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The accumulation of advanced glycation end products (AGEs) causes metabolic dysfunction and neuronal cell damage. Methylglyoxal (MG) is a major glycating agent that reacts with basic residues present in proteins and promotes the formation of AGEs. Sciadopitysin, a type of biflavonoid, exerts protective effects against neuronal cell damage; however, the underlying mechanisms have not been studied. This study aimed to investigate the mechanisms underlying the protective effects of sciadopitysin against MG-mediated cytotoxicity in SK-N-MC neuroblastoma cells. Our results demonstrated that pretreatment of SK-N-MC cells with sciadopitysin improved the cell viability that was inhibited by MG and inhibited the apoptosis induced by MG. Sciadopitysin attenuated intracellular Ca 2+ , NOX4 levels, oxidative stress, and MG-protein adduct levels, and increased nuclear Nrf2 and glyoxalase 1 levels in the presence of MG. These results suggest that sciadopitysin exerts neuroprotective effects against MG-induced death of human SK-N-MC cells via its antioxidative action. This study highlights sciadopitysin as a promising candidate for antioxidant therapy and designing natural drugs against AGE-induced neurodegenerative disorders., (© 2021 John Wiley & Sons, Ltd.)

Aims: This study was aimed to explore whether sacran polysaccharide has a therapeutic effect on atopic dermatitis (AD) and its possible mechanisms., Materials and Methods: 2, 4-dinitrochlorobenzene (DNCB)-induced AD mice were treated with 0.2% Sacran, 0.5% Sacran and 0.1% tacrolimus. Through scoring dermatitis severity, measuring ear thickness, cracking behavior, open field test, we evaluated the therapeutic effect of Sacran on DNCB-induced AD mice. CD4 + T cells and CD8 + T cells were evaluated by flow cytometry. The relative expression of Ifng and Il4 were measured by real-time quantitative PCR., Key Findings: Sacran could relieved the symptoms of DNCB-induced AD mice, such as AD score, ear thickness, and IgE release. Sacran may alleviate dermatitis by inhibiting Th2 activation and reducing IgE release., Significance: Our research further proved that polysaccharide Sacran has anti-dermatitis effects, and also clarified its mechanism of alleviating dermatitis by inhibiting the activation of Th2 cells and reducing the release of IgE, which provides a theoretical basis for the future clinical transformation of polysaccharide Sacran., (Copyright © 2021. Published by Elsevier Inc.)

Heptafluorobutyric acid (PFBA) is a synthetic chemical belonging to the per- and polyfluoroalkyl substances (PFAS) group that includes over 5000 chemicals incorporated into numerous products. PFBA is a short-chain PFAS (C4) labeled as a safer alternative to legacy PFAS which have been linked to numerous health effects. Despite the high potential for dermal exposure, occupationally and environmentally, dermal exposure studies are lacking. Using a murine model, this study analyzed serum chemistries, histology, immune phenotyping, and gene expression to evaluate the systemic toxicity of sub-chronic dermal PFBA 15-day (15% v/v or 375 mg/kg/dose) or 28-day (3.75-7.5% v/v or 93.8-187.5 mg/kg/dose) exposures. PFBA exposure produced significant increases in liver and kidney weights and altered serum chemistries (all exposure levels). Immune-cell phenotyping identified significant increases in draining lymph node B-cells (15%) and CD11b + cells (3.75-15%) and skin T-cells (3.75-15%) and neutrophils (7.5-15%). Histopathological and gene expression changes were observed in both the liver and skin after dermal PFBA exposure. The findings indicate PFBA induces liver toxicity and alterations of PPAR target genes, suggesting a role of a PPAR pathway. These results demonstrate that sustained dermal exposure to PFBA induces systemic effects and raise concerns of short-chain PFAS being promoted as safer alternatives., (Published by Elsevier Ltd.)

Protein footprinting coupled with mass spectrometry is being increasingly used for the study of protein interactions and conformations. The hydroxyl radical footprinting method, fast photochemical oxidation of proteins (FPOP), utilizes hydroxyl radicals to oxidatively modify solvent accessible amino acids. Here, we describe the further development of FPOP for protein structural analysis in vivo (IV-FPOP) with Caenorhabditis elegans. C. elegans, part of the nematode family, are used as model systems for many human diseases. The ability to perform structural studies in these worms would provide insight into the role of structure in disease pathogenesis. Many parameters were optimized for labeling within the worms including the microfluidic flow system and hydrogen peroxide concentration. IV-FPOP was able to modify several hundred proteins in various organs within the worms. The method successfully probed solvent accessibility similarily to in vitro FPOP, demonstrating its potential for use as a structural technique in a multiorgan system. The coupling of the method with mass spectrometry allows for amino-acid-residue-level structural information, a higher resolution than currently available in vivo methods.

A smart wound dressing based on carrageenan (κC), locust bean gum (LBG), and cranberry extract (CB) for monitoring bacterial wound infections was developed and characterized using UV-vis spectroscopy, FT-IR, and SEM. The mechanical, swelling, cytotoxic and pH sensor properties were also investigated. UV-vis spectra demonstrated that the obtained κC:LBG:CB hydrogel film exhibited a visible change of colors as it was immersed in PBS solution pH 5.0, 7.3 and 9.0. The spectra of FT-IR suggested that chemical interactions had occurred between κC and CB extract. The obtained κC:LBG:CB hydrogel film exhibited adequate mechanical properties and a swelling behavior dependent on pH. Cytotoxicity tests indicated that κC:LBG:CB hydrogel film had dose-dependent cytotoxicity against NIH 3T3 fibroblast cells. The in vitro studies using Staphylococcus aureus and Pseudomonas aeruginosa demonstrated that the color changes of the κC:LBG:CB hydrogel film could be observed by naked eyes, confirming the potential use of the obtained hydrogel film as a visual system for monitoring bacterial wound infections., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Many dynamic biological processes are regulated by protein-protein interactions and protein localization. Experimental techniques to probe such processes with temporal and spatial precision include photoactivatable proteins and chemically induced dimerization (CID) of proteins. CID has been used to study several cellular events, especially cell signaling networks, which are often reversible. However, chemical dimerizers that can be both rapidly activated and deactivated with high spatiotemporal resolution are currently limited. Herein, we present a novel chemical inducer of protein dimerization that can be rapidly turned on and off using single pulses of light at two orthogonal wavelengths. We demonstrate the utility of this molecule by controlling peroxisome transport and mitotic checkpoint signaling in living cells. Our system highlights and enhances the spatiotemporal control offered by CID. This tool addresses biological questions on subcellular levels by controlling protein-protein interactions.

