Effects of pre-pregnancy BMI and gestational weight gain on adverse pregnancy outcomes and complications of GDM.

To investigate the effects of pre-pregnancy BMI and gestational weight gain on adverse pregnancy outcomes and complications of gestational diabetes mellitus. 3966 pregnant women were enrolled in this study. Multivariate logistic regression analysis was conducted to estimate the relative risk between pre-pregnancy BMI, gestational weight gain, and adverse pregnancy outcome. Pre-pregnancy BMI was found to be a risk factor for preeclampsia (OR = 1.159), gestational diabetes mellitus (OR = 1.191), gestational hypertension (OR = 1.221), and macrosomia (OR = 1.165). Gestational weight gain was a risk factor for preeclampsia (OR = 1.783), placental abruption (OR = 2.209), and macrosomia (OR = 1.506). Total weight gain during pregnancy cannot be used as a predictor of GDM. Pre-pregnancy BMI is a risk factor for gestational diabetes mellitus complicated with preeclampsia, preterm delivery, gestational hypertension, and macrosomia. Impact statementWhat is already known on this subject? Obesity during pregnancy includes pre-pregnancy obesity and excessive weight gain during pregnancy. Obese pregnant women have a higher risk of pregnancy complications.What do the results of this study add? We focus on the effects of pre-pregnancy BMI on pregnancy outcomes, classified by Asian criteria. Our findings suggest for the first time that excessive weight gain during pregnancy is a risk factor for placental abruption and we specifically point out that total weight gain during pregnancy cannot be used as a predictor of GDM.What are the implications of these findings for clinical practice and/or further research? This study is helpful to monitor the risk of adverse pregnancy outcomes in the Asian population and suggest the risk of pregnancy complications, such as gestational diabetes mellitus and placental abruption.

A Novel U-Shaped Association Between Serum Magnesium on Admission and 28-Day In-hospital All-Cause Mortality in the Pediatric Intensive Care Unit.

Objective: Our purpose is to evaluate whether serum magnesium when entering the ICU is related to 28-day in-hospital all-cause mortality in the pediatric ICU. Methods: We used the PIC database to conduct a retrospective analysis to investigate the first-time serum magnesium levels of 10,033 critically ill children admitted to the pediatric ICU, and analyzed association between serum magnesium and all-cause mortality. Smoothing spline plots, subgroup analysis and segmented multivariate logistic regression analysis were conducted to estimate the relative risk between serum magnesium and all-cause mortality. The shape of the curve was used to describe the relationship between magnesium and 28-day in-hospital mortality. Results: There is a non-linear relationship between serum magnesium and 28-day in-hospital all-cause mortality. The U-type relationship between serum magnesium and all-cause mortality was observed. The optimal range of serum magnesium with the lowest risk of mortality was 0.74-0.93 mmol/L. As the serum magnesium level reaches the turning point (0.74 mmol/L), the risk of death decreases by 60% for every 0.1 mmol/L increase in serum magnesium; when the serum magnesium level exceeds 0.93, an increase of 0.1 mmol/L increases the risk of death by 38 %. Conclusion: Serum magnesium has a U-shaped relationship with 28-day in-hospital all-cause mortality. Both low and high serum magnesium can increase the risk of death. The best serum magnesium range when the risk of death is the lowest is 0.74-0.93 mmol/L.

Is serum vitamin D deficiency before gestational 20 weeks a risk factor for preeclampsia?

OBJECTIVE: Our aim was to assess whether serum vitamin D deficiency before gestational 20 weeks was associated with an increased risk of preeclampsia. METHODS: We investigated the serum levels of 25(OH)D before gestational 20 weeks, and analyzed associations between the 25(OH)D and the risk of preeclampsia. 7976 pregnant women were enrolled in this study between January 2017 and July 2019 at the Obstetrics & Gynecology Hospital of Fudan University. Adjusted smoothing spline plots, subgroup analysis and multivariate logistic regression analysis was conducted to estimate the relative risk between 25(OH)D and preeclampsia. RESULTS: After fully adjusting the confounding factors, serum vitamin D is a protective factor in preeclampsia (OR = 0.85, P = 0.04). Compared with adequate vitamin D, vitamin D deficiency (OR = 1.55, P = 0.031), deficiency (OR = 1.50, P = 0.049) and severe deficiency (OR = 2.6, P = 0.005) are independent of preeclampsia in pregnant women Risk factors. CONCLUSION: Vitamin D deficiency before gestational 20 weeks is a risk factor for preeclampsia.

