Oxidative stress-induced S100B protein from placenta and amnion affects soluble Endoglin release from endothelial cells.

Oxidative stress with elevated intracellular Ca(2+) concentration as well as endothelial dysfunction is a component of pre-eclampsia. Our aim was to investigate the oxidative stress-dependent expression of Endoglin and Ca(2+)-binding S100B protein from villous and amniotic tissue cultures, and to assess sEng expression from S100B protein-stimulated endothelial cells. We initially examined Endoglin and Hydroxy-nonenal-(HNE)-modified proteins in the placentas and amnion obtained from women with pre-eclampsia (n = 8), and healthy controls (n = 8) by immunohistochemistry. To examine oxidative stress and the S100B protein effect on sEng expression from endothelial cells, normal villous and amniotic tissue cultures were stimulated by 4-HNE, sodium fluoride and xanthine/xanthine oxidase, whereas human umbilical vein endothelial cell cultures were treated with S100B protein in a dose- and time-dependent manner at 37 degrees C in an environment of 95% air and 5% of CO(2). Culture supernatants were assessed using ELISA. Cell viability was determined using MTS assay. The concentrations of sEng and S100B protein were significantly increased in the villous and amniotic tissue culture supernatants under oxidative stress. S100B protein-stimulated endothelial cells released sEng into conditioned media with a significantly higher expression levels at a concentration of 200 pM-20 nM S100B by 2 h, whereas treated with 200 nM of S100B endothelial cells significantly expressed sEng by 12 h and stimulated the cell proliferation by the same period of time. Our findings show that oxidative stress affects sEng and S100B protein expression from villous and amniotic tissues, and picomolar and low nanomolar concentrations of S100B protein significantly up-regulate sEng release from endothelial cells leading to endothelial dysfunction.

Combination chemotherapy of intraperitoneal carboplatin and intravenous paclitaxel in suboptimally debulked epithelial ovarian cancer.

The objective of this study was to retrospectively assess the efficacy and safety of combination chemotherapy of intraperitoneal (IP) carboplatin and intravenous (IV) paclitaxel in suboptimally debulked ovarian cancer. Between March 1998 and March 2006, 44 patients with histologically confirmed epithelial ovarian carcinoma or peritoneal carcinoma with a residual mass greater than 1 cm received combination chemotherapy of IV paclitaxel and IP carboplatin. Administration of IV paclitaxel at 175 mg/m(2) immediately followed by IP carboplatin at an area under the curve of 6 was scheduled every 3 weeks for at least six cycles. The diagnosis and stage were ovarian carcinoma stage II in 8, III in 25, and IV in 6 cases, and peritoneal carcinoma stage III in 5 cases. Eighty-three percent of patients completed more than six cycles of chemotherapy. The incidences of grade 3/4 hematologic toxicities were 41 (93%) for neutrocytopenia, 10 (41%) for thrombocytopenia, and 18 (23%) for anemia. Observed grade 3/4 nonhematologic toxicities were 1 (2%) for allergy, 1 (2%) for fatigue, 1 (2%) for vomiting, 1 (2%) for liver dysfunction, and 4 (9%) for peripheral neuropathy. Two patients (5%) encountered catheter problems (one obstruction and one infection). Overall response rate was 80% (16 complete response, 19 partial response, 3 stable disease, and 6 progressive disease). Median progression-free survival and overall survival were 24 and 31 months, respectively. Combination chemotherapy of IP carboplatin and IV paclitaxel is effective and safe in suboptimally debulked ovarian cancer, and further evaluation is warranted.

The human tumor-associated antigen RCAS1 in pregnancies complicated by pre-eclampsia.

The human tumor-associated antigen RCAS1 (receptor-binding cancer antigen expressed on SiSo cells) is considered to play a role in the escape of tumor cells from immune surveillance and, at the same time, participates in the inhibition of the maternal immune response during pregnancy. The aim of our study was to investigate the expression of tumor-associated RCAS1 protein in the placenta and amniotic membranes and to assess and compare its concentration in amniotic fluid, maternal and cord blood sera in pregnancies complicated by pre-eclampsia. Samples were obtained from women with pre-eclampsia (N=9), pre-eclampsia with IUGR (N=4), normotensive IUGR (N=7) and healthy term controls (N=25) after delivery. Placentas were studied by immunohistochemistry, Western blot analysis and real-time (RT)-PCR. For assessment of RCAS1 protein concentrations in biological fluids, ELISA was performed. RCAS1 mRNA expression in the placentas of pre-eclamptic patients was significantly lower than in controls (p<0.01). The maternal blood serum RCAS1 protein concentration in the pre-eclampsia cases was also significantly lower than in controls (p=0.0207). The other study groups did not differ significantly. This study reveals the possible role of the RCAS1 protein in the development of pre-eclampsia through an immunological pathway.

