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

PURPOSE: 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. METHODS: 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. RESULTS: The areas of endometriotic lesions in the LPS group (10.8 8.6 mm2) were significantly larger than those in the control group (3.1 3.7 mm2).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. CONCLUSIONS: LPS promotes development of the early stages of murine endometriotic lesions. J. Med. Invest. 66 : 70-74, February, 2019.

A case of placental polyp treated with an intraoperative injection of prostaglandin F2α followed by hysteroscopic resection.

A 27-year-old woman had massive genital bleeding after an artificial abortion. Color Doppler ultrasonography showed a hypervascular mass. Hysteroscopy revealed a placental polyp. Serum hemoglobin level was decreased to 7.7 g/dl. Although uterine artery embolization (UAE) followed by hysteroscopic resection has been used for treatment of a placental polyp, UAE may not be an ideal option for patients with intent for future pregnancy because of the risk of ovarian function failure. This report presents a case of a placental polyp managed successfully with intracervical injection of prostaglandin F2α, as an alternative UAE, followed by hysteroscopic resection.

Effects of raloxifene on the production of cytokines in stimulated whole blood in ex vivo and in vitro studies.

PURPOSE: The aims of this study were to determine the effects of raloxifene therapy on production of cytokines and in vitro effects of raloxifene on production of cytokines by whole blood cultures. METHODS: We obtained samples of peripheral blood from 6 postmenopausal women with osteopenia at baseline and after 3 and 6 months of raloxifene therapy and 10 postmenopausal women who did not receive raloxifene therapy. Whole blood from raloxifene-treated women was stimulated with lipopolysaccharide (LPS) or phytohemeagglutinin (PHA). Whole blood from postmenopausal women who were not treated with raloxifene was preincubated with raloxifene at concentrations of 10(-10)-10(-7) M and then stimulated with LPS or PHA. Concentrations of IL-1ß, IL-4, IL-6, IL-12p40, IL-12p70, TNF-α and IFN-γ in the supernatant were measured by respective ELISAs. RESULTS: In ex vivo cultures, raloxifene therapy inhibited LPS-stimulated production of IL-1ß, IL-6, IL-12p40, IL-12p70 and TNF-α, but not PHA-stimulated production of IL-4 and IFN-γ. In in vitro cultures, raloxifene at a concentration (10(-9) M) inhibited LPS-stimulated production of IL-1ß, IL-6 and IL-12p40 and PHA-stimulated production of IFN-γ. CONCLUSIONS: Raloxifene therapy decreases the production of IL-1ß, IL-6, IL-12 and TNF-α but not that of IL-4 and IFN-γ, suggesting that modulation of cytokines could play a role in the mechanisms of the osteoprotective effect of raloxifene.

The role of blood in early endometrial-peritoneal interactions in a syngeneic mouse model of endometriosis.

PURPOSE: The aim of this study was to clarify the role of blood in the early stage of development of endometriotic lesions by developing a syngeneic transplantation model using immunocompetent mice. METHODS: Endometriotic lesions were induced in C57BL/6 mice by an intraperitoneal injection of endometrial fragments plus saline or endometrial fragments plus blood. Some endometrial fragments plus blood were injected with heparin, hirudin or tissue plasminogen activator (tPA). Endometriotic lesions on days 1, 3 and 5 were evaluated by gross and microscopic findings. RESULTS: The areas of endometriotic lesions in the blood group (6.4 ± 1.7 mm2) were significantly larger than those in the saline group (0.5 ± 0.3 mm2). The areas of endometriotic lesions were significantly reduced by the addition of heparin, hirudin or tPA. On day 1, endometriotic lesions in the blood group were observed on the peritoneum in five of the six mice. Endometriotic lesions on days 3 and 5 were significantly larger than those on day 1. On day 5, endometriotic lesions appeared cystic in all the mice. CONCLUSIONS: Blood accelerates the early stage of development of endometriotic lesions when endometrial fragments plus blood are injected. Blood property might be involved in early endometrial-peritoneal interactions.

