Is the Human Chorionic Gonadotropin Level on Day 10 After Cleaved Embryo Transfer a Predictor of Perinatal Outcome?: A Retrospective Cohort Study.

This study aimed to determine whether human chorionic gonadotropin (hCG) levels at day 10 after day 2 cleaved embryo transfer can predict pregnancy and perinatal outcomes. Patients who underwent oocyte retrieval with minimal stimulation or natural cycles and fresh or vitrified-warmed transfer of a single, day 2 cleaved embryo at our clinic between November 2018 and December 2020 were included in this study. Patients were classified into four age groups for oocyte retrieval and into ten groups based on the hCG level on day 10 after embryo transfer; pregnancy complications and delivery and neonatal outcomes were examined. Of the 5,840 cycles, 3,722 (63.7%) and 2,118 (36.3%) were fresh-cleaved and vitrified-warmed embryo transfers, respectively. The mean hCG level was 24.8 mIU/mL and the clinical pregnancy and live birth rates per transfer were 29.6% and 23.4%, respectively. Maternal age at the time of oocyte retrieval, husband's age, treatment cycle, embryo type and grade, cell number, and hCG levels were correlated with pregnancy and delivery outcomes in the univariate analysis. Conversely, only maternal age and hCG levels were correlated with the outcomes in the multivariate analysis. hCG levels on day 10 post-transfer are a useful predictor of pregnancy and delivery outcomes after cleaved embryo transfer. Live birth rates vary with maternal age, even when hCG levels are the same, but they do not vary according to the treatment cycle or type of embryo transferred. Low hCG levels may be associated with vasa previa but did not affect delivery outcomes.

Association between endometrial thickness before ovulation, live birth, and placenta previa rates in clomiphene citrate-treated cycles.

BACKGROUND: Although a recent study reported that the pregnancy outcomes in the first trimester were more correlated with endometrial thickness on the day of the trigger than with endometrial thickness on the day of single fresh-cleaved embryo transfer, it remains unclear whether endometrial thickness on the day of the trigger can predict live birth rate after a single fresh-cleaved embryo transfer. OBJECTIVE: This study aimed to examine whether endometrial thickness on the trigger day is associated with live birth rates and whether modifying the single fresh-cleaved embryo transfer criteria to reflect endometrial thickness on the trigger day improved the live birth rate and reduced maternal complications in a clomiphene citrate-based minimal stimulation cycle. STUDY DESIGN: This was a retrospective study of the outcomes of 4440 treatment cycles of women who underwent single fresh-cleaved embryo transfer on day 2 of the retrieval cycle. From November 2018 to October 2019, single fresh-cleaved embryo transfer was performed when endometrial thickness on the day of single fresh-cleaved embryo transfer was ≥8 mm (criterion A). From November 2019 to August 2020, single fresh-cleaved embryo transfer was conducted when endometrial thickness on the day of the trigger was ≥7 mm (criterion B). RESULTS: A multivariate logistic regression analysis revealed that increased endometrial thickness on the trigger day was significantly associated with an improvement in the live birth rate after single fresh-cleaved embryo transfer (adjusted odds ratio, 1.098; 95% confidence interval, 1.021-1.179). The live birth rate was significantly higher in the criterion B group than in the criterion A group (22.9% and 19.1%, respectively; P=.0281). Although endometrial thickness on the day of single fresh-cleaved embryo transfer was sufficient, the live birth rate tended to be lower when endometrial thickness on the trigger day was <7.0 mm than when endometrial thickness on the day of the trigger was ≥7.0 mm. The risk for placenta previa was reduced in the criterion B group when compared with the criterion A group (4.3% and 0.6%, respectively; P=.0222). CONCLUSION: This study demonstrated an association of decreased endometrial thickness on the trigger day with low birth rate and a high incidence of placenta previa. A modification of the criteria for a single fresh-cleaved embryo transfer based on endometrial thickness may improve pregnancy and maternal outcomes.

Letrozole-induced endometrial preparation improved the pregnancy outcomes after frozen blastocyst transfer compared to the natural cycle: a retrospective cohort study.