Transient local overexpression of human vascular endothelial growth factor (VEGF) in mouse feto-maternal interface during mid-term pregnancy lowers systemic maternal blood pressure.

The effect on maternal circulation of transient human vascular endothelial growth factor (VEGF) (165) cDNA transfection into the mouse feto-maternal interface at day 14.5 post coitus (p.c.) using a hemagglutinating virus of Japan-envelope (HVJ-E) vector system is reported. On day 15.5 p.c., Western blotting clearly showed overexpression of 18 kD VEGF protein in the uterus. After VEGF transfection, the blood pressure was significantly lowered for 48 hours. On day 17.5 p.c., the blood pressure returned to the control level. Proteinuria was not observed after VEGF transfection. No preterm birth was observed during the course of pregnancy after the transfection procedure. After 24 hours of transfection, human VEGF was not detectable and the mouse VEGF level was similar to that in peripheral blood. However, the soluble fms-like tyrosine kinase (Flt)-1 concentration was significantly lower in VEGF-transfected mice. These results suggest that extraamniotic VEGF overexpression lowered the systemic blood pressure without altering the VEGF concentration in the peripheral blood. Local overexpression of VEGF may become a novel treatment for pregnancy-related disorders such as hypertension complicated-pregnancy and preeclampsia.

S100B protein expression in the amnion and amniotic fluid in pregnancies complicated by pre-eclampsia.

Our aim was to investigate the expression of S100B protein in the amnion and to assess the amniotic fluid concentration in pregnancies complicated by pre-eclampsia. Samples were obtained from women who developed pre-eclampsia (n = 7), pre-eclampsia with intrauterine growth retardation (IUGR) (n = 4), normotensive IUGR (n = 7) and gestational hypertension (n = 4) during pregnancy and healthy controls who delivered at term (n = 35). To determine the difference in the expression of S100B in the amnion, we performed immunohistochemistry, western blot analysis and RT-PCR. Using enzyme-linked immunosorbent assay (ELISA), we assessed the S100B concentration in amniotic fluid. The S100B mRNA expression in the amnion of pre-eclamptic patients and patients with pre-eclampsia with IUGR was significantly higher than that in the control. The amniotic fluid S100B protein concentration of the pre-eclampsia and normotensive IUGR cases was significantly higher than that of the control. This study shows that amnion could be a source responsible for the increased concentration of S100B in amniotic fluid. In pre-eclampsia, reactive oxygen species (ROS) are generated by oxidative stress. Some pathological conditions that develop during pregnancy and are related to hypoxic stress can affect the elevation of S100B concentration in the amnion.

The expression of fractalkine in the endometrium during the menstrual cycle.

OBJECTIVE: The objective of the study was to evaluate the presence of fractalkine in the endometrium of the uterus and the change of fractalkine protein levels during menstrual cycle. METHODS: Twelve samples of endometrium of the uterus were obtained from gynecological patients who underwent total hysterectomy. Western blotting, RT-PCR and immunohistochemistry were performed. RESULTS: Fractalkine protein was detected in the endometrium of the uterus. Positive staining was confirmed in the epithelial cells and grandular cells in the endometrium. Expression levels of fractalkine protein and mRNA in the endometrium during secretory phase were significantly higher than those during proliferative phase. Immunohistochemical analysis using an anti-CX3CR1 antibody demonstrated positive staining in the glandular cells of the endometrium of the uterus. CONCLUSION: Fractalkine was expressed in the endometrium and its production was up-regulated during secretory phase.

Effect of coffee intake on blood flow and maternal stress during the third trimester of pregnancy.

OBJECTIVE: To investigate the effects of maternal ingestion of an ordinary dose of coffee on maternal stress and placental and fetal blood circulation during the third trimester of pregnancy. METHODS: We performed a Doppler blood flow analysis for 10 women in the third trimester of pregnancy before and after they drank a cup of coffee. Salivary samples were collected from the 10 pregnant women and 14 nonpregnant controls just before coffee intake and 30 min later. Salivary cortisol levels and chromogranin A titers were determined. RESULTS: Coffee intake had no effect on maternal or fetal blood flow. Among the pregnant women, Salivary cortisol levels were significantly reduced after coffee intake but salivary chromogranin A concentration was not significantly different before and after coffee intake. CONCLUSION: The reduced salivary cortisol levels suggest that coffee intake decreases maternal stress during pregnancy.