Raloxifene reduces circulating levels of interleukin-7 and monocyte chemoattractant protein-1 in postmenopausal women.

OBJECTIVE: The aim of the present study was to determine the effects of raloxifene on changes in circulating levels of cytokines and chemokines in relation to changes in lipid profiles and markers of inflammation in postmenopausal women. METHODS: Fifty-three postmenopausal women aged 45-65 years old were randomly assigned in open, parallel-group fashion to a control group or raloxifene group. Twenty-six women received oral administration of 60 mg raloxifene every day and 27 women did not receive any drugs for 12 months. Serum cytokines levels were simultaneously measured using a multiplexed human cytokine assay. RESULTS: Serum IL-7 concentrations in women who received raloxifene were decreased significantly (p=0.014), and serum monocyte chemoattractant protein (MCP)-1 concentrations in women who received raloxifene were decreased significantly (p=0.0003) at 12 months. In the control group, serum levels of MCP-1 and IL-7 did not show significant changes. There were significant differences (p=0.032 and p=0.0024, respectively) in percentage changes in IL-7 and MCP-1 in the control group and in the raloxifene group. Levels of low-density lipoprotein cholesterol (LDL-C) and E-selectin were decreased significantly in women who received raloxifene, but the percentage changes in LDL-C and E-selectin over a period of 12 months were not significantly correlated with percentage changes in IL-7 and MCP-1 over the same period. CONCLUSION: Circulating levels of IL-7 and MCP-1 decrease in postmenopausal women who received raloxifene.

Evaluation of the potential of synthetic peptides of 80 kDa human sperm antigen (80 kDaHSA) for the development of contraceptive vaccine for male.

80 kDaHSA has been demonstrated to be responsible for inducing immunoinfertility. Synthetic peptides NT, 1, 2 and 4 of 80 kDaHSA are immunogenic and immunobiologically mimic the native protein. Peptides 1 and NT being highly immunogenic their potential for contraceptive vaccine development was evaluated. Active immunization of male rabbits with peptide-1 and -NT induced reversible infertility in 100% and 60% of animals, respectively and subsequently active immunization of non-human primate model, male marmosets with peptide-1 induced reversible infertility in six out of seven high antibody titer animals. The present study suggests the potential of peptide-1 of 80 kDaHSA for the development of contraceptive vaccine.

Association of serum cytokine concentrations with psychological symptoms in midlife women.

OBJECTIVE: The purpose of the present study was to clarify the association of serum cytokine concentrations, determined using a multiplexed cytokine assay, with psychological symptoms in midlife women. METHODS: Fifty-three peri- and post-menopausal women with and without psychological symptoms in Greene's climacteric scale were enrolled in this study. Levels of 17 cytokines in serum samples were measured simultaneously using a multiplexed human cytokine assay. RESULTS: Serum interleukin (IL)-6 concentration in women with psychological symptoms (2.71+/-047 pg/ml) was significantly (p=0.009) higher than that in women without psychological symptoms (0.98+/-0.18 pg/ml). Serum IL-8 concentration in women with psychological symptoms (33.4+/-8.17 pg/ml) was also significantly (p=0.022) higher than that in women without psychological symptoms (7.87+/-1.64 pg/ml). In addition, serum IL-10 concentration in women with psychological symptoms (0.74+/-0.26 pg/ml) was significantly (p=0.048) higher than that in women without psychological symptoms (0.07+/-0.04 pg/ml). Tumor necrosis factor (TNF)-alpha in serum was detected only in women with psychological symptoms. Serum IL-2 concentration in women with psychological symptoms tended (p=0.066) to be higher than that in women without psychological symptoms. No significant differences were found between levels of other cytokines in women with and without psychological symptoms. CONCLUSION: Psychological stress manifested as climacteric symptoms in midlife women may be associated with increases in serum concentrations of IL-6, IL-8, IL-10, and TNF-alpha.

Diagnostic performance of serum total testosterone for Japanese patients with polycystic ovary syndrome.

