benzyloxycarbonylvalyl-alanyl-aspartyl-fluoromethyl-ketone and 6-hydroxy-2-5-7-8-tetramethylchroman-2-carboxylic-acid

This study describes the mechanism of trolox and tiron induced potentiation of cytotoxicity caused by Ery5, an analog of magnolol, in human myeloid leukemia HL-60 cells. Ery5 induced cytotoxicity in HL-60 cells by involving activation of bax and cleavage of caspase 3, which contributed towards activation of both apoptotic and autophagic pathways. Trolox and tiron, even at non-toxic concentrations, contributed to the cytotoxicity of Ery5 by activation of autophagic proteins like ATG7, ATG12 and LC3-II. Z-VAD-fmk mediated reduction in the cytotoxicity and expression of autophagic proteins, further suggested that autophagy induced by Ery5 is largely dependent upon caspases. Interestingly, Ery5 induced autophagy was accompanied by the downregulation of PI3K/AKT pathway whereas, trolox and tiron strongly enhanced this effect. In addition to that treatment of cells with Ery5, trolox and tiron individually, displayed a marked upregulation of Bax. The involvement of Bax in trolox and tiron induced enhancement of the cytotoxicity of Ery5 was confirmed, when siRNA induced silencing of Bax led to increased viability of the cells and exerted a strong inhibitory effect on LC3-II accumulation and p62 degradation in case of cells treated by the combination of Ery5 with trolox or tiron. Additionally, an important role of PARP in Ery5 mediated cell death has been suggested by PARP silencing experiments, however, potentiation of autophagic cytotoxicity by trolox and tiron did not seem to be dependent on PARP-1. Therefore, Bax seems to play a vital role in trolox and tiron mediated potentiation of autophagic cell death by Ery5 in HL-60 cells.

Topics: 1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt; Amino Acid Chloromethyl Ketones; Antineoplastic Agents; Apoptosis; Autophagy; Autophagy-Related Protein 12; Autophagy-Related Protein 7; bcl-2-Associated X Protein; Biphenyl Compounds; Caspases; Chromans; Drug Synergism; Gene Expression Regulation, Neoplastic; HL-60 Cells; Humans; Lignans; Microtubule-Associated Proteins; Phenols; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-ret; Signal Transduction; Small Ubiquitin-Related Modifier Proteins; Ubiquitin-Activating Enzymes

The objective was to determine the effects of growth factor treatment on dental pulp cell sensitivity to toxicity of 2 composite restoration materials, Flow Line and Durafill VS, and a calcium hydroxide pulp capping material, Dycal.. Toxicity of the dental materials to cultures of primary dental pulp cells was determined by the MTT metabolism assay. The ability of 6 different growth factors to influence the toxicity was tested.. A 24-hour exposure to either Flow Line or Durafill VS caused approximately 40% cell death, whereas Dycal exposure caused approximately 80% cell death. The toxicity of Flow Line and Durafill VS was mediated by oxidative stress. Four of the growth factors tested (bone morphogenetic protein [BMP]-2, BMP-7, epidermal growth factor [EGF], and transforming growth factor [TGF]-beta) decreased the basal MTT values while making the cells resistant to Flow Line and Durafill VS toxicity except BMP-2, which made the cells more sensitive to Flow Line. Treatment with fibroblast growth factor-2 caused no change in basal MTT metabolism, prevented the toxicity of Durafill VS, but increased the toxicity of Flow Line. Treatment with insulin-like growth factor-I (IGF-I) increased basal MTT metabolism and made the cells resistant to Flow Line and Durafill VS toxicity. None of the growth factors made the cells resistant to Dycal toxicity.. The results indicated that growth factors can be used to alter the sensitivity of dental pulp cells to commonly used restoration materials. The growth factors BMP-7, EGF, TGF-beta, and IGF-I provided the best profile of effects, making the cells resistant to both Flow Line and Durafill VS toxicity.

