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Endometriosis is a benign tumor that affect 6-10% women of reproductive age. To date, it is suggested that the aberrant microRNA (miRNA) expressions play important roles in the pathogenesis of endometriosis. Reviewing the literature, we found nine overexpressed miRNAs, which were thoroughly investigated in the context of endometriotic tissues and cells. Most of the overexpressed miRNAs induced endometriosis-specific characteristics including inhibition of apoptosis and decidualization, upregulation of fibrogenesis, invasion, migration, cell proliferation, attachment to extracellular matrix, inflammation, and angiogenesis in the endometriotic cells. Then, we found that the downstream target molecules of these miRNAs, such as early growth response protein-1, extracellular signal-regulated kinase, matrix metallopeptidase 1, signal transducer and activator of transcription 3, cyclooxygenase-2, phosphoinositide 3-kinase, AKT, mammalian target of rapamycin, and vascular endothelial growth factor-A are promising for the therapeutic targets of endometriosis. Recent findings suggest that complex molecular mechanisms leading to development and progression of endometriosis by miRNAs may exist in endometriosis. The meticulous balance between tumorigenic miRNAs and tumoristatic miRNAs may destine the natural course and response to the surgical, medical, and hormonal treatments of this disease. Further investigations into endometriosis-associated miRNAs may elucidate the pathogenesis of endometriosis and help to develop novel therapeutics.

Background To investigate the role of adenosine monophosphate (AMP)-activated protein kinase (AMPK) on the production of interleukin (IL)-8, monocyte chemoattractant protein (MCP)-1, prostaglandin E2 and F2α induced by IL-1β in endometrial stromal cells (ESCs) following treatment with 5-aminoimidazole-4- carboxamide ribonucleoside (AICAR). Methods Endometrial specimens were obtained and cultured. We examined the effects of IL-1β, IL-1 ra and AICAR on the production of IL-8, MCP-1, PGE2 and PGF2α in human ESCs. The phosphorylations of AMPK, IκB, 4EBP-1, p70S6K and S6 ribosomal protein were analyzed by Western immunoblotting. Results Following stimulation by IL-1β, the production of IL-8, MCP-1, PGE2 and PGF2α showed significant increases, and these increases were suppressed by AICAR. The expression of cyclooxygenase-2 (COX-2) induced by IL-1β and suppressed by AICAR. The phosphorylation of IκB, 4EBP-1, p70S6K and S6 ribosomal protein were inhibited via an AMPK-dependent signal transduction. Conclusions The production of IL-8, MCP-1, PGE2 and PGF2α induced by IL-1β in ESCs were involved in the negative regulatory mechanisms of AMPK. The substances that activate AMPK may be promising agents for the treatment of pathological problems such as dysmenorrhea.

Endometriosis is a common, estrogen-dependent benign tumor that affect 3–10% women of reproductive age, and is characterized by the ectopic growth of endometrial tissue, which is found primarily in the rectovaginal septum, ovaries, and pelvic peritoneum. To date, accumulating evidence suggests that various epigenetic aberrations, including the expression of aberrant microRNAs (miRNAs), play definite roles in the pathogenesis of endometriosis. This review summarizes the recent findings on the aberrantly repressed miRNAs, as well as their potential roles regarding the pathogenesis of endometriosis.

Background: Endometrial receptivity issues represent a potential source of implantaion failure. It is very important to investigate the expression of endometrial receptive markers in the endometrium during implantation. Therefore, we examined whether it would be possible to analyze endometrial receptivity using cells from embryo transfer catheters. In addition, we analyzed the relationship between the gene expression profile associated with pregnancy from endometrial cells taken during embryo transfer.Methods: A total of 88 cycles from 88 consenting patients were enrolled in this study. The tip of the embryo transfer (ET) catheter was cut and immersed in a dedicated reagent. Confirmation of cell distribution was carried out using a Papanicolaou stain (n=6) and immunocytochemistry (n=3). Protein expression was carried out by immunocytochemistry (n=12). Total RNA was extracted, and the expression of endometrial receptive markers (estrogen receptor α, progesterone receptor, and homeobox A10) were analyzed using quantitative reverse transcription polymerase chain reaction (n=67). we analyzed the relationship between the gene expression profile associated with pregnancy from endometrial cells. Results: Samples collected from the ET catheter showed clear staining for endometrial cells. Most of the cells were endometrial epithelial cells. Cervical cells were not included. The protein expression of endometrial receptive markers in cells was also confirmed. Three genes were analyzed that are associated with endometrial receptivity. Progesterone receptor expression was 1.5-fold and homeobox A10 expression was 2-fold higher in patients who became non-pregnant group, compared to the pregnant group. Both increases were statistically significant (p < 0.05). Estrogen receptor α expression tended to be higher in the non-pregnant group, but there was no significant difference. Conclusion: Our results suggest that endometrial receptivity can be evaluated using cells obtained from the ET catheter. This method may be useful for elucidating the cause of implantation failure by comparing a receptive and non-receptive endometrium at the time of ET.

Decidualization of the endometrium and endometriosis involves the morphological and biochemical reprogramming of the estrogen-primed proliferative stromal compartment under the continuing influence of progesterone. Here, we evaluated the involvement of microRNA in the decidualization processes of normal endometrial stromal cells (NESCs) and endometriotic cyst stromal cells (ECSCs). In vitro decidualization of NESCs and ECSCs was induced by long-term culture with a combination of 0.5 mmol/L of dibutyryl cyclic adenosine monophosphate and 100 nmol/L of dienogest. We investigated the effect of in vitro decidualization on the microRNA and messenger RNA (mRNA) expression profiles of the NESCs and ECSCs using global microarray techniques and an Ingenuity Pathways Analysis. Decidualization differentially enhanced the miR-30a-5p expression in the NESCs and the miR-210 expression in the ECSCs. The enhanced miR-30a-5p expression in the NESCs correlated with the increased mRNA expression of Krüppel-like factor 9 and period circadian clock 3 as well as the decreased mRNA expression of tolloid-like 1, tolloid-like 2, and paired-like homeodomain 1. The enhanced expression of miR-210 in the ECSCs correlated with the decreased mRNA expression of growth hormone receptor and thymidine kinase 1. Although there is no direct evidence, we speculate that the loss of miR-30a-5p-mediated mechanisms of decidualization and the acquisition of miR-210-mediated mechanisms of decidualization may be involved in the progesterone resistance in endometriosis. Further investigations are necessary to test this speculation.

