In this study, in order to further understanding of function of Mullerian inhibiting substance (MIS) and the ontogeny of the production profile of biologically active MIS and MIS type II receptor (MISR II), the patterns of their localization according to the follicular development in 21 ovarian specimens from women in reproductive age were studied by immunohistochemical staining. The flattened granulosa cells in primordial follicles failed to stain for MIS and MISR II, but the first staining was detected in the cuboidal granulosa cells in primary follicles. MIS and MISR II were detected specifically and exclusively in the cytoplasm of granulosa cells. The granulosa cells of both single and multiple layered growing preantral follicles showed strong specific staining for MIS and MISR II. Among the growing follicles, large follicle stained more intensely than small one. Within the multiple layers of granulosa cells, the innermost cells, closer to the oocyte, stained more intensely for MIS than those near the basement membrane, but MISR II was evenly distributed. In antral follicles, expression of the MIS was only seen in the granulosa cells, but MISR II was seen in the granulosa cells and theca cells. In large antral follicles, cumulus cells and periantral granulosa cells stained more intensely for MIS than those in the periphery. MIS staining waned in the mature follicles just before ovulation and could not be found in atretic follicles, corpus luteum, and corpus albicans. The expression levels of MISR II in mature follicles was lower than those in growing follicles and were even further reduced, but still detectable, in corpus luteum. There was a decreased level of MISR II expression when follicles become atretic and eventually lost from atretic follicles. The MIS and MISR II staining were not found in primordial follicles, oocytes, interstitial cells, ovarian epithelium, and corpus albicans. It is concluded that actions of MIS via MISR II are autocrine and paracrine in nature. The pattern of MIS and MISR II expression according to the menstrual cycles and development suggest that MIS may act as an intraovarian regulator of follicle maturation, selection and ovulation during the adult reproductive cycle.
Adult ; Anti-Mullerian Hormone* ; Basement Membrane ; Corpus Luteum ; Cumulus Cells ; Cytoplasm ; Epithelium ; Female ; Granulosa Cells ; Humans* ; Menstrual Cycle* ; Oocytes ; Ovarian Follicle ; Ovary* ; Ovulation ; Theca Cells

Sex cord tumors with annular tubules are known to originate from the sex cord of embryonic gonads that synthesize Sertoli cells, Leydig cells, granulosa cells, and theca cells of the ovarian stroma, while ovarian small cell carcinoma of the hypercalcemic type is a type of neuroendocrine tumor. Both these tumors are uncommon, potentially malignant neoplasms in children. We report the case of a sex cord tumor with annular tubules in an 11-year-old girl and a case of small cell carcinoma of the hypercalcemic type in a 10-year-old girl. We also discuss the prognosis and management of these tumors.
Carcinoma, Small Cell ; Child* ; Female ; Gonads ; Granulosa Cells ; Humans ; Hypercalcemia ; Leydig Cells ; Male ; Neuroendocrine Tumors ; Ovary* ; Prognosis ; Sertoli Cells ; Theca Cells

Human ovarian follicles reduce rapidly in number throughout fetal and adult life. Throughout the menstrual cycles, primordial follicles grow into mature follicles and then ovulate to form corpus luteum. Apoptosis has been implicated in several events that occur during the process of follicular growth, atresia and the regression of the corpus luteum. By the use of immunohistochemistry, we clarified the involvement of apoptosis in the human ovary during follicular growth, regression and atresia by investigating the expression of Fas, Fas-ligand, Bcl-2 and Bad in primordial follicles, primary follicles and mature follicles. Fas immunostaining was present in primordial oocytes, both oocytes and granulosa cells of primary follicles, preantral follicles and all follicular cells of mature follicles. Fas-ligand and Bad immunostaining patterns were similar to those of Fas except for theca cells. Bcl-2 immunostaining was present in both oocytes and granulosa cells of primary, preantral and mature follicles. In corpus luteum, Fas, Fas-ligand, Bcl-2 and Bad immunostaining were observed and decreased in the regressing corpus luteum. In postmenopausal ovary, Fas, Fas-ligand, Bcl-2 and Bad immunostaining were entirely negative. Bad immunostaining was observed but Bcl-2 was not in atretic follicle. These results suggest that Fas, Fas-ligand, Bcl-2 and Bad may play important roles in human ovary during follicular growth, regression and atresia simultaneously. Further studies should be required to elucidate the underlying mechanism and apoptosis of the disease associated with normal and abnormal ovarian aging.
Adult ; Aging ; Apoptosis ; Corpus Luteum ; Female ; Granulosa Cells ; Humans* ; Immunohistochemistry ; Menstrual Cycle ; Oocytes ; Ovarian Follicle* ; Ovary ; Theca Cells