Background: To investigate and compare the cytotoxicity of indocyanine green (ICG), brilliant blue G (BBG) and trypan blue (TB) using ARPE-19 cells that have been pre-treated/post-treated with balanced salt solution (BSS) or foetal bovine serum (FBS)., Methods: The cultured human retina pigment epithelium ARPE-19 cells were pre-treated/post-treated with BSS or FBS (represent the autologous serum in clinic) in parallel with cells being soaked with various concentrations of ICG, BBG and TB. The cells were then assessed for viability, growth rate, reactive oxygen species (ROS) level, mitochondrial membrane potential (Δψ) and mitochondrial mass as cytotoxic indices. For the FBS pre-treated cells, only ROS was examined., Results: Using the MTT assay, cytotoxicity seemed to appear when the dye concentration was above 2.5 mg/mL for ICG but no cytotoxicity for BBG and TB at the concentrations used. Cell growth was arrested at a concentration 1 mg/mL when ICG or BBG were present but no arrest at any of the tested concentrations was found for TB with the cell-growth curve was slowest for ICG. Cellular ROS levels increased at all concentrations of all dyes, but the increasing slopes were decreased after FBS post-treatment washout., Conclusions: As a rinse buffer FBS performs much better than BSS in terms of cell rescue, which agrees with a clinical report when autologous whole blood was applied to macular hole surgery. However, FBS pre-treatment seems to be much better than FBS use as washout buffer in post-treatment., (© 2018 Royal Australian and New Zealand College of Ophthalmologists.)

This paper provides compound-specific toxicology limits for 20 widely used synthetic reagents and common by-products that are potential impurities in drug substances. In addition, a 15 μg/day class-specific limit was developed for monofunctional alkyl bromides, aligning this with the class-specific limit previously defined for monofunctional alkyl chlorides. Both the compound- and class-specific toxicology limits assume a lifetime chronic exposure for the general population (including sensitive subpopulations) by all routes of exposure for pharmaceuticals. Inhalation-specific toxicology limits were also derived for acrolein, formaldehyde, and methyl bromide because of their localized toxicity via that route. Mode of action was an important consideration for a compound-specific toxicology limit. Acceptable intake (AI) calculations for certain mutagenic carcinogens assumed a linear dose-response for tumor induction, and permissible daily exposure (PDE) determination assumed a non-linear dose-response. Several compounds evaluated have been previously incorrectly assumed to be mutagenic, or to be mutagenic carcinogens, but the evidence reported here for such compounds indicates a lack of mutagenicity, and a non-mutagenic mode of action for tumor induction. For non-mutagens with insufficient data to develop a toxicology limit, the ICH Q3A qualification thresholds are recommended. The compound- and class-specific toxicology limits described here may be adjusted for an individual drug substance based on treatment duration, dosing schedule, severity of the disease and therapeutic indication., (Copyright © 2018. Published by Elsevier Inc.)

The aim of this work was to evaluate a fungal DNA extraction procedure with the lowest inputs in terms of time as well as of expensive and toxic chemicals, but able to consistently produce genomic DNA of good quality for PCR purposes. Two types of fungal biological material were tested - mycelium and conidia - combined with two protocols for DNA extraction using Sodium Dodecyl Sulphate (SDS) and Cetyl Trimethyl Ammonium Bromide as extraction buffers and glass beads for mechanical disruption of cell walls. Our results showed that conidia and SDS buffer was the combination that lead to the best DNA quality and yield, with the lowest variation between samples. This study clearly demonstrates that it is possible to obtain high yield and pure DNA from pigmented conidia without the use of strong cell disrupting procedures and of toxic reagents., Significance and Impact of the Study: There are numerous methods for DNA extraction from fungi. Some rely on expensive commercial kits and/or equipments, unavailable for many laboratories, or make use of toxic chemicals such as chloroform, phenol and mercaptoethanol. This study clearly demonstrates that it is possible to obtain high yields of pure DNA from pigmented conidia without the use of strong and expensive cell disrupting procedures and of toxic reagents. The method herein described is simultaneously inexpensive and adequate to DNA extraction from several different types of fungi., (© 2017 The Society for Applied Microbiology.)

Objective: To determine the role of opioid, β-adrenergic, and metabotropic glutamate 5 receptors in sacral neuromodulation of bladder overactivity., Material and Methods: In α-chloralose anesthetized cats, intravesical infusion of 0.5% acetic acid (AA) irritated the bladder and induced bladder overactivity. Electric stimulation (5 Hz, 0.2 ms, 0.16-0.7V) of S1 or S2 sacral dorsal roots inhibited the bladder overactivity. Naloxone, propranolol, or MTEP were given intravenously (i.v.) to determine different neurotransmitter mechanisms., Results: AA significantly (p < 0.05) reduced bladder capacity to 7.7 ± 3.3 mL from 12.0 ± 5.0 mL measured during saline infusion. S1 or S2 stimulation at motor threshold intensity significantly (p < 0.05) increased bladder capacity to 179.4 ± 20.0% or 219.1 ± 23.0% of AA control, respectively. Naloxone (1 mg/kg) significantly (p < 0.001) reduced the control capacity to 38.3 ± 7.3% and the bladder capacity measured during S1 stimulation to 106.2 ± 20.8% of AA control, but did not significantly change the bladder capacity measured during S2 stimulation. Propranolol (3 mg/kg) significantly (p < 0.01) reduced bladder capacity from 251.8 ± 32.2% to 210.9 ± 33.3% during S2 stimulation, but had no effect during S1 stimulation. A similar propranolol effect also was observed in naloxone-pretreated cats. In propranolol-pretreated cats during S1 or S2 stimulation, MTEP (3 mg/kg) significantly (p < 0.05) reduced bladder capacity and naloxone (1 mg/kg) following MTEP treatment further reduced bladder capacity. However, a significant inhibition could still be induced by S1 or S2 stimulation after all three drugs were administered., Conclusions: Neurotransmitter mechanisms in addition to those activating opioid, β-adrenergic, and metabotropic glutamate 5 receptors also are involved in sacral neuromodulation., (© 2016 International Neuromodulation Society.)

Hypochlorite (ClO - ) and glutathione (GSH) have been reported to closely correlate with oxidative stress and related diseases; however, a clear mechanism is still unknown, mainly owing to a lack of accurate analytical methods for live cells. Herein we create a novel surface-enhanced Raman scattering (SERS) nanoprobe, 4-mercaptophenol (4-MP)-functionalized gold flowers (AuF/MP), for imaging and biosensing of ClO - and GSH in RAW 264.7 macrophage cells upon oxidative stress. The SERS spectra of AuF/MP change with the reaction between ClO - and 4-MP on AuFs within 1 min and then recover after reaction with GSH, resulting in the ratiometric detection of ClO - and GSH with high accuracy. The single SERS probe also shows high selectivity for ClO - and GSH detection against other reactive oxygen species and amino acids which may exist in biological systems, as well as remarkable sensitivity ascribed to a larger amount of hot spots on AuFs. The significant analytical performance of the developed nanoprobe, together with good biocompatibility and high cell-permeability, enables the present SERS probe imaging and real-time detection of ClO - and GSH in live cells upon oxidative stress.