Fractalkine in the peritoneal fluid of women with endometriosis.

OBJECTIVE: The objective of this study was to evaluate the presence of fractalkine in the ascites and the association between fractalkine levels in the ascites and endometriosis. METHODS: Peritoneal fluids and peripheral blood samples were obtained from patients undergoing laparoscopy for infertility work-up or laparoscopic cystectomy. Three samples of peritoneum were obtained from patients undergoing hysterectomy. Western blotting, RT-PCR and immunohistochemistry were performed. RESULTS: Fractalkine protein was detected in the ascites. Positive staining was confirmed in peritoneal surface cells and perivascular cells of the peritoneum. CX3CR1 positive cells were present in the cells in the peritoneal fluid. The fractalkine concentrations in the ascites of patients with endometriosis were lower than those without endometriosis. There was no significant difference between serum fractalkine levels in patients with and without endometriosis. CONCLUSION: The decreased level of fractalkine found in the peritoneal fluid of patients with endometriosis may contribute to the pathogenesis of endometriosis.

Helicobacter pylori infection and platelet counts during pregnancy.

OBJECTIVE: To investigate whether there is any correlation between Helicobacter pylori infection and platelet counts during pregnancy. METHODS: One hundred and twenty pregnant women without any complications were evaluated. Platelet counts were determined during pregnancy and postpartum. H. pylori infection was assessed by the detection of serum antibodies at the 3rd day postpartum. Statistically significant differences of platelet count between H. pylori-positive and -negative patients were determined. RESULTS: H. pylori infection was found in 29 of 120 (24.2%). Platelet counts in the 3rd trimester were 22.7+/-4.7 x 10(3)/microl in H. pylori-positive group and 22.8+/-5.6 x 10(3)/microl in negative group (p=0.98). Platelet count in the 3rd trimester in positive and negative patients were 93+/-17% and 94+/-20%, respectively, of that of the 1st trimester (p=0.92). The incidence of hyperemesis gravidarum in both groups was not significant (p=0.28). CONCLUSION: A correlation between H. pylori infection and thrombocytopenia during pregnancy was not found in this study.

Fetal cerebellum: US appearance with advancing gestational age.

PURPOSE: To evaluate the change in ultrasonographic (US) appearance of the fetal cerebellum with advancing gestation. MATERIALS AND METHODS: A total of 291 normal fetuses of uncomplicated pregnancies were evaluated at gestational ages (GAs) between 15 and 41 weeks with a 3.75-MHz transabdominal curvilinear probe. After the transcerebellar view was obtained, the transverse cerebellar diameter (TCD) was measured and the images were stored. On hard-copy US images, cerebella were assigned three grades of appearance. These grades were analyzed in relation to GA and TCD. Inter- and intraobserver variations were assessed in 91 randomly selected cases. RESULTS: Cerebella in 137 (47.1%), 71 (24.4%), and 83 (28.5%) of 291 subjects were classified as grade I (hypoechoic, "eyeglass" shape), grade II (intermediate echogenicity, "dumbbell" outline), and grade III (hyperechoic, "fan" shape), respectively. With advancing gestation, the dominant grade changed from I to III gradually and progressively. The median GA and TCD, respectively, were 22 weeks and 22 mm for grade I, 29 weeks and 35 mm for grade II, and 36 weeks and 46 mm for grade III. These differences were statistically significant (P <.001). The agreements within inter- and intraobserver estimations were 96% (87 of 91) and 95% (86 of 91), respectively. CONCLUSION: A gradual change in US appearance of the fetal cerebellum is seen with advancing gestation.

[A case of papillary adenocarcinoma of the ovary that responded favorably to irinotecan hydrochloride and cisplatin after the administration of paclitaxel and carboplatin].

