Aberrant effects of altrenogest and exposure to exogenous gonadotropins on follicular cysts appearance in gilts.

Research was conducted to determine the effect of altrenogest and exposure to exogenous gonadotropins on ovarian function in prepubertal and mature gilts. Crossbred, presumably sexually mature gilts (n = 51), were fed with altrenogest for 18 consecutive days and the day after the last feeding with altrenogest, gilts were treated with eCG and 72 hours later challenged with hCG. Animals were slaughtered on Days 10 to 13 of their gonadotropins synchronized estrous cycle. Ovaries were examined for the number of CL, number of follicular cysts, and presence of corpora albicantia. Gilts were divided into two groups: those possessing corpora albicantia (group A-mature; n = 36) and those without corpora albicantia (Group W-prepubertal; n = 15) on their ovaries. In addition, each group was divided into two subgroups depending on the presence of follicular cysts (AC and WC) or their absence (AO and WO). There was no difference between the number of CL in group A and group W. Presence of corpora albicantia determined percentage of gilts possessing follicular cysts (13.9% group A vs. 66.7% group W). Gilts without follicular cysts (AO plus WO; n = 36) had higher number of CL (P < 0.01) than gilts bearing cysts (AC plus WC; n = 15). Comparison AO-AC did not show significant difference (P = 0.075) between CL number in mature cyst-free and cysts bearing gilts. A prepubertal gilts not bearing follicular cysts (WO) had higher (P < 0.02) number of CL than gilts bearing cysts. A significant negative correlation between the number of CL and number of follicular cysts was found (r = -0.664; P = 0.007). There were no differences in blood plasma progesterone and estradiol concentration between cyst-free and cyst-bearing gilts. These results indicate: (1) a higher follicular cysts appearance in prepubertal than mature gilts challenged with altrenogest and exposed to exogenous gonadotropins and (2) a negative effect of follicular cysts on the number of CL (ovulations) in prepubertal gilts.

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.

Exogenous androstenedione induces formation of follicular cysts and premature luteinization of granulosa cells in the ovary.

OBJECTIVE: To investigate the effects of androstenedione on ovarian follicle development. DESIGN: Experimental study. SETTING: University research laboratory. ANIMAL(S): Female Wistar-Imamichi rats and BDF1 mice. INTERVENTION(S): Rats were injected with androstenedione. Ovarian follicles of mice were cultured in the presence of androstenedione. MAIN OUTCOME MEASURE(S): Ovarian morphology; ovarian cell types undergoing apoptosis; ovarian expression of cytochrome P450 aromatase (P450arom), cytochrome P450 side-chain cleavage (P450scc), and cyclin-dependent kinase inhibitor p27(kip1); serum levels of T, E(2), and P in rats; and ultrastructure of granulosa cells from cultured follicles of mice. RESULT(S): In androstenedione-treated rat ovaries, follicular cysts were formed, and apoptotic cells were found in the inner part of granulosa cell layers of antral follicles. Androstenedione administration down-regulated expression of P450arom but up-regulated expression of P450scc and p27(Kip1) in the granulosa cells of antral follicles. Serum T levels were significantly increased in androstenedione-treated rats. In mouse follicles exposed to androstenedione, the granulosa cells contained abundant lipid droplets and mitochondria with complex tubular cristae. CONCLUSION(S): Excess androgen enhances apoptosis in the inner part of granulosa cell layers of antral follicles, resulting in the formation of follicular cysts. It is also demonstrated that androgen stimulates premature luteinization of granulosa cells.

Collaborative work on evaluation of ovarian toxicity. 12) Effects of 2- or 4-week repeated dose studies and fertility study of indomethacin in female rats.