Development and validation of a nomogram for the early prediction of preeclampsia in pregnant Chinese women.

To make early predictions of preeclampsia before diagnosis, we developed and validated a new nomogram for the early prediction of preeclampsia in pregnant Chinese women. A stepwise regression model was used for feature selection. Multivariable logistic regression analysis was used to develop the prediction model. We incorporated BMI, blood pressure, uterine artery ultrasound parameters, and serological indicator risk factors, and this was presented with a nomogram. The performance of the nomogram was assessed with respect to its calibration, discrimination, and clinical usefulness. Internal validation was assessed. The signature, which consisted of 11 selected features, was associated with preeclampsia status (P < 0.1) for the development dataset. Predictors contained in the individualized prediction nomogram included BMI, blood pressure, uterine artery ultrasound parameters, and serological indicator levels. The model showed good discrimination, with an area under the ROC curve of 0.8563 (95% CI: 0.8364-0.8761) and good calibration. The nomogram still had good discrimination and good calibration when applied to the validation dataset (area under ROC curve of 0.8324, 95% CI: 0.7873-0.8775). Decision curve analysis demonstrated that the nomogram was clinically useful. The nomogram presented in this study incorporates BMI, blood pressure, uterine artery ultrasound parameters, and serological indicators and can be conveniently used to facilitate the individualized prediction of preeclampsia.

Serum markers in quadruple screening associated with adverse pregnancy outcomes: A case-control study in China.

BACKGROUND: We assessed whether the results of quadruple screening during pregnancy are associated with an increased risk of adverse pregnancy outcomes. METHODS: We measured serum marker concentrations using quadruple screening in the second trimester of pregnancy and analyzed the relationship between adverse perinatal outcomes and serum markers in 12,124 pregnant women. A multivariate logistic regression analysis was used to evaluate the relative risk of quadruple screening and adverse pregnancy outcomes. RESULTS: Compared with the control group, increased concentrations of alpha-fetoprotein (AFP) and inhibin A were risk factors for preeclampsia and preterm delivery; low concentrations of unconjugated estriol and high inhibin A were risk factors for pregnancy hypertension; an increased concentration of human chorionic gonadotropin (hCG) was a risk factor for gestational diabetes mellitus; high AFP, low hCG, and high inhibin A were risk factors for low birth weight; and low AFP and high hCG were risk factors for macrosomia. CONCLUSIONS: Quadruple screening in the second trimester of pregnancy can provide early warning signs for maternal and fetal adverse pregnancy outcomes.

Are serum levels of inhibin A in second trimester predictors of adverse pregnancy outcome?

OBJECTIVE: During pregnancy, inhibin A is mainly derived from the placenta and regulates the implantation and differentiation of embryos. Our aim was to assess whether second trimester serum inhibin A was associated with an increased risk of adverse pregnancy outcomes. METHODS: We investigated the serum levels of Inhibin A during the second trimester in pregnancy, and analyzed associations between the Inhibin A and the risk of adverse pregnancy outcome. 12,124 pregnant women were enrolled in this study between January 2017 and July 2019 at the Obstetrics & Gynecology Hospital of Fudan University. Multivariate logistic regression analysis was conducted to estimate the relative risk between Inhibin A and adverse pregnancy outcome. RESULTS: Compared with the group without adverse pregnancy outcome, during the second trimester of pregnancy, age and Inhibin A were risk factors for pre-eclampsia, gestational diabetes mellitus and preterm delivery; Inhibin A was risk factors for low birth weight. Gravidity and Inhibin A were risk factors for macrosomia; while parity was a protective factor against pre-eclampsia, gestational hypertension and low birth weight. CONCLUSION: Elevated Inhibin A levels in pregnancy are significantly associated with pre-eclampsia, GDM, macrosomia, low birth weight and preterm delivery.

Creep-Based Reliability Evaluation of Turbine Blade-Tip Clearance with Novel Neural Network Regression.

