Green tea catechins and their oxidative protection in the rat eye.

Catechins, active constituents of green tea, are well-known antioxidative natural products. It was proposed that green tea extract (GTE) consumption could benefit the eye, and the pharmacokinetics of catechins and oxidation status in rat eye were investigated after oral administration. Sprague-Dawley rats were fed GTE and sacrificed at different time intervals. Their eyes were dissected into cornea, lens, retina, choroid-sclera, vitreous humor, and aqueous humor for analysis of catechins and 8-epi-isoprostane by HPLC-ECD and GC-NCI-MS, respectively. Catechins were differentially distributed in eye tissues. Gallocatechin was present at the highest concentration in the retina, 22729.4 +/- 4229.4 pmol/g, and epigallocatechin in aqueous humor at 602.9 +/- 116.7 nM. The corresponding area-under-curves were 207,000 pmol x h/g and 2035.0 +/- 531.7 nM x h, respectively. The time of maximum concentration of the catechins varied from 0.5 to 12.2 h. Significant reductions in 8-epi-isoprostane levels were found in the compartments except the choroid-sclera or plasma, indicating antioxidative activities of catechins in these tissues.

Urgent cesarean delivery for fetal bradycardia.

OBJECTIVE: To estimate whether fetal bradycardia-to-delivery interval or decision-to-delivery interval was related to cord arterial pH according to different causes of fetal distress. METHODS: Women who delivered singleton neonates by urgent cesarean delivery because of intrapartum fetal bradycardia were retrospectively categorized into three groups according to the cause of fetal bradycardia: 1) Irreversible; 2) Potentially Reversible; and 3) Unknown (no identifiable cause). Comparisons were made between groups in regard to pH, bradycardia-to-delivery interval, and decision-to-delivery interval. Correlation analyses between pH and both intervals were then performed for different groups. RESULTS: Of 235 cases, 39, 22, and 174 were respectively categorized into the Irreversible group, Potentially Reversible group, and Unknown group. The median pH was lower in the Irreversible group (7.094; interquartile range [IQR] 6.991-7.216) than in Potentially Reversible group (7.162; IQR 7.064-7.251) or Unknown group (7.210; IQR 7.161-7.255) (P<.001). The Irreversible group's median bradycardia-to-delivery interval was 5 minutes shorter than those of the other two groups (11 compared with 16.5 and 16 minutes, respectively; P<.001), whereas its median decision-to-delivery interval was 1 minute shorter (10 compared with 11.5 and 11 minutes, respectively; P=.001). In the Irreversible group, pH decreased with the bradycardia-to-delivery interval (Spearman's rho=-0.354; P=.027) at a rate of 0.011 per minute. Cord arterial pH did not correlate with the bradycardia-to-delivery interval in the Potentially Reversible and Unknown groups. In neither group did pH correlate with decision-to-delivery interval. CONCLUSION: Cord arterial pH deteriorates with bradycardia-to-delivery interval when the underlying cause of fetal distress is irreversible, but not so otherwise. LEVEL OF EVIDENCE: II.

Glucose intolerance and cardiometabolic risk in children exposed to maternal gestational diabetes mellitus in utero.

OBJECTIVE: The goal was to examine the carbohydrate tolerance and cardiometabolic risk among children exposed to maternal gestational diabetes mellitus in utero. METHODS: In this study, 164 Chinese children whose mothers had participated in a previous study on the screening and diagnosis of gestational diabetes mellitus (63 had gestational diabetes mellitus and 101 had normal glucose tolerance during the index pregnancies) underwent follow-up evaluations at a median age of 8 years (range: 7-10 years). Children's weight, height, hip and waist circumferences, and blood pressure were measured, and weight-adjusted oral glucose tolerance tests were performed. RESULTS: Six children (3.7%) demonstrated impaired glucose regulation or diabetes mellitus at the follow-up evaluation. Children exposed to maternal gestational diabetes mellitus had significantly higher systolic (94+/-1.2 vs 88+/-0.9 mmHg) and diastolic (62+/-0.8 vs 57+/-0.6 mmHg) blood pressure values and lower high-density lipoprotein cholesterol (1.58+/-0.04 vs 1.71+/-0.03 mmol/L) levels, after adjustment for age and gender. A high (>or=90th percentile) umbilical cord insulin level at birth was associated with abnormal glucose tolerance in the offspring. CONCLUSIONS: Maternal gestational diabetes mellitus increases the offspring's cardiometabolic risk, and in utero hyperinsulinemia is an independent predictor of abnormal glucose tolerance in childhood.

Method to determine stability and recovery of carboprost and misoprostol in infusion preparations.