The peptide bond-forming reagents 1-hydroxy-7-azabenzotriazole (HOAt, CAS 39968-33-7) and O-(7-Azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU, CAS 148893-10-1) either have structural alerts, unclassified features or are considered out of domain when evaluated for potential mutagenicity with in silico programs DEREK and CaseUltra. Since they are commonly used reagents in pharmaceutical drug syntheses, they may become drug substance or drug product impurities and would need to be either controlled to appropriately safe levels or tested for mutagenicity. Both reagents were tested in the bacterial reverse mutation (Ames) test at Covance, under GLP conditions, following the OECD test guideline and ICH S2(R1) recommendations and found to be negative. Our data show that HOAt and HATU-common pharmaceutical synthesis reagents-are not mutagenic, and can be treated as ordinary drug impurities., (© 2016 Wiley Periodicals, Inc.)

Previously we described the antinociceptive effect of magnesium sulfate and dizocilpine (MK-801) in the visceral and somatic rat models of pain. In the somatic model of pain, we established the influence of selective inhibitors of neuronal and inducible nitric oxide synthase on the antihyperalgesic effects of magnesium sulfate and dizocilpine. Therefore, the objective of the present study was to determine in the rat model of visceral pain whether same mechanisms are involved in the antinociceptive action of magnesium sulfate and dizocilpine. Analgesic activity was assessed using the acetic acid-induced writhing test in rats. Subcutaneous injection of either magnesium sulfate (15 mg/kg) or dizocilpine (0.01 mg/kg) decreased the number of writhes by about 60 and 70%, respectively. The role of nitric oxide on the effects of magnesium sulfate and dizocilpine was evaluated using selective inhibitor of neuronal (N-ω-Propyl-L-arginine hydrochloride (L-NPA)) and inducible (S-methylisothiourea (SMT)) nitric oxide synthase, which per se did not affect the number of writhes. We observed that the antinociceptive effect of magnesium sulfate or dizocilpine did not change in the presence of L-NPA (2 and 10 mg/kg, i.p.) and SMT (0.015 and 10 mg/kg, i.p.). We conclude that, nitric oxide produced by neuronal and inducible nitric oxide synthase does not modulate the effects of magnesium sulfate and dizocilpine in the visceral inflammatory model of pain in the rat.

N-sulfonated oversulfated chondroitin sulfate (NS-OSCS), recently reported as a potential threat to the heparin supply, was prepared along with its intermediate derivatives. All compounds were spiked into marketplace heparin and subjected to United States Pharmacopeia (USP) identification assays for heparin (proton nuclear magnetic resonance [(1)H NMR], chromatographic identity, % galactosamine [%GalN], anti-factor IIa potency, and anti-factor Xa/IIa ratio). The U.S. Food and Drug Administration (FDA) strong-anionic exchange high-performance liquid chromatography (SAX-HPLC) method resolved NS-OSCS from heparin and OSCS and had a limit of detection of 0.26% (w/w) NS-OSCS. The %GalN test was sensitive to the presence of NS-OSCS in heparin. Therefore, current USP heparin monograph tests (i.e., SAX-HPLC and %GalN) detect the presence of NS-OSCS in heparin., (Published by Elsevier Inc.)

Primary hepatocyte cell cultures are widely used for studying hepatic diseases with alterations in hepatic glucose and lipid metabolism, such as diabetes and non-alcoholic fatty liver disease. Therefore, small interfering RNAs (siRNAs) provide a potent and specific tool to elucidate the signaling pathways and gene functions involved in these pathologies. Although RNA interference (RNAi) in vitro is frequently used in these investigations, the metabolic alterations elucidated by different siRNA delivery strategies have hardly been investigated in transfected hepatocytes. To elucidate the influence of the most commonly used lipid-based transfection reagents on cultured primary hepatocytes, we studied the cytotoxic effects and transfection efficiencies of INTERFERin(®), Lipofectamine(®)RNAiMAX, and HiPerFect(®). All of these transfection agents displayed low cytotoxicity (5.6-9.0 ± 1.3-3.4%), normal cell viability, and high transfection efficiency (fold change 0.08-0.13 ± 0.03-0.05), and they also favored the satisfactory down-regulation of target gene expression. However, when effects on the metabolome and lipidome were studied, considerable differences were observed among the transfection reagents. Cellular triacylglycerides levels were either up- or down-regulated [maximum fold change: INTERFERin(®) (48 h) 2.55 ± 0.34, HiPerFect(®) (24 h) 0.79 ± 0.08, Lipofectamine(®)RNAiMAX (48 h) 1.48 ± 0.21], and mRNA levels of genes associated with lipid metabolism were differentially affected. Likewise, metabolic functions such as amino acid utilization from were perturbed (alanine, arginine, glycine, ornithine, and pyruvate). In conclusion, these findings demonstrate that the choice of non-viral siRNA delivery agent is critical in hepatocytes. This should be remembered, especially if RNA silencing is used for studying hepatic lipid homeostasis and its regulation.

6:2 Fluorotelomer alcohol (6:2 FTOH) was evaluated for potential developmental and reproductive toxicity. 6:2 FTOH was administered by oral gavage to Sprague-Dawley rats as a suspension in 0.5% aqueous methylcellulose at dosages of 5, 25, 125, or 250 mg/kg/day. The developmental toxicity study was performed in accordance with the Organization for Economic Development (OECD) Test Guideline 414, and the one-generation reproductive toxicity study was performed in accordance with the OECD Test Guideline 415. For the developmental toxicity study, adverse maternal toxicity observed at 250 mg/kg/day included reductions in body weight parameters and food consumption. Evidence of developmental toxicity was limited to increases in skeletal variations (ossification delays in the skull and rib alterations) at 250 mg/kg/day. There were no adverse maternal or developmental effects observed at 5, 25, or 125 mg/kg/day and there were no effects on reproductive outcome or quantitative litter data at any dose level. For the one-generation reproduction toxicity study, systemic parental and developmental toxicity were observed at 125 and 250 mg/kg/day. At 250 mg/kg/day, there was increased mortality among male and female parental rats, effects on body weight parameters, food consumption, and clinical signs, and there were effects on offspring survival indices and body weights. At 125 mg/kg/day, there was an increase in mortality in parental males only, and parental toxicity was limited to effects on body weight gain, food consumption (lactation), and clinical signs. Uterine weights were decreased at 125 and 250 mg/kg/day, although there were no corroborative histopathological changes. At 125 mg/kg/day, pup mortality was increased on lactation day 1, and body weights of the offspring were decreased during the second half of lactation. There was no evidence of either parental or developmental toxicity at 5 or 25mg/kg/day, and there were no effects on reproductive outcome at any dose level. Based on these data, 6:2 FTOH is not a selective reproductive or developmental toxicant at dosages that induce clear maternal/parental toxicity. Therefore, 6:2 FTOH would not be classified for reproductive/developmental toxicity under the United Nations' Globally Harmonized System of Classification and Labeling of Chemicals., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Introduction: Mercuric chloride poisoning is rare yet potentially life-threatening. We report a case of poisoning with a potentially significant amount of mercuric chloride which responded to aggressive management., Case Report: A 19-year-old female presented to the Emergency Department with nausea, abdominal discomfort, vomiting of blood-stained fluid, and diarrhea following suicidal ingestion of 2-4 g of mercuric chloride powder. An abdominal radiograph showed radio-opaque material within the gastric antrum and the patient's initial blood mercury concentration was 17.9 μmol/L (or 3.58 mg/L) at 3 h post-ingestion. Given the potential toxicity of inorganic mercury, the patient was admitted to the intensive care unit and chelation with dimercaprol was undertaken. Further clinical effects included mild hemodynamic instability, acidosis, hypokalemia, leukocytosis, and fever. The patient's symptoms began to improve 48 h after admission and resolved fully within a week., Discussion: Mercuric chloride has an estimated human fatal dose of between 1 and 4 g. Despite a reported ingestion of a potentially lethal dose and a high blood concentration, this patient experienced mild to moderate poisoning only and she responded to early and appropriate intervention. Mercuric chloride can produce a range of toxic effects including corrosive injury, severe gastrointestinal disturbances, acute renal failure, circulatory collapse, and eventual death. Treatment includes close observation and aggressive supportive care along with chelation, preferably with 2,3-dimercapto-1-propane sulfonate or 2,3-meso-dimercaptosuccinic acid.