BACKGROUND: Letrozole treatment is considered an effective option in endometrial preparation for frozen embryo transfers in patients with ovulation disorders or irregular menstruation; however, the effectiveness of letrozole-induced endometrial preparation remains unclear in ovulatory patients. Furthermore, there is no comparative study reporting on pregnancy complications and congenital anomalies after frozen embryo transfers comparing natural and letrozole-assisted cycles. This study examined whether letrozole-induced endometrial preparation affected pregnancy outcomes, perinatal outcomes, and congenital anomalies after single vitrified-warmed blastocyst transfers (SVBTs) in ovulatory patients, as compared with the natural cycle. METHODS: This historic cohort study included only patients with unexplained infertility. Overall, 14,611 patients who underwent SVBTs between July 2015 and June 2020, comprising both natural and letrozole-assisted cycles, were included. Multiple covariates that impact outcomes were used for propensity score matching; 1,911 patients in the letrozole group were matched to 12,700 patients in the natural group, and the clinical records of 1,910 patients in each group were retrospectively analysed. Cycle characteristics, pregnancy outcomes (clinical pregnancy, ongoing pregnancy, and live birth), and incidence of pregnancy complications and congenital anomalies were statistically compared between the two groups. RESULTS: Multivariate logistic regression analysis showed that letrozole administration during SVBT cycles significantly improved the live birth rate (P = 0.0355). Gestational age, birth length, birth weight, and infant sex, as well as the incidence of pregnancy complications and birth defects, were statistically comparable between the two groups. Furthermore, multivariate logistic regression analysis revealed that the perinatal outcomes were not affected by letrozole-induced endometrial preparation. CONCLUSIONS: Letrozole-induced endometrial preparation improved the live birth rate compared with the natural cycle, without adverse effects on perinatal outcomes and congenital anomalies after SVBTs. Therefore, letrozole-induced endometrial preparation might be a safe and more effective strategy, especially for patients with insufficient luteal function.

Analysis of the risk factors and treatment for repeated implantation failure: OPtimization of Thyroid function, IMmunity, and Uterine Milieu (OPTIMUM) treatment strategy.

PROBLEM: What are the pregnancy outcomes after the OPtimization of Thyroid function, Immunity, and Uterine Milieu (OPTIMUM) treatment strategy in patients with repeated implantation failure (RIF)? METHOD OF STUDY: Infertile women with a history of RIF after more than three embryo transfer (ET) cycles underwent implantation testing, including a hysteroscopy, endometrial biopsy for CD138 immunostaining and bacterial culture, and serum 25-hydroxyvitamin D3 , interferon-γ-producing helper T (Th1) cell, IL-4-producing helper T (Th2) cell, thyroid-stimulating hormone, thyroid peroxidase antibody, and thrombophilia screening between April 2017 and August 2018. We treated chronic endometritis with antibiotics, aberrant high Th1/Th2 cell ratios with vitamin D and/or tacrolimus intake, overt/subclinical hypothyroidism with levothyroxine, and thrombophilia with low-dose aspirin. Of the 116 RIF women, 88 women with 133 ET cycles were recruited from a questionnaire-based survey regarding pregnancy outcomes. Fifty-nine consecutive RIF patients without the OPTIMUM treatment strategy were also recruited as a control. RESULTS: The 116 women with RIF after the OPTIMUM treatment strategy were 38.3 ± 3.8 years old and had an implantation failure history over 5 (3-19) ET cycles. Implantation testing identified impaired intrauterine circumstances in 75 women (64.7%), an aberrant elevated Th1/Th2 cell ratio in 56 women (48.3%), and thyroid abnormalities in 33 women (28.4%). Cumulative ongoing pregnancy rates including spontaneous pregnancy in the patients aged < 40 and ≥ 40 years were 72.7% and 45.5% within two ET cycles, respectively. The pregnancy outcomes in the OPTIMUM group were significantly higher than those in the control. CONCLUSIONS: The OPTIMUM treatment strategy improved pregnancy outcomes in patients with RIF.

Effects of gonadotropin administration on clinical outcomes in clomiphene citrate-based minimal stimulation cycle IVF.