Problem Decidual cells are thought to be involved in the maintenance of pregnancy. We conducted this study to evaluate the cellular function of endometrial stromal cells (ESCs) transitioning to decidualization. Methods of study Normal endometrial specimens were obtained from premenopausal patients who had undergone hysterectomies for subserosal leiomyomas. Decidualization of the ESCs (DSCs) was induced by incubating subconfluent cells in media containing medroxyprogesterone acetate and dibutyryl-cyclic adenosine monophosphate. We first analyzed the expression profile of protease-activated receptor-1 (PAR-1) between ESCs and DSCs. To investigate the intracellular signal transduction system in the DSCs, we incubated cells with thrombin receptor activator peptide 6 (TRAP-6). The levels of IL-8, monocyte chemo-attractant protein-1, matrix metalloproteinase (MMP)-1, and vascular endothelial growth factor in the culture medium were measured by enzyme-linked immunosorbent assays. The activation of the MAP kinase signaling pathway was detected by a Western blot analysis. The activation was evaluated for the expression of p21. Results PAR-1 receptor expression is upregulated in DSCs. The productions of chemokine and MMP-1 increased in the DSCs with the addition of TRAP-6. The activity of both the ERK-1 and ERK-2 isoforms was increased by 5-15 minute after TRAP-6 treatment. p70 S6 kinase showed the strongest expression after 1 hour. p21 was strongly observed in ESCs compared to the DSCs. Conclusion Our results suggest that cell function is changed by decidualization in association with increasing PAR-1 expression. The upregulation of PAR-1 may have some influence on pregnancy in the decidua.

Sclerosing stromal tumours (SSTs) are rare benign tumours and are generally observed in individuals in their teens to 20 s. Many cases are suspected as malignant tumours pre-operatively, owing to their high vascularity on diagnostic imaging. Herein, we aimed to evaluate the expressions of various angiogenic factors in SSTs. The expressions of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and hepatocyte growth factor (HGF) were examined by immunohistochemical staining. Upon immunohistochemistry, overexpressions of VEGF, bFGF, and HGF in the ovarian stroma were observed in SSTs. In the normal ovary, the expressions were strong around the vessels and weak in the stroma of the ovary, while a Sertoli-Leydig cell tumour showed weak staining with VEGF, locally strong staining with bFGF and negative staining with HGF. This is the first report about angiogenic factors such as bFGF, and HGF in SST. Impact statement What is already known on this subject? Sclerosing stromal tumour (SST) is a rare benign tumour with high vascularity. The high vascularity of SSTs are reported to be related to the overexpression of vascular endothelial growth factor (VEGF). But there is no study on the expressions of other angiogenic factors in SST. What do the results this study add? In the immunohistochemical analysis, VEGF, bFGF and HGF were found to be widely stained in SSTs. This results suggest the possibility that the high vascularity seen on diagnostic imaging, and the many small vessels observed microscopically may be related to the expressions of VEGF, bFGF and HGF in SSTs. What are the implications of these findings for clinical practice and/or further research? Our results suggest the possibility that the combined expression pattern of VEGF, bFGF, and HGF may be used as marker of SSTs.

Objective: Although uterine didelphys per se is not associated with an impaired ability to conceive, the association between uterine anomalies and gestational trophoblastic neoplasia (GTN) remains unclear. The management of chemotherapy-resistant GTN in women with uterine didelphys raises a new issue regarding whether to perform a hemi-hysterectomy. Case report: A 23-year-old, gravida 1, para 0 Japanese woman was referred with a failed intermittent cervical dilatation for hematometra. Four years previously, she developed a GTN Stage III, score 5. As two cycles of chemotherapy with methotrexate (MTX) and one cycle of EMA-CO (etoposide, MTX, actinomycin D, cyclophosphamide and vincristine) did not result in remission, we performed an abdominal hemi-hysterectomy. After a canalization procedure and cervicoplasty were performed, the patient conceived naturally and prematurely delivered by cesarean section twice. Conclusion: A hemi-hysterectomy should be considered for fertility preservation when GTN develops on either side of a didelphic uterus and adjuvant chemotherapy does not result in remission. Keywords: Adjuvant chemotherapy, Fertility preservation, Gestational trophoblastic disease, Hysterectomy, Anomalies

The function of granulosa cells is regulated by various hormones and growth factors. Our aim is to clarify the regulation of vascular endothelial growth factor (VEGF) production induced by heparin-binding EGF-like growth factor (HB-EGF) and amphiregulin (AR) in a human granulosa cell line, KGN. KGN cells were cultured and incubated for 24 h with HB-EGF and AR. The levels of VEGF in the culture media were measured by an enzyme-linked immunosorbent assay. The activation of MAP kinase in KGN cells was detected by Western blot analysis. VEGF production was significantly increased by HB-EGF or AR alone in a dose-dependent manner, whereas it was decreased by AG1478 or U0126. The MAP kinase activity was increased by treatment with HB-EGF or AR. The results suggested that VEGF is induced by HB-EGF and AR through mechanisms involving MAP kinase. The increase in VEGF may contribute to neovascularization, which in turn would promote various ovulation phenomena as well as follicular growth.

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.

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.

The aim of the present study was to evaluate the effects of interleukin (IL)-1beta on the production of vascular endothelial growth factor (VEGF) and soluble fms-like tyrosine kinase-1 (sFlt-1) in the human fallopian tube. Human oviductal epithelial cells (OECs) and oviductal stromal fibroblasts (OSFs) were isolated from ten premenopausal patients. The secretion of VEGF and sFlt-1 by cultured OECs and OSFs in response to IL-1beta and IL-1 receptor antagonist (IL-1RA) was measured using an enzyme-linked immunosorbent assay. The secretion of VEGF and sFlt-1 was detected in cultured OECs and OSFs under untreated conditions; secretion of these angiogenic modulators was significantly stimulated with IL-1beta administration in these cells. IL-1beta-induced production of VEGF and sFlt-1 by these cells was significantly inhibited by the addition of IL-1RA. The present findings suggest that IL-1beta in the local environment may stimulate oviductal vascular permeability by inducing the production of VEGF by oviductal cells via both autocrine and paracrine mechanisms. Simultaneous upregulation of sFlt-1 secretion by these cells after IL-1beta stimulation may prevent an excessive upregulation of vascular permeability. The modulation of the ratio of VEGF and sFlt-1 in the fallopian tube may contribute to the normal and pathological processes of oviductal fluid secretion by regulating oviductal vascular permeability during the menstrual cycle and during the peri-implantation period.