Introduction: Phosgene is a rare exposure with strong clinical implications. We report a phosgene exposure that resulted in the patient's death., Case Report: A 58 year-old man arrived to the emergency department 1 hour after exposure to phosgene with complaints of a sore throat. Initial vital signs were blood pressure 175/118 mmHg, heart rate 98/min, respirations 12/min, and oxygen saturation of 93% on room air. Physical exam revealed few scattered rhonchi, without signs of distress. Initial arterial blood gases (ABG's) revealed pH 7.42, pCO2 43 mmHg, pO2 68 mmHg, HCO3 27 meq/L, and oxygen saturation of 93% on room air. Initial chest x-ray 2 hours after the exposure demonstrated clear lung fields. Approximately 2.5 hours after the exposure, he began complaining of dyspnea, restlessness and his oxygen saturation dropped below 90%. He received nebulized albuterol, 1 gram intravenous methylprednisolone, and 100 % oxygen via face mask. Minimal improvement was noted and he was intubated. The post intubation chest x-ray, 3.5 hours after the exposure, revealed diffuse alveolar infiltrates. Acetylcysteine, terbutaline, and IV steroids were administered without improvement. The patient died 30 hours after exposure., Discussion: There are many misunderstandings concerning phosgene due to its rare presentation. Traditional treatment modalities are often unproven in human trials and were unsuccessful in this case., Conclusion: This case highlights the significant toxicity that results from phosgene exposure and the challenges of the limited treatment modalities. There is concern for the use of this agent in chemical terrorism.

A new poly(aminoester) (EPAE-FA) containing folic acid and amino groups in the backbone and side chain was synthesized. EPAE-FA self-assembled readily with the plasmid DNA (pCMV-βgal) in HEPES buffer and was characterized by dynamic light scattering, zeta potential, fluorescence images, and XTT cell viability assays. To evaluate the transfection effect of graft ratio of FA on the EPAE system, EPAE-FA polymers with two different graft ratios (EPAE-FA12k and EPAE-FA14k) were also prepared. This study found that all EPAE-FA polymers were able to bind plasmid DNA and yielded positively charged complexes with nano-sized particles (< 200 nm). To assess the transfection efficiency mediated by EPAE and EPAE-FA polymers, we performed in vitro transfection activity assays using FR-negative (COS-7) and FR-positive (HeLa) cells. The EPAE-FA12k/DNA and EPAE-FA14k/DNA complexes were able to transfect HeLa cell in vitro with higher transfection efficiency than PEI25k/DNA at the similar weight ratio. These results demonstrated that the introduction of FA into EPAE system had a significant effect on transferring ability for FR-positive cells (HeLa). Examination of the cytotoxicity of PEI25k and EPAE-FA system revealed that EPAE-FA system had lower cytotoxicity. In this paper, EPAE-FA seemed to be a novel cationic poly(aminoester) for gene delivery and an interesting candidate for further study.

Neutral red (Nr) is relatively non-toxic and is widely used as indicator dye in many biological test systems. It absorbs visible light and is known to act as a photosensitizer, involving the generation of reactive oxygen species (type-I reaction) and singlet oxygen (type-II reaction). The mutagenicity of Nr was determined in the Ames test (with Salmonella typhimurium strains TA1535, TA97, TA98, TA98NR, TA100, and TA102) with and without metabolic activation, and with and without photo-activation on agar plates. Similarly to the situation following metabolic activation, photo-mutagenicity of Nr was seen with all Salmonella strains tested, albeit with different effects between these strains. To our knowledge, Nr is the only photo-mutagen showing such a broad action. Since the effects are also observed in strains not known to be responsive to ROS, this indicates that ROS production is not the sole mode of action that leads to photo-genotoxicity. The reactive species produced by irradiation are short-lived as pre-irradiation of an Nr solution did not produce mutagenic effects when added to the bacteria. In addition, mutagenicity in TA98 following irradiation was stronger than in the nitroreductase-deficient strain TA98NR, indicating that nitro derivatives that are transformed by bacterial nitroreductase to hydroxylamines appear to play a role in the photo-mutagenicity of Nr. Photo-genotoxicity of Nr was further investigated in the comet assay and micronucleus test in L5178Y cells. Concentration-dependent increases in primary DNA damage and in the frequency of micronuclei were observed after irradiation., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Two series of compounds namely, 4-aryl/heteroaryl hydrazino-3-acetyl-6-methyl-2H-pyran-2-ones (4a-4j) and pyrano[4,3-c]pyrazoles (6a-6e and 6g) were synthesized starting from 3-acetyl-4-chloro-6-methyl-2H-pyran-2-one (2). Estimation of pharmacotherapeutic potential, possible molecular mechanism of action, toxic/side effects and interaction with drug-metabolizing enzymes were made for the synthesized compounds on the basis of prediction of activity spectra for substances (PASS) prediction results and their analysis by PharmaExpert software. COX inhibition predicted by PASS was confirmed by experimental evaluation and validated via docking studies. Out of all the compounds, compounds 4h, 4j, 6e, 6g exhibited good anti-inflammatory activity, whereas compounds 4b, 4c, 4h, 4i, 4j, 6b, 6e, 6g showed excellent analgesic activity compared with standard drug Diclofenac sodium., (Copyright © 2012 Elsevier Masson SAS. All rights reserved.)

Female F344 rats were exposed to 4,4'-methylenebis(N,N'-dimethyl)aniline (MDA) by dietary feed at concentrations of 0, 50, 200, 375, 500, or 750 ppm for 5 d, 2 wk, 4 wk, and 13 wk duration. Endpoints evaluated included clinical observations, body weights, thyroid weights, serum thyroid hormones, blood MDA, gross pathology, and thyroid histopathology. There were no MDA exposure-related clinical signs of toxicity. Mean body weight decreased 5% compared to control in the 750 ppm group during study wk 6 through 13. Serum TSH increased and serum T4 and T3 levels decreased with increasing feed concentrations of MDA and time of exposure. Thyroid weight increases were both concentration- and exposure time-dependent and statistically significant at ≥375 ppm. Incidence and severity of decreased colloid, follicular cell hypertrophy and follicular cell hyperplasia were also related to MDA concentration and exposure time. A no-observed-adverse-effect level (NOAEL) of 200 ppm was selected based on the statistically significant increase in incidence of follicular cell hyperplasia at concentrations ≥375 ppm.