OBJECTIVE: To understand the physiologic effects and secretion pattern of inhibin A and inhibin B during menstrual cycle and menopausal transition, inhibin A and inhibin B levels were measured. And to detect any changes in expression of inhibins in human ovary with age, we examined immunohistochemical staining of alpha, beta A, and beta B subunits of inhibin in ovarian tissues. This study was also designed to investigate whether or not inhibin is an early marker for menopausal transition. METHODS: Inhibin A and inhibin B levels were measured in 320 samples from normal reproductive women, in 60 from perimenopausal women, and in 20 from menopausal women by ELISA. And we examined the immunohistochemical staining of alpha, beta A, and beta B subunits of inhibin in ovarian tissues of 35 normal reproductive, 20 perimenopausal, and 5 menopausal women, respectively. RESULTS: In the normal reproductive women, inhibin A begins to increase in the late proliferative phase (16.53 +/- 1.57 pg/ml), reaches the peak in the mid-secretory phase (45.85 +/- 2.08 pg/ml), and subsequently decreases. Inhibin B begins to increase in the early proliferative phase (65.40 +/- 4.08 pg/ml), reaches the peak in the ovulatory phase (110.74 +/- 9.83 pg/ml), and thereafter declines rapidly. In the perimenopausal women, mean inhibin A serum concentration was 6.68 +/- 0.53 pg/ml during proliferative phase and 21.78 +/- 3.61 pg/ml during secretory phase, which were significantly lower than that of the same phase in the normal reproductive women (P<0.01). Mean inhibin B serum concentration was 52.16 +/- 7.46 pg/ml during proliferative phase and 22.41 +/- 6.73 pg/ml during secretory phase, which were significantly lower than that of the same phase in the normal reproductive women (P<0.01, P=0.025). In the menopausal women, both inhibin A and inhibin B were not detected. In the normal reproductive women, we observed strong immunostaining for alpha subunit in granulosa cells, theca cells, and corpus luteum. Immunostaining for beta A subunit was observed in corpus luteum, but not in growing follicles. Immunostaining for beta B subunit was observed in primary follicle, granulosa and theca cells of growing follicle, and mature follicle, but less strong than immunostaining for alpha subunit. No staining for beta B subunit was observed in the corpus luteum. In the perimenopausal women, immunostaining for inhibin subunits were observed in the same pattern as that of the normal reproductive women, but weaker. Stronger immunostaining was observed in theca cells than in granulosa cells. In the menopausal women, none of the immunostaining of inhibin subunits were observed. CONCLUSION: It is concluded that inhibin A is associated with the luteal function and inhibin B, the follicular function. The secretion of inhibins decreased rapidly in the perimenopausal transition period and were not detected in the menopausal period. Inhibin A and inhibin B are associated with the follicular maturation and development. It suggests that the inhibin A and inhibin B are good candidates as markers for perimenopausal transition.
Corpus Luteum ; Enzyme-Linked Immunosorbent Assay ; Female ; Granulosa Cells ; Humans ; Inhibins* ; Menstrual Cycle ; Ovary* ; Theca Cells

