Synthesis and evaluation of 2,5-di(4-aryloylaryloxymethyl)-1,3,4-oxadiazoles as anti-cancer agents

A series of 2,5-di(4-aryloylaryloxymethyl)-1,3,4-oxadiazoles 9a-j were obtained via multistep synthesis from hydroxybenzophenones 4a-e. The cytotoxicity of compounds 9a-j was evaluated against human leukemia cell lilies (K562 and CEM). The compounds exhibited moderate to good anti-cancer activity with compounds 9b and 9i having a chloro group exhibiting the best activity (IC50 = 10 mu M). Compound 9i exhibited activity against both the cell lines and 9b only exhibited activity against CEM. Further, a lactate dehydrogenase (LDH) assay and DNA fragmentation studies of the compounds 9a-j were also performed. (C) 2013 Elsevier Masson SAS. All rights reserved.

Osteoblast and monocyte responses to 444 ferritic stainless steel intended for a Magneto-Mechanically Actuated Fibrous Scaffold

The rationale behind this work is to design an implant device, based on a ferromagnetic material, with the potential to deform in vivo promoting osseointegration through the growth of a healthy periprosthetic bone structure. One of the primary requirements for such a device is that the material should be non-inflammatory and non-cytotoxic. In the study described here, we assessed the short-term cellular response to 444 ferritic stainless steel; a steel, with a very low interstitial content and a small amount of strong carbide-forming elements to enhance intergranular corrosion resistance. Two different human cell types were used: (i) foetal osteoblasts and (ii) monocytes. Austenitic stainless steel 316L, currently utilised in many commercially available implant designs, and tissue culture plastic were used as the control surfaces. Cell viability, proliferation and alkaline phosphatase activity were measured. In addition, cells were stained with alizarin red and fluorescently-labelled phalloidin and examined using light, fluorescence and scanning electron microscopy. Results showed that the osteoblast cells exhibited a very similar degree of attachment, growth and osteogenic differentiation on all surfaces. Measurement of lactate dehydrogenase activity and tumour necrosis factor alpha protein released from human monocytes indicated that 444 stainless steel did not cause cytotoxic effects or any significant inflammatory response. Collectively, the results suggest that 444 ferritic stainless steel has the potential to be used in advanced bone implant designs. © 2011 Elsevier Ltd.

Influence of nutrient intake on antioxidant capacity, muscle damage and white blood cell count in female soccer players

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Running Pace Decrease during a Marathon Is Positively Related to Blood Markers of Muscle Damage

Background: Completing a marathon is one of the most challenging sports activities, yet the source of running fatigue during this event is not completely understood. The aim of this investigation was to determine the cause(s) of running fatigue during a marathon in warm weather. Methodology/Principal Findings: We recruited 40 amateur runners (34 men and 6 women) for the study. Before the race, body core temperature, body mass, leg muscle power output during a countermovement jump, and blood samples were obtained. During the marathon (27 uC; 27% relative humidity) running fatigue was measured as the pace reduction from the first 5-km to the end of the race. Within 3 min after the marathon, the same pre-exercise variables were obtained. Results: Marathoners reduced their running pace from 3.5 6 0.4 m/s after 5-km to 2.9 6 0.6 m/s at the end of the race (P,0.05), although the running fatigue experienced by the marathoners was uneven. Marathoners with greater running fatigue (. 15% pace reduction) had elevated post-race myoglobin (1318 6 1411 v 623 6 391 mg L21; P,0.05), lactate dehydrogenase (687 6 151 v 583 6 117 U L21; P,0.05), and creatine kinase (564 6 469 v 363 6 158 U L21; P = 0.07) in comparison with marathoners that preserved their running pace reasonably well throughout the race. However, they did not differ in their body mass change (23.1 6 1.0 v 23.0 6 1.0%; P = 0.60) or post-race body temperature (38.7 6 0.7 v 38.9 6 0.9 uC; P = 0.35). Conclusions/Significance: Running pace decline during a marathon was positively related with muscle breakdown blood markers. To elucidate if muscle damage during a marathon is related to mechanistic or metabolic factors requires further investigation.