Topics: Adult; Amino Acid Chloromethyl Ketones; Antioxidants; Bone Morphogenetic Protein 2; Bone Morphogenetic Protein 7; Calcium Hydroxide; Caspase Inhibitors; Cell Death; Cell Survival; Cells, Cultured; Chromans; Coloring Agents; Composite Resins; Dental Materials; Dental Pulp; Drug Tolerance; Epidermal Growth Factor; Fibroblast Growth Factor 2; Humans; Insulin-Like Growth Factor I; Intercellular Signaling Peptides and Proteins; Materials Testing; Minerals; Oxidative Stress; Tetrazolium Salts; Thiazoles; Time Factors; Transforming Growth Factor beta

Fas-mediated caspase-dependent cell apoptosis has been well investigated. However, recent studies have shown that Fas can induce nonapoptotic caspase-independent cell death (CICD) when caspase activity is inhibited. Currently, the molecular mechanism of this alternative cell death mediated by Fas remains unclear. In this study, we investigated the signaling pathway of Fas-induced CICD in mouse embryonic fibroblasts (MEFs) whose caspase function was disrupted by the pan-caspase inhibitor Z-VAD-FMK and its coupling to inflammatory responses. Our results revealed that receptor-interacting protein 1 and tumor necrosis factor receptor-associated factor 2 play important roles in FasL-induced CICD. This death is associated with intracellular reactive oxygen species (ROS) production from mitochondria, as a ROS scavenger (BHA), antioxidants (trolox, NAC), and a mitochondrial respiratory chain uncoupler (rotenone) could prevent this event. Furthermore, delayed and sustained JNK activation, mitochondrial membrane potential breakdown, and loss of intracellular GSH were observed. In addition to CICD, FasL also induces cyclooxygenase-2 and MIP-2 gene upregulation, and both responses are attributed to ROS-dependent JNK activation. Taken together, these results demonstrate alternative signaling pathways of Fas upon caspase inhibition in MEFs that are unrelated to the classical apoptotic pathway, but steer cells toward necrosis and an inflammatory response through ROS production.

Topics: Amino Acid Chloromethyl Ketones; Animals; Apoptosis; Caspase Inhibitors; Caspases; Cell Line; Chemokine CXCL2; Chromans; Cyclooxygenase 2; Fas Ligand Protein; fas Receptor; Fibroblasts; Gene Expression Regulation; Inflammation; Membrane Potential, Mitochondrial; Mice; Mitochondria; Reactive Oxygen Species; Signal Transduction

5-fluorouracil (5-FU) is a widely used anticancer drug. One of the adverse effects of this drug is selective cerebral white matter injury, but to the authors' knowledge its mechanism has not been well documented. The current study was performed to investigate the mechanism of cerebral white matter injury caused by 5-FU and to develop the intervention to attenuate its injury in vitro.. Mixed oligodendrocyte/astrocyte cells were dissociated from specimens taken from approximately 2-day-old postnatal mouse cortex and cultured for 3-4 weeks. The culture cells were exposed to 5-FU, cycloheximide, emetine, Z-VAD-fmk, 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)-quinoxaline (NBQX), Trolox, and epigallocatechin gallate. Oligodendrocyte cell death was assessed by counting the number of viable galactocerebroside-positive cells per x 100 field.. Mixed oligodendrocyte/astrocyte culture cells that were exposed to 5-FU (at doses of 10 microM, 30 microM, and 100 microM) for 24 hours ensued concentration-dependent oligodendrocyte death. The majority of oligodendrocytes, but few astrocytes, were injured by 100 microM 5-FU. Trolox, a vitamin E analog antioxidant, as well as cycloheximide (a protein synthesis inhibitor) and Z-VAD-fmk (a caspase inhibitor), significantly attenuated the 5-FU-induced oligodendrocyte death. However, NBQX, an alpha-amino-2,3-dihydro-5-methyl-3-oxo-4-isoxazolepropionic acid (AMPA) receptor antagonist, did not appear to effect the 5-FU-induced oligodendrocyte death.. The findings of the current study suggested that 5-FU led to oligodendrocyte death rather than astrocyte death by way of the apoptotic process, whereas antioxidants may prevent the 5-FU-induced oligodendrocyte cell death in vitro.

