Expression of TGFBR1, TGFBR2, TGFBR3, ACVR1B and ACVR2B is altered in ovaries of cows with cystic ovarian disease.

The objective of this study was to examine the expression of transforming growth factor beta receptor (TGFBR)1, TGFBR2, TGFBR3, activin receptor (ACVR)1B and ACVR2B in ovaries of cows with cystic ovarian disease (COD). The expression of the selected receptors was determined by immunohistochemistry in sections of ovaries from cows with ACTH-induced and spontaneous COD. Expression of TGFBR1 and TGFBR3 was higher in granulosa cells of cysts from cows with spontaneous COD than in tertiary follicles from the control group. Additionally, TGFBR3 expression was higher in granulosa cells of cysts from cows with ACTH-induced COD than in those from the control group and lower in theca cells of spontaneous and ACTH-induced cysts than in tertiary control follicles. There were no changes in the expression of TGFBR2. ACVR1B expression was higher in granulosa cells of tertiary follicles of cows with spontaneous COD than in the control group, whereas ACVR2B expression was higher in cysts of the spontaneous COD group than in tertiary follicles from the control group. The alterations here detected, together with the altered expression of the ligands previously reported, indicate alterations in the response of the ligands in the target cells, modifying their actions at cellular level.

Role of Glucocorticoids in Cystic Ovarian Disease: Expression of Glucocorticoid Receptor in the Bovine Ovary.

The aim of this study was to characterize the expression of glucocorticoid receptor (GR) in the components of normal bovine ovary and in animals with cystic ovarian disease (COD). Changes in the protein and mRNA expression levels were determined in control cows and cows with COD by immunohistochemistry and real-time PCR. GR protein expression in granulosa cells was higher in cysts from animals with spontaneous COD and adrenocorticotropic hormone-induced COD than in tertiary follicles from control animals. In theca interna cells, GR expression was higher in cysts from animals with spontaneous COD than in tertiary follicles from control animals. The increase in GR expression observed in cystic follicles suggests a mechanism of action for cortisol and its receptor through the activation/inactivation of specific transcription factors. These factors could be related to the pathogenesis of COD in cattle.

Characterization of persistent follicles induced by prolonged treatment with progesterone in dairy cows: an experimental model for the study of ovarian follicular cysts.

Cystic ovarian disease (COD) is a major factor contributing to poor reproductive efficiency of lactating dairy cows. The objective of the present study was to analyze the endocrine profile, growth dynamics, and histologic characteristics of persistent ovarian follicles-cysts developing in response to long-term administration of intermediate levels of progesterone. To this end, after synchronization of cows, a low dose of progesterone was administered for 5, 10, and 15 days after the expected day of ovulation in treated cows (groups P5, P10, and P15, respectively), using an intravaginal progesterone-releasing device. A significant increase in diameter was detected on Day 11 of progesterone treatment and thereafter (P < 0.05), and at Day 15 of persistence, the diameter of the persistent follicle reached a mean of 23 ± 0.6 mm. Microscopically, the persistent follicles had a complete granulosa, an intensely vascularized theca interna, and a collagenous theca externa layer. Temporal changes in the serum concentrations of estradiol, progesterone, and FSH were detected (effects of time, P < 0.01). Progesterone treatment completely inhibited the LH preovulatory surge in treated cows and affected the basal concentration of LH. The pulse frequency remained high at 5 and 10 days of persistence and declined (P < 0.05) after 15 days of persistence. The LH pulse concentration and pulse amplitude had a significant reduction (P < 0.05) during follicular persistence. Changes in the serum levels of estradiol, progesterone, 17-hydroxyprogesterone, and testosterone in serum and follicular fluid were also observed. In serum, estradiol increased gradually from proestrus to Day 10 of follicular persistence (P < 0.05), progesterone showed an increase (P < 0.05) at Day 5 of follicular persistence, 17-hydroxyprogesterone showed a significant decrease at 5 days of follicular persistence in relation to proestrus, and testosterone showed a significant increase (P < 0.05) from proestrus and Day 5 of persistence through Day 15 of follicular persistence. Correlation between serum and follicular fluid steroid concentrations was significant for testosterone (P < 0.0001) and not significant for estradiol and progesterone. These findings indicate that ovarian cysts in COD are similar in many ways to the persistent follicles induced by progesterone, with an analogous hormonal and morphologic context, thus confirming a local role of subluteal levels of progesterone in COD pathogenesis and in the regulatory mechanisms of the ovarian function.

Altered expression of transforming growth factor-beta isoforms in bovine cystic ovarian disease.

