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International Journal of Molecular... Dec 2021The delayed effects of acute intoxication by organophosphates (OPs) are poorly understood, and the various experimental animal models often do not take into account...
The delayed effects of acute intoxication by organophosphates (OPs) are poorly understood, and the various experimental animal models often do not take into account species characteristics. The principal biochemical feature of rodents is the presence of carboxylesterase in blood plasma, which is a target for OPs and can greatly distort their specific effects. The present study was designed to investigate the nephrotoxic effects of paraoxon (O,O-diethyl O-(4-nitrophenyl) phosphate, POX) using three models of acute poisoning in outbred Wistar rats. In the first model (, POX2x group), POX was administered twice at doses 110 µg/kg and 130 µg/kg subcutaneously, with an interval of 1 h. In the second model (, CBPOX group), 1 h prior to POX poisoning at a dose of 130 µg/kg subcutaneously, carboxylesterase activity was pre-inhibited by administration of specific inhibitor cresylbenzodioxaphosphorin oxide (CBDP, 3.3 mg/kg intraperitoneally). In the third model (), POX was administered subcutaneously just once at doses of LD16 (241 µg/kg), LD50 (250 µg/kg), and LD84 (259 µg/kg). Animal observation and sampling were performed 1, 3, and 7 days after the exposure. Endogenous creatinine clearance (ECC) decreased in 24 h in the POX2x group ( = 0.011). Glucosuria was observed in rats 24 h after exposure to POX in both M1 and M2 models. After 3 days, an increase in urinary excretion of chondroitin sulfate (CS, = 0.024) and calbindin ( = 0.006) was observed in rats of the CBPOX group. Morphometric analysis revealed a number of differences most significant for rats in the CBPOX group. Furthermore, there was an increase in the area of the renal corpuscles ( = 0.0006), an increase in the diameter of the lumen of the proximal convoluted tubules (PCT, = 0.0006), and narrowing of the diameter of the distal tubules ( = 0.001). After 7 days, the diameter of the PCT lumen was still increased in the nephrons of the CBPOX group ( = 0.0009). In the model, histopathological and ultrastructural changes in the kidneys were revealed after the exposure to POX at doses of LD50 and LD84. Over a period from 24 h to 3 days, a significant ( = 0.018) expansion of Bowman's capsule was observed in the kidneys of rats of both the LD50 and LD84 groups. In the epithelium of the proximal tubules, stretching of the basal labyrinth, pycnotic nuclei, and desquamation of microvilli on the apical surface were revealed. In the epithelium of the distal tubules, partial swelling and destruction of mitochondria and pycnotic nuclei was observed, and nuclei were displaced towards the apical surface of cells. After 7 days of the exposure to POX, an increase in the thickness of the glomerular basement membrane (GBM) was observed in the LD50 and LD84 groups ( = 0.019 and 0.026, respectively). Moreover, signs of damage to tubular epithelial cells persisted with blockage of the tubule lumen by cellular detritus and local destruction of the surface of apical cells. Comparison of results from the three models demonstrates that the nephrotoxic effects of POX, evaluated at 1 and 3 days, appear regardless of prior inhibition of carboxylesterase activity.
Topics: Animals; Biomarkers; Bowman Capsule; Creatinine; Kidney; Kidney Tubules, Proximal; Male; Nephrons; Paraoxon; Rats; Rats, Wistar
PubMed: 34948422
DOI: 10.3390/ijms222413625 -
Scientific Reports Nov 2023Organophosphorus poisoning kills individuals by causing central apnea; however, the underlying cause of death remains unclear. Following findings that the pre-Bötzinger...
Organophosphorus poisoning kills individuals by causing central apnea; however, the underlying cause of death remains unclear. Following findings that the pre-Bötzinger complex impairment alone does not account for central apnea, we analyzed the effect of paraoxon on the brainstem-spinal cord preparation, spanning the lower medulla oblongata to phrenic nucleus. Respiratory bursts were recorded by connecting electrodes to the ventral 4th cervical nerve root of excised brainstem-spinal cord preparations obtained from 6-day-old Sprague-Dawley rats. We observed changes in respiratory bursts when paraoxon, neostigmine, atropine, and 2-pyridine aldoxime methiodide were administered via bath application. The percentage of burst extinction in the paraoxon-poisoning group was 50% compared with 0% and 18.2% in the atropine and 2-pyridine aldoxime methiodide treatment groups, respectively. Both treatments notably mitigated the paraoxon-induced reduction in respiratory bursts. In the neostigmine group, similar to paraoxon, bursts stopped in 66.7% of cases but were fully reversed by atropine. This indicates that the primary cause of central apnea is muscarinic receptor-mediated in response to acetylcholine excess. Paraoxon-induced central apnea is hypothesized to result from neural abnormalities within the inferior medulla oblongata to the phrenic nucleus, excluding pre-Bötzinger complex. These antidotes antagonize central apnea, suggesting that they may be beneficial therapeutic agents.