PURPOSE: Exogenous gonadotropins (EGn) have been used occasionally in clomiphene citrate (CC)-based minimal stimulation cycles to compensate insufficient secretion of endogenous gonadotropin; however, the effectiveness of EGn supplementation remains unknown. In the present study, we assessed whether EGn improved pregnancy outcomes in CC-based minimal stimulation cycles. METHODS: A total of 223 patients treated with CC and EGn (CC-EGn group) were matched one to one to patients treated with CC only (CC group) by propensity score matching. Embryonic and pregnancy outcomes were retrospectively compared between the groups. RESULTS: The numbers of retrieved oocytes, fertilized oocytes, cleaved embryos, and cryopreserved blastocysts were increased in the CC-EGn group compared with the CC group. However, the cumulative live birthrate was comparable between the two groups. Although the increased number of retrieved oocytes was correlated significantly with improvement of the cumulative live birthrate in both groups, the correlation tended to be lower in the CC-EGn group than in the CC group (odds ratio, 1.193 vs 1.553). CONCLUSIONS: In CC-based minimal stimulation cycles, the stimulation should be started with CC only, and EGn administration should be scheduled only if insufficient secretion of endogenous gonadotropin is observed in the late follicular phase.

Does the endometrial thickness on the day of the trigger affect the pregnancy outcomes after fresh cleaved embryo transfer in the clomiphene citrate-based minimal stimulation cycle?

PURPOSE: Thin endometrium is often observed after clomiphene citrate (CC) administration for follicular development and is one of the reasons for embryo transfer (ET) cancelation or implantation failure. We retrospectively analyzed whether the endometrial thickness (EMT) on the days of the maturation trigger and ET are predictive factors of pregnancy outcomes after fresh cleaved ET in a CC-based minimal stimulation cycle (CC-cycle). METHODS: A total of 746 CC-cycles in vitro fertilization (IVF), followed by fresh cleaved ET, from November 2018 to March 2019 were analyzed. Associations between the pregnancy outcomes and EMT on the days of the trigger and ET were statistically evaluated. RESULTS: Although the EMT on the day of ET was not significantly associated with the ongoing pregnancy rate (adjusted odds ratio [AOR], 1.043; P = .3251), a decreased EMT on the day of the trigger was significantly associated with a low ongoing pregnancy rate (AOR, 1.154; P = .0042). Furthermore, the clinical pregnancy rate was significantly lower when the EMT was <7 mm on the day of the trigger during the CC-cycle. CONCLUSIONS: These results suggest that measurement of the EMT on the day of the trigger could be effective for predicting the pregnancy outcomes after fresh cleaved ET during the CC-cycle.

Endometrial thickness on the day of the LH surge: an effective predictor of pregnancy outcomes after modified natural cycle-frozen blastocyst transfer.

