当院におけるART妊娠者でのNIPT受検率について

当院におけるART妊娠者でのNIPT受検率について


Effects of activation with a Ca ionophore and roscovitine on the development of human oocytes that failed to fertilize after ICSI.

Sequential treatment with an Ca ionophore and roscovitine activates oocytes that remain unfertilized after ICSI. In TESE-ICSI, the activation rate tended to be increased by the co-administration of roscovitine with a Ca ionophore. J. Med. Invest. 70 : 321-324, August, 2023.

The Journal of Medical Investigation : JMI


Approach to medical education using laparoscopic simulator in covid-19 pandemic

コロナ禍における腹腔鏡シミュレーターを用いた医学教育への取り組み


徳島大学病院における不妊治療の取り組み

徳島大学病院における不妊治療の取り組み


症例報告: 血栓症を発症した卵巣癌合併妊娠の2例

症例報告: 血栓症を発症した卵巣癌合併妊娠の2例

Modern Trends in Obstetrics & Gynecology


Ultrasonographic changes in quadriceps femoris thickness in women with normal pregnancy and women on bed rest for threatened preterm labor.

This study aimed to evaluate the changes in quadriceps femoris muscle thickness during the pregnancy and postpartum periods and to elucidate the effect of bed rest for threatened preterm labor on muscle thickness. In 26 women with normal pregnancy, quadriceps femoris thickness was measured at 11-13, 26, 30, and 35 weeks' gestation, and at 3-5 days and 1 month postpartum using ultrasonography. In 15 pregnant women treated with bed rest for threatened premature labor, quadriceps femoris thickness was measured at 30 and 35 weeks' gestation and postpartum. In women with normal pregnancy, quadriceps femoris thickness increased, peaking at 35 weeks' gestation, followed by a postpartum decrease. In women on bed rest, quadriceps femoris thickness showed no significant change during the pregnancy and postpartum periods, and the muscle was significantly thinner at 35 weeks' gestation than that in women with normal pregnancy. In conclusion, a significant increase in quadriceps femoris muscle thickness during normal pregnancy was found using ultrasonography. Meanwhile, in pregnant women on bed rest treatment, the quadriceps femoris was significantly thinner in the late third trimester than that in normal pregnant women. Prolonged bed rest can affect normal changes in the quadriceps femoris muscle thickness during the pregnancy and postpartum periods.


当院における精巣内精子採取術・顕微授精(testicular sperm extraction-intracytoplasmic sperm injection: TESE-ICSI)の治療成績と周産期予後の検討


Extreme leanness, lower skeletal muscle quality, and loss of muscle mass during treatment are predictors of poor prognosis in cervical cancer treated with concurrent chemoradiation therapy

Human papillomavirus vaccination is not widespread in Japan, and the low screening rates result in many cases of locally advanced cervical cancer. We investigated the prognostic significance of sarcopenia in patients with cervical cancer to guide healthcare policies to improve treatment outcomes. This retrospective study included 83 patients with cervical cancer without distant metastasis who underwent primary concurrent chemoradiotherapy between 2013 and 2018. We analyzed the indicators of physical condition and muscle quantity using the SYNAPSE VINCENT software. Muscle mass and the relationship between treatment toxicity and prognosis were evaluated. The patients' median age was 60 (range 33-80) years. Cancer stage distribution was as follows: cT2b or higher, 84.3%; N1, 65.1%; and MA, 27.7%. The overall sarcopenia (skeletal muscle index [SMI] < 38.5) rate was 30.1%, and the rate was 33.9 and 22.2% in patients aged < 64 and ≥ 65 years, respectively. No correlation was observed between clinical stage and musculoskeletal indices. Treatment resulted in decreased body weight and SMI; after treatment, the sarcopenia rate increased to 37.3%. A higher intramuscular adipose tissue content (IMAC) reduced the number of chemotherapy cycles needed. Treatment-associated SMI decreases of ≥ 7% indicated poor prognosis, with significant differences in progression-free survival and overall survival (p = 0.013 and p = 0.012, respectively). Patients who were very lean (body mass index < 18.5 kg/m) before treatment had a poor prognosis (p = 0.016 and p < 0.001). Our findings emphasize the importance of assessing original nutritional status and maintaining muscle mass and quality during the treatment of patients with cervical cancer.

International Journal of Clinical Oncology


A novel PCOS rat model and an evaluation of its reproductive, metabolic, and behavioral phenotypes.

Although animal models of PCOS have been used in many studies, none of them can reproduce both the reproductive and metabolic phenotypes of PCOS. In addition, behavioral parameters have not been evaluated in PCOS animal models. We tried to produce an improved rat model of PCOS, and the reproductive, metabolic, and behavioral phenotypes of the model rats were evaluated. Female rats were implanted with silicon tubes containing oil-dissolved dihydrotestosterone (Oil-DHT) as a new PCOS model. Their phenotypes were compared with those of conventional PCOS model rats (DHT), into which tubes containing crystalline DHT were implanted, and non-DHT-treated rats (control). Both the Oil-DHT and DHT rats showed greater body weight gain, food intake, and fat depot weight than the control rats. Furthermore, these groups showed fewer estrous stages and increased numbers of cystic follicles. The DHT rats exhibited lower ovarian and uterine weights than the control rats, whereas no such changes were observed in the Oil-DHT rats. The Oil-DHT and DHT rats showed less locomotor activity in the light phase than the control rats. Our proposed PCOS model reproduced both the reproductive and metabolic phenotypes of PCOS and may have potential for PCOS research.


Correlations of Androstenediol with Reproductive Hormones and Cortisol According to Stages during the Menopausal Transition in Japanese Women. J

Associations of androstenediol, which has both androgenic and estrogenic activities, with circulating reproductive hormones and stress hormone in women during the menopausal transition may be different depending on the menopausal stage. The aim of this study was to determine the changes in circulating androstenediol during the menopausal transition in Japanese women and the associations of androstenediol with estrogen, androgen and cortisol for each stage of the menopausal transition. We divided the 104 subjects into 6 stages by menstrual regularity and follicle-stimulating hormone level: mid reproductive stage, late reproductive stage, early menopausal transition, late menopausal transition, very early postmenopause and early postmenopause. Levels of dehydroepiandrosterone sulfate (DHEAS), estradiol, estrone, testosterone (T), free T, androstenedione and cortisol were measured. Serum androstenediol concentration was measured by using liquid chromatography mass spectrometry. There were no significant differences in androstenediol levels among the 6 stages. Levels of DHEA-S and testosterone showed significant and positive correlations with androstenediol in all stages. Estradiol levels showed negative correlations with androstenediol levels in the late menopausal transition and very early postmenopause (r=-0.452, p = 0.052 and r=-0.617, p = 0.006, respectively). Cortisol levels showed significant and positive correlations with androstenediol levels in the mid and late reproductive stages (r = 0.719, p = 0.003 and r = 0.808, p < 0.001, respectively).The associations of androstenediol with estradiol and cortisol were different depending on the stage of the menopausal transition. Androstenediol may play a compensatory role for estrogen deficiency from late menopausal transition to very early postmenopause.

The Journal of Steroid Biochemistry and Molecular Biology


Sphingosine 1-Phosphate (S1P) in the Peritoneal Fluid Skews M2 Macrophage and Contributes to the Development of Endometriosis.

Sphingosine 1-phosphate (S1P), an inflammatory mediator, is abundantly contained in red blood cells and platelets. We hypothesized that the S1P concentration in the peritoneal cavity would increase especially during the menstrual phase due to the reflux of menstrual blood, and investigated the S1P concentration in the human peritoneal fluid (PF) from 14 non-endometriosis and 19 endometriosis patients. Although the relatively small number of samples requires caution in interpreting the results, S1P concentration in the PF during the menstrual phase was predominantly increased compared to the non-menstrual phase, regardless of the presence or absence of endometriosis. During the non-menstrual phase, patients with endometriosis showed a significant increase in S1P concentration compared to controls. In vitro experiments using human intra-peritoneal macrophages (MΦ) showed that S1P stimulation biased them toward an M2MΦ-dominant condition and increased the expression of IL-6 and COX-2. An in vivo study showed that administration of S1P increased the size of the endometriotic-like lesion in a mouse model of endometriosis.


Effects of gonadal status and the estrogen milieu on hypothalamic oxytocin gene expression and serum oxytocin levels in female rats.

Oxytocin (OT) and its receptor (OTR) play various roles in the central and peripheral regulation of appetite and body weight. Previously, we have shown that the administration of OT markedly decreased appetite and body weight gain in ovariectomized (OVX) obese rats. In addition, recent studies have shown that the endogenous OT system is also affected by endogenous or exogenous estrogen. In this study, we showed that ovariectomy decreased rats' hypothalamic OT/OTR mRNA and serum OT levels, but did not affect their visceral fat OTR mRNA levels. The chronic administration of estradiol (E2) abrogated these ovariectomy-induced changes; i.e., it increased the rats' hypothalamic OT/OTR mRNA and serum OT levels, and may be associated with reductions in food intake and body weight gain. In addition, acute E2 administration increased the rats' hypothalamic OTR mRNA and serum OT levels, but did not affect their hypothalamic OT mRNA levels. Taken together, these results suggest that endogenous OT and/or OTR expression might be positively regulated by E2 and that the suppressive effects of E2 on appetite and body weight gain might be mediated, at least in part, by the OT system. Thus, we consider that OT might be a target hormone to pursue subsequent interventions of menopause for menopause-induced metabolic disorders.


Monochorionic Dizygotic Twins Conceived Spontaneously Showed Chimerism in Karyotype and Blood Group Type

We report a very rare case of monochorionic dizygotic twins conceived spontaneously. The fetuses were sex-discordant in ultrasonography despite being monochorionic twins. After birth, the girl and boy showed normal phenotypes but they showed blood chimerism in karyotype and blood group type.


The reduction in sexual behavior of adult female rats exposed to immune stress in the neonatal period is associated with reduced hypothalamic progesterone receptor expression.