2-week and 4-week general toxicity studies of indomethacin, a nonselective inhibitor of cyclooxygenase 1 and 2, were performed using rats. A female fertility study was also conducted to compare the results to those of ovarian histopathological findings. The main purposes of the present studies are to assess whether a precise histopathological examination, taking the morphological changes the female reproductive organs undergo during each estrus phases into account, can evaluate toxicity to the ovaries, and to determine the optimal administration period for detecting ovarian toxicity. Indomethacin was administered on a daily basis to female Sprague-Dawley rats at doses of 0, 0.4, 1.3, or 4 mg/kg in the both the general toxicity studies and the female fertility study. In the general toxicity studies, unruptured follicles or luteinized cysts were observed histopathologically in the 4 mg/kg group in both the 2-week and 4-week studies. In addition, follicular cysts were found in the 4 mg/kg group in the 4-week study. Estrous cyclicity was not disturbed in both studies. There were no histopathological changes in the ovaries of the 1.3 mg/kg group in general toxicity studies. In the female fertility study, no toxic effects on female fertility parameters were detected in the 0.4 and 1.3 mg/kg group treated with indomethacin, but 8 of 10 rats in the 4 mg/kg group died or were sacrificed before completion of the dosing period. These results demonstrated that 2 weeks of indomethacin treatment is sufficient to detect unruptured follicles or luteinized cyst in the ovary. In addition, 4 weeks of dosing maybe required for induction of follicular cysts, although we could not clearly show that these histopathological changes would affect female fertility functions. These present studies suggest that a precise histopathological examination may be able to predict the effect of test articles on female reproductive functions.

Collaborative work on evaluation of ovarian toxicity. 17) Two- or four-week repeated-dose studies and fertility study of sulpiride in female rats.

To find the appropriate dosing period to detect ovarian toxicity, sulpiride, a D2 antagonist was orally dosed to female rats at dose levels of 1, 10, and 100 mg/kg/day daily for 2 or 4 weeks in repeated-dose toxicity studies. In addition, sulpiride at the same dose levels was given to female rats daily during the pre-mating period, mating period, and Days 0-7 of gestation to assess its effect on fertility. In ovarian histology in the 2-week study, increases in atretic follicle were seen at 1 mg/kg or more and increases in follicular cysts at 10 mg/kg or more. In the 4-week study, these findings were seen at 1 mg/kg or more, and a decrease in large follicles was seen at 10 mg/kg or more. Increased body weight gain was observed at 10 mg/kg or more in the 2- and 4-week studies. The females in these groups exhibited development of mammary alveolus by sulpiride-induced hyperprolactinemia. In the fertility study, sulpiride-treated females showing persistent diestrus resulted in successful mating, and almost all females got pregnant. However, increased implantation loss was observed at 10 mg/kg or more, which was considered to be caused by the adverse effect of sulpiride on oocyte development. From these results, sulpiride-induced ovarian toxicity was seen at 1 mg/kg or more in the 2- and 4-week repeated-dose toxicity studies, and the observed ovarian changes were considered to be related to adverse effects on female fertility.

Does exogenous progesterone and oestradiol treatment from the mid-luteal phase induce follicular cysts in goats?

The purpose of the present study was to investigate the effects of exogenous ovarian steroid treatment, which is known to induce follicular cyst experimentally in cows, on ovarian activity in goats. Eleven female Shiba goats with the length of the normal oestrous cycle (approximately 21 days) received subcutaneously either 1 ml of ethanol (control group, n=4) or 4 mg of progesterone and 2mg of oestradiol (treatment group, n=7) daily for 7 days beginning on day 14 of the oestrous cycle (day 0=ovulation). Ultrasonographic images of the ovary and blood samples were collected daily to monitor the ovarian activity. Ovulation was observed before 1 day after the end of treatment in the control group. In the treatment group, no detectable structures of follicles or corpus luteum (static ovarian condition) were found for 6.0+/-1.4 days (mean+/-S.D.) after the end of treatment. Then, detectable follicles appeared and ovulation was observed in all animals of the treatment group. There was no significant difference in the maximum diameter of the ovulatory follicle between the control and treatment group (4.7+/-0.4mm versus 5.1+/-0.7 mm). The large non-ovulatory follicles, which grew more than 10mm in diameter were observed after the static ovarian condition in one goat of the treatment group, whereas no turnover of the cystic follicular structures was found. The length of the inter-ovulatory intervals in the treatment group was significantly longer than that in the control group (38.4+/-7.4 days versus 20.3+/-0.5 days, P<0.05). The present results demonstrated that the exogenous treatment of progesterone and oestradiol, which was adapted from the follicular cyst model in cows, did not induce follicular cysts in goats, suggesting that there is/are different mechanism(s) mediating the occurrence of follicular cysts between cows and goats.

