Detailed multigrade evaluation of fetal disorders with the quantified actocardiogram.

AIMS: To evaluate fetal disorders using detailed quantitative values from the actocardiogram (ACG) involving simultaneous tracing of ultrasonic Doppler fetal movement bursts and fetal heart rate (FHR). METHODS: Duration of FHR accelerations and fetal movement bursts were measured manually in 20 common fetal disorders. The severity of the fetal disorder was estimated using the FHR acceleration duration to movement burst ratio (A/B ratio) and 10-0 clinical severity ranks derived from the A/B ratio. The correlation of the A/B ratio and 1 and 5 min Apgar scores, as well as numerically expressed long-term outcomes were studied. RESULTS: A/B ratios were significantly correlated with the 1 and 5 min Apgar scores and the numerically evaluated long-term outcomes. Controversial cases of FHR pattern were more easily understood using the A/B ratio. The 10-0 severity derived from the A/B ratio was useful in clinical fetal studies. CONCLUSION: Common fetal disorders were evaluated quantitatively and in more detail using the A/B ratio from the actocardiogram than when using common binary good or bad evaluation. The A/B ratio was useful in outcome estimation, where the prognostic capability of the A/B ratio was confirmed by significant correlation with 1 and 5 min Apgar scores and long-term outcomes of fetal disorders.

Involvement of p38 MAP kinase in lipopolysaccharide-induced production of pro- and anti-inflammatory cytokines and prostaglandin E(2) in human choriodecidua.

Chorioamnionitis is implicated in the pathogenesis of preterm delivery. However, the detailed mechanisms by which infection induces preterm labor are not well understood. This study has assessed the involvement of mitogen-activated protein (MAP) kinases in lipopolysaccharide (LPS)-induced pro- and anti-inflammatory cytokine and prostaglandin (PG) production in human choriodecidua. Samples of choriodecidua were collected before the onset of labor from women undergoing elective cesarean sections at term for breech presentation, previous cesarean delivery or cephalopelvic disproportion. Concentrations of TNFalpha, IL-10, PGE(2) and PGF(2)alpha in culture supernatants were measured by ELISA. Expression of COX-2 protein was analyzed by Western blotting. In human choriodecidual explants, LPS induced TNFalpha and IL-10 production in a dose- and time-dependent manner. LPS also up-regulated COX-2 expression and PG synthesis. Phosphorylations of extracellular signal-regulated kinase (ERK) 1/2, p38 MAPK, and c-Jun N-terminal kinases (JNK) were also confirmed by Western blotting. Furthermore, the effect of MAPK inhibitors was examined on LPS-induced pro- and anti-inflammatory cytokines and PG synthesis. Among the MAPK inhibitors examined, the p38 MAPK inhibitor, SB202190, significantly suppressed LPS-induced cytokine and PG production. SB202190 most profoundly suppressed the TNFalpha to IL-10 ratio, demonstrating that p38 MAPK inhibitor reduced predominantly TNFalpha other than IL-10 production. Phospho-p38 MAPK immunostaining was intense in extravillous trophoblast cells. The p38 MAPK seems to be most involved in signaling mechanisms when infection and inflammation cause preterm labor through PG synthesis. Novel therapeutic modalities targeting p38 MAPK may prevent to arrest preterm labor.

Macrophage-activating lipopeptide-2 induces cyclooxygenase-2 and prostaglandin E(2) via toll-like receptor 2 in human placental trophoblast cells.