We examined the mechanism by which neonatal immune stress reduces the sexual behavior of female rats in adulthood. Neonatal female rats were randomly divided into 3 groups: control (n = 11), postnatal day 10 lipopolysaccharide (PND10LPS) (n = 23), and PND25LPS (n = 11) groups, which received intraperitoneal injections of LPS (100 μg/kg) or saline on PND10 and 25. Daily inspections of the vaginal opening (VO) were performed from PND27 to PND37. Thereafter, the frequency of estrus was assessed for 15 days. Female rats (at 11-12 weeks of age) were placed in a cage with male rats, and their sexual behavior was monitored for 30 min. The hypothalamic mRNA expression levels of factors related to sexual behavior were examined via real-time PCR. VO occurred later and the frequency of estrus was lower in the PND10LPS group compared to the control group. The number of lordosis behaviors and the total number of mounts performed by male partners were lower in the PND10LPS and PND25LPS groups than in the control group. Acceptability: The lordosis quotient and lordosis rating were lower in the PND10LPS group than in the control group. Proceptive behavior: the number of ear wiggling events was lower in the PND10LPS group than in the other groups, and the number of hops/darts was lower in the PND10LPS group than in the control group. The hypothalamic mRNA expression level of progesterone receptors (PR)A + B was lower in the PND10LPS group than in the control group, and the hypothalamic PRB mRNA expression level was lower in the PND10LPS and PND25LPS groups than in the control group. Neonatal immune stress impeded sexual behavior and hypothalamic PR mRNA expression in female rats. Decreased progesterone activity in the hypothalamus might explain the reduction in sexual behavior seen in these rats.


The influence of psychological stress in early life on sexual maturation and sexual behavior in male and female rats.

We studied the influence of psychological stress during the early neonatal period on sexual maturation and sexual behavior in rats. Neonatal male and female rats were divided into control (C) and maternal separation (MS) groups (n = 20-24 per group). The pups in the MS groups were placed in isolation cages for 240 minutes/d from postnatal days 2-11. Vaginal opening (VO) in females and preputial separation (PS) in males (indicators of sexual maturation) were monitored, as was the estrous cycle in females. Thereafter, sexual behavior was monitored twice at 13 and 15 weeks of age. As for sexual maturation, the onset of PS occurred significantly earlier in the MS group than in the C group, whereas the onset of VO did not differ between the groups. The length of the estrous cycle did not differ between the groups. The frequencies of sexual behaviors did not differ between the groups in either sex. In conclusion, early-life psychological stress induced by MS advanced sexual maturation in male rats, whereas it did not affect sexual maturation in female rats. On the other hand, early-life psychological stress might not affect sexual behavior in adulthood in either sex.


Oxytocin treatment reduced food intake and body fat and ameliorated obesity in ovariectomized female rats.

Recent studies have shown that oxytocin reduces food intake and body weight gain and promotes lipolysis in some species, including humans. Interestingly, these effects of oxytocin are more marked in obese individuals. Although the menopausal loss of ovarian function induces increased visceral adiposity and some metabolic disorders, no safe medical interventions for these conditions have been established. In this study, we evaluated the effects of oxytocin on appetite, body weight, and fat mass in ovariectomized rats. Six-day oxytocin treatment attenuated cumulative food intake and body weight gain, and reduced visceral and subcutaneous fat weight and adipocyte cell area in ovariectomized rats. Blood examinations indicated that 6-day oxytocin treatment did not alter renal or hepatic functions. Instead, it might prevent ovariectomy-induced liver damage. In addition, acute oxytocin treatment did not affect body temperature or locomotor activity. These results indicate that oxytocin might be useful for treating or preventing menopause-induced metabolic disorders, without causing any adverse effects.


Activin effects on follicular growth in in vitro preantral follicle culture.

As the follicular environment transits from being activin dominant to inhibin dominant during folliculogenesis, it is assumed that activin plays an important role in the early stage of follicular growth. We examined the effects of activin on morphological, biochemical and molecular changes in isolated preantral follicles. Preantral follicles were mechanically isolated from 14-day old female C57BL/6 mice. Each follicle was cultured and observed for 14 days usingan in vitro follicle culture system containing FSH, FSH + activin A and FSH + inhibin in the culture medium. We subsequently examined FSH receptor (FSH-R) mRNA expression in isolated follicle cultures with or without activin on days 0 and 2. Activin was observed to significantly stimulate follicle enlargement on days 2, 4, 6 and 8, accelerate morphological changes and increase estradiollevels in culture medium on days 4, 12 and 14. In contrast, inhibin did not alter follicular growth. Additionally, activin stimulated the expression of FSH-R mRNA in isolated granulosa cells. It was demonstrated that activin stimulated the growth of preantral follicles, mainly during the early stage of folliculogenesis, by inducing FSH-R expression, in an isolated follicle culture system. J. Med. Invest. 66 : 165-171, February, 2019.

The Journal of Medical Investigation : JMI


Lipopolysaccharide promotes early endometrial-peritoneal interactions in a mouse model of endometriosis.

The aims of this study were to clarify the effects of lipopolysaccharide (LPS) on the early development of endometriosis and on the production of cytokines and chemokines in the murine peritoneal cavity. Endometriotic lesions were induced in C57BL/6J adult female mice by intraperitoneal injection of endometrial fragments plus blood or endometrial fragments plus blood with LPS. On day 7, endometriotic lesions were assessed by gross and microscopic evaluations. Time-dependent changes in the secretion of TNF-α,IL-6,and CXCL2/MIP-2 in peritoneal lavage fluid after the intraperitoneal injection of LPS (50 µg/body) were measured by their respective enzyme-linked immunosorbent assays. The areas of endometriotic lesions in the LPS group (10.8 8.6 mm) were significantly larger than those in the control group (3.1 3.7 mm).The levels of TNF-α and IL-6 peaked within 2 hours and the level of MIP-2 reached a maximum on day 1 after the injection of LPS. LPS promotes development of the early stages of murine endometriotic lesions. J. Med. Invest. 66 : 70-74, February, 2019.


Current status of chromosomal analysis in couples with recurrent miscarriage at Tokushima University Hospital

徳島大学病院産科婦人科における不育症患者に対する染色体検査の実施状況

Modern Trends in Obstetrics & Gynecology


Prenatal undernutrition suppresses sexual behavior in female rats.

Infectious, psychological and metabolic stresses in the prenatal and early neonatal period induce long-lasting effects in physiological function and increase the risk of metabolic disorders later in life. We examined the sexual behavior of female rats that were subjected to undernutrition in the prenatal period. Eight pregnant rats were divided into two groups: a maternal normal nutrition group (mNN; n = 4) and a maternal undernutrition group (mUN; n = 4), which received 50% of the daily food intake amount of the mNN group from gestation day 13 to delivery. Nine and seven female offspring were randomly selected from the mNN and mUN groups, respectively. Vaginal opening (VO), estrous cycle length, sexual behavior and mRNA expression levels of the factors that regulate sexual behavior were observed. In the mUN group, VO day was later, the estrous cycle was longer, and the lordosis quotient and lordosis rating were lower than in the mNN group; such differences were not seen in other sexual performances, such as ear wiggles, darts, kick bouts and box. The hypothalamic mRNA expression level of progesterone receptor (PR) A + B and oxytocin (OT) were significantly lower in the mUN group than in the mNN group. These findings indicated that prenatal undernutrition disrupted puberty onset, the estrous cycle, sexual behavior and hypothalamic mRNA expression of PR and OT in female rat pups.


11-oxygenated C19 steroids as circulating androgens in women with polycystic ovary syndrome.

11-oxygenated C19 steroids (11oxC19s) are newly specified human androgens. Although median serum levels of 11oxC19 were reported to be higher in patients with polycystic ovary syndrome (PCOS) than in unaffected women, inter-individual variations in androgen levels among PCOS patients have poorly been investigated. Here, we quantified four 11oxC19s, i.e., 11-ketotestosterone (11KT), 11β-hydroxytestosterone (11OHT), 11β-hydroxyandrostenedione (11OHΔ4A), and 11-ketoandrostenedione (11KΔ4A), in blood samples of 28 PCOS patients and 31 eumenorrheic women using liquid chromatography-tandem mass spectrometry. We referred to our previous data of classic androgens in these individuals. We found that 11OHT levels were higher in the PCOS group than in the eumenorrheic group. Moreover, although the median values of 11KT, 11KΔ4A, and 11OHΔ4A were comparable between the two groups, these steroids were markedly increased in some patients. Of the 28 patients, 8 had high levels of both 11oxC19s and classic androgens, whereas 4 had an increase only in 11oxC19 levels, and 12 had an increase only in classic androgen levels. Intragroup variations in androgen levels were relatively large in the PCOS group. Levels of 11OHT and 11KT were significantly higher in overweight/obese patients than in normal weight patients and correlated with body mass indexes. These results highlight the clinical significance of 11oxC19s as circulating androgens in PCOS patients and indicate that the accumulation of 11oxC19s and/or classic androgens is an essential feature of PCOS. The profiles of circulating androgens appear to vary among patients. In particular, overweight/obesity likely enhances the 11oxC19s accumulation in PCOS, although this notion awaits further validation.


The effects of chronic testosterone administration on hypothalamic gonadotropin-releasing hormone regulatory factors (Kiss1, NKB, pDyn and RFRP) and their receptors in female rats.

The effects of androgens on gonadotropin-releasing hormone (GnRH) secretion in females have not been fully established. To clarify the direct effects of androgens on hypothalamic reproductive factors, we evaluated the effects of chronic testosterone administration on hypothalamic GnRH regulatory factors in ovariectomized (OVX) female rats. Both testosterone and estradiol reduced the serum luteinizing hormone levels of OVX female rats, indicating that, as has been found for estrogen, testosterone suppresses GnRH secretion via negative feedback. Similarly, the administration of testosterone or estradiol suppressed the hypothalamic mRNA levels of kisspeptin and neurokinin B, both of which are positive regulators of GnRH, whereas it did not affect the hypothalamic mRNA levels of the kisspeptin receptor or neurokinin-3 receptor. On the contrary, the administration of testosterone, but not estradiol, suppressed the hypothalamic mRNA expression of prodynorphin, which is a negative regulator of GnRH. The administration of testosterone did not alter the rats' serum estradiol levels, indicating that testosterone's effects on hypothalamic factors might be induced by its androgenic activity. These findings suggest that as well as estrogen, androgens have negative feedback effects on GnRH in females and that the underlying mechanisms responsible for these effects are similar, but do not completely correspond, to the mechanisms underlying the effects of estrogen on GnRH.


Neurokinin B receptor agonist and Dynorphin receptor antagonist stimulated luteinizing hormone secretion in fasted male rodents.

