FIGURE 1 - uploaded by Galina F. Makhaeva
Content may be subject to copyright.
Dose-related NTE inhibition in brain, lymphocytes, and whole blood of hens 24 h after injection of the neuropathic OP compound, O,O-di-1-propyl O-2,2-dichlorvinyl phosphate (PrDChVP). Results are percent of control values for each tissue expressed as mean ± SEM, n = 3. Open square, dashed line = brain NTE, dosing experiment I (NTE assayed colorimetrically in brain and lymphocytes; both tissues assayed fresh). Open circle, dashed line = lymphocyte NTE, experiment I. Closed square, solid line = brain NTE, dosing experiment II (NTE assayed colorimetrically in brain and amperometrically in blood; both tissues assayed after freezing and thawing). Closed circle, solid line = blood NTE, dosing experiment II. Control NTE activities, nmol/(min × mg protein), mean ± SEM, n = 3: dosing experiment I, brain = 30.9 ± 2.8, lymphocyte = 9.0 ± 1.4; dosing experiment II, brain = 16.8 ± 0.88, whole blood = 0.107 ± 0.013.
Source publication
Neuropathy target esterase (NTE) is the target protein for neuropathic organophosphorus (OP) compounds that produce OP compound-induced delayed neurotoxicity (OPIDN). Inhibition/aging of brain NTE within hours of exposure predicts the potential for development of OPIDN in susceptible animal models. Lymphocyte NTE has also found limited use as a bio...
Contexts in source publication
Context 1
... Experiment I Inhibition of NTE in hen brain and lymphocytes was studied 24 h after injecting hens with increasing doses of the neuropathic OP compound, PrDChVP. NTE inhibition was measured in brain and lymphocytes using the colorimetric assay for both tissues (Figure 1). ...
Context 2
... Experiment II Inhibition of NTE in hen brain and whole blood was studied 24 h after injecting hens with increasing doses of PrDChVP. NTE inhibition was measured in brain by the colorimetric assay and in whole blood using the amperometric biosensor assay (Figure 1). ...
Context 3
... dosing experiment II, NTE inhibition was measured in brain by the colorimetric assay and in whole blood using the amperometric biosensor assay. Data from both experiments are presented in Figure 1, showing a similar pattern and degree of dose-responsive inhibition of brain, lymphocyte, and whole blood NTE. ...
Similar publications
Inhibition of acetylcholinesterase (AChE) versus inhibition and aging of neuropathy target esterase (NTE) by organophosphorus (OP) compounds in vivo can give rise to distinct neurological consequences: acute cholinergic toxicity versus OP compound-induced delayed neurotoxicity (OPIDN). Previous work has shown that the relative potency of an OP comp...
Citations
... These compounds often inhibit plasma butyrylcholinesterase (BChE) as well, providing an additional biomarker of exposure (Strelitz et al., 2014). Neuropathic OP compounds inhibit neuropathy target esterase (NTE; formerly known as neurotoxic esterase) in neural tissue; this enzyme is also found in blood lymphocytes and platelets, thereby furnishing a means of assessing exposure (Bertoncin et al., 1985;Dudek and Richardson, 1982;Makhaeva et al., 2003). However, the lifetimes of OP compounds and their metabolites in blood or urine are relatively short, and levels of inhibition of target enzymes provide no information about the identity of the inhibitors. ...
... NTE has also been found in circulating lymphocytes and platelets (Bertoncin et al., 1985;Dudek and Richardson, 1982), where its inhibition has found limited use as a biomarker of exposure to neuropathic OP compounds. Moreover, by using an amperometric biosensor (Sigolaeva et al., 2001), the NTE assay be carried out on samples of whole blood, thus obviating the need to prepare separate fractions of lymphocytes or platelets (Makhaeva et al., 2003). ...
Organophosphorus (OP) compounds include trivalent and pentavalent phosphorus; this article covers only the more
common pentavalent phosphorus compounds. These chemicals are often thought of as insecticides or nerve agents whose acute toxicity arises from inhibition of acetylcholinesterase (AChE), but OP compounds are also widely used as flame retardants, fuel additives, lubricants, plasticizers, and pharmaceuticals. In addition to their acute effects, some anti-AChE OP compounds have been associated with the intermediate syndrome. Finally, OP compounds that inhibit and age neuropathy target esterase produce OP compound-induced delayed neurotoxicity, a distal degeneration of sensory and motor axons in peripheral nerves and spinal cord.