Objective To evaluate the involvement of transforming growth factor (TGF)-β isoforms (TGF-β1, TGF-β2, and TGF-β3) on endometrial tissue remodeling during the perimenstrual period. Design The effects of TGF-β isoforms on the cell proliferation, motility, and contractivity of cultured human endometrial stromal cells (ESCs) were investigated. Setting Research laboratory at a medical school. Patient(s) Nine endometrial specimens in the late secretory phase were used. Intervention(s) Endometrial stromal cells were incubated with recombinant human recombinant TGF-β1, TGF-β2, and TGF-β3. Main Outcome Measure(s) The cell proliferation, motility, and contractivity of ESCs were accessed by a modified methylthiazoletetrazolium assay, in vitro wound repair assay, transwell invasion assay, and collagen gel contraction assay. Result(s) All three isoforms of TGF-β significantly inhibited the cell proliferation of ESCs in a dose-dependent manner. In vitro wound repair assay and transwell invasion assay demonstrated that the TGF-β isoforms significantly inhibited the motility of ESCs. However, the TGF-β isoforms were shown to have a clear effect on the collagen gel contractivity of ESCs. Conclusion(s) These results suggest that TGF-β isoforms may promote endometrial tissue repair through the inhibition of the proliferation, expansion, and migration of ESCs, and through the stimulation of the contraction of the collagen gel matrix by these cells. Transforming growth factor-β may be involved in the protection of the endometrium from extensive fibrosis and scarring by regulating ESC function during the perimenstrual period.

Mucosal T cells are essential to immune tolerance in the intestine, an organ constantly exposed to large amounts of dietary and bacterial Ags. We investigated whether local fibroblasts affect mucosal T cell survival, which is critical for maintenance of immune tolerance. Coculture with autologous fibroblasts significantly increased viability of mucosal T cells by inhibiting IL-2 deprivation- and Fas-mediated apoptosis, an effect that was both contact- and secreted product-dependent. Investigation of antiapoptotic factors in the fibroblast-conditioned medium (FCM) revealed the presence of IL-10 and PGE2, but not IFN-β, IL-2, or IL-15. Although recombinant IFN-β, but not PGE2, effectively prevented T cell apoptosis, neutralizing Ab studies showed that only IL-10 blockade significantly increased T cells apoptosis, whereas neutralizing IFN-β or IFN-α failed to inhibit the antiapoptotic effect of FCM. To confirm that fibroblast-derived IL-10 was responsible for preserving mucosal T cell viability, IL-10 mRNA was demonstrated in fibroblasts by Southern blotting and RT-PCR. When FCM was submitted to HPLC fractionation, only the peak matching rIL-10 contained the antiapoptotic activity, and this was eliminated by treatment with an IL-10-neutralizing Ab. Finally, when fibroblasts were transiently transfected with IL-10 antisense oligonucleotides, the conditioned medium lost its T cell antiapoptotic effect, whereas medium from fibroblasts transfected with IFN-β antisense oligonucleotides displayed the same antiapoptotic activity of medium from untransfected fibroblasts. These results indicate that local fibroblast-derived IL-10 is critically involved in the survival of mucosal T cells, underscoring the crucial importance of studying organ-specific cells and products to define the mechanisms of immune homeostasis in specialized tissue microenvironments like the intestinal mucosa.

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.

Objective To measure the level of interleukin 6 (IL-6), IL-8, and macrophage colony-stimulating factor (M-CSF) induced by IL-1α in endometrial stromal cells (ESC) following treatment with ceramide analogues. Design The effects of IL-1α, 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-1α, 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-1α, 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-1α plus C2-ceramide as compared with IL-1α alone. Production of both IL-8 and M-CSF was statistically significantly increased by IL-1α plus C6-ceramide as compared with IL-1α alone; however, IL-6 production was not increased. Conclusion(s) The results suggest that IL-1α 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.

To clarify the role of platelet-activating factor (PAF) in parturition, the effects of PAF on the secretion of PAF-acetylhydrolase (PAF-AH), a PAF-inactivating enzyme, by decidual macrophage populations were examined. The cells were isolated from human decidual tissue by enzymatic digestion, Ficoll-Paque centrifugation, or flow cytometric sorting. The nonhydrolyzable agonist of PAF, carbamyl-PAF (C-PAF), inhibited the secretion of PAF-AH by either decidual cells or flow cytometrically purified decidual macrophages. A specific PAF receptor antagonist, WEB 2086, blocked the C-PAF-induced inhibition. Lyso-PAF, a metabolite of PAF, had no effect on the enzyme secretion. An intracellular calcium channel blocker, bis-(o-aminophenoxy)-ethane-N,N,N′,N′-tetraacetic acid, tetra(acetoxymethyl)-ester, partially blocked the inhibition by C-PAF, whereas extracellular calcium channel blockers, nifedipine and verapamil, were without effect. The inhibitory effect of C-PAF was also partially blocked by protein kinase C (PKC) inhibitors, sphingosine and H-7. A PKC activator, 12-O-tetradecanoylphobol 13-acetate, decreased the secretion of PAF-AH. The decrease was abolished by the addition of sphingosine and H-7. It is suggested that PAF inhibits the PAF-AH secretion by decidual macrophages and that the inhibitory action is mediated by a signal transduction mechanism involving intracellular calcium and PKC.

Objective To evaluate the effects of leptin on the production of interleukin (IL)-6 family cytokines and chemokines by human endometrial stromal cells (ESC) and epithelial cells. Design The effects of leptin on the production of IL-6, IL-11, leukemia inhibitory factor (LIF), IL-8, growth-regulated oncogene (GRO)-α, monocyte chemoattractant protein (MCP)-1, and macrophage inflammatory protein (MIP)-3α by ESC and the endometrial epithelial cell line HHUA were investigated. Setting Research laboratory at a medical university. Patient(s) Eight endometrial specimens in the late proliferative phase were used for the isolation of ESC. Intervention(s) ESC and HHUA were incubated for 24 hours with recombinant human leptin. Main outcome measure(s) The concentration of IL-6, IL-11, LIF, IL-8, GRO-α, MCP-1, and MIP-3α were measured using ELISAs. Result(s) Unstimulated ESC and HHUA constitutively secreted IL-6, IL-11, LIF, IL-8, GRO-α, MCP-1, and MIP-3α. The increase in levels of IL-6, IL-8, GRO-α, MCP-1, and MIP-3α in the culture media of ESC and HHUA paralleled the addition of increasing amounts of leptin. In contrast, the levels of IL-11 and LIF were not affected by leptin administration. Conclusion(s) The present findings suggest that leptin may be an additional modulator of IL-6 and chemokine expression in the endometrium. Leptin may contribute to the normal and pathological processes of human reproduction by the regulation of these cytokines in the local environment.