To reveal the effect of high-temperature creep on the blade-tip radial running clearance of aeroengine high-pressure turbines, a distributed collaborative generalized regression extremum neural network is proposed by absorbing the heuristic thoughts of distributed collaborative response surface method and the generalized extremum neural network, in order to improve the reliability analysis of blade-tip clearance with creep behavior in terms of modeling precision and simulation efficiency. In this method, the generalized extremum neural network was used to handle the transients by simplifying the response process as one extremum and to address the strong nonlinearity by means of its nonlinear mapping ability. The distributed collaborative response surface method was applied to handle multi-object multi-discipline analysis, by decomposing one "big" model with hyperparameters and high nonlinearity into a series of "small" sub-models with few parameters and low nonlinearity. Based on the developed method, the blade-tip clearance reliability analysis of an aeroengine high-pressure turbine was performed subject to the creep behaviors of structural materials, by considering the randomness of influencing parameters such as gas temperature, rotational speed, material parameters, convective heat transfer coefficient, and so forth. It was found that the reliability degree of the clearance is 0.9909 when the allowable value is 2.2 mm, and the creep deformation of the clearance presents a normal distribution with a mean of 1.9829 mm and a standard deviation of 0.07539 mm. Based on a comparison of the methods, it is demonstrated that the proposed method requires a computing time of 1.201 s and has a computational accuracy of 99.929% over 104 simulations, which are improvements of 70.5% and 1.23%, respectively, relative to the distributed collaborative response surface method. Meanwhile, the high efficiency and high precision of the presented approach become more obvious with the increasing simulations. The efforts of this study provide a promising approach to improve the dynamic reliability analysis of complex structures.

Probabilistic Fatigue/Creep Optimization of Turbine Bladed Disk with Fuzzy Multi-Extremum Response Surface Method.

To effectively perform the probabilistic fatigue/creep coupling optimization of a turbine bladed disk, this paper develops the fuzzy multi-extremum response surface method (FMERSM) for the comprehensive probabilistic optimization of multi-failure/multi-component structures, which absorbs the ideas of the extremum response surface method, hierarchical strategy, and fuzzy theory. We studied the approaches of FMERSM modeling and fatigue/creep damage evaluation of turbine bladed disks, and gave the procedure for the fuzzy probabilistic fatigue/creep optimization of a multi-component structure with FMERSM. The probabilistic fatigue/creep coupling optimization of turbine bladed disks was implemented by regarding the rotor speed, temperature, and density as optimization parameters; the creep stress, creep strain, fatigue damage, and creep damage as optimization objectives; and the reliability and GH4133B fatigue/creep damages as constraint functions. The results show that gas temperature T and rotor speed ω are the key parameters that should be controlled in bladed disk optimization, and respectively reduce by 85 K and 113 rad/s after optimization, which is promising to extend bladed disk life and decrease failure damages. The simulation results show that this method has a higher modeling accuracy and computational efficiency than the Monte Carlo method (MCM). The efforts of this study provide a new useful method for overall probabilistic multi-failure optimization and enrich mechanical reliability theory.

Fuzzy Multi-SVR Learning Model for Reliability-Based Design Optimization of Turbine Blades.

The effectiveness of a model is the key factor of influencing the reliability-based design optimization (RBDO) of multi-failure turbine blades in the power system. A machine learning-based RBDO approach, called fuzzy multi-SVR learning method, was proposed by absorbing the strengths of fuzzy theory, support vector machine of regression (SVR), and multi-response surface method. The model of fuzzy multi-SVR learning method was established by adopting artificial bee colony algorithm to optimize the parameters of SVR models and considering the fuzziness of constraints based on fuzzy theory, in respect of the basic thought of multi-response surface method. The RBDO model and procedure with fuzzy multi-SVR learning method were then resolved and designed by multi-objective genetic algorithm. Lastly, the fuzzy RBDO of a turbine blade with multi-failure modes was performed regarding the design parameters of rotor speed, temperature, and aerodynamic pressure, and the design objectives of blade stress, strain, and deformation, and the fuzzy constraints of reliability degree and boundary conditions, as well. It is revealed (1) the stress and deformation of turbine blade are reduced by 92.38 MPa and 0.09838 mm, respectively. (2) The comprehensive reliability degree of the blade was improved by 3.45% from 95.4% to 98.85%. (3) It is verified that the fuzzy multi-SVR learning method is workable for the fuzzy RBDO of complex structures just like a multi-failure blade with high modeling precision, as well as high optimization, efficiency, and accuracy. The efforts of this study open a new research way, i.e., machine learning-based RBDO, for the RBDO of multi-failure structures, which expands the application of machine learning methods, and enriches the mechanical reliability design method and theory as well.