Kisspeptin/neurokinin B (NKB)/dynorphin (Dyn) (KNDy) neuron in hypothalamic arcuate nucleus plays a key role in GnRH/LH pulsatile secretion. We aimed to determine whether stimulation of NKB/neurokinin 3 receptor (NK3R) signaling and inhibition of Dyn/kappa-opioid receptor (KOR) signaling recover LH secretion that is suppressed by acute fasting in male rats. Furthermore, we determined dose dependent effect of NKB/NK3R signaling on serum LH level under acute fasting condition in male mice. Mature male rats were injected saline (0.1 mL) and senktide (20 μg/kg), a NK3R agonist, or nor-BNI (800 μg/kg), a KOR antagonist intraperitoneally (ip) after 72 h fasting. And mature male mice were injected multiple doses of senktide, ip after 48 h fasting. Blood and brain sample were collected 90 min after injections for LH measurement and hypothalamic mRNA expressions. All three studies showed significantly lower LH concentration in fasted groups than non-fasted groups. Senktide did not recover LH suppressed by acute fasting in male rats, whereas nor-BNI injected male rats showed significantly higher LH than 72 h fasted male rats (p < 0.05). Mice study showed significantly higher LH concentration in higher doses senktide groups than 48 h fasted group and one of lower doses senktide group. These results suggest that stimulation of NKB/NK3R signaling and attenuation of Dyn/KOR signaling could recover suppressed LH secretion under acute fasting condition in male rodents.


Effects of dihydrotestosterone administration on the expression of reproductive and body weight regulatory factors in ovariectomized and estradiol-treated female rats.

To clarify the direct effects of androgens, the changes in the hypothalamic levels of reproductive and appetite regulatory factors induced by chronic dihydrotestosterone (DHT) administration were evaluated in female rats. DHT treatment increased the BW and food intake of the ovariectomized rats, but not the estradiol (E2)-treated rats. DHT administration suppressed the expression of a hypothalamic anorexigenic factor. Although the kisspeptin (Kiss1) mRNA levels of the anterior hypothalamic block (the anteroventral periventricular nucleus, AVPV) were increased in the E2-treated rats, DHT administration did not affect the Kiss1 mRNA levels of the AVPV in the ovariectomized or E2-treated rats. Conversely, DHT administration reduced the Kiss1 mRNA levels of the posterior hypothalamic block (the arcuate nucleus, ARC) in the ovariectomized rats. Although the Kiss1 mRNA levels of the posterior hypothalamic block (ARC) were decreased in the E2-treated rats, DHT administration did not affect the Kiss1 mRNA levels of the ARC in these rats. Serum luteinizing hormone levels of these groups exhibited similar patterns to the Kiss1 mRNA levels of the ARC. These results showed that DHT affects the production of hypothalamic reproductive and appetite regulatory factors, and that these effects of DHT differ according to the estrogen milieu.


The roles of kisspeptin and gonadotropin inhibitory hormone in stress-induced reproductive disorders.

Several kinds of stress suppress the hypothalamic-pituitary-gonadal (HPG) axis and reproductive behavior in humans and animals. These changes can eventually cause diseases and disorders, such as amenorrhea and infertility. In previous studies, it has been shown that stress-related factors, e.g., corticotropin-releasing hormone, cortisol, and pro-inflammatory cytokines, promote the stress-induced suppression of the HPG axis. However, these mechanisms are not sufficient to explain how stress suppresses HPG axis activity, and it has been suggested that some other factors might also be involved. In the early 21st century, novel neuroendocrine peptides, kisspeptin and gonadotropin inhibitory hormone (GnIH)/RFamide-related peptide 3 (RFRP-3), which directly regulate GnRH/gonadotropin synthesis and secretion, were newly discovered. Growing evidence indicates that kisspeptin and GnIH/RFRP-3 play pivotal roles in the stress-induced disruption of the HPG axis and reproductive behavior in addition to their physiological functions. This review summarizes what is currently known about the roles of kisspeptin and GnIH/RFRP-3 in stress-induced reproductive disorders.


The effects of ovariectomy and lifelong high-fat diet consumption on body weight, appetite, and lifespan in female rats.

In females, ovarian hormones play pivotal roles in metabolic, appetite, and body weight regulation. In addition, it has been reported that ovarian hormones also affect longevity in some species. Recently, it was found that the consumption of a high-fat diet aggravates ovariectomy-associated metabolic dysregulation in female rodents. The aim of this study was to investigate the hypothesis that long-term high-fat diet consumption and ovariectomy interact to worsen body weight regulation and longevity in female rats. At 21days of age, female rats were weaned and randomly divided into two groups, one of which was given the high-fat diet, and the other was supplied with standard chow. At 23weeks of age, each group was further divided into ovariectomized and sham-operated groups, and then their body weight changes, food intake, and longevity were measured until 34months of age. The sham - high-fat diet rats exhibited greater body weight changes and higher feed efficiency than the sham - standard chow rats. On the other hand, the ovariectomized - high-fat diet and ovariectomized - standard chow rats displayed similar body weight changes and feed efficiency. The sham - high-fat diet and ovariectomized - standard chow rats demonstrated similar body weight changes and feed efficiency, indicating that the impact of ovariectomy on the regulation of body weight and energy metabolism might be similar to that of high-fat diet. Contrary to our expectations, ovariectomy and high-fat diet consumption both had small favorable effects on longevity. As the high-fat diet used in the present study not only had a high fat content, but also had a high caloric content and a low carbohydrate content compared with the standard chow, it is possible that the effects of the high-fat diet on body weight and longevity were partially induced by its caloric/carbohydrate contents. These findings indicate that the alterations in body weight and energy metabolism induced by ovariectomy and high-fat diet might not directly affect the lifespan of female rats.


Prenatal undernutrition disrupted the sexual maturation, but not the sexual behavior, in male rats.

Exposure to various stressors, including psychological, metabolic, and immune, in the perinatal period induces long-lasting effects in physiological function and increase the risk of metabolic disorders in later life. In the present study, sexual maturation and sexual behavior were assessed in prenatally undernourished mature male rats. All the pregnant rats were divided into the maternal normal nutrition (mNN) group and the maternal undernutrition (mUN) group. The mUN mothers received 50% of the amount of the daily food intake of the mNN mothers. Preputial separation and sexual behavior were observed in randomly selected pups of the mNN and mUN groups. The body weight of the mothers was significantly lighter in the mUN group than in the mNN group. Similarly, the pups in the mUN group showed a significantly lower body weight than those in the mNN group from postnatal day (PND) 1 to PND 15. The preputial separation day was significantly delayed in the mUN group, compared to the mNN group. Sexual behavior did not show any significant difference between the two groups. These findings indicated that prenatal undernutrition delayed sexual maturation, but did not suppress sexual behavior, in mature male rats.


Intra-follicular kisspeptin levels are related to oocyte maturation and gonadal hormones in patients who are undergoing assisted reproductive technology.

To assess the kisspeptin concentrations in follicular fluid and their relationship with clinical outcomes during assisted reproductive technology. Thirty-nine patients who were aged 24-40 years and underwent oocyte retrieval for in vitro fertilization/intracytoplasmic sperm injection participated in this study In 65 follicular fluid samples that had been obtained from 30 patients and their blood samples, the kisspeptin levels were measured in order to investigate the correlations with their gonadal hormone levels. Venous blood samples were collected from 14 patients to investigate their plasma kisspeptin levels across different phases of assisted reproductive technology. The follicular fluid kisspeptin level was significantly higher than that of the plasma level and was positively associated with the follicular fluid estradiol concentration and with the serum estradiol and number of mature oocytes. In the plasma, the maximum concentration of kisspeptin was observed on the day of ovum pick-up and on the day of embryo transfer during ovarian stimulation for assisted reproductive technology. Kisspeptin was present in the follicular fluid and the plasma kisspeptin concentration was affected by ovarian stimulation. Kisspeptin appears to affect oocyte maturation and ovulation.


The effects of chronic testosterone administration on body weight, food intake, and fat weight were age-dependent.

Previously, we showed that chronic testosterone administration increased body weight (BW) and food intake (FI), but did not alter fat weight, in young female rats. To examine our hypothesis that the effects of androgens on BW, FI and body composition might be age-dependent, the effects of chronic testosterone administration were evaluated in rats of different ages; i.e., young and middle-aged rats. Although chronic testosterone administration increased BW gain, FI, and feed efficiency in both young and middle-aged rats, it increased visceral fat weight in middle-aged rats, but not in young rats. Therefore, it is possible that testosterone promotes the conversion of energy to adipose tissue and exacerbates fat accumulation in older individuals. In addition, although the administration of testosterone increased the serum leptin level, it did not alter hypothalamic neuropeptide Y mRNA expression in middle-aged rats. On the contrary, the administration of testosterone did not affect the serum leptin levels of young rats. Thus, testosterone might induce hypothalamic leptin resistance, which could lead to fat accumulation in older individuals. Testosterone might disrupt the mechanisms that protect against adiposity and hyperphagia and represent a risk factor for excessive body weight and obesity, especially in older females.


Developmental changes in hypothalamic SF-1, POMC, and ERα mRNA expression and their sensitivity to fasting in male and female rats.

Estrogen plays pivotal roles in body weight regulation through its effects on central estrogen receptor-α (ERα) expression. ERα is found on neurons that express the hypothalamic anorexigenic factors steroidogenic factor-1 (SF-1) and pro-opiomelanocortin (POMC) mediate these effects of estrogen. As the gonadal hormonal milieu is drastically altered during the developmental period, the expression levels of SF-1 and POMC might also change during this period. In this study, we showed that hypothalamic SF-1 and ERα mRNA expression did not change during the neonatal to pre-pubertal period (from postnatal day 10 to 30), and there were no significant differences in the hypothalamic mRNA expression levels of these molecules between males and females at any examined age. On the other hand, hypothalamic POMC mRNA expression and the serum estradiol (E2) level increased during development in both males and females. Significant positive correlations were detected between the serum E2 level and hypothalamic POMC mRNA expression in both males and females. Hypothalamic ERα and POMC mRNA expression were decreased by fasting in male rats at all examined ages, whereas fasting had no effect on hypothalamic ERα or POMC mRNA expression in the female rats. These results indicate that the regulatory system involving E2 and hypothalamic POMC expression might already be established in the neonatal period and that the roles of POMC and ERα in body weight regulation during development might differ slightly between males and females.


Development-related changes in the expression of the ovarian Kiss1 and Kiss1r genes and their sensitivity to human chorionic gonadotropin in prepubertal female rats.

Kisspeptin, which is encoded by the Kiss1 gene, and its receptor, the G protein-coupled receptor 54 (Kiss1r), play important roles in the regulation of reproductive functions in mammals. Several studies have shown that the Kiss1 and Kiss1r genes are expressed in the rat, primate, and human ovaries, and that the ovarian kisspeptin system plays a pivotal role in ovulation at the proestrous stage in adulthood. The purpose of this study was to evaluate development-related changes in the expression of ovarian Kiss1 and Kiss1r genes and in kisspeptin levels, and to identify the regulatory factors for these genes during the prepubertal period. The serum kisspeptin level was also measured to examine whether ovarian kisspeptin affects serum kisspeptin levels. Variations in the ovarian Kiss1 and Kiss1r mRNA levels were observed during the prepubertal period in female rats, with levels peaking around postnatal days 20 and 15, respectively. Nevertheless, the ovarian kisspeptin content per total protein level was stably maintained. Serum kisspeptin levels at postnatal days 30 and 35 were higher than those at earlier postnatal days. The pattern of the ovarian Kiss1 mRNA levels was similar to that of the serum luteinizing hormone (LH) levels, and the ovarian Kiss1 mRNA level increased after injection with human chorionic gonadotropin (HCG) on postnatal day 20, but not on postnatal days 10 and 30. These data indicate that ovarian Kiss1 and Kiss1r mRNA levels are increased on postnatal days 20 and 15, respectively, and that changes in the serum LH level and the ovarian sensitivity to LH may be involved in the alteration of ovarian Kiss1 mRNA levels.