The method prescribed in the 8th edition of Japan's Specifications and Standards for Food Additives (JSSFA) for the quantitative analysis of thiabendazole was improved by eliminating the use of toxic reagents such as mercuric acetate and chromium trioxide. For exclusion of mercuric acetate, a nonaqueous titration was performed using four types of solvent systems, including acetic acid:acetic anhydride (1:5), acetic acid:acetic anhydride (3:7), acetic acid alone, and formic acid:acetic acid (1:10), that did not contain mercuric compounds. Because precipitates were formed in titrations using acetic acid alone and formic acid:acetic acid (1:10), we considered that it was difficult to determine the purity using these solvent systems. However, it was confirmed that the purity of thiabendazole dissolved in the two acetic acid:acetic anhydride solvent systems can be determined using either a visual indicator or potentiometry. Specifically, the purity of thiabendazole was determined to be 99.9% (relative standard deviation (RSD) = 0.07%) for acetic acid:acetic anhydride (1:5) and 99.7% (RSD = 0.13%) for acetic acid:acetic anhydride (3:7) With respect to chromium trioxide, it was determined that chromium trioxide can be excluded using acetic acid, which conforms to the JIS K8001 standard for nonaqueous titrations. Therefore, in this study, an improved method for the quantitative determination of thiabendazole was developed without the use of toxic reagents.

Purpose: To determine the toxicity in ARPE-19 human retinal pigment epithelium cells of trypan blue (TB) at 0.15% and 0.25% concentration, brilliant blue G (BBG) at 0.025% and 0.05%, their combination, and the effect of the addition of 4% polyethyleneglycol (PEG), as an additive for increasing the density and thus improving the staining in internal limiting membrane removal, on the individual dyes and their combinations, and compare the toxicity of the dyes to that of clinically used preparations., Methods: Cells were exposed for 5 and for 30 minutes to the different preparations. Cell viability was measured with the WST-1 assay measuring intracellular dehydrogenase activity., Results: Solutions containing PEG with BBG (0.025%), TB (0.15%), and mixtures of BBG (0.025%) with TB (0.15% and 0.25%) were the least toxic of the preparations as well as preparations of BBG at 0.025% in phosphate-buffered saline solution, while TB at 0.25% in phosphate-buffered saline solution was the most toxic. The addition of PEG reduced the toxicity of preparations containing TB either alone or in combination with BBG. These results were seen only after an incubation for 30 minutes; for a 5-minute incubation, no toxicity was seen for any of the preparations., Conclusions: For short incubation times, all dyes appear equally safe. For longer incubation times, TB preparations were more toxic than BBG preparations. The toxicity of TB was reduced by the addition of PEG. Further studies are required to determine the clinical impact of this finding.

A liquid chromatography-tandem mass spectrometry-based isotope dilution method was developed for the analysis of the triazine compounds melamine (MEL), ammeline (AMN), ammelide (AMD) and cyanuric acid (CYA) in infant formula samples purchased in Canada in 2008 for the purpose of a combined exposure and risk assessment. Infant formula samples were extracted with 1:1 acetonitrile-water, cleaned up on disposable ion-exchange solid-phase extraction cartridges, and analysed by ultra-high-performance liquid chromatography-tandem mass spectrometry. MEL and CYA were detected in almost all infant formula products: the highest concentrations observed were 0.32 mg kg(-1) MEL and 0.45 mg kg(-1) CYA. Samples that were relatively high in MEL in this survey tended to be low in CYA, and vice versa. Concentrations of AMN and AMD were very low in all samples. The total of MEL-related compounds (sum of all four analytes) in all samples was below the interim standard of 0.5 mg kg(-1) for infant formula products established by Health Canada.

This study investigated the effects of royal jelly (RJ) on acetic acid-induced colitis in rats. Twenty adult female Wistar albino rats were divided into four treatment groups of 5 animals each, including a control group (Group I); Group II was treated orally with RJ (150 mg kg(-1) body weight); Group III had acetic acid-induced colitis; and Group IV had acetic acid-induced colitis treated orally with RJ (150 mg kg(-1) body weight) for 4 weeks. Colitis was induced by intracolonic instillation of 4% acetic acid; the control group received physiological saline (10 mL kg(-1)). Colon samples were obtained under deep anaesthesia from animals in all groups. Tissues were fixed in 10% formalin neutral buffer solution for 24 h and embedded in paraffin. Six-micrometre-thick sections were stained with Mallory’s triple stain and toluidine blue in 1% aqueous solution at pH 1.0 for 5 min (for Mast Cells). RJ was shown to protect the colonic mucosa against the injurious effect of acetic acid. Colitis (colonic damage) was confirmed histomorphometrically as significant increases in the number of mast cells (MC) and colonic erosions in rats with acetic acid-induced colitis. The RJ treatment significantly decreased the number of MC and reduced the area of colonic erosion in the colon of RJ-treated rats compared with rats with untreated colitis. The results suggest that oral treatment with RJ could be used to treat colitis.

Toxicity of the environmental carcinogen chromate is known to involve sulfur starvation and also error-prone mRNA translation. Here we reconcile those facts using the yeast model. We demonstrate that: (i) cysteine and methionine starvation mimic Cr-induced translation errors, (ii) genetic suppression of S starvation suppresses Cr-induced mistranslation, and (iii) mistranslation requires cysteine and methionine biosynthesis. Therefore, Cr-induced S starvation is the cause of mRNA mistranslation. This establishes a single, novel pathway mediating the toxicity of chromate., (Copyright 2010 Elsevier Ltd. All rights reserved.)

Antinociceptive activity of methanolic extract of leaves of A. aspera was studied by peripheral/non-narcotic model of nociception like acetic acid induced writhing syndrome test and central/narcotic models like hot plate and tail flick tests. The methanolic extract of the plant, administered orally (@ 300, 600 and 900 mg/kg, body weight) and the standard drug (piroxicam; 10 mg/kg body weight, po) produced significant analgesic activity in acetic acid induced writhing syndrome as compared to the vehicle treated control group. In the hot plate analgesic test, in A. aspera at the above doses and the standard drug treated group (morphine sulphate @ 1.5 mg/kg, ip), the duration of reaction time (sec) increased dose dependently and significantly compared to the control group. In the tail flick test, the plant extract produced dose dependant increase in reaction time which was significantly higher in the test and standard group compared to the control group. The plant possesses significant antinociceptive property as evidenced in all the animal models of nociception. It might possibly exert its effect through diverse mechanism that may involve both central and peripheral pathways. The preliminary phytochemical investigation revealed the presence of steroids, alkaloids and triterpene in the methanolic extract of leaves of A. aspera which may be responsible for its antinociceptive activity.