Simultaneous extraction and determination of mono-/polyglutamyl folates using high-performance liquid chromatography-tandem mass spectrometry and its applications in starchy crops.

Folates are typically present in polyglutamyl form in organisms. In traditional extraction methods, polyglutamyl folates are hydrolyzed to monoglutamates, sacrificing valuable information. To advance folate metabolism research, we developed an accurate, sensitive, and reproducible extraction method for polyglutamyl folate species in maize, the main crop in most parts of the world. Twelve folates, including six polyglutamyl folates, were simultaneously determined in maize for the first time using high-performance liquid chromatography-tandem mass spectrometry. The glutamation states of the folates were protected by boiling, which inactivated the native conjugases. α-Amylase and protease were added to obtain better recoveries and decrease difficulties in centrifugation and filtration. The recoveries (n = 5) of six polyglutamyl folates were between 80.5 and 101%. All calibration curves showed good linear regression (r2 ≥ 0.994) within the working range. The instrumental limits of detection and quantitation ranged from 0.070 to 2.4 ng/mL and 0.22 to 8.0 ng/mL, respectively. Intra- and inter-day precision was below 7.81% and 11.9%, respectively (n = 5). Using this method, changes in poly- and monoglutamyl folates during maize germination were determined for the first time. The results suggest that folates were largely synthesized as germination initiated, and 5-methyltetrahydrofolate was the most abundant species. Tetraglutamyl 5-methyltetrahydrofolate contributed more than 50% of the 5-methyltetrahydrofolate species. Inverse changes in contents of 5,10-methenyltetrahydrofolate, and 10-formyl folic acid, monoglutamate, and diglutamate of 5-formyltetrahydrofolate were also observed, indicating potential regulation. Additionally, polyglutamyl folates in sweet potatoes were determined using this method, indicating its applications in starchy crops.

Endobronchial Ultrasound Elastography Differentiates Intrathoracic Lymph Nodes: A Meta-Analysis.

BACKGROUND: In this study, we investigated the clinical significance of endobronchial ultrasound elastography for differentiating malignant and benign intrathoracic lymph nodes. METHODS: A meta-analysis was performed to evaluate the sensitivity and specificity of endobronchial ultrasound elastography in diagnosing intrathoracic lymph nodes. Publications before October 1, 2017, were included for analysis. Sensitivity, specificity, and other variables were pooled using the bivariate mixed-effects regression model. RESULTS: Seven studies met the inclusion criteria and were included. The pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, and diagnostic odds ratio was 0.93 (95% confidence interval [CI], 0.85 to 0.97), 0.85 (95% CI, 0.78 to 0.90), 6.3 (95% CI, 4.2 to 9.2), 0.08 (95% CI, 0.04 to 0.18), and 74 (95% CI, 33 to 168), respectively. The summary receiver operating characteristic curve was 0.93 (95% CI, 0.91 to 0.95). CONCLUSIONS: The results revealed endobronchial ultrasound elastography is a new technique with high sensitivity and specificity. It has a fine performance in diagnosing intrathoracic lymph nodes.

MiR-124 inhibits the migration and invasion of human hepatocellular carcinoma cells by suppressing integrin αV expression.

Tumor metastasis is the major cause of cancer-related death especially in human hepatocellular carcinoma (HCC). Although microRNAs have been implicated in tumor development, the roles of miR-124 in HCC metastasis are still not well understood. We conducted functional analysis in this study to investigate miR-124. We observed that miR-124 significantly retarded the wound healing and migration of HCC SMMC-7721 and BEL-7404 cells. Further analysis indicated miR-124 directly targeting the transcriptional factor Sp1 which is an important transcription factor for the integrin αV subunit gene transcription. Co-transfection of miR-124 with the luciferase reporter containing Sp1 3' untranslated region (UTR) significantly suppressed the luciferase activities. While mutation of the binding site of miR-124 in Sp1 mRNA 3'UTR completely abrogated the suppression of miR-124. Overexpression of miR-124 resulted in robust downregulation of Sp1 and integrin αV expression at either mRNA or protein level. Ectopic expression of miR-124 in HCC dramatically repressed the wound healing and migration in vitro and tumor metastasis in mouse experiments. Our findings demonstrated that miR-124 played as an important role in regulation of integrin αV expression in HCC, and reintroduction of miR-124 might be an alternative therapeutic strategy for controlling integrin αV expression in HCC.