Involvement of nitric oxide and the ovarian blood follicle barrier in murine follicular cyst development.

OBJECTIVE: To test the involvement of nitric oxide in murine ovarian follicular cysts. DESIGN: Controlled animal study. SETTING: Academic research environment. ANIMAL(S): Immature female B6D2F1 mice at 23 +/- 2 days old. Ovarian cysts were induced by implanting miniosmotic pumps that delivered and maintained constant levels of hCG. Nitric oxide studies included the delivery of nitric oxide synthase (NOS) inhibitors, N(G)-nitro-L-arginine methyl ester (L-NAME), or N(G)-nitro-D-arginine methyl ester, by the same method. Ovulation assays measured cumulus oocyte complexes and blood follicle barrier (BFB) function. RESULT(S): Chronic treatment with hCG induced enlarged ovaries containing multiple follicular cysts, which were approximately double the size of follicles in sham-operated mice. These cysts enclosed few, if any granulosa cells, secreted high levels of testosterone, and had impaired ovarian BFB function. Inhibition of NOS by L-NAME during ovarian cyst formation reduced the size of follicular cysts, sustained normal testosterone levels, and maintained hormonal BFB reactivity in cystic follicles. CONCLUSION(S): Nitric oxide was found to be involved in the formation of hCG-induced murine follicular cysts and complications associated with these cysts were ameliorated by the NOS inhibitor L-NAME.

A GnRH/LH surge without subsequent progesterone exposure can induce development of follicular cysts.

Our hypothesis was that follicular cysts would develop if cows experienced an estradiol-induced GnRH LH surge in the absence of an ovulatory follicle. Further, we hypothesized that estradiol would fail to induce a subsequent GnRH/LH surge in these cows until they were treated with progesterone. In experiment 1, seven cows were synchronized with a controlled internal drug releasing device (CIDR) for 9 d and each received 500 microg of cloprostenol on d 7. All follicles (> or = 5 mm in diameter) were aspirated at the time of CIDR removal using transvaginal follicular aspiration. Two days after aspiration, cows were treated with 5 mg of estradiol benzoate (EB) to induce a GnRH/LH surge in the absence of an ovulatory-sized follicle. All cows had an LH surge following the estradiol treatment and three of seven developed an anovulatory condition that resembled follicular cysts. The four cows that did not develop follicular cysts luteinized remaining cells from one aspirated follicle each. Thus, all cows with a progesterone elevation after the estradiol/GnRH/LH surge had subsequent ovulatory cycles, whereas the absence of progesterone was followed by follicular cysts. After 49 d, the anovulatory cows were induced back to normal cyclicity by insertion of a CIDR for 7 d. In two subsequent experiments, nine of 26 cows were induced to have follicular cysts by follicular aspiration followed by 5 mg of EB. After 26 d of observation, all cystic cows received a second treatment with 5 mg of EB and none of the cows showed an LH surge or ovulation. Cystic cows were untreated (n = 4 controls) or treated for 7 d with a CIDR (n = 5). All cystic cows were subsequently treated for a third time with 5 mg of EB. All CIDR-treated cows had an LH surge and ovulated, whereas none of the control cows had an LH surge or ovulation after the estradiol treatment. Thus, a large follicle anovulatory condition, similar to follicular cysts, can be induced by estradiol induction of a GnRH/LH surge in the absence of subsequent luteinization, and this condition prevents a GnRH/LH surge in response to high doses of estradiol. Progesterone eliminates this condition by reinitiation of GnRH/LH surges in response to estradiol.

Ultrasonographic monitoring of ovarian follicles in women using norethisterone for contraception.

OBJECTIVE: The purpose of this study was to examine the effect of intranasal and oral norethisterone (NET) on ovarian folliculogenesis. METHODS: Sixteen healthy, sterilized women with regular menstrual cycles were recruited to the study. NET 300 micrograms per day was administered orally (n = 8) or intranasally (n = 8) for two consecutive menstrual cycles. Serial pelvic ultrasonography was performed to monitor ovarian follicular growth. RESULTS: Ultrasonographic evidence of normal follicular growth and ovulation was observed in 10 cycles whilst 22 cycles were anovulatory. Formation of follicular cysts was seen in 14 cycles, 13 of which were anovulatory and in one ovulation was observed in the opposite ovary. The size of the cysts varied between 27 and 44 mm. The cysts disappeared when NET treatment was discontinued. A positive correlation between cyst size and estradiol levels was observed with intranasal NET in 50% of cyst cycles. In three cycles, although normal follicular growth and endocrine profile were observed, the follicles failed to rupture. These were classified as luteinized unruptured follicles. Immature follicles < 10 mm were seen in six cycles. CONCLUSION: The study showed that NET administered either orally or intranasally evidently disturbs normal follicular growth and rupture.

