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Cell Metabolism Jun 2011The lipid droplet (LD) is the major site of cholesterol storage in macrophage foam cells and is a potential therapeutic target for the treatment of atherosclerosis....
The lipid droplet (LD) is the major site of cholesterol storage in macrophage foam cells and is a potential therapeutic target for the treatment of atherosclerosis. Cholesterol, stored as cholesteryl esters (CEs), is liberated from this organelle and delivered to cholesterol acceptors. The current paradigm attributes all cytoplasmic CE hydrolysis to the action of neutral CE hydrolases. Here, we demonstrate an important role for lysosomes in LD CE hydrolysis in cholesterol-loaded macrophages, in addition to that mediated by neutral hydrolases. Furthermore, we demonstrate that LDs are delivered to lysosomes via autophagy, where lysosomal acid lipase (LAL) acts to hydrolyze LD CE to generate free cholesterol mainly for ABCA1-dependent efflux; this process is specifically induced upon macrophage cholesterol loading. We conclude that, in macrophage foam cells, lysosomal hydrolysis contributes to the mobilization of LD-associated cholesterol for reverse cholesterol transport.
Topics: Animals; Atherosclerosis; Autophagy; Autophagy-Related Protein 5; Bone Marrow Cells; Cells, Cultured; Chloroquine; Cholesterol; Foam Cells; Gene Knockout Techniques; Lipid Metabolism; Lipolysis; Lipoproteins, LDL; Lysosomes; Mice; Mice, Inbred C57BL; Mice, Knockout; Microtubule-Associated Proteins; Paraoxon; Sterol Esterase
PubMed: 21641547
DOI: 10.1016/j.cmet.2011.03.023 -
Molecules (Basel, Switzerland) Mar 2020Organophosphates (OPCs), useful agents as pesticides, also represent a serious health hazard. Standard therapy with atropine and established oxime-type enzyme...
AIMS
Organophosphates (OPCs), useful agents as pesticides, also represent a serious health hazard. Standard therapy with atropine and established oxime-type enzyme reactivators is unsatisfactory. Experimental data indicate that superior therapeutic results can be obtained when reversible cholinesterase inhibitors are administered before OPC exposure. Comparing the protective efficacy of five such cholinesterase inhibitors (physostigmine, pyridostigmine, ranitidine, tacrine, or K-27), we observed best protection for the experimental oxime K-27. The present study was undertaken in order to determine if additional administration of K-27 immediately after OPC (paraoxon) exposure can improve the outcome.
METHODS
Therapeutic efficacy was assessed in rats by determining the relative risk of death (RR) by Cox survival analysis over a period of 48 h. Animals that received only pretreatment and paraoxon were compared with those that had received pretreatment and paraoxon followed by K-27 immediately after paraoxon exposure.
RESULTS
Best protection from paraoxon-induced mortality was observed after pretreatment with physostigmine (RR = 0.30) and K-27 (RR = 0.34). Both substances were significantly more efficacious than tacrine (RR = 0.67), ranitidine (RR = 0.72), and pyridostigmine (RR = 0.76), which were less efficacious but still significantly reduced the RR compared to the no-treatment group (paraoxon only). Additional administration of K-27 immediately after paraoxon exposure (posttreatment) did not further reduce mortality. Statistical analysis between pretreatment before paraoxon exposure alone and pretreatment plus K-27 posttreatment did not show any significant difference for any of the pretreatment regimens.
CONCLUSIONS
Best outcome is achieved if physostigmine or K-27 are administered prophylactically before exposure to sublethal paraoxon dosages. Therapeutic outcome is not further improved by additional oxime therapy immediately thereafter.
Topics: Animals; Cholinesterase Inhibitors; Cholinesterase Reactivators; Male; Organophosphates; Oximes; Paraoxon; Physostigmine; Post-Exposure Prophylaxis; Pre-Exposure Prophylaxis; Proportional Hazards Models; Pyridostigmine Bromide; Ranitidine; Rats; Rats, Wistar; Survival Analysis; Tacrine
PubMed: 32230733
DOI: 10.3390/molecules25071521 -
Acta Medica (Hradec Kralove) 2022Organophosphorus compounds induce irreversible inhibition of acetylcholinesterase, which then produces clinically manifested muscarinic, nicotinic and central effects....