Introduction: In 1991, the US National Institute for Occupational Safety and Health (NIOSH) reported an increased bladder cancer risk in a cohort of 1749 workers potentially exposed to o-toluidine and aniline at a chemical manufacturing plant. As additional information showed that workers in certain departments had been misclassified regarding o-toluidine exposure, we therefore conducted a reanalysis of the data using updated exposure categories., Methods: We updated exposure categories based on information ascertained during a plant walkthrough, documents on file at the plant, interviews with current and former employees, and answers provided by company and union officials to specific questions. Bladder cancer incidence was determined through 31 December 1988 and mortality through 31 December 1994., Results: Thirteen cases of bladder cancer were observed versus 3.57 expected (New York State rates excluding New York City) (standardised incidence ratio (SIR) 3.64, 95% CI 1.94 to 6.23). Among workers classified as definitely exposed, increasing risks were observed for longer duration of employment (for > or = 10 years, standardised rate ratio (SRR) 6.07, 95% CI 0.77 to 48.17) and time since first employment in the exposed departments (for > or = 20 years, SRR 3.39, 95% CI 0.40 to 29.03). One bladder cancer death was observed among those definitely exposed., Conclusions: These findings are comparable to the results reported earlier by NIOSH, and confirm that workers in this plant have an increased risk of bladder cancer.

Linear cyclen-based polyamine (LCPA, M(w) = 7392, M(w)/M(n) = 1.19) as a novel non-viral gene vector was designed and synthesized from 1,7-diprotected 1,4,7,10-tetraazacyclododecane (cyclen), bis(beta-hydroxylethyl)amine and epichlorohydrin. Agarose gel retardation and fluorescent titration using ethidium bromide showed the good DNA-binding ability of LCPA. It could retard pDNA at an N/P ratio of 4 and form polyplexes with sizes around 250-300 nm from an N/P ratio of 10 to 60 and relatively lower zeta-potential values (< +3 mV) even at the N/P ratio of 60. The cytotoxicity of LCPA assayed by MTT is much lower than that of 25 kDa PEI. In vitro transfection against A549 and 293 cells showed that the transfection efficiency of LCPA/DNA polyplexes is close to that of 25 kDa PEI at an N/P ratio of 10-15, indicating that the new material could be a promising non-viral polycationic reagent for gene delivery.

The parasites Tritrichomonas foetus and Trichomonas gallinae present veterinary and economic importance since they cause bovine and avian trichomonosis, respectively. The absence of a specific treatment and the necessity of effective and safe drugs against these etiologic agents have stimulated the search for new antiprotozoal drugs with high activity, low toxicity to the animal, and low cost. Screening of potential antiprotozoal molecules is currently a common practice and different kinds of organic solvents and surfactant vehicles are used, since many bioactive compounds have low water solubility. Thus, it is important to determine the toxicity and to provide the minimal inhibitory concentration (MIC) values of the most common solubilization vehicles used in biological activity in vitro evaluation: ethanol, methanol, isopropanol, acetone, DMSO, tween 20 and tween 80. The assays were conducted employing the resazurin microtiter assay, which demonstrated a rapid, safe, and quantitative method for the in vitro determination of T. foetus and T. gallinae trophozoites viability. In summary, all solvents and surfactants, except ethanol, could be used in cytotoxicity assays against T. foetus, and acetone, tween 20 and tween 80 are the preferential vehicles for in vitro analysis of potential bioactive compounds against T. gallinae, though these must be used with caution.

Aim: Nicotine is a well-known agent among 4000 chemicals in cigarettes. About 70 to 80% of nicotine is converted to cotinine, a major metabolite. The aim of the present study is to investigate the effect of cotinine on neural tube development in a chick embryo model., Material and Methods: Sixty fertile, specific pathogen free eggs were divided into 6 groups for this study. In the first group, a fixed cotinine concentration for each egg was calculated just to simulate the concentration of a smoker's blood level. A second experimental group was designed at a higher cotinine concentration. Embryos that succeeded to reach Hamburger-Hamilton stage 12 from each group were then embedded into paraffin for permanent sections. These two groups were compared with eggs subjected to vehicle (standard alcohol and ten times more alcohol concentration) and control groups (saline and sham groups)., Results: Embryos of the cotinine (regular dose), vehicle and control groups were normal, but embryos subjected to higher cotinine concentrations were malformed at the cranial part of the thoracic neural tube., Conclusion: Association of cotinine with neural tube defects was demonstrated in the present study. Cigarette smoking may induce hazardous effects on neural tube development.

Resazurin dye has been broadly used as indicator of cell viability in several types of assays for evaluation of the biocompatibility of medical and dental materials. Mitochondrial enzymes, as carriers of diaphorase activities, are probably responsible for the transference of electrons from NADPH + H+ to resazurin, which is reduced to resorufin. The level of reduction can be quantified by spectrophotometers since resazurin exhibits an absorption peak at 600 etam and resorufin at 570 etam wavelengths. However, the requirement of a spectrophotometer and specific filters for the quantification could be a barrier to many laboratories. Digital cameras containing red, green and blue filters, which allow the capture of red (600 to 700 etam) and green (500 to 600 etam) light wavelengths in ranges bordering on resazurin and resorufin absorption bands, could be used as an alternative method for the assessment of resazurin and resorufin concentrations. Thus, our aim was to develop a simple, cheap and precise method based on a digital CCD camera to measure the reduction of resazurin. We compared the capability of the CCD-based method to distinguish different concentrations of L929 and normal Human buccal fibroblast cell lines with that of a conventional microplate reader. The correlation was analyzed through the Pearson coefficient. The results showed a strong association between the measurements of the method developed here and those made with the microplate reader (r(2) = 0.996; p < 0.01) and with the cellular concentrations (r(2) = 0.965; p < 0.01). We concluded that the developed Colorimetric Quantification System based on CCD Images allowed rapid assessment of the cultured cell concentrations with simple equipment at a reduced cost.

Numerous industrial applications for man-made nanoparticles have been proposed. Interactions of nanoparticles with agents in the atmosphere may impact human health. We tested the postulate that in vitro exposures of respiratory epithelial cells to airborne magnetic nanoparticles (MNP; Fe(3)O(4)) with and without a secondary organic aerosol (SOA) and an inorganic acid could affect iron homeostasis, oxidative stress, and interleukin (IL)-8 release. Cell iron concentrations were increased after exposures to MNP and values were further elevated with co-exposures to either SOA or inorganic acid. Increased expression of ferritin and elevated levels of RNA for DMT1, proteins for iron storage and transport respectively, followed MNP exposures, but values were significant for only those with co-exposures to inorganic acid and organic aerosols. Cell iron concentration corresponded to a measure of oxidative stress in the airway epithelial cells; MNP with co-exposures to SOA and inorganic acid increased both available metal and indices of oxidant generation. Finally, the release of a proinflammatory cytokine (i.e. IL-8) by the exposed cells similarly increased with cell iron concentration. We conclude that MNP can interact with a SOA and an inorganic acid to present metal in a catalytically reactive state to cultured respiratory cells. This produces an oxidative stress to affect a release of IL-8.