Identification of Iron Homeostasis Genes Dysregulation Potentially Involved in Retinopathy of Prematurity Pathogenicity by Microarray Analysis.

Retinopathy of prematurity (ROP) is a serious disease of preterm neonates and there are limited systematic studies of the molecular mechanisms underlying ROP. Therefore, here we performed global gene expression profiling in human fetal retinal microvascular endothelial cells (RMECs) under hypoxic conditions in vitro. Aborted fetuses were enrolled and primary RMECs were isolated from eyeballs. Cultivated cells were treated with CoCl2 to induce hypoxia. The dual-color microarray approach was adopted to compare gene expression profiling between treated RMECs and the paired untreated control. The one-class algorithm in significance analysis of microarray (SAM) software was used to screen the differentially expressed genes (DEGs) and quantitative RT-PCR (qRT-PCR) was conducted to validate the results. Gene Ontology was employed for functional enrichment analysis. There were 326 DEGs between the hypoxia-induced group and untreated group. Of these genes, 198 were upregulated in hypoxic RMECs, while the other 128 hits were downregulated. In particular, genes in the iron ion homeostasis pathway were highly enriched under hypoxic conditions. Our study indicates that dysregulation of genes involved in iron homeostasis mediating oxidative damage may be responsible for the mechanisms underlying ROP. The "oxygen plus iron" hypothesis may improve our understanding of ROP pathogenesis.

Stress-induced early flowering is mediated by miR169 in Arabidopsis thaliana.

Plants interact with their environment and they often flower earlier under stress conditions, but how such stress-induced flowering is regulated remains poorly understood. Here evidence is presented that the miR169 family plays a key role in stress-induced flowering in plants. The microRNA (miRNA) miR169 family members are up-regulated in Arabidopsis, maize, and soybean under abiotic stresses. Overexpression of miR169d in Arabidopsis results in early flowering, and overexpression of the miR169d target gene, AtNF-YA2, especially a miR169d-resistant version of AtNF-YA2, results in late flowering. The results suggest that the miR169 family regulates stress-induced flowering by repressing the AtNF-YA transcription factor, which in turn reduces the expression of FLOWERING LOCUS C (FLC), allowing for the expression of FLC target genes such as FLOWERING LOCUS T (FT) and LEAFY (LFY) to promote flowering. It was shown that the expression of genes or miRNAs involved in the other flowering pathways, namely the photoperiod (CO), ambient temperature (SVP), ageing (miR156), and gibberelin (SOC1) pathways, was not affected in miR169d-overexpressing plants, suggesting that stress-induced early flowering is a novel signalling pathway mediated by miR169.

Regulation of integrin αV subunit expression by sulfatide in hepatocellular carcinoma cells.

Integrin is important in migration and metastasis of tumor cells. Changes of integrin expression and distribution will cause an alteration of cellular adhesion and migration behaviors. In this study, we investigated sulfatide regulation of the integrin αV subunit expression in hepatoma cells and observed that either exogenous or endogenous sulfatide elicited a robust upregulation of integrin αV subunit mRNA and protein expression in hepatoma cells. This regulatory effect occurred with a corresponding phosphorylation (T739) of the transcription factor Sp1. Based on the electrophoretic mobility shift assay, sulfatide enhanced the integrin αV promoter activity and strengthened the Sp1 complex super-shift. The results of chromatin immunoprecipitation analysis also indicated that sulfatide enhanced Sp1 binding to the integrin αV promoter in vivo. Silence of Sp1 diminished the stimulation of integrin αV expression by sulfatide. In the early stage of sulfatide stimulation, phosphorylation of Erk as well as c-Src was noted, and inhibition of Erk activation with either U0126 or PD98059 significantly suppressed Sp1 phosphorylation and integrin αV expression. We demonstrated that sulfatide regulated integrin αV expression and cell adhesion, which was associated with Erk activation.

Regulation of growth factor receptor degradation by ADP-ribosylation factor domain protein (ARD) 1.