Topics: Rats; Animals; Antidotes; Paraoxon; Rats, Sprague-Dawley; Neostigmine; Sleep Apnea, Central; Atropine; Pralidoxime Compounds; Pyridines
PubMed: 37990100
DOI: 10.1038/s41598-023-47745-x -
Journal of Lipid Research Oct 2014Hydrolysis of intracellular cholesteryl ester (CE) is the rate-limiting step in the efflux of cholesterol from macrophage foam cells. In mouse peritoneal macrophages...
Hydrolysis of intracellular cholesteryl ester (CE) is the rate-limiting step in the efflux of cholesterol from macrophage foam cells. In mouse peritoneal macrophages (MPMs), this process is thought to involve several enzymes: hormone-sensitive lipase (Lipe), carboxylesterase 3 (Ces3), neutral CE hydrolase 1 (Nceh1). However, there is some disagreement over the relative contributions of these enzymes. To solve this problem, we first compared the abilities of several compounds to inhibit the hydrolysis of CE in cells overexpressing Lipe, Ces3, or Nceh1. Cells overexpressing Ces3 had negligible neutral CE hydrolase activity. We next examined the effects of these inhibitors on the hydrolysis of CE and subsequent cholesterol trafficking in MPMs. CE accumulation was increased by a selective inhibitor of Nceh1, paraoxon, and two nonselective inhibitors of Nceh1, (+)-AS115 and (-)-AS115, but not by two Lipe-selective inhibitors, orlistat and 76-0079. Paraoxon inhibited cholesterol efflux to apoA-I or HDL, while 76-0079 did not. These results suggest that Nceh1 plays a dominant role over Lipe in the hydrolysis of CE and subsequent cholesterol efflux in MPMs.
Topics: Animals; Biological Transport, Active; Carboxylesterase; Cholesterol Esters; Enzyme Inhibitors; HEK293 Cells; Humans; Hydrolysis; Macrophages, Peritoneal; Mice; Mice, Knockout; Sterol Esterase
PubMed: 24868095
DOI: 10.1194/jlr.M047787 -
The Analyst Mar 2016We report the first use of a paper-based device coated with nanoceria as a simple, low-cost and rapid detection platform for the analysis of organophosphate (OP)...
We report the first use of a paper-based device coated with nanoceria as a simple, low-cost and rapid detection platform for the analysis of organophosphate (OP) pesticides using an enzyme inhibition assay with acetylcholinesterase (AChE) and choline oxidase (ChOX). In the presence of acetylcholine, AChE and ChOX catalyze the formation of H2O2, which is detected colorimetrically by a nanoceria-coated device resulting in the formation of a yellow color. After incubation with OP pesticides, the AChE activity was inhibited, producing less H2O2, and a reduction in the yellow intensity. The assay is able to analyze OP pesticides without the use of sophisticated instruments and gives detection limits of 18 ng mL(-1) and 5.3 ng mL(-1) for methyl-paraoxon and chlorpyrifos-oxon, respectively. The developed method was successfully applied to detect methyl-paraoxon in spiked vegetables (cabbage) and a dried seafood product (dried green mussel), obtaining ∼95% recovery values for both sample types. The spiked samples were also analyzed using LC-MS/MS as a comparison to the developed method and similar values were obtained, indicating that the developed method gives accurate results and is suitable for OP analysis in real samples.
Topics: Alcohol Oxidoreductases; Animals; Biosensing Techniques; Cerium; Cholinesterase Inhibitors; Color; Colorimetry; Environmental Pollutants; Food Analysis; Hydrogen Peroxide; Limit of Detection; Nanoparticles; Paper; Pesticides
PubMed: 26842266
DOI: 10.1039/c5an02403j -
Neurotoxicology May 2020Organophosphates (OPs) are widely used as pesticides and have been employed as warfare agents. OPs inhibit acetylcholinesterase, leading to over-stimulation of...