FTY720 attenuates excitotoxicity and neuroinflammation

Background: FTY720 (fingolimod, Gilenya(TM)), a structural analog of sphingosine-1-phosphate (S1P), is the first oral drug approved for treatment the relapsing-remitting form of multiple sclerosis (MS), and its efficacy has been related to induced lymphopenia and consequent immunosuppression via modulation of S1P(1) receptors (S1P(1)R). However, due to its lipophilic nature, FTY720 crosses the blood brain barrier (BBB) and could act directly on neural cells. In this study, we investigated the effectiveness of FTY720 as a neuroprotective agent using in vitro and in vivo models of excitotoxic neuronal death and examined if FTY720 exerts a direct action on neurons, or/and an indirect modulation of inflammation-mediated neurodegeneration as a possible mechanism of neuroprotection. Methods: Primary neuronal and organotypic cortical cultures were treated with N-methyl-D-aspartic acid (NMDA) to induce excitotoxic cell death (measured by lactate dehydrogenase (LDH) assay or propidium iodide uptake, respectively). The effects of FTY720 treatment (10, 100 and 1,000 nM) on neuronal survival were examined. As an in vivo model of neuronal death and inflammation, we used intracerebroventricular (icv) administration of kainic acid (KA; 0.5 mu g/2 mu l) in Sprague-Dawley rats. FTY720 was applied icv (1 mu g/2 mu l), together with KA, plus intraperitoneally (ip; 1 mg/kg) 24 h before, and daily, until sacrifice 3 days after icv. Rats were evaluated for neurological score, neuronal loss in CA3 hippocampal region and activation of microglia at the lesion site. In addition, we tested FTY720 as a modulator of microglia responses using microglial cell cultures activated with lipopolysaccharide (LPS) and its effects in stress signalling pathways using western blotting for p38 and JNK1/2 mitogen-activated protein kinases (MAPKs). Results: FTY720 was able to reduce excitotoxic neuronal death in vitro. Moreover, in vivo repeated FTY720 administration attenuated KA-induced neurodegeneration and microgliosis at the CA3 lesion site. Furthermore, FTY720 negatively modulates p38 MAPK in LPS-activated microglia, whereas it had no effect on JNK1/2 activation. Conclusions: These data support a role for FTY720 as a neuroprotective agent against excitotoxin-induced neuronal death and as a negative modulator of neuroinflammation by targeting the p38 MAPK stress signalling pathway in microglia.

Hybridization between two serranids, the coney (Cephalopholis fulva) and the creole-fish (Paranthias furcifer), at Bermuda

Intergeneric hybridization between the epinepheline serranids Cephalopholis fulva and Paranthias furcifer in waters off Bermuda was investigated by using morphological and molecular characters. Putative hybrids, as well as members of each presumed parent species, were analyzed for 44 morphological characters and screened for genetic variation at 16 nuclear allozyme loci, two nuclear (n)DNA loci, and three mitochondrial (mt)DNA gene regions. Four of 16 allozyme loci, creatine kinase (CK-B*), fumarase (FH*), isocitrate dehydrogenase (ICDH-S*), and lactate dehydrogenase (LDH-B*), were unique in C. fulva and P. furcifer. Restriction fragments of two nuclear DNA intron regions, an actin gene intron and the second intron in the S7 ribosomal protein gene, also exhibited consistent differences between the two presumed parent species. Restriction fragments of three mtDNA regions—ND4, ATPase 6, and 12S/16S ribosomal RNA—were analyzed to identify maternal parentage of putative hybrids. Both morphological data and nuclear genetic data were found to be consistent with the hypothesis that the putative hybrids were the result of interbreeding between C. fulva and P. furcifer. Mean values of 38 morphological characters were different between presumed parent species, and putative hybrids were intermediate to presumed parent species for 33 of these characters. A principal component analysis of the morphological and meristic data was also consistent with hybridization between C. fulva and P. furcifer. Thirteen of 15 putative hybrids were heterozygous at all diagnostic nuclear loci, consistent with F1 hybrids. Two putative hybrids were identified as post-F1 hybrids based on homozygosity at one nuclear locus each. Mitochondrial DNA analysis showed that the maternal parent of all putative hybrid individuals was C. fulva. A survey of nuclear and mitochondrial loci of 57 C. fulva and 37 P. furcifer from Bermuda revealed no evidence of introgression between the parent species mediated by hybridization.