Organophosphorus compounds induce irreversible inhibition of acetylcholinesterase, which then produces clinically manifested muscarinic, nicotinic and central effects. The aim of the study was to analyse the clinical signs of acute paraoxon poisoning in rats and to determine the relationship between the intensity of signs of poisoning and the dose of paraoxon and/or the outcome of poisoning in rats. Animals were treated with either saline or atropine (10 mg/kg intramuscularly). The median subcutaneous lethal dose (LD50) of paraoxon was 0.33 mg/kg and protective ratio of atropine was 2.73. The presence and intensity of signs of poisoning in rats (dyspnoea, lacrimation, exophthalmos, fasciculations, tremor, ataxia, seizures, piloerection, stereotypic movements) were observed and recorded for 4 h after the injection of paraoxon. Intensity of these toxic phenomena was evaluated as: 0 - absent, 1 - mild/moderate, 2 - severe. Fasciculations, seizures and tremor were more intense at higher doses of paraoxon and in non-survivors. In unprotected rats piloerection occurred more often and was more intense at higher doses of paraoxon as well as in non-survivors. In atropine-protected rats, piloerection did not correlate with paraoxon dose or outcome of poisoning. The intensity of fasciculations and seizures were very strong prognostic parameters of the poisoning severity.
Topics: Acetylcholinesterase; Animals; Atropine; Fasciculation; Paraoxon; Rats; Seizures; Tremor
PubMed: 35793503
DOI: 10.14712/18059694.2022.10 -
Chemical Research in Toxicology Sep 2022Exposure to organophosphorus pesticides (OP) can have chronic adverse effects that are independent of inhibition of acetylcholinesterase, the classic target for acute OP...
Exposure to organophosphorus pesticides (OP) can have chronic adverse effects that are independent of inhibition of acetylcholinesterase, the classic target for acute OP toxicity. In pure proteins, the organophosphorus pesticide chlorpyrifos oxon induces a cross-link between lysine and glutamate (or aspartate) with loss of water. Tubulin is particularly sensitive to OP-induced cross-linking. Our goal was to explore OP-induced cross-linking in a complex protein sample, MAP-rich tubulin from and to test 8 OP for their capacity to promote isopeptide cross-linking. We treated 100 μg of MAP-rich tubulin with 100 μM chlorpyrifos, chlorpyrifos oxon, methamidophos, paraoxon, diazinon, diazoxon, monocrotophos, or dichlorvos. Each sample was separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and stained with Coomassie blue. Five gel slices (at about 30, 50, 150, and 300 kDa, and the top of the separating gel) were removed from the lanes for each of the eight OP samples and from untreated control lanes. These gel slices were subjected to in-gel trypsin digestion. MSMS fragmentation spectra of the tryptic peptides were examined for isopeptide cross-links. Sixteen spectra yielded convincing evidence for isopeptide cross-linked peptides. Ten were from the chlorpyrifos oxon reaction, 1 from dichlorvos, 1 from paraoxon, 1 from diazinon, and 3 from diazoxon. It was concluded that catalysis of protein cross-linking is a general property of organophosphorus pesticides and pesticide metabolites. Data are available via ProteomeXchange with identifier PXD034529.
Topics: Acetylcholinesterase; Aspartic Acid; Chlorpyrifos; Diazinon; Dichlorvos; Glutamates; Lysine; Monocrotophos; Organophosphorus Compounds; Paraoxon; Peptides; Pesticides; Sodium Dodecyl Sulfate; Trypsin; Tubulin; Water
PubMed: 36048166
DOI: 10.1021/acs.chemrestox.2c00194 -
International Journal of Molecular... Nov 2021Organophosphorus hydrolase (OPH) is a metalloenzyme that can hydrolyze organophosphorus agents resulting in products that are generally of reduced toxicity. The best OPH...
