This work was supported by a grant from the Korean Health Technology R&D Project, Ministry of Health & Welfare, Republic of Korea (A111539). None of the authors has any conflicts of interest to declare.

Gonadotropins including follicle-stimulating hormone (FSH) and luteinizing hormone (LH) play a crucial role in human-assisted reproduction techniques. Despite wide use of recombinant gonadotropins in clinical practice, the efficacy of urinary gonadotropins and the dosage of LH component have not yet been elucidated. This study was designed to investigate the difference of follicle culture outcomes according to various compositions of gonadotropins during in vitro culture of mouse preantral follicles. Ovaries were obtained from the 14-day-old C57BL/6 mice, and preantral follicles were isolated and cultured in culture media supplemented with human menopausal gonadotropin (hMG) 200 mIU/mL (group 1), recombinant FSH and LH (rFSH þ rLH) 200 mIU/mL each (group 2), rFSH 200 mIU/mL þ rLH 100 mIU/mL (group 3), or rFSH 200 mIU/mL þ rLH 20 mIU/mL (group 4). Follicle survival rate was significantly lower in group 4. Antral follicles in lower doses of LH (groups 3, 4) showed a statistically significant larger diameter and tended to have a higher antral formation rate. However, follicles in group 1 tended to have a higher oocyte maturation rate. Estradiol concentration from conditioned media from 2:1 FSH/LH (group 3) was significantly higher than those from 1:1 FSH/LH (group 2) or 10:1 FSH/LH (group 4). Half dose of rLH to rFSH facilitated upregulation of growth differentiation factor 9 (Gdf9) expression in granulosa cells when compared to 1:1 FSH/LH or 10:1 FSH/LH. Conclusively, recombinant gonadotropins provided a comparable condition to hMG, and half dose of rLH to rFSH seems to be more suitable for follicular development during in vitro culture.

The administration of kainic acid (KA) causes seizures and produces neurodegeneration in hippocampal CA3 pyramidal cells. The present study investigated a possible role of acupuncture in reducing hippocampal cell death and inflammatory events, using a mouse model of kainic acid-induced epilepsy. Male C57BL/6 mice received acupuncture treatments at acupoint HT8 or in the tail area bilaterally once a day for 2 days and again immediately after an intraperitoneal injection of KA (30 mg/kg). HT8 is located on the palmar surface of the forelimbs, between the fourth and fifth metacarpal bones. Twenty-four hours after the KA injection, neuronal cell survival, the activations of microglia and astrocytes, and mRNA expression of two proinflammatory cytokines, interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α), were measured in the hippocampus. Acupuncture stimulation at HT8, but not in the tail area, significantly reduced the KA-induced seizure, neuron death, microglial and astrocyte activations, and IL-1β mRNA expression in the hippocampus. The acupuncture stimulation also decreased the mRNA expression of TNF-α, but it was not significant. These results indicate that acupuncture at HT8 can inhibit hippocampal cell death and suppress KA-induced inflammatory events, suggesting a possible role for acupuncture in the treatment of epilepsy.

BACKGROUND: Thin or damaged endometrium causes uterine factor-derived infertility resulting in a failure of embryonic implantation. Regeneration of endometrium is a major issue in gynecology and reproductive medicine. Various types of cells and scaffolds were studied to establish an effective therapeutic strategy. For this type of investigations, production of optimal animal models is indispensable. In this study, we tried to establish various murine uterine damage models and compared their features. METHODS: Three to ten-week-old C57BL/6 female mice were anesthetized using isoflurane. Chemical and mechanical methods using ethanol (EtOH) at 70 or 100% and copper scraper were compared to determine the most efficient condition. Damage of uterine tissue was induced either by vaginal or dorsal surgical approach. After 7-10 days, gross and microscopic morphology, safety and efficiency were compared among the groups. RESULTS: Both chemical and mechanical methods resulted in thinner endometrium and reduced number of glands. Gross morphology assessment revealed that the damaged regions of uteri showed various shapes including shrinkage or cystic dilatation of uterine horns. The duration of anesthesia significantly affected recovery after procedure. Uterine damage was most effectively induced by dorsal approach using 100% EtOH treatment compared to mechanical methods. CONCLUSION: Taken together, murine uterine damage models were most successfully established by chemical treatment. This production protocols could be applied further to larger animals such as non-human primate.

Human embryonic stem cells are derived from the inner cell mass of preimplantation embryo at the blastocyst stage and their differentiation occurs through an intermediate step involving the formation of embryoid bodies (EBs), which are aggregates of embryonic stem cells. The EBs seem to be a powerful tool for investigating the development of embryos, as they can mimic the initial stages of embryonic development. In this study, we aimed to investigate the effect of estrogen compounds on the proliferation and differentiation of short-term and long-term cultured EBs in vitro. For this study, 10-day-old (short-term cultured) and 30-day-old (long-term cultured) EBs were subjected to estradiol (E2), estriol (E3), selective estrogen receptor modulator (raloxifene [RLX]), bisphenol A, and 1,3,5-tris(4-hydroxyphenyl)-4-propyl-1H-pyrazole for 7 days. To confirm the effects of estrogen treatment, ICI-182780 was added to the respective EBs for additional 7 days following estrogen treatment. Quantitative reverse transcription-polymerase chain reaction was performed to analyze the relative expression of differentiation marker genes representing the 3 germ layers. The expression of 7 marker genes, which included α-fetoprotein, hepatocyte nuclear factor (HNF)-3β, HNF-4α (endoderm), brachyury, cardiac actin ([cACT]; mesoderm), nestin (ectoderm), and Oct-4 (undifferentiated), was measured. Significantly, lower expression of HNF-4α in both short-term and long-term cultured EBs was observed after treatment of estrogen compounds compared to control. The expression of HNF-3β in short-term cultured EBs has been positively affected by E2, E3, and RLX. Regarding cACT, higher expression was observed after treatment of E2 (10(-7) mol/L) and E3 (10(-9) mol/L) in short-term cultured EBs, but opposite effects were demonstrated in long-term cultured EBs. The lower expressions of HNF-4α by E2 and RLX were negated by ICI-182780 treatment, although these findings were not statistically significant in E3-treated group. These findings suggest that estrogen compounds have effects on endodermal and mesodermal differentiation of human EBs.