Effect of organophosphate, diazinon on some immunophysiological, haematological and histological variable of Rutilus frisii kutum (Kamensky,1901)

The acute toxicity and effects of diazinon on some haematological parameters of kutum (Rutilus frisii kutum, Kamensky, 1901) weighing 613.33 g±157.06 g were studied under static water quality conditions at 15°C ± 2ºC in winter and spring 2009. The effective physical and chemical parameters of water were pH= 7-8.2, dh= 300mg/L (caco3), DO= 7 ppm and T= 15°C±2ºC. The first test was primarily to determine the effects of acute toxicity (LC5096 h) of the agricultural toxicant diazinon (emulsion 60%) on kutum male brood stocks. For this purpose, 4 treatments were used to test toxicity; each treatment was repeated in 3 tanks with 9 fish per treatment and with 180 litres water capacity. After obtaining the final results, the information was analysed statistically with Probit version 1.5 (USEPA, 1985), and we determined the LC10, LC50 and LC90 values at 24 hours, 48 hours, 72 hours and 96 hours; the maximum allowable concentration value (LC5096 h divided by 10) (TRC, 1984); and the degree of toxicity. The second stage of testing consists of four treatments: LC0= 0 as experimental treatment, treatment A with a concentration of LC1= 0.107 mg/L, treatment B with concentration of LC5= 0.157 mg/L, treatment C with concentration of MAC value= 0.04 mg/L. Male brood stocks of kutum were treated with these concentrations for 45 days. Experiments were carried out under static conditions based on the standard TRC, 1984 method over 45 days. Our results show that long-term exposure to diazinon causes a decrease in the erythrocyte count (RBC), haemoglobin (Hb), haematocrit (PCV), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), leucocyte count (WBC), lymphocyte, testosterone, iron (Fe), sodium (Na), lactate dehydrogenase (LDH), and cholinesterase (CHeS). In addition, diazinon also causes an increase in prolymphocyte, aspartate aminotransferase (AST), cholesterol, alkaline phosphatase (ALP) and adrenaline (P<0.05). There are no significant effects on monocyte, eosinophil, magnesium (Mg), chloride (Cl), glucose (BS), urea (BUN), uric acid (U.A), triglyceride (TG), calcium (Ca), albumin (Alb), total protein (TP), cortisol, noradrenaline and high density lipoprotein (HDL) levels in kutum male brood stocks (P>0.05). Pathology results showed toxin diazinon no effect on average weight and fish body length, the average weight of heart, brain, spleen, liver, kidney and liver index but caueses decrease of gonad weigth and gonad index and also, cause complications of tissue necrosis, vascular congestion, inflammation in the liver, a sharp reduction in the number of glomeruli, necrosis, vascular congestion and haemorage in the kidney, capsule thickening and fibrosis, atrophy, vascular congestion, macrophages release increased, increasing sediment Hemosiderine and thickening of artery walls in the spleen, atrophy, fibrosis and necrosis in testis , vascular congestion, increased distance between the myocardium and fibrous string in heart and neuronal loss, vascular congestion and edema in the brain of kutum male brood stocks.

