Although 60–80% of the women with polycystic ovary syndrome (PCOS) ovulate with clomiphene citrate (CC), the rest are CC-resistant. Recently, the use of insulin-sensitizing agents such as metformin and pioglitazone have been proposed for inducing ovulation in CC-resistant women with PCOS, and we have reported that administration of bezafibrate, a lipid-lowering fibrate, in addition to CC, successfully induced ovulation in CC-resistant women with PCOS and dyslipidemia. Both pioglitazone and bezafibrate are peroxisome proliferator-activated receptor-γ (PPAR-γ) agonists. This paper reviews the evidence for the direct effects of the drugs, which are PPAR-γ agonists, on follicle development and steroidogenesis, collected using an in vitro mouse preantral follicle culture system. We used the in vitro follicle culture system with the addition of tumor necrosis factor-alpha (TNF-α), which plays a role in insulin resistance, as a model for studying follicle development in women with PCOS. TNF-α inhibited FSH-induced follicle development and steroidogenesis in the follicle culture system. Both pioglitazone and bezafibrate prevented TNF-α-mediated inhibition of FSH-induced follicle development and steroidogenesis through the PPAR-γ-stimulating pathway. Our results suggest that insulin-sensitizing drugs, especially PPAR-γ agonists, may directly influence follicle development and steroidogenesis in women with PCOS.

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Polycystic ovary syndrome (PCOS) is a common endocrine disorder in reproductive-age women. Hyperandrogenism, chronic anovulation, and infertility are the main features of this heterogeneous condition. The diagnosis of PCOS is based on a combination of clinical, biological, and ultrasound criteria that have been used variably to define PCOS. The usual clinical presentation of PCOS in Asia is slightly different from that in the United States and Europe, with less frequently encountered cases of hyperandrogenism. Moreover, non-obese PCOS is typical in Asian women. The Japanese Society of Obstetrics and Gynecology has recently proposed new, revised diagnostic criteria. Growth arrest of ovarian follicles in the non-obese PCOS is assumed to be associated with an abnormal endocrine environment involving chronically elevated LH. In vitro studies currently demonstrate that LH promotes follicular growth during preantral-early antral transition via the increased synthesis and growth promoting action of androgen. However, chronic LH stimulation impairs FSH-dependent antral follicle growth by suppressing FSH receptor expression in granulosa cells, via the modulation of intraovarian regulators. Therefore, the adverse effect(s) of chronically elevated LH on the theca cell/androgen system should also be considered for improved care of non-obese PCOS patients.

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Insulin resistance is one of the key factors in the pathogenesis of polycystic ovary syndrome (PCOS). Obesity, visceral fat accumulation and excessive serine phosphorylation of the insulin receptor are factors responsible for insulin resistance in PCOS. Insulin resistance induces compensatory hyperinsulinemia, which stimulates androgen synthesis in the theca cells of the PCOS ovary. Therefore, hyperinsulinemia results in hyperandrogenemia in PCOS patients who tend to have androgen-producing ovaries. In Japanese PCOS, 25% of patients are obese and 30% of patients have insulin resistance. Improvement of insulin resistance by weight loss, exercise and insulin sensitizing drugs such as metformin can recover reproductive function. Meformin has the potential to induce ovulation in PCOS, and 58% of clomiphene resistant Japanese PCOS patients resume ovulation after combined clomiphene metformin treatment. Clomiphene-metformin therapy is simple and has low risks of multiple pregnancy and ovarian hyperstimulation syndrome. Metformin use should be considered fertility treatment, especially as a second line therapy for the clomiphene resistant PCOS.

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Polycystic ovary syndrome (PCOS) is a heterogeneous group of disorders characterized by ovulation disorder, hyperandrogenism, and polycystic ovarian morphology (PCOM). Several diagnostic criteria suggest that hyperandrogenism is a core symptom of PCOS. Androgens are believed to cause preantral follicle growth and arrest the growth of antral follicles. This results in accumulation of small antral follicles in the ovaries, thus forming PCOM. Observational studies of patients with female-to-male transsexualism or congenital adrenal hyperplasia indicate that androgen administration to these patients does not produce typical PCOS-like features. However, endogenous androgen exposure in early life may lead to some traits of PCOS in adulthood. To reveal the association between the timing of excess androgen exposure and reproductive function, various animal models have been investigated using androgen administration. Rhesus monkeys exposed to excess androgen during the early fetal period show a PCOS-like phenotype, including metabolic and hypothalamic-pituitary characteristics. This finding implies that exposure to excess androgen during this critical period programs the hypothalamic-pituitary-ovary axis and metabolic organs. Although findings obtained in animal studies will not necessarily be replicated in humans, prenatal androgen excess is the dominant PCOS hypothesis.