Cystic ovarian disease (COD) is one of the main causes of infertility in dairy cattle. It has been shown that intra-ovarian factors may contribute to follicular persistence. Transforming growth factor-beta (TGFB) isoforms are important paracrine and autocrine signalling molecules that regulate ovarian follicle growth and physiology. Considering the importance of these factors in the ovarian physiology, in this study, we examined the expression of TGFB isoforms (TGFB1, TGFB2 and TGFB3) in the ovary of healthy cows and animals with spontaneous and adrenocorticotrophic hormone (ACTH)-induced COD. In the oestrous-synchronized control group, the expression of TGFB1 in granulosa and theca cells was higher in spontaneous cysts than in atretic or tertiary follicles. When we compared TGFB2 expression in granulosa cells from atretic or tertiary follicles from the oestrous-synchronized control group with that in ACTH-induced or spontaneous follicular cysts, we found a higher expression in the latter. The expression of the TGFB isoforms studied was also altered during folliculogenesis in both the spontaneous and ACTH-induced COD groups. As it has been previously shown that TGFB influences steroidogenesis, ovarian follicular proliferation and apoptosis, an alteration in its expression may contribute to the pathogenesis of this disease.

Ovarian localization of 11ß-hydroxysteroid dehydrogenase (11ßHSD): effects of ACTH stimulation and its relationship with bovine cystic ovarian disease.

Cystic ovarian disease (COD) is an important cause of infertility in cattle, and ACTH has been involved in regulatory mechanisms related to ovarian function associated with ovulation, steroidogenesis, and luteal function. Here, we examined the localization of 11ß-hydroxysteroid dehydrogenase type 1 (11ßHSD1) and 11ßHSD2 proteins in the ovary of healthy cows and animals with spontaneous and ACTH-induced COD and the in vitro response of the follicular wall exposed to ACTH. After stimulation by ACTH, we documented changes in 11ßHSD expression and cortisol secretion by the follicular wall of large antral and follicular cysts. Follicular cysts showed a higher constitutive expression of both enzymes, whereas ACTH induced an increase in 11ßHSD1 in tertiary follicles and follicular cysts and a decrease in 11ßHSD2 in follicular cysts. Moderate expression of 11ßHSD1 was observed by immunohistochemistry in granulosa of control animals, with an increase (P < 0.05) from primary to secondary, tertiary, and atretic follicles. The level of immunostaining in theca interna was lower than that in granulosa. The expression of 11ßHSD2 was lower in the granulosa of primary follicles than in that of secondary, tertiary, and atretic follicles and was lower in the theca interna than in the granulosa. In ACTH-induced and spontaneously occurring follicular cysts, differences from controls were observed only in the expression of 11ßHSD1 in the granulosa, being higher (P < 0.05) than in tertiary follicles. These findings indicate that follicular cysts may be exposed to high local concentrations of active glucocorticoids and indicate a local role for cortisol in COD pathogenesis and in regulatory mechanisms of ovarian function.

Changes in the expression of Heat Shock Proteins in ovaries from bovines with cystic ovarian disease induced by ACTH.

Cystic ovarian disease (COD), which is considered one of the most important causes of reproductive failure in dairy cattle, induces intraovarian changes in the expression of numerous genes. The purpose of this study was to analyze the changes in the expression of Heat Shock Proteins (HSPs) in ovaries from bovines with cystic ovarian disease induced by ACTH. Immunoreactivity for Heat Shock Proteins (HSPs) in ovaries of cows with induced COD showed differential expression patterns in growing follicles from the control group. The immunopositive area for Hsp27 and Hsp60 in granulosa cells showed significant differences between tertiary follicles from normal cycling animals and those from animals with induced COD. The cysts showed increased Hsp27 immunostaining in theca cells in relation to tertiary follicles from normal cycling cows. Hsp70 immunostaining was more intense in cystic follicles than in other follicular categories from animals with induced COD, in both granulosa and theca cells. In granulosa cells, tertiary follicles from the control group showed higher levels of Hsp90 than cysts. These results demonstrate that there are differences in HSP protein expression when COD is induced. In fact, HSP expression would be part of the functional response to the changes in hormones and neurotransmitters induced by stress, indicating that HSPs can control hormonal functions and vice versa.

Cell proliferation and survival mechanisms underlying the abnormal persistence of follicular cysts in bovines with cystic ovarian disease induced by ACTH.