Organophosphorus hydrolase (OPH) is a metalloenzyme that can hydrolyze organophosphorus agents resulting in products that are generally of reduced toxicity. The best OPH substrate found to date is diethyl p-nitrophenyl phosphate (paraoxon). Most structural and kinetic studies assume that the binding orientation of paraoxon is identical to that of diethyl 4-methylbenzylphosphonate, which is the only substrate analog co-crystallized with OPH. In the current work, we used a combined docking and molecular dynamics (MD) approach to predict the likely binding mode of paraoxon. Then, we used the predicted binding mode to run MD simulations on the wild type (WT) OPH complexed with paraoxon, and OPH mutants complexed with paraoxon. Additionally, we identified three hot-spot residues (D253, H254, and I255) involved in the stability of the OPH active site. We then experimentally assayed single and double mutants involving these residues for paraoxon binding affinity. The binding free energy calculations and the experimental kinetics of the reactions between each OPH mutant and paraoxon show that mutated forms D253E, D253E-H254R, and D253E-I255G exhibit enhanced substrate binding affinity over WT OPH. Interestingly, our experimental results show that the substrate binding affinity of the double mutant D253E-H254R increased by 19-fold compared to WT OPH.
Topics: Aryldialkylphosphatase; Catalytic Domain; Crystallography, X-Ray; Models, Molecular; Molecular Dynamics Simulation; Molecular Structure; Mutation; Paraoxon; Protein Conformation
PubMed: 34884430
DOI: 10.3390/ijms222312624 -
Journal of the American Chemical Society Mar 2021Melanin is a ubiquitous natural pigment found in a diverse array of organisms. Allomelanin is a class of nitrogen-free melanin often found in fungi. Herein, we find...
Melanin is a ubiquitous natural pigment found in a diverse array of organisms. Allomelanin is a class of nitrogen-free melanin often found in fungi. Herein, we find artificial allomelanin analogues exhibit high intrinsic microporosity and describe an approach for further increasing and tuning that porosity. Notably, the synthetic method involves an oxidative polymerization of 1,8-DHN in water, negating the need for multiple complex templating steps and avoiding expensive or complex chemical precursors. The well-defined morphologies of these nanomaterials were elucidated by a combination of electron microscopy and scattering methods, yielding to high-resolution 3D reconstruction based on small-angle X-ray scattering (SAXS) results. Synthetic allomelanin nanoparticles exhibit high BET areas, up to 860 m/g, and are capable of ammonia capture up to 17.0 mmol/g at 1 bar. In addition, these nanomaterials can adsorb nerve agent simulants in solution and as a coating on fabrics with high breathability where they prevent breakthrough. We also confirmed that naturally derived fungal melanin can adsorb nerve gas simulants in solution efficiently despite lower porosity than synthetic analogues. Our approach inspires further analysis of yet to be discovered biological materials of this class where melanins with intrinsic microporosity may be linked to evolutionary advantages in relevant organisms and may in turn inspire the design of new high surface area materials.
Topics: Adsorption; Biopolymers; Fungi; Melanins; Nanoparticles; Naphthols; Paraoxon; Porosity; Scattering, Small Angle; X-Ray Diffraction
PubMed: 33673734
DOI: 10.1021/jacs.1c00748 -
BMC Veterinary Research Mar 2014Paraoxonase 1 (PON1) is a widely studied enzyme based on its protective role against poisoning by organophosphate (OP) metabolites of specific OP insecticides and in... (Review)
Review
Paraoxonase 1 (PON1) is a widely studied enzyme based on its protective role against poisoning by organophosphate (OP) metabolites of specific OP insecticides and in vascular disease, as well as its use as biomarker of diseases involving oxidative stress, inflammation and liver diseases.This review provides an update about the current knowledge in the field of the analytical procedures that are used for PON1 measurements. It will be specially focused on: (a) characteristics of the different substrates used for measuring PON1, with emphasis in four aspects: toxicity, polymorphism influence, rate of hydrolysis and diagnostic performance. And (b) the technical aspects of PON1 assays, in which the reagents and reaction conditions, sources of variation, quality control systems, equipment and interferences with other esterases will be discussed.The information provided in this review can contribute to a more accurate and safe measurements of PON1 in laboratories and encourage researchers to explore the wide areas of PON1 in veterinary medicine that are still unknown.
Topics: Animals; Aryldialkylphosphatase; Humans; Insecticides; Organophosphate Poisoning; Paraoxon; Polymorphism, Genetic
PubMed: 24666514
DOI: 10.1186/1746-6148-10-74 -
PloS One 2016Organophosphates (OPs) are potentially able to affect serine proteases by reacting with their active site. The potential effects of OPs on coagulation factors such as...