This study examined the toxicological and physiological responses of a commercially important coral-reef grouper, Plectropomus leopardus (Serranidae), to injection of enriched stable-isotope barium chloride (BaCl(2)) solution. Thirty adult P. leopardus were subject to one of two (138)BaCl(2) injection treatment groups (corresponding to dosage rates of 2 and 4 mg (138)Ba kg(-1) body mass), and a control group in which fish were injected with 0.9% sodium chloride (NaCl) solution. Fish from each group were sampled at post-injection intervals of 48 h and 1, 3, 5 and 8 weeks, at which time blood and tissue samples were removed from each fish. Residual concentrations of Ba and (138)Ba:(137)Ba ratios were measured in muscle, gonad, liver and bone tissues of each experimental fish. Elevated Ba concentrations were detected in all treatment fish tissue samples within 48 h post injection. Residual Ba concentrations decreased throughout the remainder of the 8 week experimental period in all tissues except bone. The BaCl(2) injection had no significant effects on measured whole blood variables or on the plasma concentrations of steroid hormones. Enriched Ba stable isotopes can therefore be used at low dosages to mark larvae of commercially important marine fishes, without adverse effects on the health of the fishes or on humans who may consume them.

With a simple experimental set-up, common packaging materials were tested for their ninhydrin permeability. The packaging material was loaded with a ninhydrin treated paper on the inside and covered with an amino acid treated paper on the outside. If ninhydrin penetrates through the packaging film, "Ruhemann's purple" will form on the amino acid treated paper. In all, 28 packaging materials were tested over a period of 24 weeks. It turned out that all the films made of polyethylene (PE) showed ninhydrin permeability. With one exception, this permeability occurred after three days already. All packages made of other film material were not permeable to ninhydrin.

A large animal model is needed to study artificial lung attachment in a setting simulating chronic lung disease with significant pulmonary hypertension (PH). This study sought to create a sheep model that develops significant PH within 60 days with a low rate of mortality. Sephadex beads were injected in the pulmonary circulation of sheep every other day for 60 days at doses of 0.5, 0.75, and 1 g (n = 10, 10, 7). Mean pulmonary artery pressure, pulmonary capillary wedge pressure, and cardiac output were obtained every 2 weeks. In the 0.5, 0.75, and 1-g groups, 90, 70, and 14.3% of sheep completed the study, respectively, with the remainder experiencing heart failure. By the 60th day, pulmonary vascular resistance had increased (p < 0.01) from 0.89 +/- 0.3 to 3.2 +/- 0.9 mm Hg/(L/min) and from 0.9 +/- 0.3 to 4.3 +/- 3.2 mm Hg/(L/min) in the 0.5 and 0.75-g groups, respectively. Significant right ventricular hypertrophy was observed in the 0.75-g group but not in the 0.5-g group. Data from the 1-g group were insufficient for analysis due to high mortality. Thus, the 0.5 and 0.75-g groups generate significant PH, but the 0.75-g group is a better model of chronic PH in lung disease due to the development of right ventricular hypertrophy.

Luminol is a reagent that is used to enhance areas of non-visible bloodstaining and it is one of the most sensitive of such reagents available to the forensic scientist. However, its use, particularly within the UK and some other European countries, has been limited, predominantly due to concerns about the health and safety of the reagent. This paper reviews the literature currently available regarding the health and safety of luminol, and in the authors' view demonstrates that there are no significant health and safety concerns with the preparation of luminol solution and its application at the crime scene or in the laboratory, providing suitable precautions are taken.

This paper reviews the preclinical effects of brilliant blue G (BBG) on the morphology and functions of the retina, and reports on a pilot study of BBG staining and subsequent peeling of the internal limiting membrane (ILM) during vitreoretinal surgery. BBG solution was injected into rat eyes and investigated using light microscopy and electron microscopy, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining, and electroretinography (ERG). No pathological changes were caused by the BBG intravitreous injection. Although electron microscopy revealed that high doses of BBG induced vacuolization in the inner retinal cells, there was no reduction in the amplitude of the ERG waves and no detectable toxic effects. In the primate eyes, the ILM was clearly visualized by BBG staining, and peeled off easily from the retina. In the clinical study, BBG improved visualization of the ILM, allowing peeling and surgery to be performed successfully on patients with various vitroretinal diseases. Improvement of postoperative visual acuity was obtained in most cases, and no adverse effects were observed postoperatively. In conclusion, BBG has low toxicity, high staining ability, and is easy to handle, making it a good candidate dye for visualizing vitreoretinal disease surgery without adverse effects.

Unlabelled: Phenolphthalein was commonly used as a laxative for most of the 20th century. The use of phenolphthalein in laxatives has decreased since 1997 when the United States Food and Drug Administration (FDA) proposed to withdraw its classification as an over-the-counter drug (21 CFR, Part 310). Phenolphthalein has been previously evaluated in 2-year carcinogenicity studies by the National Toxicology Program (1996). The major route of human exposure to phenolphthalein is via ingestion, dermal contact, and inhalation of contaminated air originating from process units manufacturing the compound. In this study, the carcinogenic effects of phenolphthalein were studied in the haploinsufficient p16(Ink4a)/p19(Arf) mouse model as an ongoing goal of the NTP is to seek model systems for toxicology and carcinogenesis studies, especially those that can provide mechanistic information relative to understanding an agent's mode of action. Male and female haploinsufficient p16(Ink4a)/p19(Arf) mice were exposed to phenolphthalein (greater than 97% pure) in feed for 27 weeks. Genetic toxicology studies were conducted in mouse peripheral blood erythrocytes. 27-WEEK STUDY IN MICE: Groups of 15 male and 15 female mice were exposed to 0, 200, 375, 750, 3,000, or 12,000 ppm phenolphthalein (equivalent to average daily doses of approximately 35, 65, 135, 540, and 2,170 mg phenolphthalein/kg body weight to males and 50, 90, 170, 680, 2,770 mg/kg to females) in feed for 27 weeks. Survival of all exposed groups of male and female mice was similar to that of the control groups. Mean body weights of males in the 12,000 ppm group were less than those of the control group after week 11. No differences in feed consumption were noted between exposed and control groups. Atypical hyperplasia of the thymus, a premalignant change of chemically induced thymic lymphoma, occurred in exposed males and females, and the incidence was significantly increased in 12,000 ppm females. Atrophy of the seminiferous tubules in the testis, hyperplasia of the testicular interstitial (Leydig) cells, and epididymal hypospermia occurred in most 3,000 and 12,000 ppm males. Additionally, the left and right testis weights, the left epididymis weights, sperm motility, the numbers of spermatid heads per testis, and sperm heads per cauda and per gram cauda epididymis were generally significantly less in 3,000 and 12,000 ppm males than in the control group. The incidences of nephropathy were significantly increased in 3,000 and 12,000 ppm males; incidences of hypertrophy of renal tubules were significantly increased in males receiving 750 ppm or greater. Hematopoietic cell proliferation of the spleen occurred in all 12,000 ppm males, and the incidences of this lesion were significantly increased in 375, 750, and 12,000 ppm females., Genetic Toxicology: The frequency of micronucleated erythrocytes was assessed at four time points during the 27-week study in male and female haploinsufficient p16(Ink4a)/p19(Arf) mice. Significant concentration-related increases in micronucleated cells were observed at all time points in male and female mice., Conclusions: Under the conditions of this 27-week feed study, there was no evidence of carcinogenic activity of phenolphthalein in male or female haploinsufficient p16(Ink4a)/p19(Arf) mice exposed to 200, 375, 750, 3,000, or 12,000 ppm. Because this is a new model, there is uncertainty whether the study possessed sufficient sensitivity to detect a carcinogenic effect. Phenolphthalein induced atypical hyperplasia, a preneoplastic lesion of the thymus, in male and female mice, hematopoietic cell proliferation of the spleen in male and female mice, and toxicity to the kidney and reproductive system in male mice.