Transport of anti-sperm monoclonal IGA and IGC into murine male and female genital tracts from blood - Effect of sex hormones

Ab levels in the genital tract may be important in fertility and in preventing sexually transmitted diseases, In this study, I-125-labeled polymer or monomer mAb IgA (C4pIgA or C4mIgA) and IgC2b (C4IgC) to murine lactate dehydrogenase C4 and a polymer mAb IgA (npIgA) not cross-reacting with mouse sperm were intravenously injected into BALB/c mice, and the relative distribution of these Abs was determined. Polymer IgA was transported much more efficiently into the genital tract, trachea, and duodenum of both sexes than C4IgG and C4 mIgA (p < 0.01), The transport of polymer IgA (C4pIgA and npIgA) into the male genital tract greatly increased following orchiectomy (p < 0.01); this change was not affected by testosterone, suggesting that the unknown regulatory factor(s) from the testis may suppress polymer IgA transport, However, the transport of polymer IgA into female genital tissues was significantly decreased by ovariectomy (p < 0.01); this decline can be rectified by P-estradiol but not progesterone treatment, suggesting that estradiol may stimulate polymer IgA transport, Furthermore, the transport of C4IgG into tissues of the Fallopian tubes and the uterus was significantly decreased by treatment with progesterone (p < 0.01). Together, these findings indicate that serum polymer IgA can be transported selectively into the genital tracts of both sexes, that this transport is strongly under the control of gonads, and that transport of Ige into the Fallopian tubes and uterus is downregulated by progesterone.

Short-term in vitro responses of human peripheral blood monocytes to ferritic stainless steel fiber networks.

Beneficial effects on bone-implant bonding may accrue from ferromagnetic fiber networks on implants which can deform in vivo inducing controlled levels of mechanical strain directly in growing bone. This approach requires ferromagnetic fibers that can be implanted in vivo without stimulating undue inflammatory cell responses or cytotoxicity. This study examines the short-term in vitro responses, including attachment, viability, and inflammatory stimulation, of human peripheral blood monocytes to 444 ferritic stainless steel fiber networks. Two types of 444 networks, differing in fiber cross section and thus surface area, were considered alongside austenitic stainless steel fiber networks, made of 316L, a widely established implant material. Similar high percent seeding efficiencies were measured by CyQuant® on all fiber networks after 48 h of cell culture. Extensive cell attachment was confirmed by fluorescence and scanning electron microscopy, which showed round monocytes attached at various depths into the fiber networks. Medium concentrations of lactate dehydrogenase (LDH) and tumor necrosis factor alpha (TNF-α) were determined as indicators of viability and inflammatory responses, respectively. Percent LDH concentrations were similar for both 444 fiber networks at all time points, whereas significantly lower than those of 316L control networks at 24 h. All networks elicited low-level secretions of TNF-α, which were significantly lower than that of the positive control wells containing zymosan. Collectively, the results indicate that 444 networks produce comparable responses to medical implant grade 316L networks and are able to support human peripheral blood monocytes in short-term in vitro cultures without inducing significant inflammatory or cytotoxic effects.

Short-term cytotoxic and inflammatory responses of human monocytes to stainless steel fibre networks

The aim of the current work was to examine the human monocyte response to 444 ferritic stainless steel fibre networks. 316L austenitic fibre networks, of the same fibre volume fraction, were used as control surfaces. Fluorescence and scanning electron microscopies suggest that the cells exhibited a good degree of attachment and penetration throughout both networks. Lactate Dehydrogenase (LDH) and TNF-α releases were used as indicators of cytotoxicity and inflammatory responses respectively. LDH release indicated similar levels of monocyte viability when in contact with the 444 and 316L fibre networks. Both networks elicited a low level secretion of TNF-α, which was significantly lower than that of the positive control wells containing zymosan. Collectively, the results suggest that 444 ferritic and 316L austenitic networks induced similar cytotoxic and inflammatory responses from human monocytes. © 2012 Materials Research Society.