Cystic ovarian disease (COD) is an important cause of infertility that affects cattle. Alterations in the ovarian micro-environment of females with follicular cysts could alter the normal processes of proliferation and programmed cell death in ovarian cells. Thus, the objective in the present study was to evaluate apoptosis and proliferation in induced ovarian cystic follicles in cows to investigate the follicular persistence. Stage of estrous cycle was synchronized in 10 heifers and 5 were then subjected to the induction of COD by administration of ACTH. After the ovariectomy number of in situ apoptotic cells by TUNEL assay, active caspase-3, FAS/FASLG and members of the BCL2 family were compared by immunohistochemistry and multiplex PCR and cell proliferation by evaluation of Ki-67 protein and cyclin D1 and E mRNA. Significantly (p<0.05) lesser proliferative and apoptotic rates were found in cystic follicles from cows with COD compared with those with regular cycles. The relatively minimal proliferation found by immunohistochemistry with Ki-67 marker were confirmed by the gene expression of cyclin D1 and E. Lesser apoptotic rates were associated with decreased amounts of apoptotic-related proteins BAX, FASLG and caspase-3 as well as the in situ apoptosis detected by TUNEL assay, and increased amounts of the anti-apoptotic survival factor cellular BCL2 in the cystic follicles of the COD group. The BAX/BCL2 gene expression profile confirmed the immunohistochemical findings. Results from the present study indicate that cellular proliferation and apoptosis are altered in cystic follicles of cattle. The present study provides new insights into the molecular mechanisms underlying the aberrant persistence of follicular cysts and related diseases.

Ovarian cysts in users of Implanon and Jadelle subdermal contraceptive implants.

OBJECTIVE: To evaluate the prevalence of ovarian cysts in users of two contraceptive implants compared to users of intrauterine contraceptive (IUC). MATERIALS AND METHODS: A total of 344 women were enrolled at 3 months of use of the etonogestrel-releasing implant (Implanon), the levonorgestrel-releasing implant (Jadelle) or the TCu380A IUC. Bimanual pelvic examination and vaginal ultrasound were performed during routine 3-, 6- and 12-month visits of asymptomatic women. Women with ovarian cysts (or enlarged ovarian follicles > or = 25 mm) were assessed weekly until disappearance or reduction of the image [including estradiol (E2) and progesterone (P) measurement], and women with no ovarian enlargement underwent the same evaluation for the same period of time. RESULTS: Ovarian cysts were detected in 5.2%, 13.0% and 1.9% of users of Implanon, Jadelle and IUC, respectively, at 3 months. At 6 months of use, prevalence was 7.2%, 8.0% and 2.1%, whereas 12-month rates were 26.7%, 14.6% and 1.2% in the three groups, respectively. E2 levels were significantly higher among users with ovarian cysts than controls. The time until disappearance of the ovarian cyst was similar in all three groups. There were more cases of menorrhagia in users of implants who had ovarian cysts than in those with no ovarian enlargement. CONCLUSIONS: The finding of ovarian cysts or enlarged ovarian follicles during the first year of use of Implanon and Jadelle implants is common and transient and should not be interpreted as a pathologic ovarian cyst. No further medical interventions are necessary.

Effects of dehydroepiandrosterone on ovarian cystogenesis and immune function.

The purpose of the present report was to study the possible relationship between ovarian functionality and the immune response during cystogenesis induced by androgenization with dehydroepiandrosterone (DHEA). Daily injection of DHEA (6 mg/kg body weight) for 20 consecutive days induced ovarian cysts in BALB/c mice. As markers of ovarian function, serum estradiol (E) and progesterone (P) and the ovarian inmunomodulator prostaglandin E (PGE) were analyzed. In order to know how the integrity of the tissue was altered after induction of cystogenesis, the oxidative status was also evaluated. Serum E and P levels, and ovarian PGE concentration, were increased in animals with cysts compared with healthy controls. The oxidant status (quantified by malondialdehyde (MDA) formed after the breakdown of the cellular membrane by free radical mechanisms) was augmented, meanwhile the antioxidant (evaluated by the glutathione (GSH) content) diminished during the induction of cystogenesis. Both immunohistochemical and flow cytometry assays demonstrated that DHEA treatment increased the number of T lymphocytes infiltrating ovarian tissue. Therefore, while ovarian controls showed equivalent expression of CD4+ and CD8+ T cell subsets, injection of DHEA yielded a selective ovarian T cell infiltration as demonstrated by enhanced CD8+ and diminished CD4+ T lymphocyte expression. These results show that the development of cysts involves changes in ovarian function and an imbalance in the oxidant-antioxidant equilibrium. We observed also both an increased and selective T lymphocyte infiltration.