OBJECTIVE: Ovarian follicular atresia is initiated from ovarian granulosa cell apoptosis and macrophages exert their effects directly and/or indirectly on follicular atresia by phagocytosis of apoptotic bodies and secretion of various cytokines. In spite of the abundant data on ovarian macrophages, the presence of these cells within the follicles (i.e., among granulosa cells) remains controversial and the elimination methods of apoptotic bodies of atretic follicles, and the time and methods of penetration of macrophages into the follicles are not known completely. The aim of the present study is to demonstrate the presence of macrophage within the ovary as related to follicular atresia and the process of elimination of apoptotic granulosa cells by light and electron microscopy. METHODS: Using rat ovaries, immunohistochemical studies with rat macrophage monoclonal antibody ED1 for macrophages, and light and transmission electron microscopic observations were performed. RESULTS: In the rat, follicular atresia was initiated by the granulosa cell apoptosis which occured randomly within the all granulosa layers. Macrophages were observed within normal follicles, in antrum, granulosa and theca cell layers of atretic follicels, in interstium and in corpus luteum. Ultrastructurally, apoptotic granulosa cells showed characteristics, pyknotic nucleus and apoptotic body formation. Apoptotic bodies were eliminated by intact neighboring granulosa cells and macrophages. Intact granulosa cells ingested apoptotic bodies transiently, soon after they fell into the apoptosis. Finally, apoptotic bodies and degenerating oocytes were phagocytosed by macrophages. Macrophages entered the ovarian follicle at the time of initiation of granulosa cell apoptosis, and migrated with the progression of apoptosis. By elimination of theca cells, macrophages contributed the completion of follicular atresia. CONCLUSION: This study demonstrates both intact neighboring granulosa cells and macrophages in the elimination of apoptotic bodies in atretic follicles of the rat ovary. Macrophages are present within normal follicles, in atretic follicles such as antrum, granulosa and theca cell layers and in corpus luteum but are in different appearances according to their location in ovary. A number of macrophages appearing in atretic follicles and in corpora lutea suggest a role for macrophages in follicular atresia and corpus luteum differentiation. The function of macrophage according to their location in follicular development should be further studied.
Animals ; Apoptosis ; Corpus Luteum ; Cytokines ; Female ; Follicular Atresia* ; Granulosa Cells ; Macrophages* ; Microscopy, Electron ; Oocytes ; Ovarian Follicle* ; Ovary ; Phagocytosis ; Rats* ; Theca Cells

OBJECTIVES: This study was aimed to obtain information on the ontogeny of the production profile of MIS type II receptor (MISR II) and the pattern of its localization according to follicular development METHODS: Expression of MISR II were studied in 21 ovarian specimens from adult normal cycling women by RT-PCR and in situ hybridization of the MISR II mRNA and immunohistochemical staining of the MISR II. RESULTS: The first staining for MISR II and MISR II mRNA were detected in the granulosa cells in primary follicles. The granulosa cells of multiple layered growing follicles showed strong specific staining for MISR II and MISR II mRNA. Among the growing follicles, large follicle stained more intensely than small one. Expression of the MISR II and MISR II mRNA were also seen in the granulosa cells and theca cells of antral follicles. The expression levels of MISR II and MISR II mRNA in mature follicles were lower than those in growing follicles and were even further reduced, but still detectable, in corpus luteum. There was a decreased level of MISR II and MISR II mRNA expression when follicles become atretic. Both expressions were eventually lost from atretic follicles. And the MISR II and MISR II mRNA staining were not found in primordial follicles, oocytes, interstitial cells, ovarian epithelium, and corpus albicans. CONCLUSION: The production and localization of MISR II in granulosa cells, theca cells, and corpus luteum in normal reproductive ovary indicate that actions of MIS via MISR II are autocrine and paracrine in nature. The pattern of MISR II and MISR II mRNA expression according to follicular development indicate that MIS function in the ovary is turned on in primary follicles, increases to maximal levels in large growing follicles, and decreases just before ovulation. These experiments suggest that MIS may play an important role in follicle maturation and follicle selection during the adult reproductive cycle. And this study may yield important information to direct the development of newer contraceptive strategies.
Adult ; Anti-Mullerian Hormone* ; Corpus Luteum ; Epithelium ; Female ; Granulosa Cells ; Humans* ; In Situ Hybridization ; Oocytes ; Ovarian Follicle ; Ovary* ; Ovulation ; RNA, Messenger ; Theca Cells

