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High-risk human papillomavirus (HPV) is associated with the carcinogenesis of not only cervical cancer but anal, penile, vulvar, vaginal, and oropharyngeal cancers. Although molecular biological mechanisms of high-risk HPV (HR-HPV)-associated carcinogenesis is well studied, it remains unclarified why cervical cancer is the most common among these HPV-associated cancers. Two major causes are that the cervix is a susceptible site to viral infection because of its immune deficiency to protect allogenic sperm in reproductive function and that the squamocolumnar junction (SCJ) where cervical neoplastic diseases develop is composed of tissue stem cells with self-renewal and pluripotency. This specific environment of the cervix allows HPVs to be persistently infected into the cervical epithelial cells, followed by the immortalization of the HPV-infected cells. We here focused on the carcinogenesis specific to the cervix as novel therapeutic strategies for cervical cancer, targeting cancer stem cells and mucosal immunotherapy.

Background: X-ray pelvimetry is used for evaluation of pelvic inlet generally to diagnose cephalopelvic disproportion (CPD) or contracted inlet. Cesarean section delivery (C/S) is often performed for labor dystocia without CPD or contracted inlet. Here, we examined whether X-ray pelvimetry is useful to decide on mode of delivery in women with dystocia.Method: A total of 1118 pregnant women received X-ray pelvimetry before or during labor. Overall, 205 women with cesarean deliveries for indications except for dystocia were excluded. The remaining 913 women undergoing induction/augmentation were retrospectively investigated. Obstetrical and maternal variables were analyzed by univariate, multivariate or ROC analysis.Results: Among 913 women, 37 (4%), including three with contracted inlet and seven with CPD, gave birth by C/S, whereas 876 gave birth by vaginal delivery. Low maternal height, older age, small obstetrical conjugate, large weight and infant head size were significantly associated with risk of C/S for dystocia. Multivariate analysis revealed that the obstetrical conjugate was an independent variable for risk of C/S. The area under the ROC curve (AUC) and the optimal cut-off values, respectively, were as follows: obstetrical conjugate: 0.68 and 11.7 cm (odds ratio=4.27), transverse diameter: 0.59 and 11.4 cm (odds ratio=1.82), maternal height: 0.70 and 155.5 cm (odds ratio=4.33), and maternal weight before pregnancy: 0.55 and 49.7 kg (odds ratio=1.98).Conclusion: The obstetrical conjugate was an independent variable associated with risk of C/S for dystocia. Additionally, maternal height was comparable to the conjugate in term of diagnostic ability. Our data suggested that routine X-ray pelvimetry was not beneficial to identify women at risk of C/S for dystocia.

Aim Although the procedure of abdominal trachelectomy has been remarkably improved, preventing subsequent cervical stenosis remains challenging. In this study, we analyzed the clinicopathological risk factors for cervical stenosis to explore the appropriate surgical procedures for the prevention of cervical stenosis following trachelectomy. Methods Thirty-two patients who underwent abdominal extended and radical trachelectomy were assessed retrospectively (median follow-up period = 33 months). To evaluate the risk factors, the clinicopathological factors were analyzed by univariate and multivariate analyses. The reconstructed uterine length (UtL), that is, the length between the vaginal end of the neo-cervix and the uterine fundus, was measured by transvaginal ultrasound after surgery. The cut-off value for the UtL was assessed by a receiver operating characteristic (ROC) curve analysis. Results Cervical stenosis of any grade was observed in 12 patients (grade 1 = 9, grade 3b = 3). Among the various clinicopathological factors, the UtL and cervical length (CL) were significantly related to cervical stenosis following trachelectomy. The multivariate analysis revealed that the UtL, but not CL, is an independent risk factor for stenosis. The ROC curve analysis revealed that stenosis was significantly more likely to occur in patients with a UtL shorter than 53 mm (area under the ROC curve = 0.902). UtL in the patients who became pregnant was longer than that in the patients who did not. No evidence of recurrent cancer was observed during the follow-up period. Conclusion Our proposed method may provide a functional reconstructed uterus with preserving fertility by remaining UtL more than 53 mm.