We have examined whether toll-like receptor (TLR)2-mediated stimulation by macrophage-activating lipopeptide-2 (MALP-2), originally purified from Mycoplasma fermentans, induces cyclooxygenase (COX)-2 and prostaglandin (PG)E(2) in human placental trophoblast cells. The signaling mechanism by which MALP-2 exerts its effect has also been examined. Human placental trophoblast cells isolated from term placenta were used. TLR expression in trophoblast cells was confirmed by multiplex PCR and immunocytochemistry, and examined whether MALP-2 induces COX-2 and PGE(2) by Northern blotting, RT-PCR, Western blotting and ELISA, respectively. The activation of NF-kappaB and MAP kinases (ERK1/2 and p38) was examined by Western blotting. The effects of inhibitors of NF-kappaB, MEK1/2 and p38 on MALP-2-induced PGE(2) production were also evaluated. TLR2, TLR6 and TLR4 were expressed in human placental trophoblast cells. MALP-2 significantly induced COX-2 expression and enhanced PGE(2) production in a dose-dependent manner. MALP-2 induced the activation of NF-kappaB, ERK1/2 and p38 MAPK. Inhibitors of NF-kappaB, MEK1/2 and p38 blocked MALP-2-inducible PGE(2) production. TLR2-mediated stimulation by MALP-2 induces COX-2 and PGE(2) in human placental trophoblast cells via NF-kappaB and MAP kinases pathways.

Cervical Ureaplasma urealyticum colonization might be associated with increased incidence of preterm delivery in pregnant women without prophlogistic microorganisms on routine examination.

AIM: We examined whether the detection of Ureaplasma urealyticum DNA in the cervix is associated with preterm labor and delivery. METHODS: Eighty-two women (23 preterm labor cases and 59 controls) with no evidence of prophlogistic microorganisms on routine microbiologic examination were enrolled for this study. U. urealyticum colonization was examined using polymerase chain reaction of cervical swabs. RESULTS: The positivity rate of U. urealyticum DNA in preterm labor cases was significantly higher than that in the controls (87.0%vs 45.8%, P=0.0007). Women in the U. urealyticum-positive group more frequently delivered preterm compared with those in the negative group (36.2%vs 11.4%, P=0.0111). In five cases that delivered preterm with histologically confirmed chorioamnionitis, U. urealyticum DNA was detected in the placenta. CONCLUSIONS: Cervical U. urealyticum colonization might be associated with preterm labor and delivery.

Tumor necrosis factor-alpha induced the release of interleukin-6 from endometriotic stromal cells by the nuclear factor-kappaB and mitogen-activated protein kinase pathways.

OBJECTIVE: To examine the involvement of nuclear factor-kappaB (NF-kappaB) and mitogen-activated protein kinase (MAPK) in the induction of interleukin-6 (IL-6) by tumor necrosis factor-alpha (TNF-alpha) in endometriotic stromal cells (ESC). DESIGN: Prospective study. SETTING: Department of Obstetrics and Gynecology, Tottori University Hospital, Yonago, Japan. PATIENT(S): Twelve patients who underwent laparoscopic surgery. INTERVENTION(S): Endometriotic stromal cells were obtained from chocolate cyst linings of the ovary. MAIN OUTCOME MEASURE(S): We determined the effect of TNF-alpha on the production of IL-6 and the effect of inhibitors for NF-kappaB and the MAPK pathway on IL-6 production using ELISA. Western blottings and electrophoretic mobility shift assays were used to detect activation of NF-kappaB and extracellular signal-regulated kinase 1/2 (ERK1/2). RESULT(S): The addition of TNF-alpha (0.1 ng/mL) significantly increased IL-6 protein in endometriotic stromal cells. Western blottings and electrophoretic mobility shift assays revealed that incubation with TNF-alpha induced degradation of inhibitor kappaB (I kappaB) and expression of phosphorylated ERK1/2. The NF-kappaB inhibitor (TPCK) and MAPK inhibitor (U0126) blocked the TNF-alpha-induced IL-6 expression. Electrophoretic mobility shift assay revealed that U0126 attenuated activator protein-1 (AP-1) activation induced by TNF-alpha. CONCLUSION(S): These findings demonstrate that NF-kappaB and AP-1 activation is critical for TNF-alpha-induced IL-6 expression in endometriotic stromal cells. Novel therapeutic modalities targeting these molecules may be possible in the near future.