Organophosphates (OPs) are widely used as pesticides and have been employed as warfare agents. OPs inhibit acetylcholinesterase, leading to over-stimulation of cholinergic synapses and can cause status epilepticus (SE). OPs poisoning can result in irreversible brain damage and death. Despite termination of SE, recurrent seizures and abnormal brain activity remain common sequelae often associated with long-term neural damage and cognitive dysfunction. Therefore, early treatment for prevention of seizures is of high interest. Using a rat model of paraoxon poisoning, we tested the efficacy of different neuroprotective and anti-epileptic drugs (AEDs) in suppressing early seizures and preventing brain damage. Electrocorticographic recordings were performed prior, during and after injection of 4.5 LD paraoxon, followed by injections of atropine and toxogonin (obidoxime) to prevent death. Thirty minutes later, rats were injected with midazolam alone or in combination with different AEDs (lorazepam, valproic acid, phenytoin) or neuroprotective drugs (losartan, isoflurane). Outcome measures included SE duration, early seizures frequency and epileptiform activity duration in the first 24 -hs after poisoning. To assess delayed brain damage, we performed T2-weighted magnetic resonance imaging one month after poisoning. SE duration and the number of recurrent seizures were not affected by the addition of any of the drugs tested. Delayed brain injury was most prominent in the septum, striatum, amygdala and piriform network. Only isoflurane anesthesia significantly reduced brain damage. We show that acute treatment with isoflurane, but not AEDs, reduces brain damage following SE. This may offer a new therapeutic approach for exposed individuals.
Topics: Animals; Anticonvulsants; Brain; Disease Models, Animal; Isoflurane; Male; Midazolam; Paraoxon; Rats, Sprague-Dawley; Status Epilepticus
PubMed: 32084435
DOI: 10.1016/j.neuro.2020.02.007 -
Neurotoxicity Research Feb 2020Paraoxon is the bioactive metabolite of organophosphate (OP) pesticide, parathion. This study aimed to evaluate the expression of apoptosis-related genes and...
Paraoxon is the bioactive metabolite of organophosphate (OP) pesticide, parathion. This study aimed to evaluate the expression of apoptosis-related genes and histopathological changes in rat prefrontal cortex following exposure to three different doses of paraoxon. Paraoxon (0.3, 0.7, or 1 mg/kg) or corn oil (vehicle) were intraperitoneally injected to adult male Wistar rats. After 14 or 28 days, mRNA and protein levels of Bax, Bcl-2, and caspase-3 were measured in prefrontal cortex using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) and western blotting, respectively. In addition, neuronal injury and astrocyte activation were assessed using cresyl violet staining and glial fibrillary acidic protein (GFAP) immune-positive cells, respectively. Treatment with 0.7 and 1 mg/kg of paraoxon increased mRNA and protein levels of Bax and caspase-3 at 14 and 28 days post-exposure, while mRNA and protein levels of Bcl-2 decreased only in 1 mg/kg group after 14 days. Furthermore, a significant decrease in the number of neurons and a significant increase in the number of GFAP-positive cells were observed in rats receiving 0.7 and 1 mg/kg of paraoxon at both time points. Collectively, our results showed that apoptosis is a major mechanism for neuronal damage after exposure to paraoxon. Also, paraoxon-induced neuronal loss was correlated with activation of astrocytes. Since paraoxon-induced neuronal damage is closely related to convulsion, clinical management of convulsion could protect neuronal brain damage.
Topics: Animals; Apoptosis; Astrocytes; Caspase 3; Cell Survival; Cholinesterase Inhibitors; Gene Expression; Male; Neurons; Paraoxon; Prefrontal Cortex; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Wistar; bcl-2-Associated X Protein
PubMed: 31493121
DOI: 10.1007/s12640-019-00106-x -
Chemosphere Apr 2022The aim of this work is to fabricate a sensitive and novel enzymeless electrochemical sensor for the simultaneous determination of parathion and paraoxon using the...
The aim of this work is to fabricate a sensitive and novel enzymeless electrochemical sensor for the simultaneous determination of parathion and paraoxon using the Nd-UiO-66@MWCNT nanocomposite. For this purpose, Neodymium (Nd) was introduced into a Universitetet i Oslo (UiO-66) structure to construct Nd-UiO-66 and then, adding multi-walled carbon nanotubes to the Nd-UiO-66 to increase the electrocatalytic activity and surface area of the obtained composite. The Nd-UiO-66@MWCNT has numerous advantages like excellent conductivity, tunable texture, and large surface area and can be used as a distinctive structure for the construction of modified glassy carbon electrode (GCE) to enhance the charge-transfer and the efficiency of electrochemical sensors. This modified electrode showed sensitive and selective determination of paraoxon and parathion over the linear ranges of 0.7-100 and 1-120 nM, with detection limits of 0.04 and 0.07 nM, respectively. The proposed Nd-UiO-66@MWCNT/GCE sensor in this study can be applied in environmental and toxicological laboratories and field tests to detect parathion and paraoxon levels.