Organophosphates (OPs) are potentially able to affect serine proteases by reacting with their active site. The potential effects of OPs on coagulation factors such as thrombin and on coagulation tests have been only partially characterized and potential interactions with OPs antidotes such as oximes and muscarinic blockers have not been addressed. In the current study, we investigated the in vitro interactions between coagulation, thrombin, the OP paraoxon, and its antidotes obidoxime and atropine. The effects of these substances on thrombin activity were measured in a fluorescent substrate and on coagulation by standard tests. Both paraoxon and obidoxime but not atropine significantly inhibited thrombin activity, and prolonged prothrombin time, thrombin time, and partial thromboplastin time. When paraoxon and obidoxime were combined, a significant synergistic effect was found on both thrombin activity and coagulation tests. In conclusion, paraoxon and obidoxime affect thrombin activity and consequently alter the function of the coagulation system. Similar interactions may be clinically relevant for coagulation pathways in the blood and possibly in the brain.
PubMed: 27689805
DOI: 10.1371/journal.pone.0163787 -
Biomedical Papers of the Medical... Sep 2011Cholinesterases are a group of serine hydrolases that split the neurotransmitter acetylcholine (ACh) and terminate its action. Of the two types, butyrylcholinesterase... (Review)
Review
BACKGROUND
Cholinesterases are a group of serine hydrolases that split the neurotransmitter acetylcholine (ACh) and terminate its action. Of the two types, butyrylcholinesterase and acetylcholinesterase (AChE), AChE plays the key role in ending cholinergic neurotransmission. Cholinesterase inhibitors are substances, either natural or man-made that interfere with the break-down of ACh and prolong its action. Hence their relevance to toxicology and pharmacology.
METHODS AND RESULTS
The present review summarizes current knowledge of the cholinesterases and their inhibition. Particular attention is paid to the toxicology and pharmacology of cholinesterase-related inhibitors such as nerve agents (e.g. sarin, soman, tabun, VX), pesticides (e.g. paraoxon, parathion, malathion, malaoxon, carbofuran), selected plants and fungal secondary metabolites (e.g. aflatoxins), drugs for Alzheimer's disease (e.g. huperzine, metrifonate, tacrine, donepezil) and Myasthenia gravis (e.g. pyridostigmine) treatment and other compounds (propidium, ethidium, decamethonium).
CONCLUSIONS
The crucial role of the cholinesterases in neural transmission makes them a primary target of a large number of cholinesterase-inhibiting drugs and toxins. In pharmacology, this has relevance to the treatment of neurodegenerative disorders.
Topics: Acetylcholinesterase; Alzheimer Disease; Animals; Butyrylcholinesterase; Chemical Warfare Agents; Cholinesterase Inhibitors; Cholinesterases; Humans; Synaptic Transmission
PubMed: 22286807
DOI: 10.5507/bp.2011.036 -
Cognitive and Cellular Effects of Combined Organophosphate Toxicity and Mild Traumatic Brain Injury.Biomedicines May 2023Traumatic brain injury (TBI) is considered the most common neurological disorder among people under the age of 50. In modern combat zones, a combination of TBI and...
Traumatic brain injury (TBI) is considered the most common neurological disorder among people under the age of 50. In modern combat zones, a combination of TBI and organophosphates (OP) can cause both fatal and long-term effects on the brain. We utilized a mouse closed-head TBI model induced by a weight drop device, along with OP exposure to paraoxon. Spatial and visual memory as well as neuron loss and reactive astrocytosis were measured 30 days after exposure to mild TBI (mTBI) and/or paraoxon. Molecular and cellular changes were assessed in the temporal cortex and hippocampus. Cognitive and behavioral deficits were most pronounced in animals that received a combination of paraoxon exposure and mTBI, suggesting an additive effect of the insults. Neuron survival was reduced in proximity to the injury site after exposure to paraoxon with or without mTBI, whereas in the dentate gyrus hilus, cell survival was only reduced in mice exposed to paraoxon prior to sustaining a mTBI. Neuroinflammation was increased in the dentate gyrus in all groups exposed to mTBI and/or to paraoxon. Astrocyte morphology was significantly changed in mice exposed to paraoxon prior to sustaining an mTBI. These results provide further support for assumptions concerning the effects of OP exposure following the Gulf War. This study reveals additional insights into the potentially additive effects of OP exposure and mTBI, which may result in more severe brain damage on the modern battlefield.
PubMed: 37239152
DOI: 10.3390/biomedicines11051481