STUDY QUESTION: Can the endometrial thickness (EMT) on the day of the LH surge predict pregnancy outcomes after single vitrified-warmed blastocyst transfers (SVBTs) in modified natural cycles? SUMMARY ANSWER: Decreased EMT on the day of the LH surge is associated with older female age and a shortened proliferation phase and may be associated with low live birth and high chemical pregnancy rates. WHAT IS KNOWN ALREADY: The relation between EMT on the day of embryo transfer (ET) and pregnancy outcomes remains controversial; although numerous studies reported an association between decreased EMT on the day of ET and a reduced likelihood of pregnancy, recent studies demonstrated that the EMT on the day of ET had limited independent prognostic value for pregnancy outcomes after IVF. The relation between EMT on the day of the LH surge and pregnancy outcomes after SVBT in modified natural cycles is currently unknown. STUDY DESIGN SIZE DURATION: In total, 808 SVBTs in modified natural cycles, performed from November 2018 to October 2019, were analysed in this retrospective cohort study. Associations of EMT on the days of the LH surge with SVBT and clinical and ongoing pregnancy rates were statistically evaluated. Clinical and ongoing pregnancy rates were defined as the ultrasonographic observation of a gestational sac 3 weeks after SVBTs and the observation of a foetal heartbeat 5 weeks after SVBTs, respectively. Similarly, factors potentially associated with the EMT on day of the LH surge, such as patient and cycle characteristics, were investigated. PARTICIPANTS/MATERIALS SETTING METHODS: The study includes IVF/ICSI patients aged 24-47 years, who underwent their first SVBT in the study period. After monitoring follicular development and serum hormone levels, ovulation was triggered via a nasal spray containing a GnRH agonist. After ovulation was confirmed, SVBTs were performed on Day 5. The EMT was evaluated by transvaginal ultrasonography on the day of the LH surge and immediately before the SVBT procedure. MAIN RESULTS AND THE ROLE OF CHANCE: Of the original 901 patients, 93 who were outliers for FSH or proliferative phase duration data were excluded from the analysis. Patients were classified according to quartiles of EMT on day of the LH surge, as follows: EMT < 8.1 mm, 8.1 mm ≤ EMT < 9.1 mm, 9.1 mm ≤ EMT < 10.6 mm and EMT ≥ 10.6 mm. Decreased EMT on day of the LH surge was associated with lower live birth (P = 0.0016) and higher chemical pregnancy (P = 0.0011) rates. Similarly, patients were classified according to quartiles of EMT on day of the SVBT, as follows: EMT < 9.1 mm, 9.1 mm ≤ EMT < 10.1 mm, 10.1 mm ≤ EMT < 12.1 mm and EMT ≥ 12.1 mm. A decreased EMT on the day of SVBT was associated with a lower live birth rate (P = 0.0095) but not chemical pregnancy rate (P = 0.1640). Additionally, multivariate logistic regression analysis revealed a significant correlation between EMT on day of the LH surge and ongoing pregnancy; however, no correlation was observed between EMT on the day of SVBT and ongoing pregnancy (adjusted odds ratio 0.952; 95% CI, 0.850-1.066; P = 0.3981). A decreased EMT on day of the LH surge was significantly associated with greater female age (P = 0.0003) and a shortened follicular/proliferation phase (P < 0.0001). LIMITATIONS REASONS FOR CAUTION: The data used in this study were obtained from a single-centre cohort; therefore, multi-centre studies are required to ascertain the generalisability of these findings to other clinics with different protocols and/or patient demographics. WIDER IMPLICATIONS OF THE FINDINGS: This is the first report demonstrating a significant correlation between EMT on day of the LH surge and pregnancy outcomes after frozen blastocyst transfer in modified natural cycles. Our results suggest that EMT on day of the LH surge may be an effective predictor of the live birth rate. STUDY FUNDING/COMPETING INTERESTS: This study was supported by resources from the Kato Ladies Clinic. The authors have no conflicts of interest to declare.

Success rates in minimal stimulation cycle IVF with clomiphene citrate only.

PURPOSE: To determine age-adjusted overall success rates for patients undergoing clomiphene citrate only minimal stimulation cycle (mini) in vitro fertilization (IVF) without any gonadotropin administration. METHODS: Eight hundred thirty-nine women (mean age: 38.4 ± 0.1 years; 2488 cycles) underwent clomiphene citrate only mini-IVF. Their first oocyte retrieval was between January 2009 and December 2009, with follow-up until December 2014. The cumulative live birth rate (CLBR) per oocyte retrieval cycle started and live birth rate per oocyte was retrospectively analyzed. The basic CLBR was calculated as the number of women who achieved a live birth divided by the total number of women who started oocyte retrieval. RESULTS: The mean number of oocytes retrieved was 1.5. The basic CLBRs for all ages after the first and third cycles were 22.6% and 39.2%, respectively. For ≤ 34 years, 35-37 years, 38-40 years, 41-42 years, and ≥ 43 years, CLBRs after the first and third cycles were 42.5% and 70.1%, 32.9% and 49.1%, 20.0% and 38.6%, 12.6% and 25.2%, and 4.4% and 8.8%, respectively. These rates had a significant relationship with age (P < 0.01). The LBR per oocyte for all ages was 9.6%. CONCLUSION: Acceptable overall IVF success rates can be achieved in clomiphene citrate only mini-IVF, as well as acceptable LBR. The CLBRs and LBRs per oocyte are evidently influenced by women's age.

Administering human chorionic gonadotropin injections for triggering follicle maturation could impact fertility during the subsequent menstrual cycle.