The two synthetic prostaglandin analogues, carboprost and misoprostol, are used extensively in obstetric and gynaecological practice. Our recent research of these compounds' use for intra-umbilical injection to treat adherent placenta necessitated their storage in solution for 3-4 days. This raised concerns over the stability and applied dosage in the in-house infusion preparations. It requires various pharmacological preparations before administration in clinical practice. We used LCMS to develop a simultaneous, valid, fast and simple method to assess the stability and recovery of their in-house preparations in different conditions. The linearity between 0-40 microg/ml was above 0.995. The reproducibility (CV) was within 5.2%. The limit of quantitation of the method for both compounds is about 2 microg/ml. The accuracy of both compounds from 0.4-40 microg/ml is 96.4-104.3% while the precision is 0.4-7.4%. The recoveries of carboprost in the infusion were from 100.3+/-4.0 to 102.4+/-1.6% and that of misoprostol in Cytotec tablet was from 44.9+/-3.5 to 50.0+/-5.0% in water and saline at 4 degrees C and room temperature. No interference was found from the matrix and between the tested compounds. The compounds were basically stable for 6 days in water and in saline, whether they were stored at 4 degrees C or at room temperature. However, only half of the dosage of misoprostol was recovered in the solution. Therefore, misoprostol dosage should be adjusted before clinical application.

Tea epigallocatechin-3-gallate increases 8-isoprostane level and induces caudal regression in developing rat embryos.

Tea is the most common beverage after water. Concerns have been raised about the safety of tea during pregnancy, especially for embryo development. We aimed at studying the effects of active tea components on developing embryos by in vitro rat embryo culture. Rat embryos during early organogenesis were cultivated in serum supplemented with one of the tea catechins. Developmental hallmarks and malformations (Mal) in the developing embryos were compared and evaluated by a standard morphological scoring system. The embryotoxicity of each tea catechin was classified according to the European Center for the Validation of Alternative Methods. Cell viability was assessed by supervital dye staining, apoptosis by TUNEL assay, and peroxidation by the 8-isoprostane EIA method. We found that (+)-catechin had the least effect on developing embryos (Mal(50)=715.1 mg/L; IC50(Mal)=435 mg/L), whereas (-)-epigallocatechin gallate had the most adverse effect (Mal(50)=54.2 mg/L; IC50(Mal)=45.8 mg/L). The major malformation in affected embryos included caudal retardation with abnormal axial flexion and delayed hind-limb formation. All catechins were classified as nonembryotoxic except (-)-epigallocatechin gallate, which was classified as weakly embryotoxic. With (-)-epigallocatechin gallate, increased numbers of nonviable and apoptotic cells in the malformed embryos were associated with increased embryo 8-isoprostane.

Pharmacokinetic studies of green tea catechins in maternal plasma and fetuses in rats.

We carried out a pharmacokinetic study to determine the levels and profiles of catechins in pregnant rats and their fetuses after ingestion of green tea extract (GTE). We measured total catechin levels after enzyme digestions. Dams, at 15.5 days of gestation, were fed with GTE and catechins were measured in the maternal plasma, placenta, and fetus 0, 0.5, 1, 2, 3, 5, 8, 10, 12, 16, and 20 h after maternal GTE intake. The pharmacokinetic changes were analyzed by non compartmental models. We found that maternal plasma concentrations of catechins were about 10 times higher than in placenta and 50-100 times higher than in the fetus. AUC and Cmax levels of (-)-epicatechin (EC) were the highest in plasma while the levels of (-)-epigallocatechin gallate (EGCG) were the highest in the placenta and the fetus. The exposure level of catechin derivatives was higher than the gallate derivatives in maternal plasma after normalization but reversed in the placenta and fetus. The absorption of epi-isomers in plasma was found to be more favorable than their non epi-isomer counterparts. EGCG had the highest level of exposure (AUC) and the highest Cmax in the fetus, implying it may have potential for in utero antioxidant protection.

Health status function after treatment with thermal balloon endometrial ablation and levonorgestrel intrauterine system for idiopathic menorrhagia: a randomized study.