The Journal of Reproduction and Development


The effects of chronic testosterone administration on body weight, food intake, and adipose tissue are changed by estrogen treatment in female rats.

In females, estrogens play pivotal roles in preventing excess body weight (BW) gain. On the other hand, the roles of androgens in female BW, appetite, and energy metabolism have not been fully examined. We hypothesized that androgens' effects on food intake (FI) and BW regulation change according to the estrogens' levels. To evaluate this hypothesis, the effects of chronic testosterone administration in ovariectomized (OVX) female rats with or without estradiol supplementation were examined in this study. Chronic testosterone administration decreased BW, FI, white adipose tissue (WAT) weight, and adipocyte size in OVX rats, whereas it increased BW, WAT weight, and adipocyte size in OVX with estradiol-administered rats. In addition, chronic testosterone administration increased hypothalamic CYP19a1 mRNA levels in OVX rats, whereas it did not alter CYP19a1 mRNA levels in OVX with estradiol-administered rats, indicating that conversion of testosterone to estrogens in the hypothalamus may be activated in testosterone-administered OVX rats. Furthermore, chronic testosterone administration decreased hypothalamic TNF-α mRNA levels in OVX rats, whereas it increased hypothalamic IL-1β mRNA levels in OVX with estradiol-administered rats. On the other hand, IL-1β and TNF-α mRNA levels in visceral and subcutaneous WAT and liver were not changed by chronic testosterone administration in both groups. These data indicate that the effects of chronic testosterone administration on BW, FI, WAT weight, and adipocyte size were changed by estradiol treatment in female rats. Testosterone has facilitative effects on BW gain, FI, and adiposity under the estradiol-supplemented condition, whereas it has inhibitory effects in the non-supplemented condition. Differences in the responses of hypothalamic factors, such as aromatase and inflammatory cytokines, to testosterone might underlie these opposite effects.


Assisted reproductive technology in Japan: a summary report of 1992-2014 by the Ethics Committee, Japan Society of Obstetrics and Gynecology.

The Japan Society of Obstetrics and Gynecology implemented a registry report system for the clinical practice of assisted reproductive technology in 1986. The aggregated results from 1992 to 2014 are reported herein. The total number of registered treatments was 393 745 cycles, of which 66 550 were pregnancy cycles and 46 008 were cycles with a live birth. Compared to the number of registered treatments in 2008, when the cycle-based registry was newly introduced, there was a 2.07-fold increase in the total number of treatments and a 2.25-fold increase in the number of cycles with a live birth. As the average age of patients who receive assisted reproductive technology has become markedly higher year by year, the most common age of those patients who received assisted reproductive technology in 2014 was 40 years. The total numbers of both assisted reproductive technology treatments and assisted reproductive technology live births are likely to be higher in the future. In addition, the trend toward aging patients seems to be continuing into the future.


Gonadotropin-Inhibitory Hormone Plays Roles in Stress-Induced Reproductive Dysfunction.

Physical and psychological stressors suppress hypothalamic-pituitary-gonadal axis activity and sexual behavior and consequently induce reproductive dysfunction. Recently, it has been shown that gonadotropin-inhibitory hormone (GnIH), also called RFamide-related peptide 3 (RFRP) in mammals, which is a potent inhibitory regulator of gonadotropin-releasing hormone (GnRH) and gonadotropin, is involved in stress-induced reproductive dysfunction. GnIH/Rfrp (the gene coding RFRP-3) expression and activity are increased by psychological and immune stress, and this alteration suppresses GnRH and gonadotropin secretion. Glucocorticoid acts as a mediator that interacts between stress and hypothalamic GnIH/RFRP-3. GnIH/RFRP-3 also plays important roles in stress-induced suppression of sexual behavior and infertility, and genetic silencing of GnIH/Rfrp completely recovers sexual behavior and fertility. This review summarizes what is currently known about the roles of GnIH in stress-induced reproductive dysfunction.


Clinical outcome of various metformin treatments for women with polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is an ovulatory disorder and insulin resistance and diabetes are involved in its pathophysiology. Metformin, an anti-diabetic agent, has been reported to be useful to induce ovulation. Metformin treatment was classified into four types: (1) clomiphene-metformin combination treatment for clomiphene-resistant patients; (2) clomiphene-metformin combination for clomiphene-sensitive patients; (3) clomiphene-metformin combination for naïve patients; and (4) metformin monotherapy. The patients underwent physical, endocrinological, and clinical examinations for their ovulation rates, pregnancy rates, and follicular development. The ovulation rates, pregnancy rates, and single follicular development were not significantly different among the clomiphene-metformin combination treatment groups. In the Body Mass Index (BMI) subanalysis, the pregnancy rate was higher in the BMI≥30 kg/m group than in the other three groups with a BMI of ≤30 kg/m in both cycles and cases. The ovulation rates and pregnancy rates were significantly higher in the group with a fasting insulin of ≥15 μU/mL than in the groups with a fasting insulin of <15 μU/mL in both cycles and cases. Clomiphene-metformin combination treatment appears to be useful, at least for clomiphene-resistant patients, and a BMI of >30 kg/m and a fasting insulin of ≥15 μU/mL appear to be predictors of a good result with this treatment.


Relationship between serum anti-Mullerian hormone and clinical parameters in polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is an ovulatory disorder that affects 6-10% of women of reproductive age. Serum AMH level may be an additional factor, or surrogate of PCOM, in the diagnostic criteria of PCOS. We evaluated the correlations between the serum AMH level and various endocrine and metabolic features in PCOS using the latest fully automated assay. Serum AMH level was compared between 114 PCOS patient (PCOS group) and 95 normal menstrual cycle women (Control group). Correlations between serum AMH level and various endocrine and metabolic factors were analysed in PCOS group. The serum AMH level was significantly higher in the PCOS group (8.35±8.19 ng/mL) than in the Control group (4.99±3.23 ng/mL). The serum AMH level was independently affected by age and the presence of PCOS on multiple regression analysis. Ovarian volume per ovary (OPVO) showed the strongest positive correlation (r=0.62) with the serum AMH level among related factors. On receiver operating characteristic (ROC) curve analysis, the cut-off value of AMH for the diagnosis of PCOS was 7.33 ng/mL, but this value did not have high efficacy (sensitivity 44.7%, specificity 76.8%). A cut-off value of 10 ng/mL had a high specificity of 92.6%, although the sensitivity was low (24.6%). The serum AMH level was elevated and reflected ovarian size in PCOS patients. The serum AMH level could be a surrogate for ultrasound findings of the ovaries in PCOS and might be useful for estimating ovarian findings without transvaginal ultrasound in the diagnosis of PCOS.


The effects of prenatal undernutrition and a high-fat postnatal diet on central and peripheral orexigenic and anorexigenic factors in female rats.

Prenatal undernutrition and postnatal overnutrition increase the risk of some peripheral and central metabolic disorders in adulthood. We speculated that disturbances of appetite/metabolic regulatory factors might already have been established in the early stages of life and contribute to obesity later in life. The effects of a high-fat diet on the levels of peripheral and central appetite/metabolic regulatory factors were compared between the offspring of normally nourished dams and those of undernourished dams in the peri-pubertal period. In the offspring of the normally nourished dams (control), the consumption of the high-fat diet resulted in lower hypothalamic mRNA levels of orexigenic factors (neuropeptide Y (NPY) and prepro-orexin (pporexin)), whereas no such changes were seen in the offspring of the undernourished dams (subjected to intrauterine growth restriction). These results indicate that in high-energy conditions either the adaptive response does not function properly or has not been established in the offspring of undernourished dams. Because NPY and pporexin are negatively regulated by leptin, these findings suggest that in the intrauterine growth restriction group, the leptin resistance of hypothalamic functions, which is usually caused by diet-induced obesity in adulthood, had already been established in the peri-pubertal period.


Kisspeptin mRNA expression is increased in the posterior hypothalamus in the rat model of polycystic ovary syndrome.

Hypersecretion of luteinizing hormone (LH) is a common endocrinological finding of polycystic ovary syndrome (PCOS). This derangement might have a close relationship with hypothalamic kisspeptin expression that is thought to be a key regulator of gonadotropin-releasing hormone (GnRH). We evaluated the relationship between the hypothalamic-pituitary-gonadal axis (HPG axis) and kisspeptin using a rat model of PCOS induced by letrozole. Letrozole pellets (0.4 mg/day) and control pellets were placed subcutaneously onto the backs of 3-week-old female Wistar rats. Body weight, vaginal opening and vaginal smear were checked daily. Blood and tissues of ovary, uterus and brain were collected at 12-weeks of age. An hypothalamic block was cut into anterior and posterior blocks, which included the anteroventral periventricular nucleus (AVPV) and the arcuate nucleus (ARC), respectively, in order to estimate hypothalamic kisspeptin expression in each area. The letrozole group showed a similar phenotype to human PCOS such as heavier body weight, heavier ovary, persistent anovulatory state, multiple enlarged follicles with no corpus luteum and higher LH and testosterone (T) levels compared to the control group. Kisspeptin mRNA expression in the posterior hypothalamic block including ARC was higher in the letrozole group than in the control group although its expression in the anterior hypothalamic block was similar between groups. These results suggest that enhanced KNDy neuron activity in ARC contributes to hypersecretion of LH in PCOS and might be a therapeutic target to rescue ovulatory disorder of PCOS in the future.


Developmental changes in the hypothalamic mRNA expression levels of PACAP and its receptor PAC1 and their sensitivity to fasting in male and female rats.