Aim. To investigate the changes in the maternal immune system at term pregnancy, we studied the expression of natural cytotoxicity receptors (NCRs) and the cytokine production of NK cells in term placenta decidua and peripheral blood. Methods. Term decidua and peripheral blood were taken from patients undergoing elective cesarean section. The lymphocytes were separated using density gradient centrifugation (DGC) from peripheral blood and were separated from decidua using DGC after enzyme digestion. These cells were stained with FITC anti-CD56 and Per-CP anti-CD3 monoclonal antibodies, and the NCRs were stained with PE-conjugated anti-NKG2D, NKp46, NKp30, and NKp44 monoclonal antibodies. Cytokines, including IFN-γ, TNF-α, IL-10, and TGF-β, were stained and then analyzed by flow cytometry. Results. There were fewer cells positive for NKG2D, NKp46, and NKp30 among CD56+CD3- cells in deciduas than in peripheral blood, but the percentages of NKp44-positive cells in CD56+CD3- lymphocytes in deciduas tended to be higher. Conclusion. The decreased expression of some NCRs in deciduas may be related to decreased cytotoxicity at term pregnancy, but the increased expression of NKp44 may affect the increased cytokine production in the decidua. Similarly, the expression of NCRs in the decidua may be connected to the maintenance of pregnancy at term.

Implantation is essential for successful reproduction. In humans, uterine receptivity is restricted to the mid-secretory phase, days 19 to 24 of the menstrual cycle; and this period is called the window of implantation (WOI). Large populations of decidual leukocytes infiltrate the implantation site, and the levels of cytokines, prostaglandins (PGs), and leukocytes are up-regulated in the endometrium during implantation. Implantation is like a local wound healing process which is characterized by a strong Th1, pro-inflammatory response in which high levels of pro-inflammatory cytokines such as IL-6, LIF, IL-8, and TNFα are involved. In addition, cyclooxygenase (COX)-2 expression increases between days 2 to 5, suggesting that PGs are necessary for the process of stromal cell decidualization in early pregnancy. Recently, it was reported that endometrial biopsies taken during spontaneous cycles that preceding IVF treatment improved the rate of implantation, clinical pregnancies, and live births in p...

Background/Aim: The aim of this study was to evaluate the gene and protein expression of angiotensin type (AT) 1, AT2 receptors in endometriotic lesions and its relation to prostaglandin (PG) synthases. Materials and Methods: Endometriosis samples were obtained from 32 patients with endometriotic cysts. Endometrial tissues were obtained during operations for benign gynecological conditions. The expression of the AT1 and AT2 receptor mRNA and that of PG-endoperoxide synthase 2 and microsomal PGE2 synthase-1 (mPGES-1) was examined by quantitative RT-PCR. Immunohistochemical staining was performed for these receptors. Results: AT1 and AT2 receptor proteins were mostly located in endometrial glandular epithelium and some stromal cells. Immunoreactivity of the receptor proteins was observed in both the eutopic endometrium and endometriotic lesions. The AT1/AT2 ratio in endometriotic cysts (median 7.29, range 1.88-187.60) was significantly increased compared with that in the eutopic endometrium in the proliferative-phase in controls (median 1.01, range 0.37-2.09, p < 0.001). There was a relationship between the AT1 mRNA expression and that of mPGES-1 mRNA in the endometriotic cysts (r = 0.394089, p < 0.05). There was a significant relationship between the mRNA expression of the AT2 receptor and that of mPGES-1 in eutopic endometrium of non-endometriotic control (r = 0.610714, p < 0.05). Conclusion: Renin-angiotensin system may play an important role in the pathophysiology of endometriosis.

Aim: Cytokines such as tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) play an important role in maintaining pregnancy. Toll-like receptors (TLRs) have also been associated with innate immune responses during pregnancy. Placenta growth factor (PlGF) is one of the angiogenic factors and soluble vascular endothelial growth factor receptor 1 (sVEGFR1) is one of the antiangiogenic factors that regulate PlGF function. To elucidate the effects of cytokines and TLR ligands on the production of angiogenic and antiangiogenic factors in trophoblasts, we examined the production of PlGF and sVEGFR1 from trophoblasts following stimulation with cytokines and TLR ligands. Methods: Villous tissues were obtained from healthy pregnant women who had had an artificial abortion. The trophoblasts were isolated from villous tissues. Subsequently, trophoblasts were treated with TNF-α, IFN-γ and TLR ligands. The levels of PlGF and sVEGFR1 were measured by enzyme-linked immunosorbent assay. The expression of those mRNA was analyzed using quantitative reverse transcription PCR. Results: The production of PlGF in trophoblasts increased by the addition of TNF-α or IFN-γ and decreased by TLR4 ligand. sVEGFR1 levels also increased by following the stimulation with IFN-γ or TLR ligand. Conclusions: Cytokines such as TNF-α and IFN-γ and TLR ligands may contribute to the production of angiogenic and antiangiogenic factors and may affect the placental formation.