Objective: The aim is to clarify the role of platelet-activating factor and colony-stimulating factors in term and preterm parturition. Study Design: Decidual macrophage populations were obtained by an enzymic digestion, Ficoll-Paque centrifugation, or flow cytometric sorting. The effects of colony-stimulating factors on the platelet-activating factor acetylhydrolase secretion by these cells were examined. Results: Granulocyte-macrophage colony-stimulating factor inhibited the platelet-activating factor acetylhydrolase secretion by decidual macrophages. Granulocyte colony-stimulating factor also decreased the enzyme secretion but at higher concentrations than those required for granulocyte colony-stimulating factor. In contrast, macrophage colony-stimulating factor increased the enzyme secretion. These colony-stimulating factor-induced effects were specifically neutralized by the corresponding antibodies. Conclusion: Colony-stimulating factors may modulate the local concentration of platelet-activating factor in the decidua via their inhibitory or stimulatory effect on the secretion of platelet-activating factor acetylhydrolase, contributing to the regulation of term and preterm parturition at the maternal-fetal interface. (Am J Obstet Gynecol 2003;188:157-61.)

Neonatal toxic shock syndrome-like exanthematous disease (NTED) is a new entity of methicillin-resistant Staphylococcus aureus (MRSA) infection. Most of NTED cases reported previously in the literature were sporadic ones. In the present report, we describe an outbreak of NTED that occurred in a neonatal intensive care unit (NICU) between April, 1999 and April, 2000 in Japan. All MRSA strains isolated from 14 patients (6 NTED, 2 infections and 6 colonizations) in this outbreak belonged to the group of coagulase II and produced toxic shock syndrome toxin 1 (TSST-1). Of these, 14 strains produced staphylococcal enterotoxin C (SEC). No other superantigenic toxins were produced by these strains. The pulsed field gel electrophoresis (PFGE) patterns of genomic DNA digested with SmaI were indistinguishable each other due to no band shifting in all of the 13 strains except for strain O-21 and M56. Strain M56 was different from the dominant type in the positions of only 2 bands, whereas the pattern of strain O-21 had no similarity with the other pattern, suggesting that this outbreak was associated with the spread of a unique MRSA strain in the NICU. Two-dimensional electrophoresis (2-DE) analysis of exoproteins revealed that the patterns of these 14 strains were very indistinguishable to each other, and that these strains produced very large amounts of TSST-1 and SEC3 subtype superantigens, as measured with computer-assisted image analysis of the intensity of 2-DE spots. The 2-DE gel of O-21 showed the different pattern from the others. These results as well as the profiles of toxin production also supported the conclusion drawn from PFGE analysis. Based on these results, the involvement of TSST-1 and SEC3 in the pathogenesis of NTED is discussed.

Objectives : It has been reported that prostaglandin E 2 (PGE 2 ) is synthesized in the amnion and that this synthesis increases during labor. The purpose of this study was to clarify the mechanism for the expression of cyclooxygenase-2 (COX-2) mRNA and the PGE 2 synthesis of amnion-derived (WISH) cells. Study design : Cells were cultured and treated by 12- O -tetradecanoylphorbol-13-acetate (TPA) and dexamethasone (DEX). PGE 2 in the culture medium was measured by ELISA. Total RNA was extracted from the cells, and COX-2 mRNA expression was analyzed by Northern blot analysis. Results : During the time course of PGE 2 production in response to TPA stimulation, the PGE 2 production could not be detected until incubation had continued for 2h, but this production appeared to continue after 4h of incubation. PGE 2 production was significantly increased by TPA and suppressed by treatment with TPA and DEX. During the time course of COX-2 mRNA expression in response to treatment with TPA, the COX-2 mRNA band was detected after 1.5h. The strongest expression of COX-2 mRNA was observed at 2h incubation. After pre-treatment with TPA for 1h, the TPA-induced COX-2 mRNA was suppressed by treatment with DEX for 1 or 2h incubation in a dose-dependent manner. Conclusions : These results suggest that COX-2 mRNA is induced by TPA which activate protein kinase C, and suppressed by DEX in WISH cells.

PROBLEM In order to investigate the role of macrophage colony-stimulating factor (M-CSF) and macrophage chemoattractant protein-1 (MCP-1) in human ovulation, we measured the concentrations of M-CSF and MCP-1 in human follicular fluids (FFs) and correlated them with oocyte maturation. METHOD OF STUDY The oocytes were obtained from the FFs of 46 women undergoing in vitro fertilization and embryo transfer (IVF ET). The concentrations of M-CSF and MCP-1 in the FFs were measured by enzyme-linked immunosorbent assay. In addition, granulosa cells obtained from the FFs of IVF patients were cultured and treated with forskolin and 12-O-tetradecanoylphorbol 13-acetate (TPA) for 24 48 hr. RESULTS Concentrations of M-CSF and MCP-1 were significantly higher in the FFs than in the serum (P < 0.01). M-CSF concentrations tended to be higher, while MCP-1 concentrations were significantly higher in the FFs containing mature oocytes than in FFs containing immature oocytes (P < 0.05). The production of M-CSF was markedly increased over the basal level after treatment with forskolin (10 microM) for 24 (P < 0.02) and 48 hr (P < 0.01); however, the production of MCP-1 was unchanged. CONCLUSIONS Our data suggest that M-CSF and MCP-1 may play an important role in human preovulatory processes and that M-CSF, in particular, may be regulated by cyclic adenosine monophosphate. M-CSF and MCP-1 may also be valuable biochemical markers in the evaluation of oocyte maturation.

Staphylococcus aureus and S. epidermidis are common pathogens in hospitals, and care should be taken not to disseminate these organisms among patients. We have focused on human hair as a source of bacterial contamination. We treated hair with culture solutions of S. aureus and S. epidermidis, and then performed scanning electron microscopy. Bacteria were detected on the surface of the cuticles of the hair, and the attached bacteria were not completely removed even by repeated washing with detergents. These results suggested that hair could be a source of bacterial contamination and indicated the importance of decontamination of hair.

Platelet-activating factor (PAF) is involved in such reproductive processes as parturition. We investigated the effect of PAF on the expression of matrix metalloproteinase-1 (MMP-1) and that of tissue inhibitor of metalloproteinases-1 (TIMP-1) in human uterine cervical fibroblasts. Uterine cervical tissue was obtained from patients who underwent cesarean section at term. Collagenase-dispersed fibroblasts were cultured and used in the experiments. PAF receptor was identified in the uterine cervical fibroblasts by use of reverse transcription-polymerase chain reaction and Southern blot analysis. Northern blot analysis showed that PAF increased the expression of MMP-1 mRNA in a time-dependent manner, whereas expression of TIMP-1 mRNA was not affected by PAF. Concentration of MMP-1 protein in the PAF-treated culture media significantly exceeded that in control cultures. The PAF-induced production of MMP-1 protein was abolished by treatment with WEB 2170, a specific PAF receptor antagonist. Results suggest that PAF may accelerate collagenolysis in the human uterine cervix by inducing an imbalance in the activity between MMP-1 and TIMP-1, thus contributing to the cervical ripening during parturition.