It is reported that the incidence of clinical and biochemical hyperandrogenism may be lower in Japanese patients with PCOS. Hyperandrogenism is included as a referential but not as an essential factor in the diagnostic criteria of the Japanese Society of Obstetrics and Gynecology (JSOG 1993). However, some patients with the typical clinical features of PCOS are not diagnosed with PCOS using JSOG 1993 criteria because they do not have a high LH level, which is defined as essential for diagnosis. In this study, we compared total testosterone (T) levels between Japanese patients with PCOS diagnosed using the JSOG 1993 criteria and normal menstrual women (controls). Fifty controls and 46 patients with PCOS were enrolled in this study. Furthermore, we evaluated the sensitivity and specificity of each cut-off value of T. The mean T level of patients with PCOS was significantly higher than that of the control (86 +/- 48 vs 68 +/- 46, P<0.01), and the prevalence rates of hyperandrogenism (T >114 ng/dL; defined as the mean +2SD of the control) were 10.2% in patients with PCOS and 4% in controls. The area under the ROC curve of T was 0.72, and there was no decision threshold to diagnose PCOS by T alone with both high sensitivity and high specificity. If the threshold is set as 110 ng/dL in order to gain high specificity, 94% of women whose serum level passed the threshold will be patients with PCOS. Although T should not be used as an independent essential factor of Japanese PCOS, it might be useful as a complementary factor in order to diagnose patients who have typical clinical features of PCOS but does not fulfill the JSOG 1993 criteria for PCOS.

Change in serum undercarboxylated osteocalcin concentration in bilaterally oophorectomized women.

OBJECTIVE: We investigated changes in serum undercarboxylated osteocalcin (ucOC) concentrations, bone turnover markers and spine bone mineral density (BMD) in women who had undergone bilateral oophorectomy during the premenopausal period. METHODS: The study population comprised 141 bilaterally oophorectomized and 32 premenopausal women for a cross-sectional study. The longitudinal study consisted of 21 bilaterally oophorectomized women. Serum ucOC concentration, serum concentrations of intact osteocalcin (OC) and bone-specific alkaline phosphatase (BAP) as bone formation markers, urine N-telopeptide (NTx) concentration as a bone resorption marker and serum parathyroid hormone (PTH) concentration were measured. RESULTS: Serum concentration of ucOC in women at 1 month after bilateral oophorectomy was significantly (p<0.05) higher than that in premenopausal women and the high level was sustained after surgical menopause. On the other hand, serum OC concentration at 1 month after surgical menopause was not different from that in premenopausal women. In the longitudinal study, serum ucOC concentration at 1 month after surgical menopause was significantly (p<0.05) increased compared to that before bilateral oophorectomy, while serum OC concentrations before and at 1 month after surgical menopause were not significantly different. CONCLUSION: The results of this study showed that serum ucOC concentration rapidly increases in women after bilateral oophorectomy and that change in serum ucOC concentration after surgical menopause is different from change in serum OC concentration.

Changes in serum cytokine concentrations during the menopausal transition.

OBJECTIVE: The purpose of the present study was to clarify the changes in serum concentrations of 17 cytokines in healthy women during the menopausal transition by using a multiplexed cytokine assay and to clarify the associations of these cytokines with serum estradiol concentration. METHODS: Sixteen premenopausal, 54 perimenopausal and 52 postmenopausal women were enrolled in this study. Seventeen cytokines in serum samples were measured simultaneously using a Bio-Plex human cytokine 17-Plex assay. RESULTS: Serum IL-6 concentration showed a weak positive correlation with age (r=0.196, p<0.05). Postmenopausal women for whom less than 5 years had passed since menopause showed significant (p<0.05) increase in serum concentrations of IL-2, GM-CSF and G-CSF, while serum IL-4 concentration was significantly (p<0.05) increased in postmenopausal women for whom more than 5 years had passed since menopause. Serum estradiol concentration showed a significant negative correlation with serum IL-6 concentration and weak negative correlations with serum concentrations of IL-2, IL-8 and GM-CSF. CONCLUSION: We were able to simultaneously measure the levels of 17 cytokines using a highly sensitive cytokine assay, and we found that the changes in serum cytokine concentrations during the menopausal transition differed. We also found that serum IL-6 concentration during the menopausal transition was negatively correlated with serum estradiol concentration.

