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MTT Reduction (mtt + reduction)
Selected AbstractsCytotoxicity assessment of gliotoxin and penicillic acid in Tetrahymena pyriformisENVIRONMENTAL TOXICOLOGY, Issue 2 2006C. Gräbsch Abstract Various studies have documented the associations between mold exposure and effects on health. Mycotoxins, which occur in spores and mold fragments, can be involved in processes that have pathological effects, such as adynamia of the immune system, recurrent infections of the respiratory tract, or asthma. Using Tetrahymena pyriformis, a single-cell organism well established as a suitable model for human respiratory epithelium-cell functionalities, we investigated dose,response relationships of the mycotoxins gliotoxin and penicillic acid. Our study focused on the viability (cell count, MTT assay), energy levels (adenosine-5,-triphosphate content), energy-providing processes (MTT reduction per cell), and cell respiration (oxygen consumption). Both mycotoxins acted as cytotoxins in a dose-dependent manner. Gliotoxin had a stronger inhibitory effect (EC50 0.38 ,M) than did penicillic acid (EC50 343.19 ,M). The energy-providing processes were not inhibited or were only weakly inhibited under the influence of gliotoxin, whereas penicillic acid caused stimulation of the physiological parameters. Summarizing the results, it is clear that the two investigated mycotoxins must have different modes of action. They are not only different in the strength of their toxic effects but also in a variety of physiological aspects. In addition, T. pyriformis showed differences in its ability to overcome the negative effects of particular mycotoxin exposures. © 2006 Wiley Periodicals, Inc. Environ Toxicol 21: 111,117, 2006. [source] PGH2 -derived levuglandin adducts increase the neurotoxicity of amyloid ,1,42JOURNAL OF NEUROCHEMISTRY, Issue 4 2006Olivier Boutaud Abstract The body of evidence indicating that oligomers of amyloid ,1,42 (A,1,42) produce toxicity to neurons, together with our demonstration that prostaglandin H2 (PGH2) oligomerizes amyloid ,1,42, led to the examination of the neurotoxicity of amyloid ,1,42 treated with PGH2. The neurotoxic effects of A,1,42 incubated with PGH2 was examined in primary cultures of cerebral neurons of mice, monitoring the reduction of 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide (MTT) as an indicator of cell toxicity. Whereas A,1,42 itself, incubated for 24 h, has little or no effect on MTT reduction, A,1,42 24 h after exposure to PGH2 produced a marked inhibition of MTT reduction, comparable with the inhibition resulting from A,1,42 that has been oligomerized by incubation for 6 days. Similar results were obtained when A,1,42 was incubated with levuglandin E2 (LGE2), a reactive aldehyde formed by spontaneous rearrangement of PGH2. The oligomers formed from reaction of A,1,42 with LGE2 exhibit immunochemical similarity with amyloid-derived diffusible ligands (ADDLs), as determined by analysis of the products of reaction of A,1,42 with LGE2 using western blotting with an antibody that is selective for ADDLs. [source] Apolipoprotein E and ,-amyloid (1,42) regulation of glycogen synthase kinase-3,JOURNAL OF NEUROCHEMISTRY, Issue 5 2003A. Cedazo-Mínguez Abstract Glycogen synthase kinase-3, (GSK-3,) is implicated in regulating apoptosis and tau protein hyperphosphorylation in Alzheimer's disease (AD). We investigated the effects of two key AD molecules, namely apoE (E3 and E4 isoforms) and ,-amyloid (A,) 1,42 on GSK-3, and its major upstream regulators, intracellular calcium and protein kinases C and B (PKC and PKB) in human SH-SY5Y neuroblastoma cells. ApoE3 induced a mild, transient, Ca2+ -independent and early activation of GSK-3,. ApoE4 effects were biphasic, with an early strong GSK-3, activation that was partially dependent on extracellular Ca2+, followed by a