OBJECTIVE To determine whether interleukin-8 (IL-8) regulates platelet-activating factor acetylhydrolase (PAF-AH) secretion by decidual cells in vitro. METHODS Decidual macrophages were obtained from human term decidual tissue by enzymic digestion and Ficoll-Paque centrifugation. The effect of IL-8 and phorbol esters on the secretion of PAF-acetylhydrolase by these cells was examined. RESULTS IL-8 inhibited PAF-AH secretion by decidual macrophages in a dose-dependent manner. The inhibition was reversed partially by antibodies against tumor necrosis factor-alpha and IL-1 beta. IL-8-induced inhibition was blocked partially by the protein kinase C (PKC) inhibitors, sphingosine, and H-7 1-(5-isoquinoline sulfonyl)-2-methylpiperazine. A PKC activator, 12-O-tetradecanoyl phorbol 13-acetate, decreased PAF-AH secretion. CONCLUSION IL-8 may increase the PAF concentration in the decidua via its inhibitory effect on PAF-AH secretion by decidual macrophages. IL-8-induced inhibition of enzyme secretion may have been mediated in part by PKC-dependent signal transduction.

To examine the production of hepatocyte growth factor (HGF) by human endometrial stromal cells (ESC) in vitro, concentrations of HGF in the culture media of ESC were measured after the addition of various amounts of 12-O-tetradecanoylphorbol 13-acetate (TPA), forskolin, lipopolysaccharide (LPS), interleukin (IL)-1beta, IL-6, IL-8, tumor necrosis factor alpha (TNFalpha), interferon-gamma (IFNgamma), or ethynylestradiol-17alpha using an ELISA. The expression of HGF mRNA was also assayed by a reverse transcription-polymerase chain reaction. The concentration of HGF in the culture media of unstimulated ESC was below the detection level of the assay. TPA stimulated the secretion of HGF by ESC in a dose-dependent manner. TPA also induced the transcription of HGF mRNA by ESC. Forskolin, LPS, IL-1beta, IL-6, IL-8, TNFalpha, IFNgamma, or ethynylestradiol-17alpha did not alter HGF mRNA or protein levels. TPA-stimulated production of HGF was partially inhibited by the addition of 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine or sphingosine. These results suggest that a protein kinase C-dependent pathway may play an important role in the regulation of HGF production by ESC. HGF secreted by ESC may be involved in the regeneration of the endometrium during the normal menstrual cycle.

Tumor necrosis factor-alpha stimulates the secretion of vascular endothelial growth factor by cultured human oviductal epithelial cells and stromal fibroblasts. Tumor necrosis factor-alpha-stimulated vascular endothelial growth factor secretion by these cells may control oviductal fluid secretion by regulating vascular permeability.

The purpose of this investigation was to evaluate the serum level of soluble intercellular adhesion molecule-1 (sICAM-1) in patients with squamous cell carcinoma of the cervix. Serum levels of sICAM-1 were measured by enzyme-linked immunosorbent assay in patients with cervical cancer (stage 0 through IV). Expression of mRNA in tumor tissues was evaluated by reverse transcriptase-polymerase chain reaction. Serum level of sICAM-1 in patients with advanced-stage (II-IV) or recurrent cervical cancer was found to be increased significantly. Expression of ICAM-1 mRNA was observed in most tumor tissues. Results suggest that sICAM-1 is shed from the cancerous tissue in patients with squamous cell carcinoma of the cervix.

In order to investigate the role of insulin-like growth factors (IGFs) in human ovulation, we evaluated the concentrations of IGF-binding proteins (IGFBPs) in human follicular fluid (FF). The concentrations of IGFBP-1 in the FFs of 15 women undergoing in vitro fertilization and embryo transfer were measured and related to those of 17beta-estradiol (E2), progesterone and androstenedione in the FFs. IGFBP-1 levels in the FFs were positively correlated with those of E2 and progesterone. No correlation was found between the IGFBP-1 and androstenedione levels in FFs. The concentrations of IGFBP-1 were significantly increased in the FFs which contained mature oocytes compared with those of immature oocytes, whereas IGFBP-3 in FFs tended to decrease as oocytes matured. It is suggested that IGFs may play important roles in human preovulatory processes, and that IGFBP-1 may be a valuable biochemical marker in the evaluation of oocytes.

The leiomyoma of the ovary is a very rare form of ovarian neoplasia, while its uterine localization is very common. A 72-year-old woman was admitted for pelvic examination. Transvaginal ultrasonography and magnetic resonance imaging (MRI) revealed a pelvic mass (8 cm x 7 cm). At laparotomy, total hysterectomy and bilateral salpingo-oophorectomy were performed and histologic examination revealed a leiomyoma arising primarily in the ovary.

Ovarian cancer is the second most common gynecological malignancy, yet it has the highest case-fatality ratio of all gynecologic malignancies. Surgery followed by combination platinum-taxane chemotherapy is the standard approach to the management of primary epithelial ovarian cancer. However, standard treatment of patients with recurrent ovarian cancer remains poorly defined. Secondary cytoreductive surgery (SDS) at the time of relapse has been proposed as a means of improving the prognosis of recurrent ovarian cancer patients with a treatment-free interval of at least 6 months.In the present study, we retrospectively collected 16 patients with recurrent epithelial ovarian cancer who might benefit most from SDS and evaluated the impact of SDS on the outcomes for this highly select patient group.We found that SDS led to excellent outcomes, with a 73.1% 8-year overall survival rate after initial treatment, a 67.9% 5-year overall survival rate after prior SDS, and a 31.3% 5-year progression-free survival rate after prior SDS. Although the findings were not significant, these results suggest that repeated SDS might improve outcomes for this patient group.The present study may provide a platform for discussion of the impact of aggressive or repeated SDS on the survival of patients with recurrent epithelial ovarian cancer and favorable prognostic factors. Further multi-institutional studies with larger number of patients are mandatory to confirm the present findings.