Association of interleukin-8 with hot flashes in premenopausal, perimenopausal, and postmenopausal women and bilateral oophorectomized women.

OBJECTIVE: The purpose of this study was to identify serum cytokine concentrations in premenopausal, perimenopausal, and postmenopausal women and bilateral oophorectomized women with hot flashes. METHODS: Serum concentrations of 17 cytokines were simultaneously measured using a multiplexed human cytokine assay in 129 premenopausal, perimenopausal, and postmenopausal women and 50 bilateral oophorectomized women. RESULTS: Serum IL-8 concentrations in midlife women and bilateral oophorectomized women with severe hot flashes were significantly higher than the concentrations in women without hot flashes and women with mild and moderate hot flashes. Serum macrophage inflammatory protein-1beta concentration in women with severe hot flashes was significantly higher than those in women without hot flashes and women with mild and moderate hot flashes. CONCLUSION: Serum IL-8 concentrations in premenopausal, perimenopausal, and postmenopausal women and bilateral oophorectomized women with hot flashes were significantly higher than those in women without hot flashes. IL-8 may be associated with peripheral vasodilation in women with hot flashes.

Different effects of oral conjugated equine estrogens and transdermal estrogen on undercarboxylated osteocalcin concentration in postmenopausal women.

OBJECTIVE: Undercarboxylated osteocalcin (ucOC) is a sensitive marker of vitamin K status, and triglyceride (TG) has been shown to be the main transporter of vitamin K. In the present study, we examined the difference between ucOC concentrations in postmenopausal women receiving hormone therapy (HT) with oral conjugated equine estrogens (CEE) and transdermal estradiol (TE2). We also examined the associations of ucOC concentration with estradiol concentration and TG. DESIGN: Ninety-two postmenopausal women were recruited for this study. Serum concentrations of ucOC, intact osteocalcin, estradiol, and TG were measured before and after 12 months of HT. Forty-six women received oral administration of 0.625 mg of CEE and 2.5 mg of medroxyprogesterone acetate daily, and 46 women received transdermal administration of 50 mug of 17beta-estradiol twice weekly and 2.5 mg of medroxyprogesterone acetate daily. RESULTS: The ucOC concentration in women during HT with oral CEE was significantly (P < 0.01) lower than that in women during HT with TE2. Serum estradiol concentrations during HT with CEE showed a significant inverse correlation with ucOC concentrations and the ratio of ucOC/OC during HT (P < 0.05 and P < 0.01, respectively). In addition, the serum ucOC concentration in women with an increased percentage of change in TG was significantly (P < 0.01) lower than that in women with a decreased percentage of change in TG during HT with oral CEE. CONCLUSION: The effect of HT with TE2 on ucOC concentration in women is weaker than the effect of HT with oral CEE. Suppression of ucOC concentration in postmenopausal women during HT with oral CEE might be associated with the effect of vitamin K through increased TG induced by oral CEE.

Myeloid-related protein-8/14 is associated with proinflammatory cytokines in cervical mucus.

Myeloid-related protein-8 (MRP-8), MRP-14, and MRP-8/14 are found in a variety of inflammatory conditions and are involved in the host defense system. The objective of this study was to determine the concentrations of MRP-8, MRP-14, and MRP-8/14 in human cervical mucus and the associations between MRP-8/14 and proinflammatory cytokines. Samples of cervical mucus were obtained using a syringe from sexually active women (n=97) during the preovulatory phase. Samples from seven women were obtained using a swab placed in the cervical canal during the proliferative, preovulatory, and luteal phases. Concentrations of MRP-8, MRP-14, MRP-8/14, IL-1alpha, IL-6, IL-8, and granulocyte elastase were measured using an ELISA. The mean levels of MRP-8, MRP-14, and MRP-8/14 in cervical mucus were 1.87, 0.46, and 23.90microg/ml, respectively. The concentration of MRP-8/14 showed positive correlations with concentrations of IL-1alpha (p<0.0001), IL-8 (p<0.0001), and granulocyte elastase (p<0.0001). However, there were no significant differences in MRP-8/14 levels in the cervical mucus of each patient during the menstrual cycle. MRP-8/14 was mainly detected in human cervical mucus and showed a positive correlation with proinflammatory cytokines. The MRP-8/14 level in cervical mucus may be useful as a marker of inflammation of the uterine cervix.