Aim The anti-β2-GPI antibody (aβ2-GPIAb) has been detected in recurrent fetal loss with strong pathogenic activity. The effects of aβ2-GPIAb on cytokine production and aβ2-GPIAb binding sites in first-trimester trophoblast cells were evaluated. Methods First-trimester trophoblast cells were cultured in 24-well tissue culture plates with immunoglobulin G (IgG) obtained from aβ2-GPIAb-positive and aβ2-GPIAb-negative serum. Cytokines in the cultured supernatant were measured using the suspension array system and enzyme-linked immunosorbent assays. To identify potential binding sites for aβ2-GPIAb, such as toll-like receptors (TLR) 2 or TLR4, we used mouse monoclonal anti-TLR2 and/or anti-TLR4 antibodies to inhibit TLR and then measured cytokine production. Results The production of cytokines, such as interleukin-6 and interleukin-8, increased more in response to aβ2-GPIAb-positive IgG than to aβ2-GPIAb-negative IgG in trophoblast cells. The secretion of cytokines from trophoblast cells decreased when the TLR were blocked with mouse monoclonal anti-TLR2 and anti-TLR4 antibodies. Conclusion We suspect that aβ2-GPIAb might increase cytokine production by binding to TLR2 or TLR4. The increased cytokine production in response to aβ2-GPIAb might play a role in the increased inflammatory response in the placenta.

Preservation of fertility has been recommended for cancer-bearing patients of reproductive age before undergoing cancer treatment. However, there are many considerations and it is difficult to preserve fertility for all patients undergoing therapy for malignancies. Female cancer survivors had lower pregnancy and live birth rates compared with others that underwent assisted reproductive technologies (ARTs). We should continue to consider the issue of infertility in patients who underwent therapies for malignancies. This is the first report of a successful live birth in a patient with a cranial tumor who underwent radiotherapy and chemotherapy after implantation of an autologous embryo. The patient was a 27-year-old Japanese woman. She was diagnosed with suprasellar germinoma at 13 years of age, and she developed panhypopituitarism after radiotherapy and chemotherapy. At 27 years of age, she began infertility treatment with in-vitro fertilization (IVF). The level of anti-Mallerian hormone (AMH) was 4.29 ng/ml. After ovarian stimulation by high purified human menopausal gonadotropin (HP-hMG), she obtained two blastocysts and became pregnant by implantation of a cryopre- served blastocyst. At 37 gestational weeks, she delivered a healthy female baby by cesarean section.

To investigate the effects of the anti- β2-glycoprotein I (anti-β2-GPI) antibody on the tro- phoblast by evaluating the effects of the anti-β2-GPI antibody on the expression of toll-like receptor (TLR) mRNA in choriocarcinoma cells and primary trophoblast cells.We cul- tured the choriocarcinoma cells (BeWo) and the pri- mary first trimester tropho- blast with the IgGs taken from anti-β2-GPI antibody-positive and -negative sera. Four hours later we purified the RNAs from those cells. We measured the expressions of TLR mRNA in cells using real-time. PCR.The expression of TLR mRNA increased in BeWo cells and primary trophoblast cells cultured with the IgGs taken from anti-β2-GPI antibody-positive women. Specifically, the expression of TLR1, 2, and 4 in BeWo cells -and TLRI, 3, 4, and 9 in first trimester trophoblast cells increased significantly.The anti-β2-GPI antibody-positive IgG increased the TElR mRNA expression in choriocarcino- ma cells and primary trophoblast cells. We suggest that anti-β2-GPI antibodies may bind to trophoblast and increase the expression of TLR mRNA.

Background/Aim: We demonstrated that AT1 and AT2 are expressed and both pathways balance the renin-angiotensin system in endometriosis. MAS1, a specific receptor of angiotensin (1–7), opposes AT1 pathway-associated tissue remodelling. It is not known whether MAS1 has an effect on the pathogenesis of endometriosis or not. Materials and Methods: Ovarian endometriotic tissues (endo-Ov) and eutopic endometrial tissues (endo-Em) were obtained from 29 patients with endometrial cysts. Normal endometrial tissues (cont-Em) were obtained from patients without endometriosis. Immunohistochemical staining was performed for MAS1, AT1 and AT2 in the endometriosis-associated tissues. The mRNA levels of these receptors were examined by quantitative reverse transcription PCR. Results: MAS1 was immune-positive at the apical side of the glandular epithelium in the endometriotic lesions. The MAS1 mRNA levels in endo-Ov were increased significantly, irrespective of the menstrual cycle phase. The MAS1 mRNA levels were significantly higher in the proliferative-tissues of the endometriosis patients than in those of the controls. The ratio of the MAS1 to the AT1 mRNA in the proliferative tissues was increased predominantly in the endometriosis patients compared with that in the controls. Conclusion: High MAS1 expression in the endometrium might promote the initiation of endometriosis via migration of proliferative tissue.