Objective: To study the expression of c-erb B-2 in gynecologic malignancies, especially in malignant mixed mullerian tumors (MMTs). Methods: Using immunohistochemical techniques, we examined 6 cases of primary MMT, 6 cases of leiomyosarcoma (LMS), 7 cases of endometrial adenocarcinomas, and 10 cases of normal endometria. Results: The expression of c-erb B-2 was observed in the carcinomatous area of all 6 cases of MMT (100%), the sarcomatous area of 5 of 6 cases of MMT (83.3%), in 1 of the 6 cases of LMS (16.7%), in all 7 cases of adenocarcinoma (100%), and in all cases of normal epithelial cells (100%), but was not observed in any of the cases of normal stromal cells (0%). Conclusion: The results suggest that the carcinomatous and sarcomatous elements of MMT are similar in their expression of c-erb B-2. MMT differed immunohistochemically from pure sarcoma cells and normal stromal cells, but resembled pure carcinoma cells and normal epithelial cells of the female genital tract.

In order to investigate the roles of epidermal growth factor (EGF) and transforming growth factor (TGF)-α in ovulation, we studied the production of interleukin (IL)-8 and growth-regulated oncogene (GRO)-α in cultured human granulosa-lutein cells.Granulosa-lutein cells obtained from the follicular fluids of in vitro fertilization and embryo transfer patients were cultured and treated with EGF, TGF-α, tumor necrosis factor (TNF)-α or 12-O-tetradecanoylphorbol 13-acetate (TPA). An immortalized granulosa cell line (GC1a) was also cultured and treated with EGF, TGF-α or mitogen-activated protein kinase kinase inhibitor. The supernatants were collected, and IL-8 and GRO-α were measured by ELISA.The levels of IL-8 and GRO-α were significantly increased after treatment with EGF, TGF-α, TNF-α and TPA by primary cultured granulosa-lutein cells. The levels of IL-8 and GRO-α were also significantly increased after treatment with EGF or TGF-α in a dose-dependent manner by GC1a. When GC1a was treated with EGF, TGF-α or U0126, the levels of IL-8 and GRO-α were significantly decreased.Our data indicate that the production of IL-8 and GRO-α is upregulated by EGF and TGF-α. It is suggested that EGF and TGF-α may play an important role in luteinization processes involving IL-8 and GRO-α production.

BACKGROUND: In order to investigate the regulation of chemokines [interleukin-8 (IL-8), growth-regulated oncogene (GRO)α, monocyte chemoattractant protein-I (MCP-I)) induced by thrombin in endometrial stromal cells (ESCs), the effects of thrombin, a protease activated receptor (PAR)-I antagonist (PPACK), mitogen-activated protein kinase kinase inhibitor (U0126), phospholipase C inhibitor (U-73122), an antagonist of the intracellular InsP3 receptor (2-aminoethoxy-diphenylborate (2-APB)] and a protein kinase C inhibitor (GF-109203X) on the production of chemokines by ESCs were evaluated. METHODS: ESCs from eight endometrial specimens in the secretory phase were cultured and incubated for 24 h with thrombin and PPACK, U0126, U-73122, 2-APB or GF-109203X. The levels of IL-8, GROα and MCP-I in the culture medium were measured by means of ELISA. The activation of MAP kinase was detected by western blot analysis using anti-phosphorylated MAP kinase (ERK I /2) antibody. RESULTS: Following stimulation by thrombin, the production of IL-8, GROα and MCP-I increased significantly in a dose-dependent manner. PPACK, U0126, U-73 122, 2-APB or GF-109203X suppressed the increases in production of IL-8, GROα and MCP-I induced by thrombin (P < 0.001, P < 0.001 and P < 0.001, respectively). MAP kinase activities were induced by treatment with thrombin, and were suppressed by PPACK, U0126, U-73122, 2-APB or GF-109203X. CONCLUSIONS: Our results suggest that thrombin stimulates the production of IL-8, GROα and MCP-I via PAR-I by a mechanism involving the MAP kinase system. The increases in IL-8, GROα and MCP-I may contribute to the maintenance of implantation involving leukocyte chemotaxis.

Primary amenorrhea caused by the hyperprolactinemia is a rare condition characterized by the onset of thelarche and pubarche at appropriate ages but arrest of pubertal development before menarche. Hyperprolactinemia might be found in a few women with primary amenorrhea, yet relevant experience has apparently not been reported. We report a 16-year-old patient with hyperprolactinemia caused by a pituitary microadenoma. Her only symptom was delayed puberty without galactorrhea. Bromocriptine therapy was useful in order to induce the ovulation and cause the menarche.

Background: Metallothionein (MT) is known to bind to metals with high affinity. The potential for MT-1 mRNA expression in endometrial stromal cells (ESC) and amniotic cells in response to cytokines and cadmium chloride (CdCl2) was evaluated. Methods: Human ESC were cultured and treated with interleukin-1α, 12-O-tetradecanoylphobol 13-acetate (TPA), forskolin, transforming growth factor-β, and CdCl2. Amnion-derived (WISH) cells were also cultured and treated with the same reagents. The levels of MT mRNA were evaluated by Northern blot analysis in ESC and WISH cells. Results: In response to treatment with CdCl2 (0.01–10 µM), the expression of MT mRNA markedly increased in ESC and WISH cells in a dose-dependent manner. On the other hand, the expression of MT mRNA did not increase after treatment with interleukin-1α (1 nM), TPA (10 nM), forskolin (1 µM) or transforming growth factor-β (1 nM) in these cells. Conclusion: These findings indicate that MT expression in ESC and WISH cells is sensitive to the CdCl2 concentration, which is known to be evaluated in cigarette smokers. The present results suggest that increased levels of MT may affect metal metabolism at the feto-maternal interface.

Objective The aim of this study is to evaluate the effects of interleukin (IL)-13, a Th2 cytokine, on the production of vascular endothelial growth factor (VEGF) and soluble fms -like tyrosine kinase-1 (sFlt-1) in human oviductal cells in vitro . Study design Human oviductal epithelial cells (OECs) were isolated from five premenopausal patients. The secretion of VEGF 165 and sFlt-1 by cultured OECs in response to IL-13 was measured using an enzyme-linked immunosorbent assay. Results The secretion of VEGF 165 and sFlt-1 was detected in cultured OECs under untreated conditions. IL-13 enhanced the secretion of VEGF 165 and sFlt-1 by OECs in a dose-dependent manner. Conclusion The present findings suggest that IL-13 is a regulatory factor of VEGF and sFlt-1 production in the human fallopian tubes. IL-13 in the local environment may stimulate oviductal vascular permeability by inducing the production of VEGF by oviductal cells. The modulation of VEGF secretion by IL-13 secreted by the peri-implantation embryo may contribute to the normal and pathological processes of human reproduction during the peri-implantation period.