ADP-ribosylation factor domain protein 1 (ARD1) is a 64-kDa protein containing a functional ADP-ribosylation factor (GTP hydrolase, GTPase), GTPase-activating protein, and E3 ubiquitin ligase domains. ARD1 activation by the guanine nucleotide-exchange factor cytohesin-1 was known. GTPase and E3 ligase activities of ARD1 suggest roles in protein transport and turnover. To explore this hypothesis, we used mouse embryo fibroblasts (MEFs) from ARD1-/- mice stably transfected with plasmids for inducible expression of wild-type ARD1 protein (KO-WT), or ARD1 protein with inactivating mutations in E3 ligase domain (KO-E3), or containing persistently active GTP-bound (KO-GTP), or inactive GDP-bound (KO-GDP) GTPase domains. Inhibition of proteasomal proteases in mifepristone-induced KO-WT, KO-GDP, or KO-GTP MEFs resulted in accumulation of these ARD1 proteins, whereas KO-E3 accumulated without inhibitors. All data were consistent with the conclusion that ARD1 regulates its own steady-state levels in cells by autoubiquitination. Based on reported growth factor receptor-cytohesin interactions, EGF receptor (EGFR) was investigated in induced MEFs. Amounts of cell-surface and total EGFR were higher in KO-GDP and lower in KO-GTP than in KO-WT MEFs, with levels in both mutants greater (p = 0.001) after proteasomal inhibition. Significant differences among MEF lines in content of TGF-ß receptor III were similar to those in EGFR, albeit not as large. Differences in amounts of insulin receptor mirrored those in EGFR, but did not reach statistical significance. Overall, the capacity of ARD1 GTPase to cycle between active and inactive forms and its autoubiquitination both appear to be necessary for the appropriate turnover of EGFR and perhaps additional growth factor receptors.

3'-Sulfo-Le(x) is important for regulation of integrin subunit alphaV.

Carbohydrate structures with a 3'-sulfo betaGal linkage, such as 3'-sulfo-Le(x), can be synthesized by Gal:3-O-sulfotransferase-2 (Gal3ST-2) catalysis, but little is known about their roles in many biological processes. To investigate the role of Gal3ST-2 and its product 3'-sulfo-Le(x), we depleted Gal3ST-2 via siRNA and added exogenous Lewis-x trisaccharide 3'-sulfate sodium salt in human SMMC7721 hepatoma cells. After siRNA transfection, a striking morphological change in SMMC7721 hepatoma cells from polygon to shuttle shape and a significant decrease in the level of adhesion to sL-selectin, HUVEC, fibronectin, vitronectin, and fibrinogen were observed. The expression of integrin subunit alphaV was markedly downregulated, and 3'-sulfated subunit alphaV almost disappeared in the transfectants. The level of cell surface integrin alphaVbeta3 was reduced simultaneously, although total subunit beta3 underwent almost no change. After treatment with exogenous Lewis-x 3'-sulfate, cellular integrin subunit alphaV was upregulated and the level of cell surface integrin alphaVbeta3 was elevated. Interestingly, knockdown of Gal3ST-2 expression effectively inhibited cell proliferation, and the result was significantly correlated with the decrease in the levels of ILK, phosphorylated AKT, and ERK. On the other hand, treatment with Lewis-x trisaccharide 3'-sulfate sodium salt greatly upregulated the phosphorylation of AKT and ERK. Our results also indicated that downregulation of Gal3ST-2 via siRNA transfection was associated with the decrease in the level of expression of anti-apoptotic protein, Bcl-2, with a consequent decrease in the ratios for Bcl-2 to Bax. By exposure to Lewis-x trisaccharide 3'-sulfate sodium salt, the apoptotic response of cells was inhibited. Therefore, Gal3ST-2 and its product, 3'-sulfo-Le(x), were involved in regulation of integrin subunit alphaV and might be associated with cancer cell regulation.

Ischemia-induced release of cytochrome c from mitochondria and up-regulation of Bcl-2 expression in rat hippocampus.