Interleukin-1beta-induced expression of IL-6 and production of human chorionic gonadotropin in human trophoblast cells via nuclear factor-kappaB activation.

PROBLEM: The cytokine, interleukin-6 (IL-6), stimulates the production of human chorionic gonadotropine (hCG) in chorionic cells. The purpose of this study was to examine the role of nuclear factor-kappaB (NF-kappaB) during the induction of IL-6 by IL-1beta in human trophoblast cells. METHOD OF STUDY: Reverse transcription-polymerase chain reaction was used to determine the gene expressions of IL-1, IL-1 receptor (IL-1R), IL-6R and gp130 in a human choriocarcinoma cell line, BeWo. The BeWo cells were cultured for 24 hr with IL-1beta (0-10 ng/mL), IL-1beta (10 ng/mL) together with IL-1 receptor antagonist (IL-1Ra) or NF-kappaB inhibitor, N-tocyl-l-phenylalanine chloromethyl ketone (TPCK). The concentrations of IL-6 in culture supernatants were measured by enzyme-linked immunosorbent assay. RESULTS: The gene expressions of IL-1R, IL-6R and gp130, but not IL-1beta, were observed. The concentration of IL-6 was enhanced by IL-1beta in a dose-dependent fashion. The addition of IL-1Ra neutralized the effects of IL-1beta on IL-6 secretion. IL-1beta induced expression of phosphorylated IkappaB and the activation of NF-kappaB. The addition of TPCK reduced the IL-1beta-induced IL-6 expression. Adding IL-1beta increased beta-hCG production in a dose-dependent manner, and the addition of TPCK neutralized the effect of IL-1beta. CONCLUSION: IL-1beta may induce the IL-6 expression and hCG production in human BeWo cells via NF-kappaB.

A combination of interleukin-6 and its soluble receptor impairs sperm motility: implications in infertility associated with endometriosis.

BACKGROUND: We previously reported that the level of interleukin (IL)-6 is increased in the peritoneal fluid of women with endometriosis. This study was undertaken to assess the effects of IL-6 and soluble IL-6 receptor (sIL-6R) on in vitro sperm motility. METHODS: Sperm (n = 20) were cultured with IL-6 or sIL-6R, or with a combination of both. After 24 h cultures, sperm motility was evaluated using a computer-assisted semen analysis system. Gene and protein expressions of IL-6, IL-6 receptor (IL-6R), and glycoprotein 130 (gp130) were examined in sperm by RT-PCR analysis and western blot analysis. RESULTS: Addition of IL-6 or sIL-6R individually to the culture media had no affect on sperm motion. However, adding a combination of IL-6 and sIL-6R dose-dependently reduced the percentage of motile and rapidly moving sperm. Adding anti-IL-6R antibody abolished these adverse effects. Sperm expressed the gp130 gene and protein, but not IL-6 or IL-6R. CONCLUSIONS: A combination of IL-6 and sIL-6R may be associated with gp130 expressed in the sperm and reduce sperm motility. IL-6 and sIL-6R may contribute to the pathogenesis of endometriosis-associated infertility.

Use of the LAP DISK (abdominal wall sealing device) in laparoscopically assisted myomectomy.

OBJECTIVE: To evaluate the efficacy of an abdominal wall sealing device (the LAP DISK) used during laparoscopically assisted myomectomy (LAM). DESIGN: Retrospective study. SETTING: Tottori University Hospital, Yonago, Japan. PATIENT(S): All 43 patients who underwent LAM using the LAP DISK. INTERVENTION(S): Ultrasonography and magnetic resonance imaging. MAIN OUTCOME MEASURE(S): Treatment strategy, operative outcome, and postoperative pregnancy rate. RESULT(S): Weight and size of the myomas removed ranged from 40-700 g (mean: 208.0 g) and 2-10 cm (mean: 5.4 cm). Mean blood loss was 42.3 mL. Half of the 18 patients who had been diagnosed with primary infertility for >2 years became pregnant without postoperative assisted reproductive techniques. CONCLUSION(S): The LAP DISK, a useful device for LAM, allows surgeons to remove myomas safely and repair uterine defects effectively while minimizing blood loss and trauma.

