Satomi Nakamura

The production and clinical evaluation of macrophage colony-stimulating factor and macrophage chemoattractant protein-1 in human follicular fluids.

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 μM) 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.

Changes in vascular endothelial growth factor production associated with decidualization by human endometrial stromal cells in vitro

Background. 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.

The effect of epidermal growth factor on vascular endothelial growth factor secretion by endometrial stromal cells

Abstract Endometrial stromal cells undergo morphological and functional changes to facilitate oocyte implantation under regulation of various hormones and growth factors. We studied physiological induction by epidermal growth factor (EGF) of vascular endothelial growth factor (VEGF) in these cells. In human endometrial stromal cells, the effect of EGF, genistein, tryphostin AG1478 (a tyrosine kinase inhibitor), and wortmannin (a phosphatidylinositol 3-kinase inhibitor) on production of VEGF was examined: Total RNA was extracted and VEGF mRNA expression was quantified by Northern analysis. EGF induced production of VEGF by stromal cells in a time-dependent manner; the effect became significant after 12 h and increased further between 24 and 48 h (P<0.05). Dose dependency was also significant (P<0.01). Genistein, tryphostin AG1478, and wortmannin partially suppressed the increase in production induced by EGF (P<0.01, P< 0.01, P<0.01), respectively. Production of EGF by fertilized oocytes and trophoblasts has been reported in early pregnancy. VEGF is believed to be induced by EGF through mechanisms involving tyrosine kinase and phosphatidylinositol 3-kinase. The increase in VEGF may contribute to neovascularization that promotes proliferation of endometrium and placentation.

Synergistic effect of interleukin-1alpha and ceramide analogue on production of prostaglandin E2 and F2alpha by endometrial stromal cells.

PROBLEM: Prostaglandins (PGs) are synthesized in the endometrium. Our objective was to evaluate interleukin (IL)‐1α‐induced production of PGE2 and PGF2α in endometrial stromal cells (ESC) following treatment with ceramide analogues.
 METHODS OF STUDY: ESC were obtained from human uterine endometrium by enzymic digestion and filtration. ESC were treated with IL‐1α, IL‐1 receptor antagonist (ra), C2‐ceramide and C6‐ceramide. The concentrations of PGE2 and PGF2α in media were determined using ELISA. The induction of prostaglandin H synthase (PGHS)‐2 mRNA was also ascertained by reverse transcription‐polymerase chain reaction (RT‐PCR).
 RESULTS: The production of PGE2 and PGF2α was significantly increased by IL‐1α and suppressed by IL‐1 ra, in a dose‐dependent manner. PGF2α production was further increased by treatment with the combination of IL‐1α and C2‐ceramide as compared with IL‐1α treatment alone. There was no significant difference in PGE2 production between cells treated with IL‐1α and C2‐ceramide and those treated with IL‐1α alone. Both PGE2 and PGF2α production were significantly increased by treatment with IL‐1α and C6‐ceramide as compared with IL‐1α treatment alone. Treatment of ESC with IL‐1α stimulated PGHS‐2 mRNA. PGHS‐2 mRNA was decreased when IL‐1 ra was added to the IL‐1α‐stimulated cells.
 CONCLUSIONS: These results suggest that IL‐1α stimulates the production of PGE2 and PGF2α by a mechanism that involves the sphingomyelin‐ceramide system, and thus that ceramide may be important in increasing the production of PGE2 and PGF2α in the human endometrium.

The effect of epidermal growth factor on production of vascular endothelial growth factor by amnion-derived (WISH) cells

Our objective was to clarify the physiological role of vascular endothelial growth factor (VEGF) by amnion-derived (WISH) cells. WISH cells were cultured, and the effect of epidermal growth factor (EGF), mitogen-activated protein (MAP) kinase kinase or extracellular signal-regulated kinase (ERK) kinase (MEK) inhibitors (U0126) or phosphatidylinositol (PI) 3-kinase on the production of VEGF was examined. VEGF was assayed by ELISA. The activation 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 EGF treatment, VEGF production showed a significant increase only after 16 (p < 0.01)–32 h (p < 0.01). EGF increased the production of VEGF by WISH cells in a dose-dependent manner. The MAP kinase and akt activity were determined by treatment with EGF. VEGF production was significantly decreased following pretreatment with U0126 or wortmannin for two hours before treatment with EGF (p < 0.01, p < 0.01). WISH cells appeared to produce VEGF via a mechanism involving tyrosine kinase activation of EGF receptor and MAP kinase or PI 3-kinase. It is suggested that VEGF may contribute to the neovascularization and proliferation of the placenta and gestational tissue, and EGF may play an important role in regulation of VEGF production in the placenta.

Effect of ceramide analogs on interleukin-1α-induced production of prostaglandin E2 by amnion-derived (WISH) cells

Background. Prostaglandin E2 (PGE2) is reportedly synthesized in the amnion, and its levels are increased during labor. Our objective was to measure the level of PGE2 induced by interleukin (IL)‐1α following treatment with ceramide analogs in amnion‐derived cells.

The effect of dexamethasone on expression of mitogen-induced cyclooxygenase-2 mRNA by amnion-derived (WISH) cells

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.