Undercarboxylated osteocalcin concentration in postmenopausal women receiving hormone therapy daily and on alternate days.

OBJECTIVE: Undercarboxylated osteocalcin (ucOC) is a sensitive marker of vitamin K status. The authors examined the difference in serum ucOC concentrations in postmenopausal women receiving hormone therapy (HT) daily and on alternate days, and assessed the association between ucOC and triglyceride concentrations, which are related to the transport of vitamin K. DESIGN: Seventy-three postmenopausal women were recruited for this study. Thirty-seven women received 0.625 mg of conjugated equine estrogens (CEE) and 2.5 mg of medroxyprogesterone acetate (MPA) daily, and 36 women received 0.625 mg of CEE and 2.5 mg of MPA on alternate days. The concentrations of serum ucOC, bone turnover markers, lipid profiles, and hormones were measured before and after 12 months of HT. RESULTS: The ucOC concentration in women taking HT daily was significantly (P < 0.01) lower than that in women taking HT on alternate days. Serum ucOC concentrations during HT showed a significant (P < 0.01) inverse correlation with estradiol concentrations during HT. Serum estradiol concentrations during HT showed a significant (P < 0.01) positive correlation with triglyceride concentrations during HT. Furthermore, ucOC concentrations during HT showed a significant (P < 0.05) inverse correlation with triglyceride concentrations in women receiving HT. CONCLUSIONS: The effect of HT on alternate days on ucOC concentration was weaker than the effect of HT daily. In addition, ucOC concentration after 12 months of HT daily might be decreased due to the conversion of ucOC to carboxylated OC by the effect of vitamin K through increased triglyceride levels induced by oral CEE.

Essential role of IkappaB kinase alpha in thymic organogenesis required for the establishment of self-tolerance.

IkappaB kinase (IKK) alpha exhibits diverse biological activities through protein kinase-dependent and -independent functions, the former mediated predominantly through a noncanonical NF-kappaB activation pathway. The in vivo function of IKKalpha, however, still remains elusive. Because a natural strain of mice with mutant NF-kappaB-inducing kinase (NIK) manifests autoimmunity as a result of disorganized thymic structure with abnormal expression of Rel proteins in the thymic stroma, we speculated that the NIK-IKKalpha axis might constitute an essential step in the thymic organogenesis that is required for the establishment of self-tolerance. An autoimmune disease phenotype was induced in athymic nude mice by grafting embryonic thymus from IKKalpha-deficient mice. The thymic microenvironment that caused autoimmunity in an IKKalpha-dependent manner was associated with defective processing of NF-kappaB2, resulting in the impaired development of thymic epithelial cells. Thus, our results demonstrate a novel function for IKKalpha in thymic organogenesis for the establishment of central tolerance that depends on its protein kinase activity in cooperation with NIK.

Comparison and problems of measured values of LH, FSH, and PRL among measurement systems.