GSK-3, inactivation. ApoE4 also activated PKC-, and PKB possibly giving the subsequent GSK-3, inhibition. A,(1,42) effects were also biphasic with a strong activation dependent partially on extracellular Ca2+ followed by an inactivation. A,(1,42) induced an early and potent activation of PKC-, and a late decrease of PKB activity. ApoE4 and A,(1,42) were more toxic than apoE3 as shown by MTT reduction assays and generation of activated caspase-3. ApoE4 and A,(1,42)-induced early activation of GSK-3, could lead to apoptosis and tau hyperphosphorylation. A late inhibition of GSK-3, through activation of upstream kinases likely compensates the effects of apoE4 and A,(1,42) on GSK-3,, the unbalanced regulation of which may contribute to AD pathology. [source] Activation of phosphoinositide-3 kinase/Akt pathway by FeSO4 in rat cerebral cortex synaptic endingsJOURNAL OF NEUROSCIENCE RESEARCH, Issue 13 2007Romina M. Uranga Abstract The aim of this work was to study the involvement of the phosphoinositide-3-kinase (PI3K)/Akt pathway in synaptic endings incubated under oxidative stress conditions. Synaptosomes purified from rat cerebral cortex were exposed to FeSO4 (50 ,M) for different periods of time. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction and lactate dehydrogenate (LDH) leakage were significantly affected after 5 min of incubation in the presence of FeSO4, with respect to control conditions. In whole synaptosomes incubated in the presence of [,- 32P]ATP, phosphoinositide (PPI) labeling was increased after 5 min of Fe2+ exposure. This effect was prevented by the specific PI3K inhibitor LY294002. Anti-p85 immunoprecipitates (IPs) obtained from synaptosomes preincubated with Fe2+ (5 min) showed a PI3K activity two-fold higher than the activity recovered under control conditions. Additionally, Akt activation was temporally coincident with PI3K activation. LY294002 was not able to prevent the LDH leakage and diminution of MTT reduction induced by Fe2+. Our results demonstrate that free iron provokes the early activation of PI3K/Akt pathway, but this activation is not sufficient for protecting synaptic endings from oxidative damage. © 2007 Wiley-Liss, Inc. [source] Differential effects of redox-cycling and arylating quinones on trans-plasma membrane electron transportBIOFACTORS, Issue 3 2008An S. Tan Abstract Cytotoxicity of quinones has been attributed to free radical generation and to arylation of cellular nucleophiles. For redox-cycling quinones, cell injury is associated with mitochondrial permeability transition, whereas arylating quinones directly depolarise the mitochondrial membrane and deplete ATP. Like mitochondrial electron transport, plasma membrane electron transport (PMET), plays a multifaceted role in cellular redox homeostasis but the effects of quinones on PMET are unknown. Here we investigate the effects of redox-cycling 2,3-dimethoxy-1,4-naphthoquinone (DMNQ), arylating 1,4-benzoquinone (BQ) and mixed mechanism 2-methyl-1,4-naphthoquinone (MNQ) on PMET, viability and growth of P815 mouse mastocytoma cells. BQ and MNQ rapidly and extensively inhibited PMET as determined by WST-1 /mPMS reduction (IC50 3.5-5 ,Mat 30 min) whereas the effects of DMNQ were less pronounced. In contrast, MTT reduction (cytosolic NADH dehydrogenase activity over 30 min) was weakly inhibited by BQ (IC50 20 ,M) but not by MNQ or DMNQ and cell viability was unaffected. Inhibition of WST-1/mPMS reduction by BQ and MNQ but not DMNQ was fully reversed by NAC. Treatment with DMNQ, MNQ and to a lesser extent BQ inhibited cell proliferation as determined by MTT reduction at 48 h. The effects of BQ and MNQ were reversed by NAC through covalent bonding to BQ and MNQ, but not DMNQ. These results show that arylating quinones are more potent inhibitors of PMET than pure redox-cycling quinones, but that redox-cycling quinones are more cytotoxic. [source] | ||||||||||