To evaluate the effects of different antagonists on the release of cytochrome c from mitochondria to cytosol and the expression of Bcl-2 in mitochondria in rat hippocampus after ischemia, we examined Bcl-2 and cytochrome c expression by immunoblotting using 4-vessel occlusion (4-VO) as brain ischemia model. The results showed that after 24 h ischemia/reperfusion (I/R) cytochrome c decreased markedly in mitochondria, which was correspondingly increased in the cytosolic fraction. Bcl-2 expression was time-dependent, reaching its peak level after 6 h I/R. In all those samples, there were no alterations in the subcellular distribution of cytochrome oxidase, a mitochondrial respiratory chain protein. The decreases in Bcl-2 and cytochrome c in mitochondria were restored by pretreatment with non-competitive NMDA receptor antagonist ketamine or L-type voltage-gated Ca(2+) channel (L-VGCC) antagonist nifedipine at 20 min prior to ischemia. The results demonstrate that the release of cytochrome c from mitochondria to cytosol and the up-regulation of Bcl-2 are possibly mediated by NMDA receptors or L-VGCC following brain ischemia. Cytochrome c release may be injurious while Bcl-2 up-regulation may be protective to ischemic hippocampus.

Antioxidants attenuate reperfusion injury after global brain ischemia through inhibiting nuclear factor-kappa B activity in rats.

AIM: To investigate the effects of two antioxidants on the alterations of nuclear factor kappaB (NF-kappaB) activity and p65, p50 protein expression and phosphorylation of IkappaBalpha in rat hippocampus following global brain ischemia. METHODS: Using a 4-vessel occlusion (4-VO) as brain ischemia model, NF-kappaB protein (p65 or p50 subunit) expression was examined by Western blot analysis, and NF-kappaB activity was assayed by electrophoretic mobility shift assay (EMSA), and neuronal loss was observed by histology. RESULTS: NF-kappaB activity displayed a time-dependent manner, and p65, p50 proteins showed their peak levels after ischemia/reperfusion 6 h. NF-kappaB inductions (p65: 4.79+/-0.78, p50: 5.50+/-0.33, sham control=1) and activity (4.93+/-0.95) after 6 h of reperfusion were markedly reduced by pretreatment with antioxidants pyrrolidine dithiocarbamate (PDTC, 200 mg/kg) (p65: 1.11+/-0.74, p50: 1.38+/-0.98, activity: 2.20+/-0.86, respectively) or N-acetylcysteine (NAC, 300 mg/kg) (p65: 0.64+/-0.39, p50: 1.89+/-0.87, activity: 0.61+/-0.65), and histological observations of the pyramidal layer of CA1 also showed a reduction of neuronal loss in rat hippocampus (70 %+/-5 % or 92 %+/-4 % cells are survival, respectively). Furthermore, PDTC and NAC prevented the decrease (from 0.50+/-0.10 to 0.80+/-0.20 or 1.20+/-0.24, respectively) and phosphorylation (from 2.00+/-0.15 to 0.46+/-0.10 or 0.41+/-0.10, respectively) of IkappaBalpha protein in the cytoplasm. CONCLUSION: The protective effects of antioxidants against ischemia/reperfusion-induced injury may be mediated by down-regulation of NF-kappaB activity. NF-kappaB activation and deactivation are controlled mainly through phosphorylation and degradation of IkappaBalpha following brain ischemia.

Critical elements of the immunoglobulin heavy chain gene enhancers for deregulated expression of bcl-2.

Translocation of the bcl-2 gene to the immunoglobulin heavy chain gene is the most common alteration in follicular lymphoma. The result is the deregulated expression of bcl-2 and increased resistance to cell death. Regulation of the immunoglobulin heavy chain gene is controlled in part by four DNase I-hypersensitive regions located 3' of the gene. Here, we show that these four enhancer regions also contribute to bcl-2 up-regulation in t(14;18) cells. The enhancers are able to individually or in combination activate bcl-2 promoter activity. The HS4 enhancer region was found to impart the largest positive effect on the bcl-2 promoter, activating it by 6-fold, whereas addition of the HS1,2 region with HS4 increased promoter activity by approximately 9-fold. Nuclear factor kappaB binding sites were shown to be primarily responsible for the positive activity contributed by the HS1,2 and HS4 regions, and we observed the in vivo interaction of these factors with the human immunoglobulin heavy chain gene enhancer regions in t(14;18) cells. In addition, two Sp1 binding sites in HS4 were also found to positively influence bcl-2 activity, and Sp1 was observed to interact with the human HS4 enhancer in vivo. These results suggest that the interactions of the nuclear factor kappaB and Sp1 transcription factors with the immunoglobulin heavy chain enhancer region are important for bcl-2 deregulation in t(14;18) cells.