Problem This study investigated whether angiotensin II type 1 receptor agonistic autoantibodies (AT1-AAs) mediate the increased release of soluble endoglin (sEng) in women with preeclampsia. Method of study Serum samples were obtained from women with normal pregnancies or with preeclampsia. Human first-trimester trophoblast cells were cultured with purified IgG derived from these sera, and the sEng protein and mRNA expression levels were measured in the supernatants. We also determined the effects of the AT1-AAs on these cells following treatment with an AT1 receptor antagonist (losartan). Results Compared with the IgG isolated from the women with normal pregnancies, treatments of the preeclamptic patients markedly increased sEng production and mRNA expression in trophoblast cells. Co-treatment with losartan significantly attenuated the release of sEng and sEng mRNA expression in the trophoblast cells. Conclusion AT1-AAs may be related to the increased release of sEng observed during preeclampsia and may play important roles in the pathology of this disorder.

Aim We studied the effect of pre-eclampsia sera on the expression of placenta growth factor (PlGF), soluble fms-like tyrosine kinase-1 (sFlt-1), metal-responsive transcription factor-1 (MTF-1), heme oxygenase 1 (HO-1) and hypoxia inducible factor-1α (HIF-1α) mRNAs in JEG-3 cells (trophoblast-derived cells) and placenta from pre-eclampsia patients to investigate pre-eclampsia pathophysiology. Material and Methods Placenta and serum samples were taken from pre-eclampsia and normal pregnancy patients. JEG-3 cells were cultured with pre-eclampsia and normal pregnant sera in 24-well tissue culture plates. RNA was purified from placental trophoblast cells and JEG-3 cells 24 h after incubation. The expression of mRNA was measured using real-time polymerase chain reaction. Results The expression of sFlt-1 mRNA increased, and that of PlGF and HO-1 mRNA decreased in JEG-3 cells after incubation with pre-eclampsia sera. The expression of PlGF mRNA decreased, and that of sFlt-1mRNA increased in pre-eclampsia placenta. The expression of MTF-1 and HO-1 mRNA decreased. A correlation was found between PlGF mRNA expression and the expression of MTF-1 and HIF-1α mRNA. A correlation between sFlt-1 and HIF-1α mRNA expression was also found. Conclusion Changes in PlGF mRNA expression in pre-eclampsia placenta may relate to serum factors and the expression of MTF-1 and HIF-α mRNA. Changes in sFlt-1mRNA expression may relate to serum factors and the expression of HIF-α mRNA. We suggest that serum factors play a role in PlGF and sFlt-1 expression in pre-eclampsia placenta.

A 34-year-old, gravida 0 para 0 Japanese woman visited a regional hospital complaining of dysmenorrhea, hematuria during menstruation, and right inguinal pain. She had a history of dysmenorrhea and three prior rounds of in vitro fertilization with embryo transfer, which were all with transfers of cryopreserved-thawed single embryos in natural cycles, resulting in no pregnancy. An ultrasound revealed a large 2 × 1-cm nodule between the bladder and the anterior wall of the uterus and a 3-cm cystic lesion in the right adnexal area. A combined cystoscopic and laparoscopic resection of the bladder endometriosis and cystectomy of the right endometrioma were carried out. A single ultrasound-guided transfer of a cryopreserved-thawed embryo in the cleavage stage was performed 4 months postoperatively, which resulted in an uncomplicated pregnancy. The combined, single procedure was minimally invasive and eradicated the lesions that may have caused the infertility.