OBJECTIVE: To determine whether the use of a human chorionic gonadotropin (hCG) injection as a follicle-maturation trigger affects a patient's reproductive ability during their subsequent menstrual cycle. METHODS: Patients that were infertile undergoing natural-cycle in vitro fertilization at Kato Ladies Clinic, Japan, between March and June 2012 were enrolled in a prospective cohort study. Patients who had received fertility treatments other than hCG injections were excluded from the study. The remaining patients were divided into two cohorts; patients who had received injection-administered hCG (study group) and patients who had not received any fertility treatment (control group) during their preceding menstrual cycle. The rates of oocyte retrieval, fertilization, clinical pregnancy, and live deliveries were analyzed using a Fisher exact test. RESULTS: The rate of successful oocyte-retrieval (P<0.001) and the delivery-rate (P=0.002) were significantly lower in the study group in comparison with the control group. Additionally, the incidence of empty follicles (P<0.001) and degenerated oocytes (P=0.002) was significantly higher in the exposure group. CONCLUSION: Triggering follicle maturation with hCG during in vitro fertilization could impact patient fertility during their next cycle. Treatment with hCG injection has the potential to influence not only the cycle during which it is administered, but also the subsequent menstrual cycle.

The production of vascular endothelial growth factor and metalloproteinase via protease-activated receptor in human endometrial stromal cells.

OBJECTIVE: To measure the levels of vascular endothelial growth factor (VEGF) and matrix metalloproteinases (MMPs) induced by thrombin in endometrial stromal cells (ESC). DESIGN: Evaluation of the effects of thrombin, thrombin receptor activator peptide 6 (TRAP-6), and D-phenylalanyl-1-propyl-L arginine chloromethyl ketone (PPACK) on the production of VEGF and MMPs by ESC. SETTING: Research laboratory at the Oita University Medical School. PATIENT(S): Eight endometrial specimens in the secretory phase. INTERVENTION(S): ESC were incubated for 24 hours with thrombin, TRAP-6, and PPACK. MAIN OUTCOME MEASURE(S): The levels of VEGF, MMP-1, and active MMP-2 were measured by enzyme-linked immunosorbent assay (ELISA). The presence of protease-activated receptor-1 (PAR-1) and activation of mitogen-activated protein (MAP) kinase were detected by Western blot analysis. RESULT(S): Following stimulation by thrombin and TRP-6, the production of VEGF, MMP-1, and active MMP-2 statistically significantly increased; U0126 and PPACK statistically significantly suppressed the increases in the production of VEGF, MMP-1, and active MMP-2 induced by thrombin and TRAP-6. Activity by MAP kinase was induced by treatment with thrombin and TRAP-6 and was suppressed by PPACK. CONCLUSION(S): The results suggest that thrombin stimulates the production of VEGF and MMPs by a mechanism involving the MAP kinase system. The increases in VEGF and MMPs may contribute to neovascularization, which promotes the proliferation of endometrium and placentation.

The effects of platelet-activating factor on the secretion of interleukin-8 and growth-regulated oncogene alpha in human immortalized granulosa cell line (GC1a).

PROBLEM: To investigate the role of platelet-activating factor (PAF) in human ovulation, we studied the regulation of interleukin (IL)-8 and growth-regulated oncogene (GRO) alpha in cultured human immortalized granulosa cell line (GC1a). METHOD OF STUDY: GC1a was cultured in serum-free medium, and incubated with carbamyl-PAF (C-PAF) and/or PAF receptor antagonist (WEB 2086). The supernatants were collected, and IL-8 and GRO alpha were measured by enzyme-linked immunosorbent assay. RESULTS: After treatment with C-PAF, the levels of IL-8 and GROalpha increased in a time-dependent manner. The levels of IL-8 and GROalpha were significantly increased after treatment with C-PAF in a dose-dependent manner. However, the levels of IL-8 and GROalpha were significantly decreased by treatment with C-PAF and with increasing concentrations of WEB 2086. CONCLUSION: Our data indicated that IL-8 and GROalpha were regulated by C-PAF. The results suggested that PAF may play an important role in human pre-ovulatory processes involving IL-8 and GROalpha production.

Granulosa cell tumor associated with secondary amenorrhea and serum luteinizing hormone elevation.