The actions and responses of hypothalamic appetite regulatory and factors change markedly during the neonatal to pre-pubertal period. Pituitary adenylate cyclase-activating polypeptide (PACAP) has been found to play pivotal roles in the regulation of metabolic and nutritional status through its specific receptor PAC1. PACAP/PAC1 have anorectic roles, and their functions are regulated by leptin in adulthood. In the present study, we showed that hypothalamic PACAP mRNA expression decreases during the neonatal to pre-pubertal period (from postnatal day 10-30) in both male and female rats. During this period, hypothalamic PACAP mRNA expression was not affected by 24h fasting in either sex, while the serum leptin levels (leptin is a positive regulator of hypothalamic PACAP expression in adulthood) of both sexes were decreased by fasting. On the other hand, hypothalamic PAC1 mRNA expression did not change during the neonatal to pre-pubertal period in either sex; however, its levels were consistently higher in males than in females. Hypothalamic PAC1 mRNA expression was decreased by 24h fasting in males, but no such changes were observed in females. These results indicate while hypothalamic PACAP expression is sensitive to a negative energy state and the serum leptin level in adulthood, no such relationships are seen in the pre-pubertal period. In addition, we speculate that differences in the gonadal steroidal milieu might induce sexual dimorphism in the basal hypothalamic PAC1 mRNA level and its response to fasting. The mechanisms responsible for and the physiological effects of such changes in hypothalamic PACAP and PAC1 expression during the developmental period remain to be clarified.

International Journal of Developmental Neuroscience


The expression of orexigenic and anorexigenic factors in middle-aged female rats that had been subjected to prenatal undernutrition.

Fetal growth retardation, which affects short- and long-term fetal brain development, is associated with metabolic, hematological, and thermal disturbances, which can increase the risk of metabolic syndrome later in life. Orexigenic and anorexigenic factors regulate food intake and energy expenditure. We studied how the expression of these factors was affected by food deprivation (FD) in middle-aged female rats that had been subjected to prenatal undernutrition. Eight pregnant rats were divided into two groups, the normal nutrition (NN) (n=4) group and the undernutrition (UN) (n=4) group, which received 50% (approximately 11g) of the daily food intake of the normal nutrition rats from day 13 of pregnancy to delivery. The pups from these dams were defined as the maternal NN (mNN) and maternal UN (mUN) groups, respectively. After weaning, all of the pups were housed and allowed ad libitum access to food and water. At the age of 6 months, both groups of pups were sub-divided into three groups. One group was allowed to consume normal amounts of food (Fed), and the other two groups were subjected to 24h or 48h FD (n=7-8 per group). The rats' serum leptin levels and hypothalamic mRNA expression levels of various orexigenic or anorexigenic factors were measured. In both the mNN and mUN rats, the serum leptin levels of the 24h and 48h FD groups tended to be lower than those of the Fed group, and the serum leptin levels of the 24h FD mUN rats and the Fed mUN rats differed significantly. The hypothalamic neuropeptide Y (NPY) mRNA expression levels of the 24h and 48h FD groups were significantly higher in the mUN rats than in the mNN rats. In addition, among the mUN rats the hypothalamic NPY mRNA expression levels of the 48h FD group were significantly higher than those of the Fed group. In both the mNN and mUN rats, prepro-orexin mRNA expression was lower in the 48h FD group than in the corresponding Fed group. Among the mUN rats, the 48h FD group exhibited significantly lower hypothalamic proopiomelanocortin (POMC) mRNA expression than the Fed group, and a similar tendency was seen among the mNN rats. Among the mNN rats, the 24h FD group displayed significantly higher hypothalamic leptin receptor (OBRb) mRNA levels than the Fed group. However, no such differences were seen among the mUN rats. As a result, the hypothalamic OBRb mRNA expression levels of the mUN rats in the 24h and 48h FD groups were lower than those of the corresponding mNN rat groups. These findings indicate that rats that are subjected to prenatal undernutrition exhibit upregulated expression of orexigenic factors and are more sensitive to FD in middle age, which might increase their risk of developing metabolic disorders in later life.


Steroidogenic pathways involved in androgen biosynthesis in eumenorrheic women and patients with polycystic ovary syndrome.

The conventional Δ5 and Δ4 steroidogenic pathways mediate androgen production in females. While multiple non-conventional pathways to dihydrotestosterone (DHT) have recently been postulated in humans, the functional significance of these pathways remains to be elucidated. The aim of this study was to clarify the origin of androgens in healthy women and in patients with polycystic ovary syndrome (PCOS), a multifactorial disorder characterized by androgen overproduction. We measured 13 steroids in blood samples of 31 eumenorrheic females and 28 PCOS patients using liquid chromatography-tandem mass spectrometry and chemiluminescent enzyme immunoassay. We found that 17-hydroxy (17-OH) progesterone (17-OHP), androstenedione (Δ4A), testosterone, androstanedione, androsterone, and androstanediol levels were higher in the patient group than in the eumenorrheic group, while levels of other steroids were comparable between the two groups. In the eumenorrheic group, DHT levels were correlated with testosterone, androstanedione, and androstanediol. Quantitative correlations were also observed among 17-OH allopregnanolone, androsterone, androstanediol, and DHT, and among Δ4A, androstanedione, androsterone, and androstanediol. In the patient group, DHT levels were correlated with testosterone levels, but not with androstanedione or androstanediol levels. Δ4A and testosterone paralleled 17-OHP. Androstanedione, androsterone, androstanediol, and 17-OH allopregnanolone were quantitatively correlated. In both groups, multivariable linear regression analyses suggested relationships between androsterone and androstanedione, as well as between androsterone and 17-OH allopregnanolone. These results indicate that multiple androgen biosynthesis pathways are operating in eumenorrheic females and PCOS patients. In PCOS patients, excessive androgens are produced primarily via the conventional pathways, while two alternative pathways; i.e., an androstanedione-mediated pathway and a so-called backdoor pathway, likely serve as sources of a weak androgen and potential precursors of DHT.

The Journal of Steroid Biochemistry and Molecular Biology


Prenatal undernutrition results in greater lipopolysaccharide-induced changes in hypothalamic TNF- expression, but does not affect the equivalent changes in the serum levels of luteinizing hormone and testosterone, in adult male rats.

Immune stress can cause reproductive dysfunction. Some hypothalamic factors such as pro-inflammatory cytokines play pivotal roles in reproductive disorders under immune stress conditions. Recently, it has been reported that prenatal undernutrition affects not only metabolic functions, but also the responses of physiological functions to immune stress in adulthood. In this study, the long-term effects of prenatal undernutrition on the responses of hypothalamic pro-inflammatory cytokine (interleukin (IL)-1, tumor necrosis factor (TNF)-, and IL-6) expression; reproductive endocrine factors; i.e., the serum levels of gonadotropins and testosterone; and hypothalamic kisspeptin expression to lipopolysaccharide (LPS) were examined in male rats. Pregnant rats were divided into two groups; i.e., the normally nourished group and the undernourished (50% food restricted) group. The offspring of the normally nourished mothers (control) and undernourished mothers (the intrauterine growth restriction [IUGR] group) were sub-divided into saline-injected and LPS (500g, i.p.)-injected groups at 10 weeks of age. The rats' hypothalamic pro-inflammatory cytokine levels and serum luteinizing hormone (LH) and testosterone levels were measured and compared between the control and IUGR groups. The hypothalamic pro-inflammatory cytokine mRNA levels of the LPS-injected rats were significantly higher than those of the saline-injected rats in both the control and IUGR groups. The changes in the hypothalamic expression level of TNF-, but not those of the other cytokines, induced in response to LPS were more marked in the IUGR group than in the control group. On the other hand, although the serum LH and testosterone levels of the LPS-injected rats were significantly lower than those of the saline-injected rats in both the control and IUGR groups, their levels did not differ between the control and IUGR groups under the LPS-injected conditions. These results suggest that prenatal undernutrition results in more marked LPS-induced changes in hypothalamic TNF- expression, but does not alter the effects of LPS on the serum levels of LH or testosterone, in adult male rats.


Developmental changes in the hypothalamic mRNA levels of nucleobindin-2 (NUCB2) and their sensitivity to fasting in male and female rats.

Nesfatin-1 is a central anorectic peptide derived from the precursor protein nucleobindin-2 (NUCB2). In the present study, the changes in hypothalamic NUCB2 mRNA expression and their responses to food deprivation during the neonatal to pre-pubertal period (postnatal days 10, 20, and 30) were evaluated in male and female rats. The rats' serum leptin levels were also measured because NUCB2 mRNA expression is positively regulated by leptin. In both the female and male rats, hypothalamic NUCB2 mRNA expression tended to fall throughout development. Similarly, higher serum leptin levels were detected on postnatal day 10 than on postnatal days 20 and 30 in both sexes. Hypothalamic NUCB2 mRNA expression was positively correlated with the serum leptin level in both the female and male rats; however, the relationship was not significant in males. The hypothalamic NUCB2 mRNA levels of the fed and 24h fasted groups did not differ at any time point in either sex. On the other hand, the serum leptin levels of the 24h fasted group were significantly lower than those of the fed group at all time points in both sexes. It can be speculated that the upregulation of hypothalamic leptin activity might induce a transient increase in hypothalamic NUCB2 mRNA expression during the early postnatal period (postnatal day 10) in both sexes. However, hypothalamic NUCB2 mRNA expression does not become sensitive to a negative energy balance during the neonatal to pre-pubertal period.


The suppressive effect of immune stress on LH secretion is absent in the early neonatal period in rats.

Some physiological functions display weak responses to stress in the early neonatal period; i.e., they exhibit stress hyporesponse periods. In this study, we evaluated whether gonadotropin regulatory factors exhibit stress hyporesponsive periods in male and female rats. Rats were intraperitoneally injected with lipopolysaccharide (100g/kg) (LPS group) or saline (control group) on postnatal day (PND) 5, 10, 15, or 25. Then, their serum luteinizing hormone (LH) concentrations and hypothalamic mRNA levels of gonadotropin regulatory factors; i.e., kisspeptin (Kiss1), the kisspeptin receptor (Kiss1r), and gonadotropin-releasing hormone (GnRH), were measured at 2h after the injection. The hypothalamic mRNA levels of pro-inflammatory cytokines were also measured because they suppress gonadotropin secretion. The serum LH concentration of the LPS group was lower than that of the control group at PND25 in both sexes, but no such difference was seen at PND5, 10, or 15 in either sex. In both sexes, the hypothalamic tumor necrosis factor (TNF) and interleukin (IL)-6 mRNA expression levels of the LPS group were higher than those of the control group at PND25, but not at PND5 or 10. The hypothalamic IL-1 mRNA expression level of the LPS group was higher than that of the control group at all time points. The hypothalamic Kiss1, Kiss1r, and GnRH mRNA expression levels of the LPS and control groups did not differ at any time point in either sex. These findings suggest that gonadotropin regulatory factors exhibit stress hyporesponse periods. The hypothalamic-pituitary-gonadal axis (HPG) might become responsive to immune stress between PND15 and 25, which could be related to enhanced hypothalamic cytokine expression. The avoidance of infectious stress during the early neonatal period might be important for normal development of the HPG axis.