A 33-year-old, gravida 3, para 2, woman was transferred to our hospital, with acute abdominal pain. Abdominal computed tomography (CT) revealed a cystic lesion accompanied by ring-enhancement between the liver and right kidney with fluid collection in the pelvic cavity. Serum hCG value was 3100 mIU/mL. Transvaginal sonography revealed a pseudo-gestational sac in a thickened endometrium. With a preoperative diagnosis of ectopic pregnancy at 7 weeks of gestation, laparotomy was performed. Following careful removal of clots between the liver and right kidney that contained a gestational sac, continuous bleeding from a defect in the Gerota's fascia of the right kidney was noted. The postoperative course was uneventful and the serum hCG concentration decreased markedly. This case demonstrates that the Gerota's fascia is a possible site of ectopic pregnancy, and that CT can identify a pregnancy in the Gerota's fascia as well as in the liver and spleen.

Problem Toll-like receptors (TLRs) are innate immune receptors that mediate the pattern recognition of, and response toward, pathogens and host-derived danger signals. We reported that cyclooxygenase-2 (COX-2) and microsomal prostaglandin E synthase (mPGES) mRNA were expressed in cases of endometriosis. The relationship between COX-2, mPGES-1, and TLR4 in endometriotic lesions has yet to be determined. Method of study Endometriosis samples were obtained from 37 patients with endometrial cysts. Endometrial tissues were obtained from patients undergoing surgical procedures for benign gynecological conditions. COX-2, mPGES-1, and TLR4 mRNA expressions were examined by real-time quantitative reverse transcription PCR (qRT-PCR) and mPGES-1, and TLR4 protein localization was examined by immunohistochemistry. Results TLR4 proteins were mostly located to the glandular epithelium. The immunoreactivities of TLR4 and mPGES-1 from endometriosis lesions were significantly higher than those in eutopic endometrium in the proliferative phase. The expression levels of mPGES-1 mRNA in peritoneal endometriosis were higher than those in eutopic endometrium in the proliferative phase. The expression of TLR4 mRNA correlates with that of mPGES-1 mRNA and not with that of COX-2 in endometriotic lesions. Conclusion Relationship between TLR4 and mPGES-1 mRNA in endometriotic lesions indicate that innate immunity may play an important role in the pathogenesis of endometriosis.

Aim: Anti-β2-glycoprotein I (anti-β2-GPI) antibody has been detected in cases of recurrent abortion and intrauterine death. Placenta growth factor (PlGF) and vascular endothelial growth factor (VEGF) are growth factors that accelerate villous proliferation. Soluble VEGF receptor-1 (sVEGFR1) is a soluble receptor for PlGF and suppresses the function of PlGF. In order to study the pathological mechanism of anti-β2-GPI antibody, we evaluated the effects of anti-β2-GPI antibody on PlGF, VEGF and sVEGFR1 production from cultured choriocarcinoma cells. Methods: The sera were taken from six anti-β2-GPI antibody-positive and six anti-β2-GPI antibody-negative patients with recurrent miscarriages. Choriocarcinoma cells (JEG-3) were cultured in 24-well plates. After each serum and isolated immunoglobulin G (IgG) were added to the culture medium, the PlGF, VEGF and sVEGFR1 in the culture medium were measured by ELISA 24 h later. When the isolated IgG was added to culture medium, only the levels of PlGF were measured. Results: The anti-β2-GPI antibody-positive sera and isolated IgG significantly suppressed the production of PlGF from JEG-3 cells more strongly than the anti-β2-GPI antibody-negative sera. The suppressive effects were not changed by serum inactivation. There was no significant difference in the values of the XTT assay and the production of VEGF and sVEGFR1 between the antibody-positive and antibody-negative sera. Conclusions: Anti-β2-GPI antibody-positive sera and IgG suppress the production of PlGF from JEG-3 cells. We suggest that the anti-β2-GPI antibodies may suppress PlGF production from trophoblasts and cause the failure of placenta formation and function.

Paragangliomas are rare tumors arising from the chromaffin cells in the autonomic nervous system. While they both occur most frequently along the paraaortic chain, paraganglioma and ovarian carcinoma very rarely occur together. A 61-year-old, post-menopausal woman visited our hospital, with complaints of abdominal pain and genital bleeding. Image analysis showed a 21 x 18 x 10 cm ovarian mass, and a 38 mm tumor at the paraaortic lesion. First, she underwent bilateral salpingo-oophorectomy. Serous papillary cystadenocarcinoma of the left ovary was found, and so a second surgery was performed. The paraaortic tumor was completely eliminated in spite of fluctuating blood pressure intraoperatively. Microscopic examination revealed that the paraaortic tumor was paraganglioma. She was ultimately diagnosed as having ovarian carcinoma stage Ia (FIGO) with coincident paraganglioma. If blood pressure fluctuation is observed during dissection of the paraaortic lymph node, paraganglioma should be suspected and blood pressure must be carefully controlled.