Effect of dietary linolenic acid/linoleic acid ratio on growth performance, hepatic fatty acid profiles and intermediary metabolism of juvenile yellow catfish Pelteobagrus fulvidraco

The present study aimed to evaluate the effect of dietary linolenic acid (LNA)linoleic acid (LA) ratio on growth performance, hepatic fatty acid profile and intermediary metabolism of juvenile yellow catfish Pelteobagrus fulvidraco. Six isonitrogenous and isolipidic diets were formulated to contain incremental levels of LNA from 0 to 5% at the expense of corn oil (rich in LA), resulting in six dietary treatments with LNA to LA ratios ranging from 0.35 to 14.64. The experiment continued for 7 weeks. Best growth and feed intake were obtained in the fish fed the diets containing the LNA/LA ratios of 1.17 and 2.12 (P<0.05). In contrast, feed conversion ratio was the lowest for fish fed the diets containing the LNA/LA ratios of 1.17 and 2.12 (P<0.05). Dietary LNA to LA ratios significantly influenced viscerosomatic index and hepatosomatic index (P<0.05), but not condition factor (P>0.05). Body composition was also significantly influenced by dietary LNA to LA ratios (P<0.05). Generally, liver FA compositions reflected dietary FA profiles. Declining LA and increasing LNA contents in liver were observed with the increasing dietary LNA/LA ratios (P<0.05). Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) increased with the increasing LNA to LA ratios, suggesting that yellow catfish could elongate and desaturate C18 polyunsaturated fatty acids into highly unsaturated fatty acids. As a consequence, the n-6 fatty acids (FA) declined, and total n-3 FA and n-3/n-6 ratios increased with the dietary ratios of LNA/LA (P<0.05). Dietary LNA to LA ratios significantly influenced several enzymatic activities involved in liver intermediary metabolism (P<0.05), such as lipoprotein lipase, hepatic lipase, pyruvate kinase, succinate dehydrogenase, malic dehydrogenase and lactate dehydrogenase, suggesting that dietary LNA/LA ratios had significant effects on nutrient metabolism in the liver. To our knowledge this is the first demonstration of the effects of dietary LNA to LA ratios on the enzymatic activities of liver in fish, which provides information on diet quality and utilization, and can also be used as an indicator of the nutritional status of this fish. (C) 2009 Elsevier B.V. All rights reserved.

First Identification of the Hepatotoxic Microcystins in the Serum of a Chronically Exposed Human Population Together with Indication of Hepatocellular Damage

Hepatotoxic microcystins (MCs) are the most commonly reported cyanotoxins in eutrophic freshwaters. In 1996, human intoxications by MCs caused deaths of 76 patients at Caruaru dialysis centers in Brazil. So far, there have been no direct evidences of MC occurrence in human tissue in consequence of exposure to MC. In this study, we improved cleanup procedures for detecting MCs in serum sample using liquid chromatographymass spectrometry, and confirmed for the first time the presence of MCs in serum samples (average 0.39 ng/ml, which amounts to ca. 1/87 of the concentrations found in tissue samples of the Caruaru victims) of fishermen at Lake Chaohu. Daily intake by the fishermen was estimated to be in the range of 2.2-3.9 mu g MC-LReq, whereas the provisional World Health Organization tolerable daily intake (TDI) for daily lifetime exposure is 0.04 mu g/kg or 2-3 mu g per person. Moreover, statistical analysis showed closer positive relationships between MC serum concentrations and concentrations of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and lactate dehydrogenase than between the MC concentrations and other biochemical indicators. Thus, the data raise the question whether extended exposure in the range of the TDI or up to a factor of 10 above it may already lead to indication of liver damage. The results also demonstrate a risk of health effects from chronic exposure to MCs at least for populations with high levels of exposure, like these fishermen.

