CYP2E1 Sensitizes the Liver to LPS- and TNF α-Induced Toxicity via Elevated Oxidative and Nitrosative Stress and Activation of ASK-1 and JNK Mitogen-Activated Kinases

The mechanisms by which alcohol causes cell injury are not clear. A major mechanism is the role of lipid peroxidation and oxidative stress in alcohol toxicity. Many pathways have been suggested to play a role in how alcohol induces oxidative stress. Considerable attention has been given to alcohol elevated production of lipopolysaccharide (LPS) and TNFα and to alcohol induction of CYP2E1. These two pathways are not exclusive of each other; however, interactions between them, have not been extensively evaluated. Increased oxidative stress from induction of CYP2E1 sensitizes hepatocytes to LPS and TNFα toxicity and oxidants, activation of inducible nitric oxide synthase and p38 and JNK MAP kinases, and mitochondrial dysfunction are downstream mediators of this CYP2E1-LPS/TNFα-potentiated hepatotoxicity. This paper will summarize studies showing potentiated interactions between these two risk factors in promoting liver injury and the mechanisms involved including activation of the mitogen-activated kinase kinase kinase ASK-1. Decreasing either cytosolic or mitochondrial thioredoxin in HepG2 cells expressing CYP2E1 causes loss of cell viability and elevated oxidative stress via an ASK-1/JNK-dependent mechanism. We hypothesize that similar interactions occur as a result of ethanol induction of CYP2E1 and TNFα

N-(2-Hy­droxy­eth­yl)-5-(4-meth­oxy­phen­yl)-4H-pyrazole-3-carboxamide

In the title compound, C13H15N3O3, the dihedral angle between the benzene and pyrazole rings is 7.7 (1)° and the O—C—C—N torsion angle of the side chain is 74.1 (2)°. In the crystal, mol­ecules are linked by O—H⋯O, N—H⋯O and N—H⋯N hydrogen bonds

Ethanol Consumption bythe NursingMotherInduces Cytochrome P-4502E1in Neonatal Rat Liver1