This study evaluated the gastroprotective activity of DA-6034 against various ulcerogens including ethanol, aspirin, indomethacin, stress, and acetic acid. The basic mechanisms of DA-6034 as a defensive factor such as mucus secretion and endogenous prostaglandin E(2) synthesis were determined. Rats with gastric lesions induced by ethanol-HCl, aspirin, indomethacin, and stress that had been pretreated with DA-6034 orally showed a statistically significant decrease or decreasing tendency of the gastric lesion. In acetic acid-induced gastric lesions, repeated oral administration of DA-6034 exhibited a U-shape activity in ulcer healing, with the maximum and minimum inhibition being observed at 30 and 10 mg/kg/day, respectively. DA-6034 also increased the mucus content in the gel layer as well as endogenous prostaglandin E(2) synthesis. These results suggest that DA-6034 prevents gastric mucosal injury, and these gastroprotective activities appear to be due to the increase in the gastric defensive systems.

Inflammatory bowel disease (IBD) is a chronic condition of the intestine with unknown etiology involving multiple immune, genetic and environmental factors. We were interested in examining the effect of a total extract from Ziziphora clinopoides, an Iranian folk herbal medicine, in the prevention and control of experimental mouse IBD. Z. clinopoides was administered (75, 150, 300 mg/kg) through drinking water to mice, which dispensed a toxic dose of acetic acid intrarectally. Prednisolone was used as the standard drug for comparison. Biochemical, macroscopic and microscopic examinations of the colon were performed. Biochemical evaluation of the inflamed colon was carried out using assays of myeloperoxidase (MPO) activity and thiobarbituric acid reacting substances (TBARS) as indicators of free radical activity and cellular lipid peroxidation. Results indicated that the activity of MPO and lipid peroxidation products (TBARS) increased in acetic acid-treated groups, while recovered by pretreatment of animals with Z. clinopoides (75-300 mg/kg) and prednisolone. All doses of Z. clinopoides and prednisolone-treated groups showed significant lower score values of macroscopic and microscopic characters when compared to the acetic acid-treated group. The beneficial effect of Z. clinopoides (300 mg/kg) was comparable to that of prednisolone. It is concluded that Z. clinopoides inhibits acetic acid toxic reactions in the mouse bowel through inhibition of cellular oxidative stress. Proper clinical investigation should be carried out to confirm the same activity in human.

Nitric oxide (NO) is a non-adrenergic, non-cholinergic neurotransmitter found in the enteric nervous system that plays a role in a variety of enteropathies, including inflammatory bowel disease. Alteration of nitrergic neurons has been reported to be dependent on the manner by which inflammation is caused. However, this observed alteration has not been reported with acetic acid-induced colitis. Therefore, the purpose of the current study was to investigate changes in nitrergic neuromuscular transmission in experimental colitis in a rat model. Distal colitis was induced by intracolonic administration of 4 % acetic acid in the rat. Animals were sacrificed at 4 h and 48 h postacetic acid treatment. Myeloperoxidase activity was significantly increased in the acetic acid-treated groups. However, the response to 60 mM KCl was not significantly different in the three groups studied. The amplitude of phasic contractions was increased by Nomega-nitro-L-arginine methyl ester (L-NAME) in the normal control group, but not in the acetic acid-treated groups. Spontaneous contractions disappeared during electrical field stimulation (EFS) in normal group. However, for the colitis groups, these contractions initially disappeared, and then reappeared during EFS. Moreover, the observed disappearance was diminished by L-NAME; this suggests that these responses were NO-mediated. In addition, the number of NADPH-diaphorase positive nerve cell bodies, in the myenteric plexus, was not altered in the distal colon; whereas the area of NADPH-diaphorase positive fibers, in the circular muscle layer, was decreased in the acetic acidtreated groups. These results suggest that NO-mediated inhibitory neural input, to the circular muscle, was decreased in the acetic acid-treated groups.

Dynamic and reciprocal epithelial-mesenchymal interactions are critical for the normal morphogenesis and maintenance of epithelia. Epimorphin has been identified as a unique molecule expressed by mesenchymal cells and myofibroblasts and has putative morphogenetic effects in multiple epithelial tissues, including intestine, skin, mammary gland, lung, gallbladder, and liver. To define the in vivo role of epimorphin, we created epimorphin-null mice by targeted inactivation of the epimorphin gene. Male epimorphin-/- mice are sterile due to abnormal testicular development and impaired spermatogenesis. Intestinal growth is increased in epimorphin-/- mice due to augmented crypt cell proliferation and crypt fission during the neonatal (suckling) period, mediated at least in part by changes in bone morphogenetic protein (Bmp) and Wnt/beta-catenin signaling pathways. Colonic mucosal injury and colitis induced by dextran sodium sulfate (DSS) are ameliorated in epimorphin-/- mice, probably due to the increased proliferative capacity of the epimorphin-/- colon. These in vivo findings support the notion that epimorphin is a key stromal regulator of epithelial cell proliferation and growth in the intestine. In addition, our studies demonstrate a novel and critical role for epimorphin in regulating testicular development and growth as well as spermatogenesis.

Adverse effects of cDNA and oligonucleotide delivery methods have not yet been systematically analyzed. We introduce a protocol to monitor toxic effects of two non-viral lipid-based gene delivery protocols using CNS primary tissue. Cell membrane damage was monitored by quantifying cellular uptake of propidium iodide and release of cytosolic lactate dehydrogenase to the culture medium. Using a liposomal transfection reagent, cell membrane damage was already seen 24 hr after transfection. Nestin-positive target cells, which were used as morphological correlate, were severely diminished in some areas of the cultures after liposomal transfection. In contrast, the non-liposomal transfection reagent revealed no signs of toxicity. This approach provides easily accessible information of transfection-associated toxicity and appears suitable for prescreening of transfection reagents.