Adult granulosa cell tumors (GCTs) are the most common type of ovarian sex cord tumors. Menstrual irregularity, menorrhagia, or even secondary amenorrhea is frequently observed in premenopausal women bearing GCTs with hormonal activity. We report herein a case of GCT in a patient presenting with secondary amenorrhea and serum luteinizing hormone elevation. A 28-year-old primigravid Japanese woman was admitted complaining of secondary amenorrhea of 2 years' duration. Pelvic examination, transvaginal ultrasonography, and magnetic resonance imaging demonstrated a left ovarian tumor 4 cm in diameter. Serum hormone assays revealed a follicle-stimulating hormone level of 4.8 mIU/ml, luteinizing hormone (LH) of 35.8 mIU/ml, estradiol of 24 pg/ml, progesterone of 1.6 ng/ml, and testosterone of 40 ng/dl. A left salpingo-oophorectomy was performed. The tumor was diagnosed as an adult-type GCT stage IIb (FIGO [International Federation of Obstetricians and Gynecologists], 1988). Spontaneous menstruation occurred soon after the surgery. Serum levels of LH also decreased to normal levels and showed cyclic changes during the menstrual cycle. Subsequently, the patient conceived and delivered a healthy female baby. The tumor recurred in the pelvis 50 months after the initial conservative surgery, with elevated serum LH levels of 36.0 mIU/ml and amenorrhea. The patient was treated by hysterectomy, right salpingo-oophorectomy, omentectomy, paraaortic and pelvic lymphadenectomy, and low anterior resection of the recto-sigmoid colon. Her hormone levels progressed to the postmenopausal state after this surgery. Although LH elevation in patients with GCT is rare and its mechanism is unknown, monitoring of serum LH may provide an additional tumor marker after conservative surgery in such patients.

K252a inhibits proliferation of ovarian cancer cells by upregulating p21WAF1.

The furanosylated indolocarbazole, K252a, belongs to a family of microbial alkaloids that also includes staurosporine, which is known to inhibit proliferation, stimulate apoptosis and induce cell cycle arrest of cancer cells. To elucidate the involvement of K252a in ovarian cancer, we investigated the effects of K252a on the ovarian cancer cell line, SK-OV-3. SK-OV-3 cells were treated with K252a, and its effect on cell growth, cell cycle, and related measurements was assessed. MTT assays showed that the ovarian cancer cell line SK-OV-3 cells were sensitive to the growth inhibitory effect of K252a. Cell cycle analysis indicated that their exposure to K252a decreased the proportion of cells in the S-phase and increased the proportion of cells in the G0/G1 phase of the cell cycle. This occurred in concert with altered expression of p21WAF1 protein related to the G0/G1 phase of the cell cycle. These results raise the possibility that K252a may prove particularly effective in treatment of ovarian cancer.

Expression and regulation of thrombospondin-1 by human endometrial stromal cells.

Thrombospondin-1 (TSP-1) production was modulated by EGF, IFN-gamma, and in vitro decidualization. It is suggested that TSP-1 may contribute to the regulation of neovascularization in the endometrium and gestational tissues.

Endometriotic cells are resistant to interferon-gamma-induced cell growth inhibition and apoptosis: a possible mechanism involved in the pathogenesis of endometriosis.

In order to evaluate the involvement of cell proliferation and apoptosis in the pathogenesis of endometriosis, we investigated the effects of interferon-gamma (IFN-gamma) on cell growth inhibition and apoptosis of cultured ovarian endometriotic cyst stromal cells (ECSC), eutopic endometrial stromal cells with endometriosis (ESCwE) and normal endometrial stromal cells (NESC) by modified methylthiazoletetrazolium assay, 5-bromo-2'-deoxyuridine incorporation assay and internucleosomal DNA fragmentation assay. The expression of apoptosis-related molecules and IFN-gamma receptor 1 was also examined in ECSC, ESCwE and NESC using western blot analysis. IFN-gamma significantly inhibited cell proliferation and DNA synthesis of ESCwE and NESC, and induced apoptosis of these cells. In contrast, IFN-gamma did not show apparent effects on the viable cell number, DNA synthesis, or apoptosis of ECSC. An up-regulated expression of Bcl-2 and Bcl-X(L) proteins was observed in ECSC in comparison with ESCwE and NESC, whereas the levels of Bax, Bad, Fas and Fas ligand proteins in ECSC were similar to those in ESCwE and NESC. IFN-gamma receptor 1 expression was detected in ECSC, ESCwE and NESC. Enhanced expression of anti-apoptotic molecules in the ectopic endometrial cells may contribute to the development of endometriosis by conferring resistance to cytokine-induced apoptosis and increasing the chance that these cells will survive and implant outside the uterus. Further investigations on the regulation of cell proliferation in both the endometriotic and the normal endometrium may be important for the elucidation of the pathogenesis of endometriosis.