ABSTRACT CytochromeP-4502E1(P-4502E1) is not presentin fetalrat liver because activation of the gene occurs shortly after birth. Ethanol is an inducerof P-4502E1in adultrats.Studieswerecarriedout to evaluatewhethertransplacental induction of P-4502E1by ethanol can occur after oral consumption of ethanol by the pregnantmother. Becauseethanolcan be excreted in breast milk, the possible induction of P-4502E1 in neonatal liver when ethanol was consumed during the gestational and neonatal period by the mother was also determined. Pregnant rats re calved control or an ethanol-containing liquid diet starting on the 9th day of gestation and were killed on the 17th day or 21St day of gestation or allowed to deliver. The rats continued on their respective diets for the first 2 weeks of the neonatal period. P 4502E1 messenger RNA (mRNA), protein or catalytic activity was not detectablein fetal liverand was not inducedin the fetusesfrom the ethanol-consuming mothers. Transpiacental induction of P-4502E1 by ethanol did not occur in this model. Induction by ethanol of P-4502E1 proteinandcatalyticactivity but not mRNA occurred in maternal liver. P-4502E1 mRNA, proteinandcatalyticactivityweredetectedshortlyafterbirth and increased over the 2-week neonatal period. The P-4502E1 content and oxidation ofp-nitrophenol or dimethylnitrosamine by hepaticmicrosomes fromneonates of mothersconsuming the ethanol diet were increased 2-to 3-fold compared with controls; however, P-4502E1 mRNA levels were not elevated. These resultsindicate thatconsumption of ethanol duringthe gesta tional and neonatal period can result in induction of P-4502E1 in hepatic microsomes of neonates, suggesting that the ethanol present in the mother's milk is transferred to the newborn and is capableof inducing the P-4502E1by a post-transcriptional mechanism. Suth a mechanism of induction, which can occur immediately afterbirth,maybe of toxicological significance to the newborn in view of the catalytic properties associated with P-4502E1. activity, such as aniline and DMN Injecting pyrazole or 4-methylpyrazole into rat pups imme diately after birth or during the first 2 weeks of the neonatal period resulted in 2-to 4-fold increases in the content of P 4502E1 in hepatic microsomes (Wu and Cederbaum, 1993). These increases were associated with enhanced oxidation of DMN but not with elevated P-4502E1 mRNA levels. Ding et at. Materials and Methods Pregnant Sprague-Dawley rats, weighing about 250 to 300 g, were purchased from Zivic Miller (Allison Park, PA) and fed normal Purina lab chow (Ralston Purina PMI Feeds, Inc., St. Louis, MO) until the 9th day of gestation. The rats were divided into two groups. One group was fed a high-protein liquid diet containing 6.7% w/v ethanol (Bio Serve, Frenchtown, NJ) and the other group was fed the high-protein liquid control diet. Some rats in each group were killed on the 17th or 21st day of gestation and the livers were removed from the mothers and their fetuses. Other rats were allowed to deliver;the pups were killed either on the day of birth (generally within 12 hr after birth) or 4, 8 and 14 days after birth. The mothers were killed 14 days after delivery. During this 14-day neonatal period, the mothers continued to consume the same liquid diet that they had consumed during the gestational period. Male and female pups were pooled and not studied separately. The livers were rapidly removed and divided into two parts for each fetus, pup or mother. The livers from several fetuses and pups were pooled together to provide sufficient material for the various analyses. One part of the whole liver or the pooled livers was homoge nized and microsomes were prepared by differential centrifugation, suspended in 125 mM KC1 with 10 mM potassium phosphate, pH 7.4, and stored at â€"¿ 70'C. The second part of the liver was used to prepare total RNA, which was isolated by an acid guanidinium thiocyanate phenol-chloroform extraction method Microsomal protein was assayed by the method of Lowry et aL (1951); the content oftotal cytochrome P-450 was determined from the reduced cytochrome P-450 carbon monoxide binding spectrum (Omura and Sato, 1964).The oxidation of DMN was determined in a reaction system containing 100 mM potassium phosphate buffer, pH 7.4, 4 mM DMN, 1 mM NADPH and 1.2 mg of microsomalprotein in a final volume of 0.25 ml. Hydroxylation of PNP was determined in a reaction system containing 100 mM phosphate buffer, pH 7.2, 0.2 mM PNP, 1 mM NADPH and 0.6 mg of microsomal protein in a final volume of 0.5 ml. The reactions were initiated by the addition of NADPH and were carried out for 20 mm at 37CC. They were terminated by the addition of trichloroacetic acid and formaldehyde or p-nitrocatechol formation was determined by standard colorimetric assays Sodium dodecyl sulfate polyacrylamide gel electrophoresis was per formed using a 7.5%running gel and a 4% stacking gel. Heat-denatured microsomes (10â€"50 @g, depending on the expected P-4502E1 content) or 5 pmol of P-4502E1 purified from pyrazole-treated rats (Palakodety et a !., 1988) were loaded onto the gel and electrophoresis was carried out at 70 mA for 2 hr. The proteins were transferred onto nitrocellulose membranes and the membranes were incubated with 3% albumin-Tris buffered saline, pH 7.5, overnight. The membranes were then incubated with anti-P-4502E1 immunoglobulin G (1:2000 dilution) for 2 hr. The antibody was raised in rabbits against the P-4502E1 purified from pyrazole-treated rats Slot-blot analysis was carried out using 2 @g oftotal RNA, essentially as previously described (Winters and Cederbaum, 1992). The RNA was immobilized into nitrocellulose membrane filters by baking at 80CCin a vacuum oven for 2 hr and it was hybridized with a solution containing 50 zg/ml of salmon sperm DNA, 20% formamide, 5x SSPE (0.75M NaCl, 0.05M NaH2Po4.H20, 0.005M EDTA, pH 7.4), 2x Denhart's reagent and 0.1% sodium dodecyl sulfate at 4TC overnight. The membranes were then hybridized with a P-4502E1 oligonucleotide probe at 47CCovernight. The 51-mer oligonucleotideprobe (Johansson et al., 1988; Results The growth rate of neonates from mothers who consumed ethanol during the gestational period and continued to consume ethanol during the neonatal period was less than that of con trols (table 1). Under our reaction conditions, the total cyto chrome P-450 was not spectrally detectable in fetal liver from either the controls or the ethanol-consuming mothers until birth. 1, table 2). P-4502E1 was detected shortly after birth and progressively increased during the neonatal period. The content of P-4502E1 was elevated in the hepatic microsomes from neonates of mothers who consumed ethanol. A slight increase was observed on the day of birth and an approximate 3-fold increase was found during the neonatal period ( Hepatic microsomes from the ethanol-consuming mothers showed an approximate 2-fold induction of P-4502E1 The mechanism whereby ethanol induces P-4502E1 in adult rats is complex and oxidation of PNP and DMN were elevated in the hepatic microsomes of the pregnant rats consuming alcohol from day 9 of gestation and killed on day 17 or day 20 of gestation, P 4502E1 mRNA levels were similar in livers from the ethanol consuming pregnant rats and control

