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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.

Myelin oligodendrocyte glycoprotein (MOG) antibodies are associated with relapsing inflammatory demyelinating disease. Pregnancy complicates the disease course, potentially leading to either symptom improvement or worsening. A 28-year-old woman with MOG antibody-associated encephalomyelitis had 2 pregnancies; her disease worsened during both postpartum periods despite continuing prednisolone and levetiracetam. The umbilical cord blood was positive for MOG antibodies following her second pregnancy, but neither baby had MOG antibody-associated disease. This is the first case report of MOG antibody-associated demyelinating disease that worsened postpartum despite continuous medication. Furthermore, we observed the placental transfer of MOG antibodies for the first time.

Uterine fibroids are associated with heavy menstrual bleeding, abdominal discomfort, subfertility and a reduced quality of life. The present study reported a case series of life-threatening anemia with hemoglobin levels

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.

Background and Aims: Platelet activating factor (PAF), a potent phospholipid mediator, has been implicated in a number of reproductive processes through ovulation to parturition. To clarify the regulatory mechanism of PAF metabolism in the decidua, we have investigated the effect of activation of protein kinase C (PKC) on the secretion of PAF‐acetylhydrolase (PAF‐AH), a PAF‐inactivating enzyme, by human decidual macrophages.

The incidence of ectopic pregnancy is approximately 1.3∼2% of all pregnancies, and more than 90% of ectopic pregnancies are detected in the ampulla of the fallopian tube. Ectopic pregnancy occurring in tubal stump after tubectomy is extremely rare, and the frequency of tubal stump pregnancy is approximately 0.4% of all pregnancies. We report one of these rare cases of ectopic pregnancy in a 26-year-old Japanese woman, gravida 4, parity 1. She had undergone laparoscopic tubectomy because of a tubal pregnancy two years ago. She was presented to our hospital with a positive pregnancy test, but no gestational sac was detected in the uterus by echography, even though the level of human chorionic gonadotropin (hCG) in the blood was elevated to 8,900 mIU/mL. Laparoscopic surgery for ectopic pregnancy was performed. During surgery, the position of the pregnancy was found to be in the tubal stump, where tubectomy had already been performed, and the gestational sac was successfully removed. After the surgery, the condition of the patient uneventfully improved and she was discharged from the hospital three days after the surgery. The diagnosis of tubal stump pregnancy is more difficult than that of the more common positions of an ectopic pregnancy in the fallopian tube, and so it is more important to carefully examine the patients with suspected ectopic pregnancy. Laparoscopic surgery is one of the options for tubal stump pregnancy if diagnosed early and if the condition of the patient is stable.

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

Although parasitic leiomyoma could be spontaneous or iatrogenic in origin, port-site implantation of parasitic leiomyoma is an iatrogenic benign sequela of laparoscopic surgery. A 30-year-old, primigravida Japanese woman was referred after unresponsiveness to preoperative gonadotropin-releasing hormone for intramural fibroids. Magnetic resonance imaging showed multiple intramural fibroids and left ovarian endometrioma with no malignant features. Laparoscopic myomectomy with power morcellation and ovarian cystectomy were performed, followed by treatment with a combined oral contraceptive. Seven years after the primary surgery, she underwent abdominal myomectomy for a port-site, and peritoneal recurrence of the leiomyoma and intramural leiomyomas was detected. Microscopic examination revealed that resected specimens from the port-site demonstrated leiomyoma with lesser cell density and more prominent hyalinization than those from the uterus. Therefore, clinicians should counsel patients regarding the risks and benefits of laparoscopy with morcellation versus laparotomy. Further development of techniques for uterine tissues extraction is warranted.

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.

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.