Plasma biochemical responses of the omnivorous crucian carp (Carassius auratus) to crude cyanobacterial extracts

Healthy crucian carp (Carassius auratus) were treated by intraperitoneal (i.p.) injection of crude cyanobacterial extracts at two doses, 50 and 200 mu g MC-LR equiv kg(-1) BW. High mortality (100%) was observed within 60 h post injection in the high-dose group. In the treated fish, activities of four plasma enzymes, alanine aminotransferase (ALT), alkaline phosphatase (ALP), aspartate aminotransferase (AST), and lactate dehydrogenase (LDH), all showed substantial increases, with both dose and time-dependent effects. These increases of enzyme activity indicate severe impairment occurred in the liver of crucian carp over time. Plasma concentrations of energy-related biomolecules including glucose (GLU), cholesterol (CHO), triglyceride (TG), and total protein (TP) showed marked changes in the high-dose group, possibly a nutritional imbalance correlated with the liver injury caused by intraperitoneal exposure to crude cyanobacterial extracts.

Cytotoxicity evaluation of three pairs of hexabromocyclododecane (HBCD) enantiomers on Hep G2 cell

Hexabromocyclododecanes (HBCDs) are additive brominated flame retardants mainly used in plastics and textiles. At the present time, these compounds are found in almost all environmental and human samples. In order to evaluate the environmental safety and health risk of HBCDs, the enantiomerically pure alpha-, beta-, and gamma-HBCD were prepared using high performance liquid chromatography (HPLC) on a PM-P-CD column and the cytotoxicities of their enantiomers were evaluated in Hep G2 cells. Results from the 3-(4,5-dimethylthioazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), resazurin reduction and lactate dehydrogenase (LDH) release assays showed a good agreement that the order of cytotoxicity was gamma-HBCD >= beta-HBCD > alpha-HBCD, and that significantly lower cell viability and higher LDH release were observed in all (+)-enantiomers ((+) alpha-, (+) beta- and (+) gamma-HBCD) than the corresponding (-)-forms ((-) alpha-, (-) beta- and (-) gamma-HBCD). Additionally, the formation of reactive oxygen species (ROS) induced by these HBCD enantiomers were detected. The positive correlation between the LDH release and ROS formation demonstrated that the toxic mechanism might be mediated by oxidative damage. These results suggest that environmental and human health risks of HBCDs must be evaluated at the level of individual enantiomers. (C) 2008 Published by Elsevier Ltd.

DE-71-induced apoptosis involving intracellular calcium and the Bax-mitochondria-caspase protease pathway in human neuroblastoma cells In vitro

Polybrominated diphenyl ethers (PBDEs) are used extensively as flame-retardants and are ubiquitous in the environment and in wildlife and human tissue. Recent studies have shown that PBDEs induce neurotoxic effects in vivo and apoptosis in vitro. However, the signaling mechanisms responsible for these events are still unclear. In this study, we investigated the action of a commercial mixture of PBDEs (pentabrominated diphenyl ether, DE-71) on a human neuroblastoma cell line, SK-N-SH. A cell viability test showed a dose-dependent increase in lactate dehydrogenase leakage and 3-(4,5-dimethylthia-zol-2-yl)-2,5-diphenyl-tetrazolium bromide reduction. Cell apoptosis was observed through morphological examination, and DNA degradation in the cell cycle and cell apoptosis were demonstrated using flow cytometry and DNA laddering. The formation of reactive oxygen species was not observed, but DE-71 was found to significantly induce caspase-3, -8, and -9 activity, which suggests that apoptosis is not induced by oxidative stress but via a caspase-dependent pathway. We further investigated the intracellular calcium ([Ca2+](i)) levels using flow cytometry and observed an increase in the intracellular Ca2+ concentration with a time-dependent trend. We also found that the N-methyl d-aspartate (NMDA) receptor antagonist MK801 (3 mu M) significantly reduced DE-71-induced cell apoptosis. The results of a Western blotting test demonstrated that DE-71 treatment increases the level of Bax translocation to the mitochondria in a dose-dependent fashion and stimulates the release of cytochrome c (Cyt c) from the mitochondria into the cytoplasm. Overall, our results indicate that DE-71 induces the apoptosis of ([Ca2+](i)) in SK-N-SH cells via Bax insertion, Cyt c release in the mitochondria, and the caspase activation pathway.