Treatment responses in adult depressive patients treated with dexamethasone/corticotrophin-releasing hormone

Purpose: To study the dexamethasone/corticotrophin releasing hormones (DEX/CRH) in depressed and healthy patients and to analyse the occurrence of relapse connected to hormonal dysregulation.Methods: A total of 117 depressive patients between 20 and 70 years of age were included in the study group and 40 healthy patients between 25 and 60 years of age in the control group. Group I consisted of 59 patients who received sertraline 50 - 100 mg/day for 5 weeks along with a low dose of 30 mg T3. Group II included 58 patients who received dexamethasone 1 mg orally for 5 weeks. DEX/CRH levels were analyzed. Adrenocorticotrophic hormone and cortisol levels in the blood were analysed by immuno-radiometric assay. Cortisol levels were also analysed by kinetic assay method.Results: In group I, among the 59 patients that received sertraline 50-100 mg/day for 5 weeks with a low dose of 30 mg T3, relapse was observed in 12 (20.3 %) of them. The area under the curve (AUC) was 13.9 ± 6.4 ng.min.1000/mL, which was higher than that for healthy individuals (3.8 ± 3.6 ng.min.1000/mL). Group I patients with relapse showed an adrenocorticotrophic hormone AUC of 16.9 ± 2.4 ng.min.1000/mL, while group II patients exhibited AUC of 13.9 ± 6.4 ng.min.1000/mL.Conclusion: The results emphasizes the need to test hormonal responses to different types of antidepressants.Keywords: stress, depressive patients, hormonal response, hormonal dysregulation, sertraline, dexamethasone, corticotrophin releasing hormon

Design of UDE-based dynamic surface control for dynamic positioning of vessels with complex disturbances and input constraints

In practice, dynamic positioning (DP) vessels are subjected to complex disturbances as well as the magnitude and changing rate constraints of the thrusts and moments. This study applied a dynamic surface controller based on an uncertainty and disturbance estimator (UDE) to a DP vessel with complex disturbances and input constraints. The UDE was designed to estimate and handle the complex disturbances. An auxiliary dynamic system (ADS) and smooth switching function were employed to compensate for the input constraints and avoid the singularity phenomenon caused by the ADS, respectively. The combination of the UDE method and dynamic surface control (DSC) technology significantly simplified the design process for the control law and increased the practicability for DP vessels. The stability of the proposed control law was proved using the Lyapunov theory. The effectiveness of the control law and possibility of actually applying it to a DP vessel were verified using simulation experiments

Characterization of the Fiber Connectivity Profile of the Cerebral Cortex in Schizotypal Personality Disorder: A Pilot Study