AIM: To compare patients' health status function after treatment with thermal balloon endometrial ablation (TBEA) and levonorgestrel intrauterine system (LNG-IUS) for idiopathic menorrhagia. METHODS: Forty-four patients were recruited into a randomized trial comparing their health status after treatment with TBEA or LNG-IUS for idiopathic menorrhagia. RESULTS: At 1 year follow-up, the mean haemoglobin was significantly higher in women treated with TBEA (12.6 g/dl vs. 10.3 g/dl, p = 0.018). Iron deficiency occurred in 13.3% from the TBEA arm and in 50% from the LNG-IUS arm (p = 0.026). The women's mean Short Form 36 Questionnaire general health perception scores (54.9 vs. 40.5, p = 0.024) and mental health scores (49.5 vs. 38.3, p = 0.021) in TBEA arm were significantly higher than in the LNG-IUS arm. The mental health domain score was also significantly lower in the LNG-IUS arm (46.1 vs. 38.3, p = 0.041). CONCLUSION: TBEA appears to offer better health status function at 1 year follow-up and to be more acceptable to our Chinese population in the treatment of idiopathic menorrhagia following failed medical treatment.

Metabolomic and bioinformatic analyses in asphyxiated neonates.

OBJECTIVES: We tested the application of bioinformatic algorithms in studying the metabolomic profiles of neonatal urine samples with clinical evidence of severe asphyxia at birth and subsequent neurodevelopmental handicap. DESIGN AND METHODS: The clinical outcomes of 256 newborns that required direct admission to neonatal intensive care unit for respiratory support or did not require direct admission were studied. Urinary metabolite profiles were measured by high throughput mass spectrometry and analyzed by bioinformatic methods. RESULTS: We found a positive relationship between suppressed biochemical networks involved in macromolecular synthesis and birth asphyxia associated with significant neonatal oxidative stress and morbidity. The metabolomic discriminators between good neonatal outcome and poor neonatal outcome were established using hierarchical clustering analysis. Concentrations of eight urinary organic acids in distinct biochemical pathways were elevated and significantly associated with the prognosis of neurodevelopmental handicap with high sensitivity and specificity: ethylmalonate, 3-hydroxy-3-methylglutarate, 2-hydroxy-glutarate and 2-oxo-glutarate were associated with good neonatal outcome, whereas glutarate, methylmalonate, 3-hydroxy-butyrate and orotate were associated with poor outcome. CONCLUSIONS: The data demonstrated the potential application of bioinformatics methods in this metabolomic study and proved its clinical relevance.

Meconium-stained liquor during labor is associated with raised neonatal cord blood 8-iso-prostaglandin F2alpha concentration.

OBJECTIVE: The purpose of this study was to compare the umbilical arterial 8-iso-prostaglandin F2alpha, concentrations between pregnancies that were complicated by moderate or thick meconium-stained liquor and those with clear liquor. STUDY DESIGN: Umbilical cord arterial blood samples were collected from 247 singleton pregnancies with either moderate or thick meconium-stained liquor at any stage of labor or clear liquor at all stages of labor for the determination of the total 8-iso-prostaglandins F2alpha concentration. RESULTS: The median total 8-iso-prostaglandins F2alpha concentration of the meconium-stained liquor group was significantly higher than that of the control group (719.2 vs 115.8 pg/mL). Among the meconium-stained liquor group, those who had a change from "clear liquor" at early labor to "moderate/ thick meconium-stained liquor" at late first stage or at delivery (late meconium-stained liquor group) had higher 8-iso-prostaglandins F2alpha concentration, compared with those who had moderate/ thick meconium-stained liquor since early labor (early meconium-stained liquor group; 959.8 vs 499.9 pg/mL). With the use of multiple regression analysis, meconium-stained liquor, duration of second stage of labor, and abnormal fetal heart tracings were independent determinants of cord blood 8-iso-prostaglandins F2alpha concentration. CONCLUSION: Moderate or thick meconium-stained liquor is an independent factor for increased oxidative stress in pregnancy.

Determination of catechins and catechin gallates in tissues by liquid chromatography with coulometric array detection and selective solid phase extraction.

Catechins levels in organ tissues, particularly liver, determined by published methods are unexpectedly low, probably due to the release of oxidative enzymes, metal ions and reactive metabolites from tissue cells during homogenization and to the pro-oxidant effects of ascorbic acid during sample processing in the presence of metal ions. We describe a new method for simultaneous analysis of eight catechins in tissue: (+)-catechin (C), (-)-epicatechin (EC), (-)-gallocatechin (GC), (-)-epigallocatechin (EGC), (-)-catechin gallate (CG), (-)-epicatechin gallate (ECG), (-)-gallocatechin gallate (GCG) and (-)-epigallocatechin gallate (EGCG) (Fig. 1). The new extraction procedure utilized a methanol/ethylacetate/dithionite (2:1:3) mixture during homogenization for simultaneous enzyme precipitation and antioxidant protection. Selective solid phase extraction was used to remove most interfering bio-matrices. Reversed phase HPLC with CoulArray detection was used to determine the eight catechins simultaneously within 25 min. Good linearity (>0.9922) was obtained in the range 20-4000 ng/g. The coefficients of variance (CV) were less than 5%. Absolute recovery ranged from 62 to 96%, accuracy 92.5 +/- 4.5 to 104.9 +/- 6%. The detection limit was 5 ng/g. This method is capable for determining catechins in rat tissues of liver, brain, spleen, and kidney. The method is robust, reproducible, with high recovery, and has been validated for both in vitro and in vivo sample analysis.