Changes in plasma concentrations of gonadotropins and steroid hormones during the formation of bovine follicular cysts induced by the administration of ACTH.

Bovine follicular cysts were induced by treatments with ACTH (3 mg, im) daily for 14 days beginning in the late luteal phase. Cortisol concentrations in plasma significantly increased after ACTH treatments. During the formation of follicular cysts induced by the injections of ACTH, mean plasma concentrations of progesterone were significantly higher than those in the untreated preovulatory period, while mean plasma concentrations of estradiol-17 beta were significantly lower. During the treatment period, mean plasma concentrations of LH and FSH remained low, and the preovulatory surges of LH and FSH did not occur. Suppressed concentrations of LH and FSH might be caused by the increases in secretions of cortisol and progesterone, and by the decrease in secretion of estradiol-17 beta.

Complications of ovulation induction.

Induction of ovulation has its own risks. Since this treatment is elective the physician should be convinced that it is really indicated for the specific patient. Multiple pregnancies still occur in 4 to 15% in in vivo treatment and in 15 to 20% in assisted reproduction. Abortions occur in 20% of the pregnancies achieved. These numbers demonstrate the complexity of induction of ovulation. In recent years the average age of the treated patient has increased, but it is too early to see whether this influences the frequency of complications. The physician should be aware of the possible complications and should remain in contact with the patients at risk after completion of the treatment. The patient should be well informed about the possible complications before starting treatment. At the end of the treatment she should be able to recognize any clinical warning signs of OHSS and inform her physician, in order to be treated appropriately. Further studies of the pathogenesis of OHSS in the future will hopefully lead to more specific treatments or even prevention of this phenomenon. The increasing experience in selective fetal reduction seems to be a practical solution to high rank multifetal gestation, preventing extreme prematurity and its sequelae.

Induction of ovarian follicular cysts in the pregnant rat by human chorionic gonadotropin.

Prolonged stimulation by human chorionic gonadotropin (hCG) induces ovarian follicular cysts in progesterone-synchronized immature rats [Bogovich, Endocrinology 1989; 124:1646-1653]. To determine if unabated stimulation by hCG has a similar effect on follicular development in adult ovaries, pregnant rats were given either 0 (control), 1, or 3 IU hCG twice daily for 9 days beginning on Day 13 of pregnancy. By Day 22 of pregnancy, rats treated with 1 IU hCG possessed large antral follicles at least 1 mm in diameter: approximately 33% larger than the diameters of preovulatory follicles observed in control rats (0 IU hCG). In contrast, rats treated with 3 IU hCG displayed ovarian follicular cysts up to 5 mm in diameter, with well-developed thecae and just a remnant of granulosa cells. Progesterone, androstenedione, and estradiol accumulation was greater in follicular incubates from hCG-treated rats than in incubates from control rats. Progesterone increased in response to cAMP in incubates from all treatment groups on all days tested. Androstenedione increased in response to cAMP on Day 22 of pregnancy for follicles from control animals, on all days tested for follicles from rats treated with 1 IU hCG, and on Days 15-19 for follicles from rats treated with 3 IU hCG. Androstenedione production in the presence of 300 ng of exogenous testosterone was significantly greater in follicular incubates from animals treated with 1 and 3 IU hCG than incubates from control animals on Days 19-22 of pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)

Follicle cyst formation following long-acting gonadotropin-releasing hormone analog administration.

The incidence of follicular cyst formation is 13.6% in cycles treated with GnRH-a. A conservative mode of treatment is suggested, leading to the disappearance or regression of the cysts. Doing so, the number and quality of the oocytes and embryos, the pregnancy and abortion rates are not significantly different between the groups with and without follicular cysts.