It has been revealed that multiple cohorts of tertiary follicles develop during some animal estrous cycle and the human menstrual cycle. To reach developmental competence, oocytes need the support of somatic cells. During embryogenesis, the primordial germ cells appear, travel to the gonadal rudiments, and form follicles. The female germ cells develop within the somatic cells of the ovary, granulosa cells, and theca cells. How the oocyte and follicle cells support each other has been seriously studied. The latest technologies in genes and proteins and genetic engineering have allowed us to collect a great deal of information about folliculogenesis. For example, a few web pages (http://www.ncbi.nlm.nih.gov; http://mrg.genetics.washington.edu) provide access to databases of genomes, sequences of transcriptomes, and various tools for analyzing and discovering genes important in ovarian development. Formation of the antrum (tertiary follicle) is the final phase of folliculogenesis and the transition from intraovarian to extraovian regulation. This final step coordinates with the hypothalamic-pituitary-ovarian axis. On the other hand, currently, follicle physiology is under intense investigation, as little is known about how to overcome women's ovarian problems or how to develop competent oocytes from in vitro follicle culture or transplantation. In this review, some of the known roles of hormones and some of the genes involved in tertiary follicle growth and the general characteristics of tertiary follicles are summarized. In addition, in vitro culture of tertiary follicles is also discussed as a study model and an assisted reproductive technology model.
Animals ; Axis, Cervical Vertebra ; Cohort Studies ; Embryonic Development ; Estrous Cycle ; Female ; Genetic Engineering ; Genome ; Germ Cells ; Gonads ; Granulosa Cells ; Hand ; Humans ; Menstrual Cycle ; Mental Competency ; Oocytes ; Ovary ; Pregnancy ; Proteins ; Reproductive Techniques, Assisted ; Theca Cells ; Transcriptome ; Transplants

Testicular granulosa cell tumor(GCT) is a rare neoplasm. We report here on an incidentally discovered testicular granulosa cell tumor in a 36-year-old man. The serum tumor markers were within the normal limits. The ultrasonographic findings revealed a mass with a heterogenous hypoechoic echotexture, including multiple variable sized cystic components. The histology on the orchiectomy specimen demonstrated a gonadal stromal tumor with granulosa cell features. Testicular granulosa cell tumor of the adult type is a very rare tumor, and there have been several isolated case reports and small serial studies described in the literature.
Adult ; Female ; Gonads ; Granulosa Cell Tumor ; Granulosa Cells ; Humans ; Orchiectomy ; Testicular Neoplasms ; Testis ; Biomarkers, Tumor

OBJECTIVES: It has been identified a novel protein in porcine and bovine follicular fluid termed Gonadotropin Surge Inhibiting Factor (GnSIF) which inhibits GnRH-stimulated LH secretion by pituitary cells in culture. The purpose of this study was to investigate the production of GnSIF by porcine granulosa cell in vitro. MATERIALS AND METHODS: Granulosa cells were aspirated from different size of porcine follicles (1-3mm, 3-5mm, >6mm in diameter) and cultured in 24-well plates in Ham's F-12/DMEM at 500,000 cells/well for 24h. After plating, cells were washed and incubated in triplicate for 72h in DMEM/F-12 containing 2% FBS in the presence of 100ng/ml FSH and the media were collected. Granulosa cell-conditioned media were chromatographed on heparin-Sepharose to remove inhibin and follistatin, and assayed for GnSIF activity using a rat pituitary cell bioassay. Briefly, dispersed pituitary cells from adult female Sprague-Dawley rats were plated (500,000 cells/well) for 72h, washed, and incubated with granulosa cell-conditioned media in the presence of 10nM GnRH for 4h. The media were collected and assayed for LH by RIA. GnSIF activity is defined as the suppression of GnRH-stimulated LH secretion. RESULTS: Granulosa cell-conditioned media significantly inhibited GnRH-stimulated LH secretion by rat pituitary cells in vitro (69% of control, P<0.001, n=2 experiments). Preliminary data suggest that granulosa cells from small follicles (1-3mm) produce greater amounts of GnSIF than those obtained from larger follicles (>6mm). CONCLUSIONS: GnSIF activity is produced by porcine granulosa cells in vitro. Granulosa cell production of GnSIF, along with other gonadal proteins such as inhibin, follistatin, and activin may be an important component of the ovarian control of pituitary gonadotropin secretion.
Activins ; Adult ; Animals ; Biological Assay ; Female ; Follicular Fluid ; Follistatin ; Gonadotropin-Releasing Hormone ; Gonadotropins* ; Gonads ; Granulosa Cells* ; Humans ; Inhibins ; Rats ; Rats, Sprague-Dawley

The ectopy of testis may be due to an abnormal connection of the distal end of the gubernaculum testis and fibrous obstruction of scrotal inlet that lead the gonad to an abnormal position. We observed a case of perineal testis that is known to be rare form of ectopic testis and report it with review of literatures.
Bays ; Gonads ; Testis*