In vitro exposure to carbon dioxide induces oxidative stress in human peritoneal mesothelial cells.

BACKGROUND: The objective of this study was to verify whether in vitro exposure of human peritoneal mesothelial cells to carbon dioxide (CO(2)) influences the levels of 8-isoprostaglandin F(2alpha) (8-iso-PGF(2alpha)), a marker of oxidative stress. METHODS: Mesothelial cells were exposed to either: (i). 100% CO(2) for 4 h; (ii). 100% helium (an alternative gas with which to create hypoxic conditions) for 4 h; (iii). 100% CO(2) for 24 h; or (iv). standard conditions (control). After gas exposure, mesothelial cells were returned to standard conditions and harvested immediately (T(0)), and at 1-(T(1)) and 3 (T(3)) h afterwards. Cell viability and culture medium pH were monitored throughout the experiments. 8-iso-PGF(2alpha) was assayed by enzyme-linked immunosorbent assay (ELISA). RESULTS: Exposure to CO(2) decreased the culture medium pH whereas helium increased the pH. 8-iso-PGF(2alpha) levels in all treated groups were significantly higher than in the control group: in the 4 h CO(2) group at T(1); in the 24 h CO(2) group at T(0) and T(1); and in the 4 h helium group at T(0), T(1) and T(3). 8-iso-PGF(2alpha) levels following 4 h CO(2) exposure were significantly lower than after 24 h CO(2) exposure at T(1), and lower than following 4 h helium exposure at all time points. CONCLUSIONS: Exposure to both CO(2) and helium induces oxidative stress in mesothelial cells. Hypoxia-reoxygenation may play a role in this process.

Molecular hierarchy in neurons differentiated from mouse ES cells containing a single human chromosome 21.

Defects in neurogenesis and neuronal differentiation in the fetal brain of Down syndrome (DS) patients lead to the apparent neuropathological abnormalities and contribute to the phenotypic characters of mental retardation, and premature development of Alzheimer's disease, those being the most common phenotype in DS. In order to understand the molecular mechanism underlying the cause of phenotypic abnormalities in the DS brain, we have utilized an in vitro model of TT2F mouse embryonic stem cells containing a single human chromosome 21 (hChr21) to study neuron development and neuronal differentiation by microarray containing 15K developmentally expressed cDNAs. Defective neuronal differentiation in the presence of extra hChr21 manifested primarily the post-transcriptional and translational modification, such as Mrpl10, SNAPC3, Srprb, SF3a60 in the early neuronal stem cell stage, and Mrps18a, Eef1g, and Ubce8 in the late differentiated stage. Hierarchical clustering patterned specific expression of hChr21 gene dosage effects on neuron outgrowth, migration, and differentiation, such as Syngr2, Dncic2, Eif3sf, and Peg3.

The effect of intra-abdominal pressure on the generation of 8-iso prostaglandin F2alpha during laparoscopy in rabbits.

BACKGROUND: Carbon dioxide pneumoperitoneum induces peritoneal oxidative stress. The aim of this study was to verify the effect of intra-abdominal pressure on oxidative stress in the peritoneum and on post-operative adhesion formation. METHODS: Forty-one rabbits underwent laparoscopic surgery: either gasless, or with CO(2)-pneumoperitoneum at pressures of 5, 10 or 15 mmHg. Serial parietal peritoneal biopsies were taken at various time-points: immediately after reaching the abdominal cavity, 30, 60, 90 and 120 min afterwards, and 15 min after abdominal desufflation. 8-iso prostaglandin F(2alpha) (8-iso PGF(2alpha)), a marker of oxidative stresss, was assayed by enzyme immunoassay and adhesion formation was scored by second-look laparoscopy on day 14. RESULTS: The gasless group showed no significant changes in 8-iso PGF(2alpha). Conversely, significant changes occurred in CO(2)-pneumoperitoneum in a time- and pressure-dependent manner. Adhesions developed only in the CO(2)-pneumoperitoneum groups, and total adhesion score was correlated with the amount of CO(2) insufflated and intra-abdominal pressure, but not with 8-iso PGF(2alpha), which was correlated with intra-abdominal pressure. CONCLUSION: Intra-abdominal pressure increased 8-iso PGF(2alpha) in the parietal peritoneum in a graded fashion, whilst gasless laparoscopy had no impact. It also influenced the frequency and severity of adhesion formation, but no causal link was found between 8-iso PGF(2alpha) and post-operative adhesion formation.