Recently, the standard treatment for advanced ovarian cancer has changed from CP therapy (cyclophosphamide, cisplatin (CDDP)) to TJ therapy (paclitaxel (TXL), carboplatin (CBDCA)). Irinotecan (CPT-11) is one of the derivatives of camptotecin and has been reported to have a high efficacy for ovarian cancer. In one case of ovarian cancer, chemotherapy was applied with CBDCA and TXL. However, after 2 months of six courses of the chemotherapy, CA-125 was elevated. The elevation of tumor marker levels in serum without the recurrent focus forced us to treat the patient with CPT-11 and CDDP for the second line chemotherapy. Tumor marker levels improved at the beginning of the therapy. In conclusion, CPT-11 and CDDP was effective against the recurrence of ovarian cancer treated with TJ therapy.

Production of secretory leukocyte protease inhibitor by human amniotic membranes and regulation of its concentration in amniotic fluid.

Secretory leukocyte protease inhibitor (SLPI) is a potent inhibitor of human leukocyte elastase. SLPI is a protein found in various human fluids, including parotid secretions, cervical mucus, seminal plasma and ascites. Western blot analysis revealed that SLPI protein is detected as a 12 kDa band in both the amniotic fluid and the amniotic membrane. The amniotic fluid concentrations of SLPI were assayed by enzyme-linked immunosorbent assay. SLPI concentrations in the amniotic fluid of women in the third trimester were higher than those in the second trimester. Immunohistochemistry using an anti-SLPI polyclonal antibody revealed positive staining in epithelial cells in amniotic membranes. Reverse transcription-polymerase chain reaction demonstrated that SLPI transcripts could be detected in the amniotic membranes. To determine the mechanism of SLPI production by amniotic cells, purified amniotic cells were stimulated with various cytokines. Amniotic cells produced SLPI in a dose-dependent manner when stimulated with interleukin (IL)-1alpha, IL-1beta, and tumour necrosis factor-alpha. The present findings show that SLPI is produced by the amniotic membranes in response to cytokine concentrations. The SLPI in the amniotic fluid may contribute to immunodefence mechanisms during pregnancy.

Elevations of group II phospholipase A2 concentrations in serum and amniotic fluid in association with preterm labor.

OBJECTIVE: The purpose of this study was to determine whether the elevation of secretory group II phospholipase A(2) concentration in the serum and amniotic fluid in preterm labor is associated with intrauterine inflammation. STUDY DESIGN: Serum and amniotic fluid were collected from women with preterm delivery (<37 weeks' gestation; n = 38) and term delivery (n = 20). Phospholipase activity was measured with a highly sensitive system that was based on reverse-phase high-performance liquid chromatographic separation of 9-anthracenylmethyl derivatives of fatty acids released by phospholipase A(2). The concentrations of immunoreactive isozymes (group I or II) of secretory phospholipase A(2) were assayed with a radioimmunoassay kit with a monoclonal antibody against human pancreatic phospholipase A(2) and splenic IIA phospholipase A(2). Localization of immunoreactive group II phospholipase A(2) in the amniotic membrane was determined by immunostaining visualized with the Vectastain ABC (Vector Laboratories, Inc, Burlingame, Calif) method. RESULTS: Enzymatic activities of phospholipase A(2) in the serum and amniotic fluid specimens obtained from patients in preterm labor with chorioamnionitis were significantly higher than those in specimens from patients in term labor. Significant elevations of phospholipase A(2) activities were observed in patients with preterm labor without histologically evident chorioamnionitis. The activity of phospholipase A(2) was clearly correlated with the concentration of the immunoreactive group II phospholipase A(2). Group II phospholipase A(2) was localized in amniotic cells obtained from patients with a pathologically determined diagnosis of chorioamnionitis. The predictive value for chorioamnionitis of the group II phospholipase A(2) concentration was relatively higher than the predictive values of the concentrations of C-reactive protein and interleukins 6 and 8. CONCLUSION: Significant elevations of group II phospholipase A(2) concentrations were detected in the serum and amniotic fluid of women with preterm labor. Group II phospholipase A(2) concentration may be a useful indicator for preterm labor, and phospholipid metabolism is certainly activated both in preterm labor and in apparent inflammatory diseases.

Chorioamnionitis decreased incidence of respiratory distress syndrome by elevating fetal interleukin-6 serum concentration.