Topics: Amino Acid Chloromethyl Ketones; Animals; Antimetabolites, Antineoplastic; Antioxidants; Caspase Inhibitors; Cell Culture Techniques; Cell Death; Chromans; Cycloheximide; Cysteine Proteinase Inhibitors; Excitatory Amino Acid Antagonists; Fluorouracil; Mice; Oligodendroglia; Protein Synthesis Inhibitors; Quinoxalines; Receptors, AMPA

There have been a number of studies regarding the toxicity of orthodontic archwires, but little is known concerning the mechanism of their toxicity. This investigation used murine cortical cell cultures to examine the in vitro neurotoxicity of commonly used orthodontic metallic archwire alloys. The materials examined included 0.016 inch nickel-titanium (NiTi), copper-nickel-titanium, titanium-molybdenum, Elgiloy, and stainless steel archwire alloys. Standard sized samples of each material were placed on tissue culture inserts suspended above the cell cultures. Neuronal death was determined using the lactate dehydrogenase release assay 24 hours after exposure to the archwires. The results indicated that NiTi, copper-nickel-titanium and titanium-molybdenum alloys were not neurotoxic, while stainless steel and Elgiloy were significantly toxic. Washing the archwires for 7 days in a saline solution did not alter the toxicity. However, the free radical scavenger, trolox, blocked the toxicity of both stainless steel and Elgiloy, indicating that the death was free radical mediated. The caspase inhibitor, Z-VAl-Ala-Asp-fluoromethylketone (zVAD-FMK), blocked the toxicity of stainless steel, but not Elgiloy, suggesting that stainless steel induced apoptosis. Further evidence that stainless steel induced apoptosis was provided by propidium staining which showed nuclear chromatin condensation and fragmentation into discrete spherical or irregular shapes, characteristic of apoptosis. The specific metal responsible for the toxicity was not determined; the metals common to each of the toxic archwires were nickel, iron, and chromium.

Topics: Amino Acid Chloromethyl Ketones; Animals; Antioxidants; Apoptosis; Caspase Inhibitors; Cell Death; Cells, Cultured; Cerebral Cortex; Chromans; Chromium Alloys; Copper; Dental Alloys; Free Radicals; L-Lactate Dehydrogenase; Mice; Molybdenum; Neurons; Neuroprotective Agents; Nickel; Orthodontic Wires; Stainless Steel; Titanium

Evidence has accumulated that Zn2+ plays a central role in neurodegenerative processes following brain injuries including ischaemia or epilepsy. In the present study, we examined patterns and possible mechanisms of Zn2+ neurotoxicity. Inclusion of 30-300 microM Zn2+ for 30 min caused neuronal necrosis apparent by cell body and mitochondrial swelling in cortical cell cultures. This Zn2+ neurotoxicity was not attenuated by antiapoptosis agents, inhibitors of protein synthesis or caspase. Blockade of glutamate receptors or nitric oxide synthase showed no beneficial effect against Zn2+ neurotoxicity. Interestingly, antioxidants, trolox or SKF38393, attenuated Zn(2+)-induced neuronal necrosis. Pretreatment with insulin or brain-derived neurotrophic factor increased the Zn(2+)-induced free radical injury. Kainate or AMPA facilitated Zn2+ entry and potentiated Zn2+ neurotoxicity in a way sensitive to trolox. Reactive oxygen species and lipid peroxidation were generated in the early phase of Zn2+ neurotoxicity. These findings indicate that entry and accumulation of Zn2+ result in generation of toxic free radicals and then cause necrotic neuronal degeneration under certain pathological conditions in the brain.

Topics: 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; 6-Cyano-7-nitroquinoxaline-2,3-dione; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Amino Acid Chloromethyl Ketones; Animals; Antioxidants; Apoptosis; Brain-Derived Neurotrophic Factor; Cells, Cultured; Cerebral Cortex; Chromans; Cysteine Proteinase Inhibitors; Dizocilpine Maleate; Dopamine Agonists; Drug Synergism; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Female; Free Radicals; Hypoglycemic Agents; Insulin; Kainic Acid; Lipid Peroxidation; Mice; Microscopy, Electron; Mitochondrial Swelling; Necrosis; Nerve Degeneration; Neurons; Neurotoxins; Oligopeptides; Oxidative Stress; Pregnancy; Zinc