The effects of ovariectomy and LPS-induced endotoxemia on resistin levels in female rats.

Resistin is involved in the inflammatory response, as well as in insulin resistance. In rodents, resistin levels are partially regulated by ovarian hormones. Thus, ovariectomy-induced changes in resistin levels and their response to lipopolysaccharide (LPS)-induced septic stress were evaluated. Ovariectomized (OVX) rats exhibited higher serum resistin concentrations and visceral and subcutaneous white adipose tissue (WAT) resistin mRNA levels than sham-operated (sham) rats under the saline-injected (basal) conditions. The serum resistin levels of the gonadal intact male rats were higher than those of the sham rats, whereas the serum resistin levels of the male and OVX rats did not differ. In both the sham and OVX rats, the serum resistin concentration and the resistin mRNA levels of WAT were increased by LPS injection. At 24h after the LPS injection, no difference was detected in the serum resistin concentrations or WAT mRNA resistin levels between the sham and OVX rats. These results suggest that ovarian hormones partially regulate the basal resistin levels of female rats.


Effect of high fat diet on artificial oocyte activation following superovulation in mice.

The aim of the present study was to determine the effects of increased dietary intake and high fat diet (HFD) in mice on artificial oocyte activation by using puromycin or roscovitine. Six-week-old mice were fed as either a control diet group, an increased dietary intake group or an HFD group for 4 weeks. Oocytes were obtained following superovulation and were divided into three treatment groups (no activation treatment, calcium ionophore and puromycin treatment, and calcium ionophore and roscovitine treatment) and were incubated for 4 h. Retrieved oocytes and numbers of oocytes activated as assessed by morphological changes were compared among the three treatment groups. The proportion of degenerated oocytes in HFD mice was significantly higher than that in control diet mice. The rates of activation in oocytes treated with roscovitine were 90.3% in control diet mice, 89.8% in increased dietary intake mice and 67.9% in HFD mice. The rate of activation in oocytes treated with roscovitine in HFD mice was significantly lower than the rates in control diet mice and increased dietary intake mice. The rates of activation in oocytes treated with puromycin were 90.6% in control diet mice, 94.0% in increased dietary intake mice and 71.4% in HFD mice, and the rate of activation in oocytes treated with puromycin in HFD mice was significantly lower than the rates in control diet mice and increased dietary intake mice. HFD-induced obesity deteriorated induction of oocyte activation by roscovitine or puromycin in mice.


Prenatal undernutrition increases the febrile response to lipopolysaccharides in adulthood in male rats.

It has been reported that prenatal undernutrition affects the development of the peripheral immune system. In this study, the effects of prenatal undernutrition on the febrile response and hypothalamic innate immune system were evaluated in male rats. Pregnant rats were divided into normally nourished (NN) and undernourished groups (UN). The febrile and anorectic responses to lipopolysaccharides (LPS) were evaluated in the offspring of NN and UN dams. The hypothalamic expression levels of pro-inflammatory cytokines, toll-like receptor 4 (TLR4), and neuropeptide Y (NPY) were also evaluated. The UN rats exhibited significantly lighter body weights than the NN rats at birth; however, their mean body weight was the same as that of the NN rats by postnatal day 10. In adulthood, the UN rats exhibited significantly stronger febrile responses than the NN rats, and the anorectic responses of the UN rats also tended to be stronger than those of the NN rats. On the other hand, no differences in hypothalamic interleukin (IL)-1, IL-6, tumor necrosis factor-, TLR4, or NPY mRNA expression were detected between the NN and UN rats. These results suggest that prenatal undernutrition has long-lasting effects on the febrile response to LPS. However, the precise mechanism underlying these effects and their pathophysiological significance remain unclear.

International Journal of Developmental Neuroscience


LH and testosterone production are more sensitive to the suppressive effects of food deprivation in prenatally undernourished male rats.

Although prenatal undernutrition affects the development of metabolic, physiological, and reproductive functions, it remains unclear whether it also affects physiological responses to undernutrition in adulthood. Therefore, in this study we examined whether prenatal undernutrition alters the sensitivity of the hypothalamic-pituitary-gonadal (HPG) axis to fasting in adult male rats. The offspring of ad libitum fed dams (control) and 50% food-restricted (during the late gestational period) dams (IUGR) were sub-divided into ad libitum fed (fed) and 48 h food deprivation (FD) groups at 10 weeks of age. In each group, the serum levels of luteinizing hormone (LH), testosterone, and leptin and the hypothalamic mRNA expression levels of gonadotropin-releasing hormone (GnRH) regulatory factors were measured. The serum LH and testosterone levels of the IUGR-fed rats were significantly or tend to be higher than those of the control-fed rats, respectively. The serum LH levels of the IUGR-FD rats were lower than those of the IUGR-fed rats. Similarly, the serum testosterone levels of the IUGR-FD rats tended to be lower than those of the IUGR-fed rats. On the other hand, the serum LH and testosterone levels of the control-fed and control-FD rats did not differ. The serum leptin levels of the IUGR fed rats were higher than those of the control-fed rats. The serum leptin levels of the control-FD and IUGR-FD rats were lower than those of the control-fed and IUGR-fed rats, respectively. The hypothalamic neuropeptide Y (NPY) mRNA levels of the IUGR-FD rats were higher than those of the IUGR-fed rats. Similarly, hypothalamic NPY mRNA levels of control-FD rats were higher than those of the control-fed rats. The hypothalamic kisspeptin, kisspeptin receptor, RFamide-related peptide, GPR147, and OBRb mRNA levels of control fed rats did not differ between control-fed and IUGR-fed rats. Their mRNA levels of the fed and FD rats did not differ in the control or IUGR groups. These results suggested that prenatal undernutrition increased the basal LH and testosterone production, whereas they are easily reduced by food deprivation in male rats. Changes of serum leptin level, but not of hypothalamic reproductive related factors, might be involved in these alterations. However, the precise mechanisms responsible for these effects remain unclear.


The effects of prenatal undernutrition and postnatal high-fat diet on hypothalamic Kiss1 mRNA and serum leptin levels.

Prenatal undernutrition and postnatal overnutrition increase the risk of some metabolic disorders in adulthood, and hypothalamic leptin resistance makes an important contribution to these effects. Leptin plays important roles in the maintenance of reproductive function, and its actions might be partially mediated by kisspeptin, which is a potent positive regulator of gonadotropin-releasing hormone. In this study, the effects of prenatal undernutrition and postnatal overnutrition on reproductive parameters and sexual maturation during the peripubertal period were evaluated. Rats subjected to prenatal undernutrition (IUGR) and fed a postnatal high-fat diet (HFD) (n=7) exhibited 40% higher serum leptin levels and 30% lower hypothalamic Kiss1 (the gene encoding kisspeptin) mRNA levels than those subjected to prenatal undernutrition (IUGR) and fed a normal diet (n=7). No such HFD-induced postnatal alterations were observed in the rats fed a normal diet during the prenatal period (control) (n=7 per group). Although the consumption of the HFD did not affect the serum luteinizing hormone levels or body weight of the IUGR or control rats, it did promote vaginal opening in both groups (evaluated in 14 rats per group). These findings indicate that hypothalamic leptin resistance might occur in IUGR-HFD rats, but these changes do not influence downstream effectors of the reproductive endocrinological system. They also suggest that the relationships between nutritional conditions, body weight, reproductive factors, and sexual maturation are complex.


Developmental changes in hypothalamic oxytocin and oxytocin receptor mRNA expression and their sensitivity to fasting in male and female rats.

Oxytocin (OT) affects the central nervous system and is involved in a variety of social and non-social behaviors. Recently, the role played by OT in energy metabolism and its organizational effects on estrogen receptor alpha (ER-) during the neonatal period have gained attention. In this study, the developmental changes in the hypothalamic mRNA levels of OT, the OT receptor (OTR), and ER- were evaluated in male and female rats. In addition, the fasting-induced changes in the hypothalamic mRNA levels of OT and the OTR were evaluated. Hypothalamic explants were taken from postnatal day (PND) 10, 20, and 30 rats, and the mRNA level of each molecule was measured. Hypothalamic OT mRNA expression increased throughout the developmental period in both sexes. The rats' hypothalamic OTR mRNA levels were highest on PND 10 and decreased throughout the developmental period. In the male rats, the hypothalamic mRNA levels of ER- were higher on PND 30 than on PND 10. On the other hand, no significant differences in hypothalamic ER- mRNA expression were detected among the examined time points in the female rats, although hypothalamic ER- mRNA expression tended to be higher on PND 30 than on PND 10. Significant positive correlations were detected between hypothalamic OT and ER- mRNA expression in both the male and female rats. Hypothalamic OT mRNA expression was not affected by fasting at any of the examined time points in either sex. These results indicate that hypothalamic OT expression is not sensitive to fasting during the developmental period. In addition, as a positive correlation was detected between hypothalamic OT and ER- mRNA expression, these two molecules might interact with each other to induce appropriate neuronal development.


The responses of hypothalamic NPY and OBRb mRNA expression to food deprivation develop during the neonatal-prepubertal period and exhibit gender differences in rats.

Neuropeptide Y (NPY) is an important hypothalamic orexigenic neuropeptide that acts in the brain. It has been established that the fasting-induced up-regulation of NPY expression is mainly caused by a reduction in the activity of leptin, which is a hormone secreted by adipose tissue. We have reported that in female rats hypothalamic NPY mRNA expression does not respond to fasting during the early neonatal period, but subsequently becomes sensitive to it later in the neonatal period. In this study, we compared the developmental changes in the responses of NPY and leptin expression to fasting between male and female rats during the neonatal to pre-pubertal period. Fasting was induced by maternal deprivation during the pre-weaning period (postnatal days 10 and 20) and by food deprivation during the post-weaning period (postnatal day 30). Hypothalamic NPY mRNA expression was not affected by fasting on postnatal day 10, whereas it was increased by fasting on postnatal day 20 and 30 in both males and females. On the other hand, the serum leptin level was decreased by fasting at all examined ages in both sexes. Namely, hypothalamic NPY mRNA expression was not correlated with the reduction in the serum leptin level at postnatal day 10 in either sex. Under the fasted conditions, the hypothalamic NPY mRNA levels of the males were higher than those of the females on postnatal days 20 and 30, whereas no such differences were observed under the normal nourishment conditions. The serum leptin levels observed under the fasted conditions did not differ between males and females at any examined age. These results suggest that some hypothalamic NPY functions develop during the neonatal period and that there is no major difference between the sexes with regard to the time when NPY neurons become sensitive to fasting. They also indicate that hypothalamic NPY expression is more sensitive to under-nutrition in male rats than in female rats, at least during the pre-pubertal period.


