC) and forskolin (in LTC) enhanced progesterone production while inhibiting ETA receptor mRNA levels. Our findings may therefore suggest that, during early stages of luteinization when peak levels of both LH and insulin-like growth factor I exist, the expression of ETA receptors in the gland are suppressed. This study demonstrates physiologically relevant regulatory mechanisms controlling ETA receptor gene expression and further supports the inhibitory role of ET-1 in CL function.

Animals↗
PubMed1989

Virilizing stromal thecosis of the ovary associated with multiple corpora atretica.

Virilism can be associated with a variety of ovarian lesions, including stromal hyperplasia. Focal luteinization, a thickened tunica albuginea, and multiple cystic follicles are histologic features that have been associated with stromal hyperplasia in virilized women. This report describes the case of a 57-year-old woman who presented with a four-year history of virilization. The testosterone level was 13.8 nmol/L (3,980 ng/L). At laparotomy, the ovaries were found to be enlarged, each measuring 4 cm in maximum dimension. Salient histologic findings included diffuse stromal hyperplasia interspersed with a marked increase in the number of corpora atretica. Focal luteinization, including in some corpora atretica, was seen. Electron microscopic study demonstrated cells with increased smooth endoplasmic reticulin and mitochondria with tubular cristae as well as other features consistent with theca cells; Reinke crystals were not seen. The testosterone level normalized after removal of the ovaries, and the signs of virilism gradually abated. These results are compatible with an abnormal stimulation of the ovaries as evidenced by an increased though abortive follicular development, stromal hyperplasia, and focal luteinization.

Female↗