Platelet activating factor (PAF), a potent phospholipid mediator, has been implicated in a number of reproductive processes through ovulation to parturition. To clarify the regulatory mechanism of PAF metabolism in the decidua, we have investigated the effect of activation of protein kinase C (PKC) on the secretion of PAF-acetylhydrolase (PAF-AH), a PAF-inactivating enzyme, by human decidual macrophages. Decidual macrophage populations were isolated from human decidua by using enzymic digestion, Ficoll-Paque centrifugation, or flow cytometric sorting. The cells were treated with a PKC activator (TPA), H-7, dibutyryl cyclic adenosine monophosphate (AMP), Bis-(o-aminophenoxy)-ethane-N,N,N′,N′-tetraacetic acid, tetra (acetoxymethyl)-ester (BAPTA/AM) and/or nifedipine. The activity of PAF-AH secreted in the culture medium was assayed. The PKC activator, TPA, inhibited the PAF-AH secretion by decidual cells in a dose-dependent manner. The TPA also decreased the enzyme secretion by flow cytometrically purified macrophages. The inhibitory effect of TPA was blocked by a PKC inhibitor, H-7. Protein kinase A (PKA) activation by dibutyryl cyclic AMP was without effect on the enzyme secretion. Calcium channel blockers, BAPTA/AM and nifedipine had no effect on the PAF-AH secretion. It is suggested that the TPA-induced inhibition of PAF-AH secretion may be mediated, in part, by a PKC-dependent signal transduction, and that activation of PKC may result in the increase in the local concentration of PAF in the decidua because of its inhibitory effect on the PAF-AH secretion by decidual macrophages.

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.

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.

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.

Objective: To retrospectively compare the postsurgical pain after single-port laparoscopic surgery and conventional laparoscopic salpingectomy for the surgical treatment of tubal pregnancy. Methods: The cases of two groups of patients with ectopic pregnancies were reviewed: those who had undergone a single-port laparoscopic surgery (n=6) and those who had undergone a conventional multi-port laparoscopic surgery (n=20). We compared these groups’ surgical outcomes, including operative time, blood loss, use of analgesics, and complications. Results: There was no significant difference between the two groups regarding the surgical time, blood loss during surgery, or analgesics use after laparoscopic surgery. There were no serious complications and no need for conversion to conventional laparoscopy or laparotomy in both groups. Conclusion: Our present findings suggest that single-port laparoscopic surgery is feasible and practical for the surgical treatment of ectopic pregnancies. However, the results also indicate that the reduction of the number of laparoscopy ports did not offer further pain relief in these patients.

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.

OBJECTIVE 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 via mitogen-activated protein kinase (MAPK) induced by low level laser therapy (LLLT) in human granulosa cells. METHODS A human granulosa cell line, KGN cells, were cultured and incubated after LLLT (60mW, GaAlAs 830nm). The levels of VEGF in the culture media were determined by an enzyme-linked immunosorbent assay. The activation of MAP kinase in KGN cells was detected by western blot analysis. RESULTS VEGF production was significantly increased by LLLT in a time-dependent manner. MAP kinase activity was increased by LLLT. In addition it was enhanced by LLLT and follicle-stimulating hormone (FSH) stimulation. CONCLUSIONS The results suggested that VEGF is induced by LLLT through mechanisms involving MAPK. The increase in VEGF may contribute to neovascularization, which in turn would promote various ovulation phenomena as well as follicular growth.

Target cells of cytotoxic T lymphocytes (CTL) directed to the individual structural proteins (except for the large polymerase (L) protein) of rabies virus were established by expressing only the respective protein in murine neuroblastoma (NA) and murine macrophage (J774-1) cell lines. Mice infected with the ERA strain of rabies virus developed CTL responses to all of these rabies virus proteins. The cytotoxic activity was abrogated by pretreatment of the effector cells with anti-CD8 monoclonal antibody (MAb) and complement but not with anti-CD4 MAb. Cell lysis by CTL was blocked in the presence of anti-major histocompatibility complex (MHC) class 1 antibodies in J774-1 cell lines. Rabies virus-infected cells express these proteins at the surface, which can be recognized and lysed by the respective CTL. Mice immunized with beta-propiolactone-inactivated virus induced a CTL response against glycoprotein but not against internal viral components. This assay system might be useful for further analysis of the possible contribution of these proteins in the cell-mediated immune protection against rabies.

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.

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.

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.

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.