Effects of LPS injection on the hypothalamic and testicular mRNA expression levels of reproductive factors in male rats.

We speculate that changes in hypothalamic RFRP expression might represent a protective response aimed at attenuating LPS-induced anorectic responses.


Developmental changes in hypothalamic toll-like-receptor 4 mRNA expression and the effects of lipopolysaccharide on such changes in female rats.

Hypothalamic pro-inflammatory cytokine expression exhibits a weaker response to lipopolysaccharides (LPS) during the early neonatal period than during the later developmental period. Although toll-like receptor 4 (TLR4), which recognizes bacterial molecules, activates pro-inflammatory cytokine responses, the developmental changes in hypothalamic TLR4 expression have not been evaluated. In this study, the hypothalamic TLR4 mRNA levels of saline-injected and LPS-injected rats were measured during the neonatal, pre-pubertal, and post-pubertal periods. The rats' hypothalamic TLR4 mRNA levels gradually increased from the neonatal to pubertal period and were altered by the injection of LPS at all examined ages (postnatal day (PND) 5, 15, 25, and 42). LPS injection resulted in decreased hypothalamic TLR4 mRNA expression at PND5, whereas it increased hypothalamic TLR4 mRNA expression at PND15, 25, and 42. After the injection of LPS, the hypothalamic mRNA levels of the pro-inflammatory cytokines interleukin (IL)-1, tumor necrosis factor , and IL-6 were attenuated during the early developmental period and increased acutely on PND42. The expression profiles of these pro-inflammatory cytokines exhibited similar, but not entirely consistent, changes to those displayed by TLR4 during the developmental period. Hypothalamic TLR4 mRNA expression gradually increased throughout the developmental period, whereas the mRNA expression levels of the pro-inflammatory cytokines increased acutely at PND42. Thus, it is assumed that hypothalamic TLR4 hypoactivity contributes to the low sensitivity of pro-inflammatory cytokines to LPS during the early developmental period.


Effects of ovariectomy on the inflammatory responses of female rats to the central injection of lipopolysaccharide.

It has been reported that obesity leads to more marked inflammatory responses in a site-specific manner. As has been seen in other animal models of obesity, ovariectomized rodents exhibit obesity and exacerbated fever and anorectic responses to the systemic injection of lipopolysaccharide (LPS). Furthermore, they also display increased pro-inflammatory cytokine expression in several central and peripheral tissues. Interestingly, the alterations observed in the hypothalamus are more marked than those seen in other peripheral tissues. In this study, the effects of ovariectomy on hypothalamic inflammatory responses were evaluated using the central LPS injection method. LPS was intracerebroventricularly (i.c.v.) injected into ovariectomized and gonadally intact female rats, and the immune responses of the two groups were compared. The ovariectomized rats exhibited heavier body weights than the gonadally intact rats. In addition, the ovariectomized rats displayed stronger febrile responses than the gonadally intact rats. After the i.c.v. injection of LPS, the hypothalamic interleukin (IL)-1, IL-6, and cyclooxygenase 2 mRNA levels of the ovariectomized rats were significantly higher than those of the gonadally intact rats. The effects of estradiol supplementation on the rats' immune responses were also examined. However, the febrile responses and hypothalamic IL-1, IL-6, and TNF- mRNA levels of estradiol-supplemented ovariectomized rats and body weight matched oil-administered (control) rats did not differ after the i.c.v. injection of LPS. These results indicate that hypothalamic sensitivity to LPS is increased in ovariectomized rats and that this change is induced by the indirect effects of gonadal steroid deficiency. As is seen in other obese animal models, ovariectomy-induced obesity might play important roles in the exacerbated inflammatory responses observed in ovariectomized rats.


Changes in the responsiveness of hypothalamic PK2 and PKR1 gene expression to fasting in developing male rats.

Prokineticin (PK2) and its receptors (PKRs) are expressed in several regions of the central nervous system, including the hypothalamus. It has been reported that PK2 inhibits food intake via PKR1 and that the hypothalamic PK2 mRNA levels of adult rodents were reduced by food deprivation. However, some hypothalamic factors do not exhibit sensitivity to undernutrition in the early neonatal period, but subsequently become sensitive to it during the neonatal to pre-pubertal period. In this study, we investigated the changes in the sensitivity of hypothalamic PK2 and PKR1 mRNA expression to fasting during the developmental period in male rats. Under the fed conditions, the rats' hypothalamic PK2 and/or PKR1 mRNA levels were higher on postnatal day (PND) 10 than on PND20 or PND30. In addition, the hypothalamic PK2 and/or PKR1 mRNA levels of the male rats were higher than those of the females at all examined ages (PND10, 20, and 30). Hypothalamic PK2 mRNA expression was decreased by 24h fasting at PND10 and 30, but not at PND20. In addition, hypothalamic PKR1 mRNA expression was decreased by 24h fasting at PND10, but not at PND20 or 30. These results indicate that both PK2 and PKR1 are sensitive to nutritional status in male rats and that this sensitivity has already been established by the early neonatal period. It can be speculated that the PK2 system might compensate for the immaturity of other appetite regulatory factors in the early neonatal period.

International Journal of Developmental Neuroscience


The effects of LPS-induced endotoxemia on the expression of adiponectin and its receptors in female rats.

Adiponectin (APN), secreted by white adipose tissue (WAT), acts as a protective factor against inflammatory conditions. However, the changes in the expression levels of endogenous APN and the two types of APN receptor (AdipoR1 and AdipoR2) induced by acute inflammatory conditions have not been fully elucidated. In this study, the changes in peripheral and/or central APN and AdipoR expression caused by lipopolysaccharide (LPS)-induced sepsis were examined in gonadal-intact (Sham) and ovariectomized (OVX) female rats. As it has been reported that APN and AdipoR suppress the production of inflammatory cytokines to prevent excessive inflammation, the mRNAs of these molecules were also examined. LPS injection induced increases in visceral WAT APN mRNA without affecting the serum APN level in both the Sham and OVX rats. OVX rats exhibited higher serum APN levels than Sham rats. LPS injection increased the subcutaneous WAT APN mRNA in OVX rats. In both Sham and OVX rats, LPS injection led to a decrease in hepatic AdipoR2 mRNA and an increase in hypothalamic AdipoR2 mRNA. Hypothalamic AdipoR2 mRNA was upregulated 24 h after LPS injection in OVX but not Sham rats. Serum TNF- level at 6 h after LPS injection and hypothalamic and hepatic IL-6 and TNF- mRNA at 24 h after LPS injection were significantly higher in Sham than OVX rats. These results suggest that APN and AdipoR play roles in modulating inflammation under septic conditions in female rats.


Hypothalamic Kiss1 and RFRP gene expressions are changed by a high dose of lipopolysaccharide in female rats.

Reproductive function is suppressed by several types of stress. Hypothalamic kisspeptin, which is a product of the Kiss1 gene, and GnIH/RFRP have pivotal roles in the regulation of GnRH and gonadotropins through their receptors Kiss1r and GPR147 in many species. However, alterations of these factors under stress conditions have not been fully evaluated. This study investigated the mechanisms of immune stress-induced reproductive dysfunction, especially focusing on the changes of Kiss1 and RFRP gene expression. Serum LH levels and hypothalamic Kiss1 and GnRH mRNA levels were decreased, while hypothalamic RFRP and GPR147 mRNA levels were increased by administration of a high dose of LPS (5mg/kg) in both ovariectomized and gonadal intact female rats. In this condition, Kiss1 and/or RFRP mRNA levels were positively and negatively correlated with GnRH expression, respectively. In contrast, hypothalamic Kiss1, RFRP, and GPR147 mRNA levels were not changed by administration of a moderate dose of LPS (500g/kg) in ovariectomized rats. Rats with high-dose LPS injection showed more prolonged fever responses and severe anorexia compared with rats with moderate-dose LPS injection, indicating that more energy was used for the immune response in the former. These results suggest that the underlying mechanisms of dysfunction of gonadotropin secretion are changed according to the severity of immune stress, and that changes of some reserved factors, such as kisspeptin and RFRP, begin to participate in the suppression of GnRH and gonadotropin in severe conditions. As reproduction needs a large amount of energy, dysfunction of gonadotropin secretion under immune stress may be a biophylatic mechanism by which more energy is saved for the immune response.


Site-specific changes in cytokine response to septic dose of lipopolysaccharide in ovariectomized female rats.

These findings indicate that ovariectomy-induced site-specific changes in cytokine response under septic conditions. As hypothalamic, but not peripheral, pro-inflammatory cytokines are directly involved in the fever response, the attenuation of fever response observed in ovariectomized rats may be caused by a reduction in central cytokine responses.

Journal of Endocrinological Investigation


Successful advanced maternal age pregnancy with mosaic turner syndrome conceived after ovulation induction with clomiphene citrate: a case report.

Turner women typically experience gonadal dysfunction that results in amenorrhea and sterility. We encountered a case of mosaic Turner syndrome where conception was possible after ovulation induction with clomiphene citrate (CC). The patient's ovaries were overresponsive to induction with CC. The challenges and successful outcome are reported.

Case Reports in Obstetrics and Gynecology


Prenatal exposure to glucocorticoids affects body weight, serum leptin levels, and hypothalamic neuropeptide-Y expression in pre-pubertal female rat offspring.

Glucocorticoid secretion is a key endocrine response to stress. It has been reported that prenatal stress induces long-lasting alterations in body weight regulation systems, which persist after the stress has ceased. In this study, the long-term effects of prenatal glucocorticoid exposure on body weight changes and the expression of appetite-regulating factors were examined in female rats. Pregnant rats were given normal drinking water (control) or dexamethasone (1 g/mL) dissolved in drinking water (DEX) from day 13 of pregnancy until delivery. Then, the body weight change, serum leptin levels, and hypothalamic NPY mRNA levels of their offspring were examined. The DEX dams gained significantly less body weight during pregnancy than the control dams. The DEX dams' offspring exhibited a significantly lower birth weight than the offspring of the control dams, and the same was true for body weight at postnatal days 20 and 28. The offspring of the DEX dams displayed significantly higher serum leptin levels and significantly lower hypothalamic NPY mRNA levels compared with the offspring of the control dams. Significant inverse correlations were detected between body weight and the serum leptin level, and between the serum leptin level and the hypothalamic NPY mRNA level. On the other hand, a significant positive correlation was detected between body weight and the hypothalamic NPY mRNA level. These results indicate that leptin production is increased in a long-lasting manner in offspring exposed to glucocorticoids during the prenatal period and that this results in attenuated body weight gain and hypothalamic NPY expression during the pre-pubertal period.


Changes in leptin production/secretion induced in response to septic doses of lipopolysaccharides in gonadally intact and ovariectomized female rats.