... Two hundred frozen fly heads were homogenized (5% v/v) in a buffer (50 mM Tris-HCl and 0,2 mM EDTA (pH 8,0)). After centrifugation (9000× g; 15 min; +4 °C; Eppendorf Centrifuge 5804R, Hamburg, Germany), the 9S homogenate supernatant was assayed by the differential inhibition colorimetric method [154] with slight modifications [155] as described in detail [156,157]. The NTE-like activity is defined as the portion of phenyl valerate (PV) hydrolyzing activity that is resistant to paraoxon (40 μM, a nonneuropathic organophosphate (OP)) but sensitive to mipafox (1 mM; a neuropathic OP). ...
Various neurodegenerative disorders are associated with human NTE/PNPLA6 dysfunction. Mechanisms of neuropathogenesis in these diseases are far from clearly elucidated. Hereditary spastic paraplegia belongs to a type of neurodegeneration associated with NTE/PNLPLA6 and is implicated in neuron death. In this study, we used Drosophila melanogaster to investigate the consequences of neuronal knockdown of swiss cheese (sws)—the evolutionarily conserved ortholog of human NTE/PNPLA6—in vivo. Adult flies with the knockdown show longevity decline, locomotor and memory deficits, severe neurodegeneration progression in the brain, reactive oxygen species level acceleration, mitochondria abnormalities and lipid droplet accumulation. Our results suggest that SWS/NTE/PNPLA6 dysfunction in neurons induces oxidative stress and lipid metabolism alterations, involving mitochondria dynamics and lipid droplet turnover in neurodegeneration pathogenesis. We propose that there is a complex mechanism in neurological diseases such as hereditary spastic paraplegia, which includes a stress reaction, engaging mitochondria, lipid droplets and endoplasmic reticulum interplay.
... The discovery and characterization of NTE activity in circulating lymphocytes and platelets (Bertoncin et al., 1985;Bleecker et al., 1983;Dudek and Richardson, 1982;Maroni and Bleecker, 1986) made it possible to use blood samples for biomonitoring exposures to neuropathic OP compounds (Lotti et al., 1983;Schwab and Richardson, 1986). With the advent of electrochemical biosensors, NTE activity could be determined in whole blood (Makhaeva et al., 2003;Sigolaeva et al., 2001), thus obviating the need to isolate leukocyte fractions prior to analysis. Moreover, biosensor assays permitted real-time detection of NTE activity and could be extended for the simultaneous assay of several esterases (Kohli et al., 2007;Sigolaeva et al., 2010). ...
Systemic inhibition of neuropathy target esterase (NTE) with certain organophosphorus (OP) compounds produces OP compound-induced delayed neurotoxicity (OPIDN), a distal degeneration of axons in the central nervous system (CNS) and peripheral nervous system (PNS), thereby providing a powerful model for studying a spectrum of neurodegenerative diseases. Axonopathies are important medical entities in their own right, but in addition, illnesses once considered primary neuronopathies are now thought to begin with axonal degeneration. These disorders include Alzheimer's disease, Parkinson's disease, and motor neuron diseases such as amyotrophic lateral sclerosis (ALS). Moreover, conditional knockout of NTE in the mouse CNS produces vacuolation and other degenerative changes in large neurons in the hippocampus, thalamus, and cerebellum, along with degeneration and swelling of axons in ascending and descending spinal cord tracts. In humans, NTE mutations cause a variety of neurodegenerative conditions resulting in a range of deficits including spastic paraplegia and blindness. Mutations in the Drosophila NTE orthologue SwissCheese (SWS) produce neurodegeneration characterized by vacuolization that can be partially rescued by expression of wild-type human NTE, suggesting a potential therapeutic approach for certain human neurological disorders. This chapter defines NTE and OPIDN, presents an overview of OP compounds, provides a rationale for NTE research, and traces the history of discovery of NTE and its relationship to OPIDN. It then briefly describes subsequent studies of NTE, including practical applications of the assay; aspects of its domain structure, subcellular localization, and tissue expression; abnormalities associated with NTE mutations, knockdown, and conventional or conditional knockout; and hypothetical models to help guide future research on elucidating the role of NTE in OPIDN.