Quantifying F2-isoprostanes in umbilical cord blood of newborn by gas chromatography-mass spectrometry.

We attempted to improve the extraction procedures to determine the F(2)-isoprostanes in plasma of umbilical cord arterial and venous blood by gas chromatography mass spectrometry. Plasma samples were deproteinized and hydrolyzed; free and esterified F(2)-isoprostanes were extracted by solid-phase extraction columns with citric acid/methanol/cyclohexane and ammonia solution/methanol and then derivatized by PFBBr and BSTFA. Concentrations of total plasma F(2)-isoprostanes eluted at the retention time of an internal standard of 8-iso-prostaglandin F(2alpha)-D(4) were quantified. The absolute recovery was 83+/-1.9% (95% confidence). Intraassay precision and interassay precision were lower than 1.0%. Analytical accuracy was 99.0+/-0.4% (95% confidence). Linearity, r(2), over the concentration range of 10 to 5000 pg/ml of spiked 8-iso-prostaglandin F(2alpha) in plasma was 0.9985. The method detection limit was 21 pg/ml (99% confidence) and the limit of quantitation was approximately 4 pg/ml. Analysis of 200 neonatal cord blood samples revealed few overlapping peaks causing interference in the elution of the F(2)-isoprostanes. With the use of an autosampler and one technician, 48 samples can be completed within 24h with 6h of actual hands-on work. This method could be potentially employed for routine analysis of plasma F(2)-isoprostanes in clinical laboratories.

Effects of myo-inositol on the in-vitro maturation and subsequent development of mouse oocytes.

BACKGROUND: The study aim was to assess whether the incorporation of myo-inositol (MI) into culture medium could improve oocyte maturation in vitro. METHODS AND RESULTS: We performed a controlled prospective study using female ICR strain mice superovulated with pregnant mare's serum gonadotrophins. Cumulus-enclosed germinal vesicle (GV) oocytes were randomly cultured in medium with or without MI supplementation. The kinetics of GV breakdown after 4 h of incubation was significantly higher in oocytes incubated with 30 mmol/l of MI than in controls (P < 0.001). Accordingly, this concentration of MI was used for subsequent experiments. The proportion of metaphase II oocytes achieved after 24 h of culture, their fertilization and cleavage rates were significantly higher in the MI-treated group (P < 0.01, P < 0.05, P < 0.05 respectively). This group also demonstrated significant improvement in postimplantation development after transferring the 2-cell embryos to pseudopregnant mice. Confocal microscopy revealed spontaneous intracellular Ca(2+) oscillations within competent GV oocytes and treatment with MI caused an earlier onset of these Ca(2+) signals. CONCLUSIONS: Our results suggest that MI may affect meiotic progression of mouse GV oocytes possibly by enhancing the intracellular Ca(2+) oscillations. Supplementation of MI in culture medium may be useful for human oocyte maturation.

Comparison of peritoneal oxidative stress during laparoscopy and laparotomy.

STUDY OBJECTIVE: To identify oxidative stress in peritoneum during laparoscopic and open surgery by measuring products of lipid peroxidation, and to determine whether surgical approach influences the type of oxidative metabolite synthesized. DESIGN: Retrospective analysis (Canadian Task Force classification II-2). SETTING: University-affiliated hospital. PATIENTS: Twenty-eight consecutive women with uterine myomas or ovarian cysts. INTERVENTION: Laparoscopic or open surgery (14 patients each). MEASUREMENTS AND MAIN RESULTS: We obtained 1 x 1-cm squares of peritoneum at the beginning and end of surgical procedures away from sites of surgery. 8-Isoprostaglandin F(2alpha), hydroxyeicosatetranoic acids (HETEs), and malondyaldehyde (MDA) were measured by enzyme-immunoassay, high-performance liquid chromatography, and thiobarbituric acid adduction method, respectively. Comparisons showed significant increases in 5-HETE and 8-prostane in the laparoscopy group, which were correlated with duration of pneumoperitoneum and volume of carbon dioxide (CO(2)) insufflated, respectively. In the laparotomy group only MDA rose significantly related to duration of surgery. CONCLUSIONS: Lipid peroxidation was observed in peripheral peritoneum during laparoscopic surgery, mediated through noncyclooxygenase and lipoxygenase pathways, and appears to be due to effects of CO(2) pneumoperitoneum. Biochemical reactions were also observed in the laparotomy group, but are thought to be related to mechanisms other than lipid peroxidation.