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Morphological changes are observed during the luteinization process of granulosa cells, however mechanisms and the roles are remained unclear. In the present study, we demonstrated that Sema3C is expressed in granulosa cells and then secreted to extracellular matrix accumulated within cumulus cell and granulosa cell layers. The expression is dependent on EGF-like factors (amphiregulin and neuregulin 1) and EGF receptor dependent manner. The knockdown of Sema3c not only significantly decreased the surface area of the granulosa cells, but also significantly suppressed the cell migration of granulosa cells cultured with EGF like factors. The phosphorylation of focal adhesion kinase (FAK) was dramatically suppressed by transfection with Sema3c siRNA, with an increasing level of phosphorylated ROCK, indicating that Sema3C regulated actin remodeling in a FAK signaling pathway-dependent manner. Moreover, the expression levels of genes involved in progesterone production, Star, Cyp11a1 were significantly decreased in Sema3c siRNA-transfected granulosa cells as compared with control siRNA transfection. From these results, we conclude that Sema3C is secreted and then accumulated within granulosa cell layers and cumulus cell layers. The FAK pathway is activated by Sema3C and induces cell migration and enlarges the area of granulosa cells. These morphological changes are required for luteinization (production of progesterone) of granulosa cells.

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A time-lapse embryo image monitoring system consists of an incubator with a built-in microscope and CCD camera. Embryo assessment can be performed with this type of system without removing embryos from the incubator, which eliminates the risks of stress due to temperature changes, light exposure, high oxygen exposure, and pH changes in the culture medium. We cultured embryos to the blastocyst stage using this system and compared the results with those of a normal incubator. We compared the rates of fertilization and cryopreservation of embryos cultured using EmbryoScope^® and a conventional incubator. We then compared the pregnancy rates of EmbryoScope^® and the conventional incubator. We finally examined the relationship between the early cleavage of embryos and blastocyst formation rate. The fertilization rates (EmbryoScope^®: 74.1%, 298/402; conventional incubator: 77.4%, 202/261) and cryopreservation rates (EmbryoScope^®: 57.4%, 163/284; conventional incubator: 48.7%, 91/187) of embryos were not affected by the incubator. No significant difference was observed in clinical pregnancy rates between the groups: (EmbryoScope^®: 24.5%, 24/98; conventional incubator: 15.0%, 15/100). The first division time of embryos was significantly faster in the “2-cell” groups (Groups A and B) (P < 0.01) than in the “none 2-cell” group (Group C). The blastocyst formation rates of embryos in Groups A and B were significantly (P < 0.05) higher than those in Group C. These results indicated that EmbryoScope^® enables a detailed and dynamic analysis of the development of human embryos.

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Successful development of preimplantation embryos is essential for reproduction. Growth factors secreted by reproductive tracts are important for the development of preimplantation embryos. FMS-like tyrosine kinase 3 (FLT3) is a tyrosine kinase receptor related to colony-stimulating factor-1 receptor and c-KIT, promoting preimplantation embryo development following their ligand binding. We found expression of FLT3 ligand transcripts in the oviducts and uteri of pregnant mice. The transcripts for its receptor, FLT3, were detectable throughout the early embryonic stages with an increase after the eight-cell stage. In contrast, the expression of FLT3 ligand was negligible after the morula stage, suggesting potential paracrine actions of FLT3 ligand produced by the reproductive tracts. Treatment with FLT3 ligand promoted the development of two-cell embryos to the morula and blastocyst stages in a dose-dependent manner with increases in cell proliferation, but not inhibition of cell survival. The effects of FLT3 ligand were blocked by a FLT3 receptor inhibitor, TCS359. Studies using specific inhibitors demonstrated the potential involvement of the PI3K pathway in mediating FLT3 ligand actions. Our findings suggest that the FLT3 ligand/FLT3 signaling system plays important paracrine roles during the development of preimplantation embryos.

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Purpose: Pelvic adhesions inhibit follicular growth and ovum transport, causing female infertility. In this report, we describe successful ovum pick-up (OPU) during adhesiotomy in a woman with severe adhesions that prevented transvaginal OPU. Subsequently, in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) were attempted after transport of cumulus–oocyte complexes (COCs). Materials and methods: Clinical examination revealed a chocolate cyst on the patient’s left ovary. Her right ovary was adherent to the posterior part of the uterus. COCs were harvested during adhesiotomy and stored at 37°C in a 1.5-ml tube containing Sperm Washing Medium. The tube was transported to our clinic using hot gel bags and a portable infant incubator. Immediately upon arrival, IVF and ICSI were performed. On day 7, two ICSI-derived blastocysts were cryopreserved. Two months later, these blastocysts were transferred to the patient’s uterus. Results: The patient conceived, and a normal, healthy boy was born. Conclusions: In summary, we performed OPU during adhesiotomy, and COCs were transported to the infertility clinic on the same day; subsequently, the patient was successfully impregnated using the transported oocytes.

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