Respiratory distress syndrome (RDS) of newborns is one of the most important factors determining neonatal morbidity and mortality. The interleukin-6 (IL-6) titre in cord sera of RDS-free neonates born to mothers with histological chorioamnionitis was significantly higher than that in RDS-complicated neonates without chorioamnionitis. Maternal administration of glucocorticoid suppressed the IL-6 concentrations in the cord sera of fetuses with chorioamnionitis. The fetuses without chorioamnionitis who suffered from RDS even after maternal glucocorticoid administration showed a similar IL-6 titre to that of RDS-affected neonates without chorioamnionitis. Examination of the mechanism by which IL-6 decreased the incidence of fetal RDS revealed that H441-4, a human pulmonary adenocarcinoma cell line, stimulated with recombinant (r)-IL-6 started the synthesis of mRNA and protein of pulmonary surfactant protein (SP)-A. The present study shows that IL-6 elevation in fetuses with chorioamnionitis promotes fetal lung maturation by inducing SP-A synthesis, thereby decreasing the incidence of RDS in the preterm neonates.

Increased concentrations of secretory leukocyte protease inhibitor in peritoneal fluid of women with endometriosis.

Secretory leukocyte protease inhibitor (SLPI) is a potent inhibitor of human leukocyte elastase. We investigated whether SLPI was present in the peritoneal fluid of women with endometriosis and to clarify the role of SLPI in the pathogenesis of endometriosis. Western blot analyses revealed that SLPI protein was detected as a 12 kDa band in peritoneal fluid. The peritoneal fluid concentrations of SLPI, elastase and interleukin-6 were assayed by enzyme-linked immunosorbent assays (ELISA). SLPI concentrations and the SLPI/elastase ratio in the peritoneal fluid of women with endometriosis were higher than in samples from women without endometriosis. There was no significant correlation between concentrations of SLPI and interleukin-6 in the peritoneal fluid. Immunohistochemistry using an anti-SLPI polyclonal antibody revealed positive staining in peritoneal macrophages, but not lymphocytes. The present findings suggest that SLPI found in the peritoneal fluid of patients with endometriosis may contribute to the pathogenesis of endometriosis.

Cytokine production in chorioamnionitis.

Lymphohematopoietic cytokines play a significant role in many biological mechanisms including a number of reproductive processes such as ovulation, implantation, placentation, cervical dilation and parturition. Recent experiments have suggested that cytokines play a crucial role in the mechanisms of preterm labor and delivery, which are the leading causes of perinatal morbidity and mortality. Growing evidence suggests that infection is deeply concerned in the pathogenesis of preterm labor and delivery. Chorioamnionitis, a subset of intrauterine infection, has been identified in 20-33% of women with preterm delivery, and the inflammatory and related cytokines, interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6) and interleukin-8 (IL-8), showed substantial increases in the amniotic fluid at women with intrauterine infection. Although the precise mechanism for chorioamnionitis-driven preterm labor mediated via cytokines is still unknown, both IL-1 and TNF-alpha along with IL-6 enhance prostaglandin production by human amnion cells, chorionic cells and decidual cells. Analysis of the regulatory sequences in the 5' upstream regions of receptor gene for human oxytocin, a potent uterotonic agent, suggests a close relationship between preterm labor and inflammatory cytokines through induction at the oxytocin receptor. Prompt identification of the patients with intra-amniotic infection may be useful in clinical practice. At present, the measurement of IL-8 in maternal serum or the measurement of IL-6 in cervical secretion may be helpful as a non-invasive screening for chorioamnionitis.

Oncostatin M is produced during pregnancy by decidual cells and stimulates the release of HCG.

Oncostatin M (OSM) is a member of the interleukin-6 superfamily and a multifunctional cytokine that effects the growth and differentiation of many different cell types. OSM concentrations in the sera of pregnant women were found to be significantly higher than those of non-pregnant women. Western blot analysis revealed that the OSM protein was present in the decidua and chorionic tissue in each trimester. Throughout pregnancy, the amount of the OSM protein in the decidua was larger than that in the chorionic tissue. Immunohistochemistry using an anti-OSM monoclonal antibody demonstrated that OSM was mainly localized in the decidual glands and stroma. OSM transcripts in the decidua and the chorionic tissue were detected during each trimester by reverse transcription-polymerase chain reaction (RT-PCR). The regulation of human chorionic gonadotrophin (HCG) release by the placenta in first trimester stimulated with recombinant OSM was also investigated. Stimulation of the placenta by OSM augmented HCG release in a time- and dose-dependent manner. HCG release induced by recombinant human OSM was completely blocked by antibodies against OSM and the signal transducer, gp130, but only partially inhibited by antibodies against the leukaemia inhibiting factor (LIF) receptor. These results suggest that OSM molecules produced by decidual glands and stromal cells during pregnancy have an important role in placental endocrine function.