... Therefore, part of an effective chemical defense strategy is to develop methods for detecting DN agents and distinguishing them from conventional nerve agents. This can be done by exploiting the sensitivity and selectivity afforded by appropriate biomarkers and biosensors (Kohli et al., 2007a(Kohli et al., , 2014Makhaeva et al., 2003;Malygin et al., 2003;Srivastava et al., 2010;Sigolaeva et al., 2010Sigolaeva et al., , 2013. ...
... Because of the insidious onset and permanent debilitating effects of OPIDN, it is essential to be able to predict the potential of a given OP compound to produce this disease as opposed to causing acute cholinergic toxicity. Furthermore, it is important to develop specific and stable biomarkers and biosensors of exposure to DN agents and devise countermeasures against them (Kohli et al., 2007a;Makhaeva et al., 2003;Malygin et al., 2003;Sigolaeva et al., 2010Sigolaeva et al., , 2013. To accomplish these goals, it will be necessary to acquire a level of mechanistic understanding of interactions between DN compounds and target macromolecules akin to what has been achieved for conventional (anti-AChE) nerve agents. ...
Delayed neuropathic (DN) agents are defined here as organophosphorus (OP) compounds that produce OP compound–induced delayed neurotoxicity (OPIDN). This syndrome involves distal sensorimotor degeneration of long large-diameter axons in peripheral nerves and spinal cord with associated sensory deficits and paralysis typically occurring within 1–4 weeks after exposure. Mechanistic studies indicate that inhibition and aging of >70% of neuropathy target esterase (NTE) in neural tissue correlates with development of OPIDN and that acetylcholinesterase (AChE) is not involved. The neuropathic potential of an OP compound can be assessed by its relative inhibitory potency (RIP) against AChE versus NTE. By the convention used in this chapter, a high RIP value indicates that the compound will produce cholinergic toxicity rather than OPIDN. Lymphocyte and platelet NTE can be employed as biomarkers of exposure to DN compounds, and human recombinant NTE esterase domain has been incorporated into a biosensor for these agents.
... Inhibitory potency of progressive (time-dependent) inhibition of an OPC against an EOH is best assessed kinetically by the bimolecular rate constant of inhibition, k i . Alternatively, potency can be expressed as the less-preferred fixed-time IC 50 (inhibitor concentration required to inhibit 50% of the enzyme's activity after a fixed time of incubation (t) of the OPC with the EOH). When pseudo-first-order kinetics of inhibition are obtained, it is valid to calculate the IC 50 from the k i via the equation, IC 50 z 0.693/(k i  t). ...
... These selectivities consist of ratios of inhibitory potencies of a given OPC against a pair of EOHs. For example, as defined here, k i (NTE)/k i (AChE) or IC 50 (AChE)/ IC 50 (NTE) represents the relative inhibitory potency (RIP) of an OPC against targets for delayed neurotoxicity (NTE) and acute neurotoxicity (AChE). This ratio correlates with that between the LD 50 and the neuropathic dose, thereby serving as a convenient index of the neuropathic potential of an OPC that is subject to undergoing the aging reaction [6,17e20]. ...
... Brain NTE activity was determined in 9S supernatants by the differential inhibition colorimetric method of Johnson [48] with slight modifications [49] as described in detail [31,50]. The endpoint absorbance was measured at 486 nm (ε 486 ¼ 16470 M À1 cm À1 ) using a Bio-Rad Benchmark Plus microplate reader (Marnes-la-Coquette, France). ...
... Therefore, part of an effective chemical defense strategy is to develop methods for detecting DN agents and distinguishing them from conventional nerve agents. This can be performed by exploiting the sensitivity and selectivity afforded by appropriate biomarkers and biosensors (Makhaeva et al., 2003;Malygin et al., 2003;Kohli et al., 2007aKohli et al., , 2010Kohli et al., , 2014Sigolaeva et al., 2010Sigolaeva et al., , 2013Srivastava et al., 2010). ...
... Because of the insidious onset and permanent debilitating effects of OPIDN, it is essential to be able to predict the potential of a given OP compound to produce this disease as opposed to causing acute cholinergic toxicity. Furthermore, it is important to develop specific and stable biomarkers and biosensors of exposure to DN agents and devise countermeasures against them (Makhaeva et al., 2003;Malygin et al., 2003;Kohli et al., 2007a;Sigolaeva et al., 2010Sigolaeva et al., , 2013). To accomplish these goals, it will be necessary to acquire a level of mechanistic understanding of interactions between DN compounds and target macromolecules akin to what has been achieved for conventional (anti-AChE) nerve agents. ...