Cultured human endometrial epithelial cells produce thymus and activation-regulated chemokine with stimulation of interleukin-4 and interleukin-13.

OBJECTIVE: To evaluate the effects of T-helper (Th)1 and Th2 cytokines on the production of thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC) by cultured endometrial epithelial cells (EEC) and endometrial stromal cells (ESC). DESIGN: The effects of interleukin (IL)-2, IL-4, IL-5, IL-10, IL-12, IL-13, interferon-gamma (IFN-gamma), and tumor necrosis factor-beta (TNF-beta) on the production of TARC and MDC were investigated. SETTING: Research laboratory at a medical school. PATIENT(S): Fifteen endometrial specimens in the mid-late secretory phase were used. INTERVENTION(S): The EEC and ESC were incubated for 24 hours with recombinant human IL-2, IL-4, IL-5, IL-10, IL-12, IL-13, IFN-gamma, and TNF-beta. MAIN OUTCOME MEASURE(S): The concentrations of TARC and MDC in the culture media were measured using ELISA. RESULT(S): Small amounts of TARC were detected in the culture medium of nonstimulated EEC. The increase in levels of TARC in the culture media of EEC paralleled the addition of increasing amounts of IL-4 and IL-13. Other cytokines, however, did not affect the production of TARC by EEC. Production of TARC by ESC was not detected under either nonstimulated or cytokine-stimulated conditions. Production of MDC was not detected in the culture media of EEC and ESC. CONCLUSION(S): These results suggest that IL-4 and IL-13 secreted from the embryo during the implantation period may selectively up-regulate the production of TARC by EEC. The controlled production of TARC in the endometrium may contribute to the modulation of the immune reaction by the regulation of Th2 lymphocyte trafficking and functions.

Synergistic effect of interleukin (IL)-1alpha and ceramide analogue on the production of IL-6, IL-8, and macrophage colony-stimulating factor by endometrial stromal cells.

OBJECTIVE: To measure the level of interleukin 6 (IL-6), IL-8, and macrophage colony-stimulating factor (M-CSF) induced by IL-1alpha in endometrial stromal cells (ESC) following treatment with ceramide analogues. DESIGN: The effects of IL-1alpha, IL-1 receptor antagonist (IL-1RA), C2-ceramide, and C6-ceramide on the production of IL-6, IL-8, and M-CSF by ESC. SETTING: Research laboratory at Oita University Medical School. PATIENT(S): Eleven premenopausal women who had undergone hysterectomies for subserous myoma provided endometrial specimens in the secretory phase. INTERVENTION(S): The ESC were incubated for 24 hours with IL-1alpha, IL-1RA, C2-ceramide, and C6-ceramide. MAIN OUTCOME MEASURE(S): The levels of IL-6, IL-8, and M-CSF in the culture media were measured via enzyme-linked immunoabsorbent assay. RESULT(S): : Following stimulation by IL-1alpha, the production of IL-6, IL-8, and M-CSF showed a statistically significant increase, and they were suppressed by IL-1RA in a dose-dependent manner. Production of IL-6, IL-8, and M-CSF was not statistically significantly increased by IL-1alpha plus C2-ceramide as compared with IL-1alpha alone. Production of both IL-8 and M-CSF was statistically significantly increased by IL-1alpha plus C6-ceramide as compared with IL-1alpha alone; however, IL-6 production was not increased. CONCLUSION(S): The results suggest that IL-1alpha stimulates the production of IL-8 and M-CSF by a mechanism that involves the sphingomyelin-ceramide system. Ceramide may be important in increasing the production of IL-8 and M-CSF in the human endometrium.