The aim of the current study is to evaluate the effect of insulin-like growth factor-I (IGF-I) on the production of vascular endothelial growth factor (VEGF) in the human fallopian tube. Human oviductal epithelial cells (OEC) and oviductal stromal fibroblasts (OSF) were isolated from the ampullary segment of the fallopian tubes of six premenopausal patients in the proliferative phase of the menstrual cycle. The secretion of VEGF165 by cultured OEC and OSF in response to IGF-1 was measured using an enzyme-linked immunosorbent assay (ELISA). The secretion of VEGF165 was detected in cultured OEC and OSF under untreated conditions. The secretion of VEGF165 was significantly stimulated with IGF-I administration in these cells. The present findings suggest that IGF-I in the local environment may stimulate oviductal vascular permeability by inducing the production of VEGF by oviductal cells through autocrine and paracrine mechanisms. The modulation of the VEGF production in the fallopian tube may contribute to the normal and pathologic processes of oviductal fluid secretion by regulating oviductal vascular permeability during the menstrual cycle and in the peri-implantation period.

The purpose of the present study was to investigate the effects of an exogenously administered cell-permeable synthetic ceramide analogue, C(2)-ceramide (N-acetyl-sphingosine) on the growth, cell cycle, and death of Ishikawa human endometrial carcinoma cells. We investigated the effects of C(2)-ceramide on Ishikawa endometrial cancer cell lines in vitro. The cells were treated with C(2)-ceramide, and its effects on cell growth, cell cycle, apoptosis, and related measurements were investigated. MTT assays showed that C(2)-ceramide, a cell-permeable analogue of ceramide, significantly induced dose- and time-dependent death in human endometrial carcinoma Ishikawa cells. Cell-cycle analysis indicated that their exposure to C(2)-ceramide decreased the proportion of cells in S phase and increased the proportion in G0/G1 and/or G2/M phases of the cell cycle. Induction of apoptosis was confirmed by annexin V staining of externalized phosphatidylserine and loss of the transmembrane potential of mitochondria. This induction occurred in concert with the altered expression of genes related to cell growth, malignant phenotype, and apoptosis, including cleavage of poly-ADP ribose polymerase. Furthermore, we demonstrated that the amount of phosphorylated Akt was decreased by C(2)-ceramide. These results raise the possibility that C(2)-ceramide may prove particularly effective in the treatment of endometrial cancers.

Objective To evaluate the effect of hypoxia on the production of vascular endothelial growth factor (VEGF) and soluble fms-like tyrosine kinase-1 (sFlt-1) in the human fallopian tube. Design The secretion of VEGF and sFlt-1 by cultured oviductal epithelial cells (OECs) and oviductal stromal fibroblasts (OSFs) in response to hypoxia was investigated. Setting Research laboratory at a medical school. Patient(s) Normal oviducts obtained from seven premenopausal patients were used. Intervention(s) Oviductal epithelial cells and OSFs were incubated under normoxic (20% O 2 ) or hypoxic (2% O 2 ) conditions. Main Outcome Measure(s) The concentrations of VEGF and sFlt-1 in the culture media of OECs and OSFs were measured by enzyme-linked immunosorbent assays. Result(s) The secretion of both VEGF and sFlt-1 was detected in cultured OECs and OSFs and was found to have been stimulated under hypoxic conditions in these cells. Conclusion(s) The present findings suggest that hypoxia in the local environment may stimulate oviductal vascular permeability by inducing the production of VEGF by oviductal cells. Simultaneous up-regulation of sFlt-1 secretion by these cells under hypoxic conditions may prevent excessive up-regulation of vascular permeability. The modulation of the bias of VEGF and sFlt-1 in the fallopian tubes may contribute to the normal and pathological processes of oviductal fluid secretion by regulating oviductal vascular permeability during the menstrual cycle.

We report herein a rare case of choledochal cyst diagnosed at 14 weeks gestation and treated with percutaneous transhepatic drainage until postpartum. A 26-year-old primigravid woman at 14 weeks gestation presented with epigastric pain, slight fever, and nausea of 3 days duration. Abdominal ultrasonography revealed a 6-cm-diameter cystic mass between the porta hepatis and the pancreas head, which was diagnosed as a type I choledochal cyst. At 18 weeks of gestation, her upper abdominal pain became severe, and the size of the choledochal cyst increased to 12 cm in diameter. Laboratory data revealed a biliary obstruction. Percutaneous transhepatic drainage was performed immediately. She delivered a healthy male baby by elective cesarean section at 37 weeks of gestation. At 6 weeks postpartum, the patient underwent surgical excision of the choledochal cyst, cholecystectomy, and formation of a Roux-en-Y hepaticojejunostomy. Although choledochal cysts are rare during pregnancy, obstetricians should be familiar with the condition to ensure prompt diagnosis and adequate definitive management, given that the implications of missed or delayed diagnosis may be detrimental to both mother and fetus.

Objective To investigate the role of macrophageinflammatory protein (MIP)-3α in human ovulation. Design Study of the levels of MIP-3α in serum and follicular fluid. The effects of interleukin (IL)-1α, IL-1 receptor antagonist (RA), and tumor necrosis factor (TNF)-α on the secretion of MIP-3α 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 (IVF-ET). Intervention(s) Follicular fluid was obtained from study participants, and granulosa-lutein cells and GC1a were incubated with IL-1α, IL-1RA, or TNF-α for 4 to 32 hours. Main outcome measure(s) The concentration of MIP-3α in human follicular fluid was measured and correlated with oocyte maturation. We also cultured granulosa cells and examined the regulation of MIP-3α production. The concentrations of MIP-3α in the serum, follicular fluid, and culture medium were measured using enzyme-linked immunoabsorbent assay. Result(s) Concentrations of MIP-3α were significantly higher in the follicular fluid, but it was not detected in the serum. Concentrations of MIP-3α were statistically significantly higher in the follicular fluid containing mature oocytes than in follicular fluid containing immature oocytes. The production of MIP-3α was markedly increased over the basal level after treatment with IL-1α and TNF-α in a dose-dependent manner. The stimulatory effect of IL-1α was inhibited by IL-1RA. Conclusion(s) Our data suggest that MIP-3α was present in follicular fluid and correlated with oocyte maturation, and was regulated by IL-1α and TNF-α. Thus, MIP-3α may play an important role in the human preovulatory process.