Topics: Electrochemical Techniques; Electrodes; Metal-Organic Frameworks; Nanotubes, Carbon; Neodymium; Paraoxon; Parathion; Phthalic Acids
PubMed: 34973245
DOI: 10.1016/j.chemosphere.2021.133440 -
Chemico-biological Interactions Aug 2019Carbamates are esters of substituted carbamic acids that react with acetylcholinesterase (AChE) by initially transferring the carbamoyl group to a serine residue in the... (Review)
Review
Carbamates are esters of substituted carbamic acids that react with acetylcholinesterase (AChE) by initially transferring the carbamoyl group to a serine residue in the enzyme active site accompanied by loss of the carbamate leaving group followed by hydrolysis of the carbamoyl enzyme. This hydrolysis, or decarbamoylation, is relatively slow, and half-lives of carbamoylated AChEs range from 4 min to more than 30 days. Therefore, carbamates are effective AChE inhibitors that have been developed as insecticides and as therapeutic agents. In this report, we review recent data showing that decarbamoylation rate constants are independent of the ester leaving group for a series of carbamic acid esters with the same carbamoyl group and that decarbamoylation rate constants decreased by 800-fold when the alkyl substituents on the carbamoyl group increased in size from N-monomethyl- to N,N-diethyl-. We also review data showing that solvent deuterium oxide isotope effects for decarbamoylation decreased from 2.8 for N-monomethylcarbamoyl AChE to 1.1 for N,N-diethylcarbamoyl AChE, indicating a shift in the rate-limiting step from general acid-base catalysis to a likely conformational change in the distorted active site in N,N-diethylcarbamoyl AChE. The nature of such a conformational change is suggested from X-ray crystal structures of AChE phosphorylated by paraoxon.
Topics: Acetylcholinesterase; Carbamates; Catalytic Domain; Crystallography, X-Ray; Kinetics; Paraoxon
PubMed: 31175846
DOI: 10.1016/j.cbi.2019.06.004 -
Toxicology Mechanisms and Methods Jan 2018Organophosphate (OP) poisoning is a major global health issue; while compounds from this group have been used intensively over the last century, an effective antidote is... (Comparative Study)
Comparative Study
Organophosphate (OP) poisoning is a major global health issue; while compounds from this group have been used intensively over the last century, an effective antidote is still lacking. Oxime-type acetylcholinesterase (AChE) reactivators are used to reactivate the OP inhibited AChE. Pralidoxime is the only US Food and Drug Administration approved oxime for therapeutic use but its efficacy has been disappointing. Two novel oximes (K378 and K727) were investigated in silico and in vitro and compared with an experimental oxime (kamiloxime; K-27) and pralidoxime. In silico the molecular interactions between AChE and oximes were examined and binding energies were assessed. LogP (predicted log of the octanol/water partition coefficient) was estimated. In vitro the intrinsic ability of the oximes to inhibit AChE (IC) and their reactivation potency (R) when used in paraoxon inhibited human RBC-AChE was determined. Molecular docking revealed that K378 and K727 bind to the peripheral site(s) with high binding energies in contrast to the central binding of K-27 and pralidoxime. LogP values indicating that the novel compounds are significantly less hydrophilic than K-27 or pralidoxime. IC of K378 and K727 were comparable (0.9 and 1 µM, respectively) but orders of magnitude lower than comparators. R values revealed their inability to reactivate paraoxon inhibited AChE. It is concluded that the novel oximes K378 and K727 are unlikely to be clinically useful. The in silico and in vitro studies described allow avoidance of unnecessary in vivo animal work and contribute to the reduction of laboratory animal use.
Topics: Acetylcholinesterase; Antidotes; Binding Sites; Cholinesterase Inhibitors; Cholinesterase Reactivators; Dose-Response Relationship, Drug; GPI-Linked Proteins; Humans; Male; Molecular Docking Simulation; Organophosphate Poisoning; Oximes; Paraoxon; Pralidoxime Compounds; Protein Binding; Protein Conformation; Pyridinium Compounds; Structure-Activity Relationship
PubMed: 28722512
DOI: 10.1080/15376516.2017.1357777 -
Mutation Research. Genetic Toxicology... 2023We tested the hypothesis that the pesticides paraoxon and glyphosate cause DNA double-strand breaks (DSB) by poisoning the enzyme Type II topoisomerase (topo II)....
We tested the hypothesis that the pesticides paraoxon and glyphosate cause DNA double-strand breaks (DSB) by poisoning the enzyme Type II topoisomerase (topo II). Peripheral lymphocytes in G0 phase, treated with the pesticides, plus or minus ICRF-187, an inhibitor of Topo II, were stimulated to proliferate; induced cytogenetic damage was measured. Micronuclei, chromatin buds, nucleoplasmic bridges, and extranuclear fragments were induced by treatments with the pesticides, irrespective of the pre-treatment with ICRF-187. These results indicate that the pesticides do not act as topo II poisons. The induction of DSB may occur by other mechanisms, such as effects on other proteins involved in recombination repair.
Topics: Dexrazoxane; Paraoxon; Topoisomerase II Inhibitors; Poisons; DNA Topoisomerases, Type II; DNA; Pesticides; Glyphosate
PubMed: 37567644
DOI: 10.1016/j.mrgentox.2023.503657