In addition to its role as a regulator of energy homeostasis, leptin plays a pivotal role in certain immune/inflammatory responses. It has been reported that the synthesis and secretion of leptin are increased during immune stress in male experimental animals, whereas the changes in leptin synthesis and secretion induced by immune stress in females have not been fully elucidated. In this study, we found that during lipopolysaccharide (LPS)-induced immune stress the synthesis and secretion of leptin were decreased in gonadally intact female rats, but increased in ovariectomized rats. However, the LPS-induced increase in the serum leptin level observed in the ovariectomized rats was partially attenuated by estradiol supplementation. These results suggest that the changes in leptin synthesis and secretion induced in response to immune stress in females are affected by the gonadal steroid milieu and that estradiol and other factors are involved in these alterations.


Pre-pubertal serum leptin levels and sensitivity to central leptin injection of prenatally undernourished female rats.

It has been reported that intrauterine undernutrition is closely associated with the pathogeneses of certain diseases in adulthood; i.e., insulin resistance and diabetes, and that leptin resistance plays a pivotal role in the pathology of such intrauterine growth restriction (IUGR)-related conditions. Therefore, examinations of IUGR-induced leptin resistance in early developmental period are important for protecting against future disease. In this study, the effects of prenatal undernutrition on the serum leptin levels and central leptin responses of rats during the neonatal and/or pre-pubertal period were examined. The 50% food-restricted undernourished dams' offspring (UNO) exhibited a significantly lower birth weight than the normal nutrition dams' offspring (NNO). However, the UNO grew rapidly, and their mean body weight had caught up with that of the NNO by postnatal day 8. Thus, there were no significant differences between the body weights of the two groups at postnatal day 12, 16, 20, or 28. The serum leptin levels of the UNO were significantly higher than those of the NNO at postnatal days 20 and 28. At postnatal day 28, no significant difference in the hypothalamic mRNA level of neuropeptide Y, which is the main target of leptin, or that of ObRb, which is the leptin receptor, was detected between the NNO and UNO. The chronic intracerebroventricular injection of leptin attenuated body weight gain in both the NNO and UNO; however, there were no significant differences between the body weights of the two groups at any of the examined postnatal time points, indicating that the UNO and NNO exhibited similar central sensitivity to leptin during the pre-pubertal period. These results suggest that prenatal undernutrition induces leptin resistance until the neonatal to pre-pubertal period and that these alterations might be caused by impaired transportation of leptin to central tissues.

International Journal of Developmental Neuroscience


Changes in the responsiveness of hypothalamic prokineticin 2 mRNA expression to food deprivation in developing female rats.

Prokineticin 2 (PK2) is highly expressed in several regions of the central nervous system, including the hypothalamus. Recently, it has been suggested that PK2 plays a role in appetite regulation. In adult male rodents, the administration of PK2 decreased food intake, and PK2 mRNA expression was reduced by food deprivation. Usually, the changes in the expression levels of appetite-regulating factors induced in response to fasting are not fully established during the neonatal period. Thus, we investigated the developmental changes in hypothalamic PK2 mRNA expression and the alterations in hypothalamic PK2 mRNA expression induced by fasting during the pre-pubertal period in female rats. The changes in hypothalamic neuropeptide Y (NPY) mRNA expression were also examined because NPY is a potent appetite regulatory factor. Hypothalamic PK2 mRNA expression was extremely high during the early neonatal period (postnatal day (PND) 5) compared with that observed during subsequent periods (PND15, 25, and 42), while hypothalamic NPY mRNA expression did not differ among any of the examined periods. A fasting-induced reduction in hypothalamic PK2 mRNA expression was observed on PND5, but no fasting-induced increase in hypothalamic NPY mRNA expression was seen during the same period. In addition, the fasting-induced reduction in hypothalamic PK2 mRNA expression observed on PND5 was more marked than that seen on PND25. These results suggest that the sensitivity of hypothalamic PK2 expression to undernutrition develops during the early neonatal period, when the responses of other appetite regulatory factors to such pressures remain immature.


Improvement in diagnostic performance of the revised total testosterone measuring system in Japanese women with polycystic ovary syndrome.

Hyperandrogenemia has become an important factor in the diagnosis of polycystic ovary syndrome (PCOS), in Japan as well as Western countries, since the Japanese PCOS diagnostic criteria were revised in 2007. The revised electrochemiluminescence immunoassay (ECLIA) reagent for total testosterone (T) (ECLusys TESTO II; New T assay), which has lower cross-reactivity with DHEA-S than the previous reagent (ECLusys TESTO I; Old T assay), has recently been developed. We studied the improvement between New and Old T assays in regards to the diagnostic performance of serum T for patients with PCOS. Serum T levels were measured in both normal women (Control, n=92) and PCOS patients (n=86). The rate of elevated T using each kit was significantly higher in the PCOS group than in the Control group, and significantly higher using New T assay (38/86, 44.2%) compared with Old T assay (26/86, 30.2%). A significant correlation was found between T level and body mass index, but not LH level. Furthermore, higher detection rates of hormonal abnormalities were seen in PCOS patients using combined measurement of both T and LH (63/80) compared with single measurement of either T (37/80) or LH (36/80). Therefore, the serum T assay is essential for the diagnosis of PCOS, and New T assay combined with the LH assay may improve the diagnostic process for PCOS. J. Med. Invest. 61: 65-71, February, 2014.

The Journal of Medical Investigation : JMI


Changes in central and peripheral inflammatory responses to lipopolysaccharide in ovariectomized female rats.

Obesity leads to increases in inflammatory responses in a site-specific manner. Ovariectomized animals, usually used as menopause models, exhibit obesity; however, their inflammatory responses have not been fully examined. In the present study, we investigated whether ovariectomy had site-specific effects on inflammatory responses. First, fever and anorectic responses to systemic injections of lipopolysaccharide (LPS) (500g/kg, i.p.) were compared between ovariectomized rats (OVX) and sham-operated female rats (Sham). Inflammatory cytokines at the central and peripheral levels were also compared under saline-injected and LPS-injected conditions. Body weight in OVX was significantly higher than in Sham. The anorectic responses (reduction of body weight and food intake) to LPS were higher in OVX than in Sham. In the hypothalamus, all of the examined cytokine (IL-1, TNF- and IL-6) mRNA levels in OVX were higher than in Sham under the LPS-injected condition. On the other hand, in serum and adipose tissue, only IL-6, not IL-1 and TNF-, levels in OVX were significantly higher than those in Sham under the LPS-injected condition. Second, responses to central (intracerebroventricular) injections of LPS (500ng) were compared between OVX and Sham. The result was that the fever response in OVX was more evident than in Sham. Finally, responses to systemic injections of LPS (500g/kg, i.p.) were compared between OVX (OVX-oil) and OVX with estradiol (E) and progesterone (P) supplementation (OVX-EP). The anorectic responses and hypothalamic cytokine mRNA levels under LPS-injected condition were not different between OVX-oil and OVX-EP. These results indicate that ovariectomy enhances inflammatory responses, especially at the central level compared with the peripheral level. As supplementation of E and P could not attenuate the anorectic and cytokine responses to LPS, the deficiency of gonadal steroids might not be directly involved in the increase of inflammatory responses in OVX.


Gonadotropin-inhibitory hormone reduces sexual motivation but not lordosis behavior in female Syrian hamsters (Mesocricetus auratus).

Reproductive success is maximized when female sexual motivation and behavior coincide with the time of optimal fertility. Both processes depend upon coordinated hormonal events, beginning with signaling by the gonadotropin-releasing hormone (GnRH) neuronal system. Two neuropeptidergic systems that lie upstream of GnRH, gonadotropin-inhibitory hormone (GnIH; also known as RFamide related peptide-3) and kisspeptin, are potent inhibitory and excitatory modulators of GnRH, respectively, that participate in the timing of the preovulatory luteinizing hormone (LH) surge and ovulation. Whether these neuropeptides serve as neuromodulators to coordinate female sexual behavior with the limited window of fertility has not been thoroughly explored. In the present study, either intact or ovariectomized, hormone-treated female hamsters were implanted for fifteen days with chronic release osmotic pumps filled with GnIH or saline. The effect of GnIH on sexual motivation, vaginal scent marking, and lordosis was examined. Following mating, FOS activation was quantified in brain regions implicated in the regulation of female sexual behavior. Intracerebroventricular administration of GnIH reduced sexual motivation and vaginal scent marking, but not lordosis behavior. GnIH administration altered FOS expression in key neural loci implicated in female reproductive behavior, including the medial preoptic area, medial amygdala and bed nucleus of the stria terminalis, independent of changes in circulating gonadal steroids and kisspeptin cell activation. Together, these data point to GnIH as an important modulator of female proceptive sexual behavior and motivation, independent of downstream alterations in sex steroid production.


Ovarian torsion associated with cessation of hormonal treatment for polycystic ovarian syndrome: a case report.

Torsion of an ovary or fallopian tube (adnexal torsion) usually occurs in ovaries with tumors or functional cysts. In polycystic ovarian syndrome (PCOS), the ovaries are bilaterally enlarged, but these enlarged ovaries rarely twist. Recently, the authors encountered a PCOS patient with ovarian torsion after the cessation of Kaufmann treatment. The etiological factors were unclear, but the authors suggest that the increase in ovarian volume was due to transient hypergonadotropic feedback. Thus, more attention should be paid to adnexal torsion that may arise subsequent to transient hypergonadtropic states, in relation to the cessation of hormonal treatment, and enlarged ovaries in PCOS patients.

Clinical and Experimental Obstetrics & Gynecology


Developmental changes in the responsiveness of hypothalamic ER alpha mRNA levels to food deprivation.

Estrogen plays an important role in feeding and energy balance, and the critical role of estrogen in the control of appetite and energy balance is mediated by hypothalamic estrogen receptor (ER) alpha. In undernourished rodents, hypothalamic ER alpha mRNA expression are decreased. Responses of some hypothalamic factors to negative energy balance develop during the early neonatal period. In this study, we examined the developmental changes of fasting-induced alterations in hypothalamic ER alpha mRNA expression in female rats. ER alpha mRNA expression was reduced after a 12-h or 24-h fast at postnatal days 15 and 25, but not at day 5. Serum estradiol levels in postnatal day 25 rats were not changed by fasting. Although serum leptin levels were suppressed by fasting at all ages, hypothalamic ER alpha mRNA expression at postnatal day 25 was not changed by leptin administration after a 24-h fast. These data show that the sensitivity of hypothalamic ER alpha to negative energy balance may not be established in the early neonatal period, and that it develops by postnatal day 15. Decreased leptin levels might not be involved in the alterations of hypothalamic ER alpha mRNA expression in the undernourished condition.