Schizotypal personality disorder (SPD) is considered one of the classic disconnection syndromes. However, the specific cortical disconnectivity pattern has not been fully investigated. In this study, we aimed to explore significant alterations in whole-cortex structural connectivity in SPD individuals (SPDs) by combining the techniques of brain surface morphometry and white matter (WM) tractography. Diffusion and structural MR data were collected from twenty subjects with SPD (all males; age, 19.7 ± 0.9 yrs) and eighteen healthy controls (all males; age, 20.3 ± 1.0 yrs). To measure the structural connectivity for a given unit area of the cortex, the fiber connectivity density (FiCD) value was proposed and calculated as the sum of the fractional anisotropy of all the fibers connecting to that unit area in tractography. Then, the resultant whole-cortex FiCD maps were compared in a vertex-wise manner between SPDs and controls. Compared with normal controls, SPDs showed significantly decreased FiCD in the rostral middle frontal gyrus (crossing BA9 and BA10) and significantly increased FiCD in the anterior part of the fusiform/inferior temporal cortex (P < 0.05, Monte Carlo simulation corrected). Moreover, the gray matter volume extracted from the left rostral middle frontal cluster was observed to be significantly greater in the SPD group (P = 0.02). Overall, this study identifies a decrease in connectivity in the left middle frontal cortex as a key neural deficit at the whole-cortex level in SPD, thus providing insight into its neuropathological basis

MAPK Signaling Pathway Alters Expression of Midgut ALP and ABCC Genes and Causes Resistance to \u3cem\u3eBacillus thuringiensis\u3c/em\u3e Cry1Ac Toxin in Diamondback Moth

Insecticidal crystal toxins derived from the soil bacterium Bacillus thuringiensis (Bt) are widely used as biopesticide sprays or expressed in transgenic crops to control insect pests. However, large-scale use of Bt has led to field-evolved resistance in several lepidopteran pests. Resistance to Bt Cry1Ac toxin in the diamondback moth, Plutella xylostella (L.), was previously mapped to a multigenic resistance locus (BtR-1). Here, we assembled the 3.15 Mb BtR-1 locus and found high-level resistance to Cry1Ac and Bt biopesticide in four independent P. xylostella strains were all associated with differential expression of a midgut membrane-bound alkaline phosphatase (ALP) outside this locus and a suite of ATP-binding cassette transporter subfamily C (ABCC) genes inside this locus. The interplay between these resistance genes is controlled by a previously uncharacterized trans-regulatory mechanism via the mitogen-activated protein kinase (MAPK) signaling pathway. Molecular, biochemical, and functional analyses have established ALP as a functional Cry1Ac receptor. Phenotypic association experiments revealed that the recessive Cry1Ac resistance was tightly linked to down-regulation of ALP, ABCC2 and ABCC3, whereas it was not linked to up-regulation of ABCC1. Silencing of ABCC2 and ABCC3 in susceptible larvae reduced their susceptibility to Cry1Ac but did not affect the expression of ALP, whereas suppression of MAP4K4, a constitutively transcriptionally-activated MAPK upstream gene within the BtR-1 locus, led to a transient recovery of gene expression thereby restoring the susceptibility in resistant larvae. These results highlight a crucial role for ALP and ABCC genes in field-evolved resistance to Cry1Ac and reveal a novel trans-regulatory signaling mechanism responsible for modulating the expression of these pivotal genes in P. xylostella

Identification of RE1-Silencing Transcription Factor as a Promoter of Metastasis in Pancreatic Cancer

Pancreatic cancer is characterized by its rapid progression and early metastasis. This requires further elucidation of the key promoters for its progression and metastasis. In this study, we identified REST as the hub gene of a gene module which is closely associated with cancer stage by weighted gene correlation network analysis. Validation with the TCGA database, western blot analysis of human pancreatic cancer cell lines (AsPC-1, Capan-2, SW-1990, and PANC-1) and immunohistochemical analysis of paraffin-embedded pancreatic cancer tissue sections showed that REST was enriched in tissue samples of advanced stage and metastatic phenotype cell lines. Survival analysis with the TCGA database and our own follow-up data suggested that patients with higher expression level of REST showed worse overall survival rate. In vitro functional experiments suggested that knockdown of REST suppressed proliferation, migration, invasion and epithelial-mesenchymal transition of AsPC-1 and PANC-1 cells. In vivo experiments (a subcutaneous BALB/c nude mouse model and a superior mesenteric vein injection BALB/c nude mouse model) suggested that knockdown of REST suppressed growth and metastasis of xenograft tumor. Finally, we investigated the underlying molecular mechanism of REST and identified REST as a potential downstream target of MAPK signaling pathway. In conclusion, our results of bioinformatic analysis, in vitro and in vivo functional analysis suggested that REST may serve as a promoter of metastasis in pancreatic cancer