Production of macrophage inflammatory protein-3alpha in human follicular fluid and cultured granulosa cells.

OBJECTIVE: To investigate the role of macrophage inflammatory protein (MIP)-3alpha in human ovulation. DESIGN: Study of the levels of MIP-3alpha in serum and follicular fluid. The effects of interleukin (IL)-1alpha, IL-1 receptor antagonist (RA), and tumor necrosis factor (TNF)-alpha on the secretion of MIP-3alpha by primary cultured granulosa-lutein cells and an immortalized granulosa cell line (GC1a) were investigated. SETTING: Research laboratory at a university medical school. PATIENT(S): Forty-six patients with sterility undergoing in vitro fertilization and embryo transfer (i.v.f.-ET). INTERVENTION(S): Follicular fluid was obtained from study participants, and granulosa-lutein cells and GC1a were incubated with IL-1alpha, IL-1RA, or TNF-alpha for 4 to 32 hours. MAIN OUTCOME MEASURE(S): The concentration of MIP-3alpha in human follicular fluid was measured and correlated with oocyte maturation. We also cultured granulosa cells and examined the regulation of MIP-3alpha production. The concentrations of MIP-3alpha in the serum, follicular fluid, and culture medium were measured using enzyme-linked immunoabsorbent assay. RESULT(S): Concentrations of MIP-3alpha were significantly higher in the follicular fluid, but it was not detected in the serum. Concentrations of MIP-3alpha were statistically significantly higher in the follicular fluid containing mature oocytes than in follicular fluid containing immature oocytes. The production of MIP-3alpha was markedly increased over the basal level after treatment with IL-1alpha and TNF-alpha in a dose-dependent manner. The stimulatory effect of IL-1alpha was inhibited by IL-1RA. CONCLUSION(S): Our data suggest that MIP-3alpha was present in follicular fluid and correlated with oocyte maturation, and was regulated by IL-1alpha and TNF-alpha. Thus, MIP-3alpha may play an important role in the human preovulatory process.

Down-regulation of interleukin-1 receptor type 1 expression causes the dysregulated expression of CXC chemokines in endometriotic stromal cells: a possible mechanism for the altered immunological functions in endometriosis.

To evaluate the involvement of chemokines in the pathogenesis of endometriosis, we investigated the expression of CXC chemokines in cultured ovarian endometriotic cyst stromal cells (ECSC), endometrial stromal cells with endometriosis (ESCwE), and normal endometrial stromal cells (NESC). Using ELISA, TNF-alpha significantly enhanced the production of IL-8, growth-related oncogene alpha, and epithelial neutrophil-activating peptide-78 in all cases of ECSC (n = 10), ESCwE (n = 6), and, NESC (n = 10). IL-1beta did not affect the production of these chemokines in eight of 10 cases of ECSC. In contrast, IL-1beta significantly enhanced the expression of these chemokines in all cases of ESCwE (n = 6) and NESC (n = 10). Western blot analysis revealed down-regulation of expression of IL-1 receptor type 1 (IL-1-R1) in all cases of ECSC with low response to IL-1beta (n = 8). In contrast, significant IL-1-R1 expression was detected in all cases of NESC. Although IL-1-R1 expression was detected in all cases of ESCwE (n = 6), its expression in ESCwE tended to decrease compared with that in NESC. Moreover, phosphorylation of inhibitor kappaB-alpha was detected in all cases of ESCwE and NESC after stimulation with IL-1beta, but not in ECSC with low response to IL-1beta (n = 8). In contrast, significant IL-1-R2 expression was detected in all cases of ECSC, ESCwE, and NESC. The present findings suggest that the dysregulation of IL-1/IL-1-R system relates to immunological dysfunction in endometriosis. The alteration of the CXC chemokines expression may be important for elucidation of the pathogenesis of endometriosis.

Differential effects of interferon-alpha and -beta on the secretion of vascular endothelial growth factor by eutopic and ectopic endometrial stromal cells.

Interferon (IFN)-beta, not IFN-alpha, suppresses vascular endothelial growth factor secretion in endometriotic cyst stromal cells. It was suggested that IFN-beta may be more suitable than IFN-alpha as an anti-endometriotic agent.