Hepatocyte growth factor/Met system promotes endometrial and endometriotic stromal cell invasion via autocrine and paracrine pathways.

Endometrial stromal cells reportedly have a role in the initial invasion of endometrial tissue into the peritoneum. Hepatocyte growth factor (HGF), which is a ligand for the c-met protooncogene product (Met), stimulates proliferation and invasion of a large number of cells. In this study we investigated the role of the HGF/Met system in the pathogenesis of endometriosis. HGF concentrations in the peritoneal fluid of patients with endometriosis were significantly higher than in those without endometriosis and correlated positively with revised American Society of Reproductive Medicine scores. We showed that the peritoneum and endometriotic stromal cells may be major sources of HGF in peritoneal fluid. Endometrial and endometriotic stromal cells expressed the Met receptor, which was activated by endogenous and exogenous HGF. HGF enhanced stromal cell proliferation and invasion. We also demonstrated that the HGF-stimulated stromal cell invasion was due in part to the induction of urokinase-type plasminogen activator, a member of the extracellular proteolysis system. In conclusion, the HGF/Met system is involved in the pathogenesis of endometriosis by promoting stromal cell proliferation and invasion of shed endometria and endometrial lesions via autocrine and paracrine pathways.

Tumor necrosis factor-alpha-induced interleukin-8 (IL-8) expression in endometriotic stromal cells, probably through nuclear factor-kappa B activation: gonadotropin-releasing hormone agonist treatment reduced IL-8 expression.

Endometriosis, a common disease among women of reproductive age, is characterized by the presence of endometrial-like tissue outside the uterus. We previously reported that TNFalpha promoted proliferation of endometriotic stromal cells by inducing IL-8 gene and protein expression. We hypothesize that TNFalpha may induce IL-8 production in endometriotic cells through nuclear factor-kappa B (NF-kappa B) activation. Western blot analyses and electrophoretic mobility shift assays revealed that incubation with TNF alpha induced the expression of phosphorylated inhibitor kappa B (p-I kappa B) and activation of NF-kappa B in endometriotic stromal cells. The NF-kappa B inhibitor, N-tosyl-L-phenylalanine chloromethyl ketone, reduced TNFalpha-induced IL-8 gene and protein expression. The medical treatment of endometriosis with GnRH agonist (GnRHa) has been shown to induce hypoestrogenemia and reduce the observable number of endometriotic implants. We compare the expression of IL-8 gene and protein in endometriotic stromal cells of patients treated with GnRHa and those of patients without treatment before laparoscopic cystectomy for endometrioma. The addition of TNFalpha (0.1 ng/ml) significantly increased protein and gene expression of IL-8 in the cells of patients without GnRHa treatment, but this expression was not observed in the cells of patients with GnRHa. The addition of estradiol (E2; 10(-7) M) enhanced the expression of IL-8. However, in the cells of patients who received GnRHa treatment, TNFalpha and E2 did not show any significant effect. In endometriotic stromal cells without GnRHa treatment, TNFalpha and E2 increased the expression of p-I kappa B. In contrast, TNFalpha and E2 had no significant effect on the expression of p-I kappa B in cells that received GnRHa treatment. These findings demonstrate that NF-kappa B activation is critical for TNFalpha-induced IL-8 expression in endometriotic stromal cells. The current study showed for the first time that GnRHa treatment attenuated the expression of IL-8 by reducing TNFalpha-induced NF-kappa B activation.

Activation of mitogen-activated protein kinase pathway by keratinocyte growth factor or fibroblast growth factor-10 promotes cell proliferation in human endometrial carcinoma cells.