Problem Recently, an inducible microsomal human prostaglandin E synthase (mPGES) was identified. This enzyme converts the cyclooxygenase (COX) product, prostaglandin (PG) H2, to PGE2, an eicosanoid linked to carcinogenesis. Although elevated levels of PGE2 have been observed in many tumor types including colorectal adenomas and cancers, its role in the pathophysiology of endometriosis is unknown. We previously reported increased expression of COX-2 messenger RNA (mRNA) in local lesions of endometriosis. To further elucidate the mechanism responsible for the elevated levels of PGE2 in endometriosis, we examined the expression levels of mPGES. Method of study Samples were obtained from 28 patients, fixed in formalin, and embedded in paraffin for immunohistochemical analysis. We examined the expression of mPGES mRNA in seven cases by reverse transcriptase-polymerase chain reaction using total RNA extracted from frozen samples. Results Immunohistochemistry revealed increased mPGES immunoreactivity in endometriosis samples compared with eutopic endometria. Microsomal PGES immunoreactivity was observed in both epithelial cells and stromal or inflammatory cells of endometriosis. Increased expression of mPGES-1 mRNA was detected in most of the endometriosis samples. Conclusion Our results suggest that expression of mPGES in addition to COX-2 plays a role in increasing PGE2 production in endometriosis.

The anti-β2-GPI antibody (aβ2-GPIAb) has been detected in recurrent fetal loss with strong pathogenic activity. The effects of aβ2-GPIAb on cytokine production and aβ2-GPIAb binding sites in first-trimester trophoblast cells were evaluated.First-trimester trophoblast cells were cultured in 24-well tissue culture plates with immunoglobulin G (IgG) obtained from aβ2-GPIAb-positive and aβ2-GPIAb-negative serum. Cytokines in the cultured supernatant were measured using the suspension array system and enzyme-linked immunosorbent assays. To identify potential binding sites for aβ2-GPIAb, such as toll-like receptors (TLR) 2 or TLR4, we used mouse monoclonal anti-TLR2 and/or anti-TLR4 antibodies to inhibit TLR and then measured cytokine production.The production of cytokines, such as interleukin-6 and interleukin-8, increased more in response to aβ2-GPIAb-positive IgG than to aβ2-GPIAb-negative IgG in trophoblast cells. The secretion of cytokines from trophoblast cells decreased when the TLR were blocked with mouse monoclonal anti-TLR2 and anti-TLR4 antibodies.We suspect that aβ2-GPIAb might increase cytokine production by binding to TLR2 or TLR4. The increased cytokine production in response to aβ2-GPIAb might play a role in the increased inflammatory response in the placenta.

Introduction Placenta growth factor (PlGF) is a growth factor originated from placenta. The sFlt-1 is soluble receptor for PlGF and suppresses PlGF function. It has been reported that in preeclampsia, serum level of PlGF decreased and sFlt-1 level increased and that preeclampsia placenta is in hypoxic condition. Metal-responsive transcription factor (MTF)-1, Hemoxigenase 1 (HO-1) and Hypoxia responsive factor -1 (HIF-1) may be induced in hypoxic condition. Objectives In order to investigate pathophysiology in preeclampsia, we studied the expression of PlGF, sFlt-1, MTF-1, HO-1 and HIF-1 alpha mRNAs in placenta taken from preeclampsia and the effect of preeclampsia sera on their expression of choriocarcinoma cells and analysed the effect of placental hypoxia and serum factor on the expression of PlGF and sFlt-1 mRNA. Methods Placenta and serum samples were taken from preeclampsia and normal pregnancy with informed consent. The choriocarcinoma cells (JEG-3) were cultured in 24-well tissue culture plate. The cells were cultured with preeclampsia and normal pregnant sera. The RNAs were purified from these cells 24h after and placenta. The expressions of these mRNA were measured by using the real time PCR method (Applied Biosystems-7500). Results The expression of PlGF mRNA decreased and that of sFlt-1mRNA increased in preeclampsia placenta. The expression of MTF-1 and HO-1 mRNA decreased. The correlation was found between the expression of PlGF and MTF-1 mRNA, PlGF and HO-1 mRNA and sFlt-1 and HO-1mRNA. Moreover, expression of sFlt-1mRNA increased and HO-1mRNA decreased in JEG-3 cells after incubation of preeclampsia sera. Conclusion The changes of PlGFmRNA in preeclampsia placenta may relate to the expression of MTF-1 and HO-1 mRNA. The changes of sFlt-1mRNA may relate to the expression of HO-1 mRNA and serum factor. Not only hypoxia but also serum factor may play a role of the levels of PlGF and sFlt-1 in preeclampsia placenta.