The aim of this study was to clarify changes in the expression of vascular endothelial growth factor (VEGF) during decidualization by endometrial stromal cells (ESCs) in vitro.ESCs were separated by enzymic digestion and filtration, and were cultured with RPMI 1640 in 10% fetal calf serum (FCS) and treated with medroxyprogesterone acetate (MPA) and dibutyryl-cyclic adenosine monophosphate (db-cAMP) to induce decidualization in vitro. The levels of prolactin (PRL) in the culture media were measured by enzyme immunoassay (EIA) and the levels of VEGF were measured by enzyme-linked immunosorbent assay (ELISA). The expression of VEGF mRNA by ESCs and decidualized cells was analyzed by Northern blot analysis.In treated cells, PRL production was significantly increased due to treatment with both db-cAMP (0.5 mmol/L) and MPA (100 nmol/L). VEGF mRNA expression was detected without any stimulation by ESCs. VEGF production was also significantly greater in cells treated with db-cAMP (0.5 mmol/L) and MPA (1 nmol/L or 100 nmol/L) than in control cells. VEGF mRNA was also increased in association with decidualization in vitro.VEGF production increased in association with decidualization in this in vitro study. Decidualization of ESCs may play a role in the induction of growth in the human fetus and/or placentation. The mechanism involved may include influencing the production of angiogenic growth factors such as VEGF.

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.

Our objective was to clarify the modulation of vascular endothelial growth factor (VEGF) by amniotic cells.Amnion-derived (WISH) cells were cultured, and the effect of insulin-like growth factor (IGF), mitogen-activated protein (MAP) kinase kinase and/or extracellular signal-regulated kinase (ERK) kinase (MEK) inhibitors (U0126), and phosphatidylinositol (PI) 3-kinase inhibitors (wortmannin) on the production of VEGF was examined. VEGF was assayed using ELISA. The activations of MAP kinase and akt, which is phosphorylated by PI 3-kinase, were detected by Western blot analysis using anti-phosphorylated MAP kinase antibody and anti-phosphorylated akt antibody.In the time course of VEGF production following IGF-I treatment, VEGF production showed significant increases only at 16 and 32 h (p0.01). Also, IGF-I increased the production of VEGF by WISH cells in a dose-dependent manner. The MAP kinase and akt activities were recognized by treatment with IGF-I and suppressed by U0126 and wortmannin, respectively. When WISH cells were pretreated for 2 h with U0126 and wortmannin and then treated with IGF-I for 16 h, the production of VEGF was significantly decreased in a dose-dependent manner (p0.01, p0.01, respectively).WISH cells appeared to produce VEGF via a mechanism involving tyrosine kinase interaction with IGF-I receptor, resulting in MAP kinase and PI 3-kinase activation. It is suggested that VEGF may contribute to the neovascularization and proliferation of the placenta and gestational tissue, and IGF-I may play an important role in the modulation of VEGF production in the placenta.

Interleukin-8 (IL-8) and growth-regulated oncogene-alpha (GRO-alpha) have been proved to be important modulators of leukocyte chemotaxis in the mechanism of human ovulation. This study investigated the possible effects of IL-1alpha, tumor necrosis factor-alpha (TNF-alpha), protein kinase C (PKC) activators (TPA), and db-cyclic adenosine monophosphate (cAMP) on IL-8 and GRO-alpha production by immortalized GC1a and granulosa-lutein cells.Confluent granulosa-lutein cells were placed in serum-free medium before incubated for 8 hr with the above-mentioned test agents. Finally, we measured IL-8 and GRO-alpha levels in the culture media using an enzyme-linked immunosorbent assay (ELISA).Treatment of granulosa-lutein cells with of IL-1alpha (1 nM), TNF-alpha (1 nM), TPA (1 nM) and db-cAMP (100 microM) produced higher levels of IL-8 than untreated cells by 8 hr (2274.7 +/- 146.3, 1489.8 +/- 190.1, 1452.9 +/- 152.7, 1313.6 +/- 48.4 pg/mL, respectively; control = 457.7 +/- 38.2 pg/mL; P0.001). Treatment of granulosa-lutein cells with 1 nM of IL-1alpha, TNFalpha, TPA, and db-cAMP (100 microM) resulted in higher levels of GRO-alpha than untreated cells by 8 hr (993.7 +/- 9.5, 171.4 +/- 6.5, 147.5 +/- 6.7, 472.4 +/- 16.2 pg/mL respectively; control = 73.8 +/- 8.2 pg/mL; P0.001).Our data strongly suggests roles for IL-1alpha, TNFalpha, and PKC activators in the inflammation-like mechanism of human ovulation. Furthermore, our study suggests a positive, but still debatable, role for cAMP in the same mechanism.

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.

Hypoxia downregulated the concentration of endostatin in the culture media of human endometrial stromal cells but did not affect the messenger (m)RNA expression of collagen XVIII. Both mRNA and protein expression of vascular endothelial growth factor were upregulated in a hypoxic condition.

Recurrent ovarian cancer after front-line chemotherapy is incurable. In most institutions, chemotherapy is continued as salvage therapy after primary chemotherapy failure and despite the fact that long-term survival and complete responses are infrequent. Radiation therapy for patients with recurrent ovarian cancer has often been done with palliative intent. A patient with ovarian clear cell adenocarcinoma received irradiation with palliative intent to the whole pelvis after chemotherapy (paclitaxel, carboplatin, and irinotecan) produced no effect. Although she developed a rectovaginal fistula due to cancer invasion during radiation therapy. One year and half after the therapy, she is still alive with no evidence of disease. In an effort to maximize salvage potential and quality of life while minimizing toxicity, selected patients with ovarian cancer should be treated with radiation therapy directed to residual or recurrent sites.

We applied two-dimensional gel electrophoresis (2-DE) to the total exoproteins secreted from pathogenic MRSA strains and identified major protein spots by N-terminal amino acid sequence analysis. In approximately 300 to 500 spots visualized on each gel, various exoproteins and cell-associated proteins were identified and their sites on the gels confirmed for construction of a reference map. Major exotoxins such as enterotoxins SEA, SEB, and SEC,, toxic shock syndrome toxin-1 (TSST-1), and hemolysins were distributed in the region of pI 6.8 to 8.1 and MW 21 to 35 kDa. Although the differences between calculated and observed values of pI and MW were relatively small in each exoprotein, those of several proteins including alpha-hemolysin and SEB were considerably deviated from the positions of the expected values. Some exoproteins were detected as multiple spots. These included beta-hemolysin, enterotoxins SEA, SEB, and SEC3, glutamic acid-specific endopeptidase, glycerophosphoryl diester phosphodiesterase and triacylglycerol lipase. The multiple spots of these exoproteins may be generated by the action of own proteases. Certain similarities of 2-DE patterns among strains belonging to the same coagulase types were observed. On the basis of 2-DE image analysis, coagulase type II strains secreted somewhat larger amounts of SEB and SEC3 as well as TSST-1 than the strains belonging to other coagulase types. Taken together, 2-DE analysis of exoproteins is applicable to epidemiological studies for MRSA, as compared with pulsed field gel electrophoresis of restricted chromosomal DNA.