Fibroblast growth factors (FGFs) exert diverse effects resulting from their interaction with cognate receptors on target cells. Our current study was designed to examine the local production and action of two specific stromal-epithelial cell mediatory factors, keratinocyte growth factor (KGF) and FGF-10, in human endometrial carcinoma cells. The RT-PCR method was used to determine gene expression of KGF, FGF-10, and KGF receptor in human endometrial carcinoma cells (HEC-1) and human endometrial stromal cells. KGF mRNAs were expressed in both of these cell types. On the other hand, FGF-10 mRNA was detected only in the endometrial stromal cells, and KGF receptor mRNA was observed in the HEC-1 cells. The novel finding of the present study is that KGF is expressed in carcinoma cells and FGF-10 is expressed in human endometrial stromal cells. The distinct phosphorylation of ERK-1 and -2 (ERK1/2), which are members of the MAPK family, was observed when HEC-1 cells were treated with KGF or FGF-10. KGF and FGF-10 could induce the prompt phosphorylation of ERK1/2 and consequently stimulate DNA synthesis. KGF and FGF-10 did not activate the phosphorylation of Akt, protein kinase C, or signal transducer and activator of transcription-3. Blocking the MAPK pathway with the specific methyl ethyl ketone 1/2 inhibitor (U0126) completely neutralized the enhancement of cell proliferation induced by KGF and FGF-10. In addition, KGF and FGF-10 activated expressions of downstream nuclear transcription factors, such as Elk-1 and c-myc, but not c-fos. These results demonstrate for the first time that KGF and FGF-10 are capable of stimulating the growth of endometrial carcinoma cells via activating MAPK pathway through autocrine/paracrine fashion.

Laparoscopic surgery for the management of ovarian endometrioma.

The purpose of this study was to evaluate the effects of laparoscopic surgery on the symptoms associated with ovarian endometrioma. We also examined serum IL-6 concentrations in patients with endometrioma. Ninety-two patients who underwent laparoscopic surgery for ovarian endometrioma were enrolled in this study. The mean duration of follow-up was 27.6 months. Transvaginal ultrasound examinations revealed a recurrence of endometrioma in 13% of the cases. We evaluated the severity of dysmenorrhea using a 0-3-point verbal rating scale, and found that the dysmenorrhea score was statistically improved after the operation. Follicular growth was preserved in 94%, and the pregnancy rate was 43%. We measured serum IL-6 concentrations in 14 patients with ovarian endometrioma and 4 patients with benign gynecologic disease without endometriosis. IL-6 was significantly higher in patients with endometrioma than in those without endometriosis at the time of diagnosis. The mean serum IL-6 concentration significantly decreased after the operation. In conclusion, laparoscopic surgery is effective for alleviating pain and preserving fertility in patients with endometrioma. Measurements of serum IL-6 concentrations may be useful for the management of endometrioma.

Induction of hepatocyte growth factor in stromal cells by tumor-derived basic fibroblast growth factor enhances growth and invasion of endometrial cancer.

Tumor progression is often regulated through interactions between carcinoma cells and host stromal cells. In this study of endometrial cancer, we investigated one mechanism potentially involved in hepatocyte growth factor (HGF)-mediated cancer-stromal interactions. Endometrial cancer cells (HEC-1 and ISHIKAWA) expressed the c-met receptor, but HGF did not. HGF, however, did stimulate the proliferation and invasion of these cells. The HGF gene was expressed in stromal cells, which had been separated from primary cultures of endometrial cancers, 6.4 times more than in isolated normal endometrial stromal cells. Immunohistochemical staining revealed immunoreactive HGF in cancer stromal cells, the staining intensity being more pronounced in cancer tissue than in normal endometrium. The conditioned medium from normal epithelial cells and cancer cell lines induced HGF production in normal stromal cells. We identified basic fibroblast growth factor as an HGF inducer derived from endometrial cancer cell lines. Basic fibroblast growth factor derived from tumor cells may induce HGF in endometrial stromal cells, whereas stromal cell-derived HGF leads to the invasive growth of carcinoma cells. These interactions, mediated by HGF and HGF inducers, may play a significant role in the progression of endometrial cancer.