Problem: The placenta is one of the few non-hematopoietic tissues to express granulocyte colony stimulation factor (G-CSF). Placental G-CSF production is considered to be one of the major causes of granulocytosis during pregnancy although its physiological role in pregnancy has not yet been examined. Method of study: The effects of G-CSF on interleukin (IL)-2 and/or IL-12 induced interferon (IFN)-γ production of magnetic cell sorting (MACS) sorted decidual lymphocytes was examined by enzyme-linked immunosorbent spot-forming cell assay (ELISPOT). The effect of G-CSF on cytotoxicity of decidual lymphocytes against the choriocarcinoma cell line JEG-3 was examined by lactate dehydrogenase (LDH) release assay. Results: As previously reported by us, IL-2 and/or IL-12 activated decidual mononuclear cells were capable of killing choriocarcinoma cells. We observed that G-CSF abolished IFN-γ production and cytotoxicity of decidual mononuclear cells and MACS sorted CD56+ cells. Conclusions: In addition to its well-known trophic effects on hematopoiesis, our results suggest about new roles of G-CSF in reproductive immunology.

Problem Chemokine receptors of placental trophoblasts possibly act as co-receptors or alternative receptors of maternal fetal infection by HIV. To clarify their possible expression and the physiological roles of chemokines on human placentae, we studied chemokine chemokine receptor expression and the effects of exogenous chemokines on choriocarcinoma cell lines. Materials and methods Placental samples were obtained from 13 placentae of various gestational ages. Villous tissue was mechanically dissected from samples. Trophoblasts were enriched by anti-human chorionic gonadotropin (hCG)-coated magnetic beads. Human choriocarcinoma cell lines (JAR, BeWo, JEG-3) were maintained in RPMI 1640 media supplemented with 10% FCS. Expression of chemokine receptors was studied by RT-PCR. The effects of MIP-1alpha, RANTES, MCP-1 on hCG production were estimated by EIA. Effects of chemokines on proliferation of choriocarcinoma cell lines were examined by MTT assay. Results We observed mRNA expression of CCR-1, 2, 3, 4, 5 and CXCR-1, 2, 4 in 1st trimester placental villi, CCR-I, 2, 4 and CXCR-1, 2. 4 in 2nd trimester placental villi, CCR-1, 2, 4 and CXCR-4 in 3rd trimester placental villi. Using MACS enriched trophoblasts, we observed identical results. A choriocarcinoma cell line BeWo expressed CCR-1, 3, 4 and CXCR-1, 2, 4 while JEG-3 and JAR expressed CCR-1, 3, 4, 5 and CXCR-1, 2, 4. Expression of the CCR-5 and CXCR-4 protein in choriocarcinoma cell lines and MACS-enriched trophoblats were confirmed by flow cytometry. Chemokine MCP-3, MIP-1alpha, RANTES mRNA were expressed by the 1st, 2nd and 3rd trimester placental samples and the three choriocarcinoma cell lines examined. MCP-1 was expressed by 1st and 2nd trimester placental villi. Administration of chemokines up-regulated proliferation (10(-1) - 10 ng/mL) and hCG production (10(-1) - 10(-2)ng/ mL) of the three choriocarcinoma cell lines examined. Conclusions Our results suggest possible roles of chemokines/chemokine receptors on placental physiology and their involvement in HIV transmission as alternative receptors.