Prepubertal administration of estradiol valerate disrupts cyclicity and leads to cystic ovarian morphology during adult life in the rat: role of sympathetic innervation.

Administration of estradiol valerate (EV) to adult rats leads to anovulation and cystic ovarian morphology. Sympathetic ovarian nerve denervation (SONX) overcomes this disruption. In this study, we determined whether EV administration to juvenile rats prevents achievement of reproductive competence, disrupts cyclicity, and whether this programming is facilitated via activation of the sympathetic nerve input to the ovary. Prepubertal rats were administered 2 mg EV in corn oil or corn oil alone. One half of the animals from each group underwent SONX on d 71 of life. Rats were euthanized on d 91 for determination of serum gonadotropins, progesterone, Delta4 androstenedione, and estradiol concentrations, ovarian norepinephrine (NE), and 3beta-hydroxysteroid dehydrogenase (3beta-HSD) activities and ovarian dynamics. Results revealed that EV administration during juvenile period advanced pubertal onset, suppressed circulating LH, FSH, and Delta4 androstenedione, increased ovarian NE, estradiol, and 3beta-HSD activities, disrupted ovarian dynamics evidenced as absent corpus luteum and presence of ovarian cysts and culminated in anovulation. SONX restored cyclicity in these animals, normalized LH, estradiol, ovarian 3beta-HSD activities, and ovarian dynamics as evidenced by the disappearance of ovarian cysts and appearance of corpus luteum and restored corpus luteum function. These findings provide evidence that EV exposure during juvenile life leads to long-lasting deleterious reproductive consequences via activation of the sympathetic ovarian nerve.

Nonmenstrual adverse events during use of implantable contraceptives for women: data from clinical trials.

Contraceptive methods, including implants, do not prevent common symptoms and adverse health events that most people experience. It is difficult, therefore, to decide whether or not the occurrence of symptoms or adverse events that are common can be attributed to use of a contraceptive method or to determine if a given method changes the likelihood of their occurrence. Based on the review of the literature, no apparent differences in the frequency of adverse events are evident between the six-implant or two-rod levonorgestrel systems and the single implant etonogestrel and nomegestrol acetate systems. The most frequent adverse events reported in clinical trials that are probably related to implant use are headaches and acne. Weight gain, dizziness, and mood changes are also frequently mentioned adverse events and are possibly steroid-related. Other possibly related adverse events, although much less frequently reported, are loss of libido, fatigue, hair loss, and other skin conditions. Persistent ovarian follicles that spontaneously disappear are a common event during use of progestin-only contraceptives, and providers should be aware of this condition to avoid unnecessary interventions. Overall, the vast experience reported in the clinical studies reviewed here show that all existing implantable contraceptives are equally safe. This can probably be attributed to the low-hormonal dose delivered by progestin-implant systems.

[Abnormalities of sexual development in Puerto Rico: status report]. / Anomalías de desarrollo sexual en Puerto Rico: estado de situación.

Puerto Rico presents the highest incidence in the world of anomalous sexual development. The authors have collected over 3100 cases in the past 19 years. Clinical and laboratory studies suggest possible estrogenic contamination of meats and poultry products. Variation in diet provides protection to a significant number of patients. The possibility of mycotic contamination of food employed in animal husbandry by Fusarium sp., and mycotoxins capable of estrogenic effects have been suggested in a preliminary study. Private and government investigators are active in the study of the condition, but more fiscalizing action is needed. A detailed study of food components for possible contaminants determination is considered mandatory.

Anomalías de desarollo sexual en Puerto Rico: estado de situación / Abnormalities of sexual development in Puerto Rico: status report

Puerto Rico presenta el mayor número de casos conocidos de anomalias de desarrollo sexual en infantes, niños y adolescentes. Los autores han evaluado sobre 3100 pacientes en un período aproximado de 19 años. Diversos estudios clínicos y de laboratorio sugieren una posible contaminación de carnes, pollo y derivados de éstos con sustancias estrogénicas. Una dieta modificada, libre de estos alimentos, causa una mejoría clínica y/o de laboratorio en un número significativo de pacientes. Un estudio preliminar en alimentos utilizados en el engorde de pollos sugiere la contaminación por Fusarium sp, hongo que puede producir micotoxinas con efecto estrogénico. Al presente, diversos investigadores, privados y de gobierno, llevan a cabo diversos estudios. Sin embargo, se precisa de una acción más vigorosa y fiscalizadora. Consideramos imprescindible un estudio específico de los alimentos de engorde para determinar la presencia de contaminantes con efecto hormonal. La persistencia de estas anomalías debe deterner-se por sus posibles consecuencias a largo plazo