Several measurement systems are used for LH, FSH, and PRL in Japan, but the comparison of the measured values is difficult in some cases. In this study, we compared the measured values of 3 types of measurement systems widely used in Japan, SPAC-S, ARCHITECT, and Centaur, in females with a normal menstrual cycle and with various ovulatory disorders. Variant LH was discriminated by the criterion, SPAC-S LH/ARCHITECT LH ratio <0.5. Excluding the variant LH, the correlation of the measured LH values among the measurement systems was high, 0.94-0.99, and the value measured by ARCHITECT LH and Centaur LH was slightly higher than that measured by SPAC-S LH. As for FSH, the correlation was also high, 0.94-1.00, and the value measured by ARCHITECT FSH and Centaur FSH was slightly lower than that measured by SPAC-S FSH. The important judgment criterion of polycystic ovary syndrome (PCOS), a high level of LH relative to FSH, has been set to 1.0 or higher LH/FSH using SPAC-S. When the regression line of the LH/FSH ratio among the measurement systems was investigated, it can be said that 1.25 or higher by ARCHITECT and 1.40 or higher by Centaur are appropriate for the judgment criteria of a high level of LH relative to FSH. As for PRL, the correlation was high, 0.96-0.99, but the value showed large differences among the systems. For the diagnosis of hyperprolactinemia, the higher limit of the normal range may be 15 ng/mL for SPAC-S, 30 ng/mL for ARCHITECT, and 24 ng/mL for Centaur.

Sperm-zona pellucida interaction and immunological infertility.

Immune reactions against gametes appear to be physiologically important for the maintenance of homeostasis in reproduction. In contrast, aberration of the immune homeostasis might give rise to 'immunological infertility'. Antisperm antibodies cause infertility by blocking fertilization. The mechanism can be explained as inhibiting the acrosome reaction of sperm by their blocking effect on capacitation through inhibiting an increase of fluidity of the sperm membrane. Autoantibodies against zona pellucida also cause infertility by blocking sperm-zona pellucida interaction, though the definitive mechanism has not been elucidated. Pretreatment of spermatozoa with D-mannnose completely inhibited sperm penetration through, but not binding to, the zona pellucida. Furthermore, very rapid kinetics between sperm extracts and D-mannnose by a BIAcore apparatus suggest that a D-mannose ligand of the sperm surface is easy to bind to and dissociate from a D-mannose residue in the sperm receptor site on the zona pellucida. Thus, D-mannnose on the human zona pellucida might be an essential molecule acting as a second sperm receptor, through which sperm penetrate into the zona pellucida. Because these antibodies appear to not cause any deleterious clinical symptoms, sperm and zona pellucida antigens are promising candidates in the development of an immunocontraceptive. (Reprod Med Biol 2006; 5: 95-104).

Effects of postmenopausal hormone therapy every day and every other day on lipid levels according to difference in body mass index.

OBJECTIVE: The objective of this study was to determine the effects of postmenopausal estrogen and progestogen therapy (EPT) every day and every other day on lipid levels, particularly triglyceride (TG) levels, according to difference in body mass index (BMI). DESIGN: Ninety-nine postmenopausal women (mean age, 53.9 +/- 5.6 years; mean BMI, 22.8 +/- 2.8 kg/m) were randomly treated with EPT every other day or every day for 1 year. Fifty women received oral administration of 0.625 mg of conjugated equine estrogen (CEE) and 2.5 mg of medroxyprogesterone acetate (MPA) every other day, and 49 women received oral administration of 0.625 mg of CEE and 2.5 mg of MPA every day. Blood samples were collected at baseline and after 1 year of therapy for measurement of fasting TG, total cholesterol, high-density lipoprotein-cholesterol (HDL-C), and apolipoproteins. Data from 88 of the 99 postmenopausal women were used for analysis. RESULTS: In women whose BMI was 25 kg/m or higher, TG levels during EPT every day increased by 26.8%, while TG levels during EPT every other day decreased by 12.3%. There was a significant (P < 0.05) difference between percentage changes in TG during EPT every day and every other day. In women whose BMI was less than 25 kg/m, TG levels during EPT every day increased by 21.7%, while during EPT every other day TG levels did not change. The mean levels of estradiol during EPT every day in women whose BMI was less than 25 kg/m and in women whose BMI was 25 kg/m or higher were 28.5 and 38.7 pg/mL, respectively, the difference between these levels was significant (P < 0.01). On the other hand, there was no significant difference between levels of estradiol during EPT every other day in these two BMI groups. CONCLUSIONS: Triglyceride levels during EPT every day with conventional doses of CEE and MPA increased more in overweight and obese postmenopausal women in association with increased estrogen levels.