PROBLEM: The mechanism of immunotherapy for patients with recurrent spontaneous abortions is not well understood. In order to investigate the suppressor mechanism of paternal lymphocyte immunization, we examined peripheral blood lymphocyte subpopulations and the repertoire of T-cell receptor (TCR) gene segments. METHOD OF STUDY: Twelve patients with recurrent miscarriage were treated with immunization with paternal lymphocyte vaccinations three times during 12-14 weeks. Before and 2 weeks after the final inoculation, lymphocyte subsets and intra-cellular interferon (IFN)-gamma and/or interleukin (IL)-4 production were examined by flow cytometry. TCR V beta and V gamma repertoires were examined by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: We found no significant difference in CD4/CD8 ratios, prevalence of CD56+CD3+ or CD57+CD3+ cells (possible extrathymic T cells), gamma(delta)T cells, and CD5+ CD19+ (B-1) cells. However, by in vitro activation with phorbol 12-myristate 13-acetate (PMA) and ionomycin, peripheral CD4 cells demonstrated a significant decrease of IFN-gamma-producing T helper 1 (Th1) cells and an increase of IL-4-producing T helper 2 (Th2) cells after immunotherapy. Seven of nine patients who exhibited remarkable decreases in Th1/Th2 ratios became pregnant within 6 months after three courses of immunotherapy, and four women delivered healthy babies, while none of the three patients who exhibited an increased or unchanged Th1/Th2 ratio had full-term pregnancies (chi2 < 0.0001). Further, changes in usage of TCR V beta and V gamma gene segments were observed after immunotherapy in six patients examined. CONCLUSION: Our findings suggest a shift of Th1-dominant to Th2-dominant status by vaccination might play important roles in maintaining successful pregnancies. Induction of some T cells that utilize different TCR repertoires possibly suppresses maternal rejection reactions.

A 34-year-old, gravida 0 para 0 Japanese woman visited a regional hospital complaining of dysmenorrhea, hematuria during menstruation, and right inguinal pain. She had a history of dysmenorrhea and three prior rounds of in vitro fertilization with embryo transfer, which were all with transfers of cryopreserved-thawed single embryos in natural cycles, resulting in no pregnancy. An ultrasound revealed a large 2 × 1-cm nodule between the bladder and the anterior wall of the uterus and a 3-cm cystic lesion in the right adnexal area. A combined cystoscopic and laparoscopic resection of the bladder endometriosis and cystectomy of the right endometrioma were carried out. A single ultrasound-guided transfer of a cryopreserved-thawed embryo in the cleavage stage was performed 4 months postoperatively, which resulted in an uncomplicated pregnancy. The combined, single procedure was minimally invasive and eradicated the lesions that may have caused the infertility.

A 33-year-old, gravida 3, para 2, woman was transferred to our hospital, with acute abdominal pain. Abdominal computed tomography (CT) revealed a cystic lesion accompanied by ring-enhancement between the liver and right kidney with fluid collection in the pelvic cavity. Serum hCG value was 3100 mIU/mL. Transvaginal sonography revealed a pseudo-gestational sac in a thickened endometrium. With a preoperative diagnosis of ectopic pregnancy at 7 weeks of gestation, laparotomy was performed. Following careful removal of clots between the liver and right kidney that contained a gestational sac, continuous bleeding from a defect in the Gerota's fascia of the right kidney was noted. The postoperative course was uneventful and the serum hCG concentration decreased markedly. This case demonstrates that the Gerota's fascia is a possible site of ectopic pregnancy, and that CT can identify a pregnancy in the Gerota's fascia as well as in the liver and spleen.

Anti-β2-glycoprotein I (anti-β2-GPI) antibody has been detected in cases of recurrent abortion and intrauterine death. Placenta growth factor (PlGF) and vascular endothelial growth factor (VEGF) are growth factors that accelerate villous proliferation. Soluble VEGF receptor-1 (sVEGFR1) is a soluble receptor for PlGF and suppresses the function of PlGF. In order to study the pathological mechanism of anti-β2-GPI antibody, we evaluated the effects of anti-β2-GPI antibody on PlGF, VEGF and sVEGFR1 production from cultured choriocarcinoma cells.The sera were taken from six anti-β2-GPI antibody-positive and six anti-β2-GPI antibody-negative patients with recurrent miscarriages. Choriocarcinoma cells (JEG-3) were cultured in 24-well plates. After each serum and isolated immunoglobulin G (IgG) were added to the culture medium, the PlGF, VEGF and sVEGFR1 in the culture medium were measured by ELISA 24 h later. When the isolated IgG was added to culture medium, only the levels of PlGF were measured.The anti-β2-GPI antibody-positive sera and isolated IgG significantly suppressed the production of PlGF from JEG-3 cells more strongly than the anti-β2-GPI antibody-negative sera. The suppressive effects were not changed by serum inactivation. There was no significant difference in the values of the XTT assay and the production of VEGF and sVEGFR1 between the antibody-positive and antibody-negative sera.Anti-β2-GPI antibody-positive sera and IgG suppress the production of PlGF from JEG-3 cells. We suggest that the anti-β2-GPI antibodies may suppress PlGF production from trophoblasts and cause the failure of placenta formation and function.