... However, experiments with conventional NTE knockouts (Winrow et al., 2003;Moser et al., 2004;Mühlig-Versen et al., 2005) and conditional NTE knockouts (Akassoglou et al., 2004), along with demonstrations of a likely role of NTE in membrane lipid metabolism (vanTienhoven et al., 2002;Quistad et al., 2003;Zaccheo et al., 2004), indicate that lethality or neuropathology can result from a loss of function of NTE (Glynn, 2006). Nevertheless, while the physiological and pathogenic roles of NTE are being deciphered, the fact that an excellent correlation exists between inhibition/ aging of NTE and induction of OPIDN is sufficient to enable the pragmatic use this information for the development of biomarkers and biosensors for DN agents (Makhaeva et al., 2003Makhaeva et al., , 2007). ...
Delayed neuropathic (DN) agents are defined here as organophosphorus (OP) compounds that produce OP compound–induced delayed neurotoxicity (OPIDN). This syndrome involves distal sensorimotor degeneration of long, large-diameter axons in peripheral nerves and spinal cord with associated sensory deficits and paralysis typically occurring within 1–4 weeks after exposure. Mechanistic studies indicate that inhibition and aging of more than 70% of neuropathy target esterase (NTE) in neural tissue correlates with development of OPIDN and that acetylcholinesterase (AChE) is not involved. The neuropathic potential of an OP compound can be assessed by its relative inhibitory potency (RIP) against AChE versus NTE. By the convention used in this chapter, a high RIP value indicates that the compound will produce cholinergic toxicity rather than OPIDN. Lymphocyte and platelet NTE can be used as biomarkers of exposure to DN compounds, and human recombinant NTE esterase domain (NEST) has been incorporated into a biosensor for these agents.
... Thus, CPS served as an OP control for CPO in our in vitro experiments. CPO is a predominantly cholinergic OP compound, whereas DFP, MIP and PrDChVP are intermediate between cholinergic and neuropathic, but each capable of producing OPIDN (Kropp and Richardson, 2003;Makhaeva et al., 1995Makhaeva et al., , 2003Makhaeva et al., , 2009b. PrDChVP was intensively studied in our previous work in experiments in vitro and in vivo using both hens and rats (Makhaeva et al., 1995(Makhaeva et al., , 2009b. ...
... The fixed-time IC 50 values calculated for the hen brain enzymes in the present work are consistent with previously reported values by either falling within the range or being within 1 standard error for MIP (Kropp and Richardson, 2003;Richardson et al., 1993), DFP (Atkins and Glynn, 2000;Lotti and Johnson, 1978); CPO (Atkins and Glynn, 2000;Correll and Ehrich, 1987;Kropp and Richardson, 2003;Lotti and Johnson, 1978) and PrDChVP (Lotti and Johnson, 1978;Makhaeva et al., 1995Makhaeva et al., , 2003. ...
... Moreover, we found that the RIP of DFP is slightly less than 1, indicating neuropathicity at doses near the LD 50 , and that the RIP for MIP and PrDChVP is slightly higher than 1, which signifies that these compounds are neuropathic. These results correspond with previously published data (Kropp and Richardson, 2003;Lotti and Johnson, 1978;Makhaeva et al., 2003). Furthermore, earlier studies of other OP compounds in vitro have shown similar correlations of neuropathic potential with RIPs determined from human and hen brain tissues as well as neuroblastoma cell lines of both human and murine origin (Ehrich et al., 1997;Lotti and Johnson, 1978). ...
Inhibition and aging of neuropathy target esterase (NTE) by neuropathic organophosphorus (OP) compounds triggers OP compound-induced delayed neuropathy (OPIDN), whereas inhibition of acetylcholinesterase (AChE) produces cholinergic toxicity. The neuropathic potential of an OP compound is defined by its relative inhibitory potency toward NTE vs. AChE assessed by enzyme assays following dosing in vivo or after incubations of direct-acting compounds or active metabolites with enzymes in vitro. The standard animal model of OPIDN is the adult hen, but its large size and high husbandry costs make this species a burdensome model for assessing neuropathic potential. Although the mouse does not readily exhibit clinical signs of OPIDN, it displays axonal lesions and expresses brain AChE and NTE. Therefore, the present research was performed as a further test of the hypothesis that inhibition of mouse brain AChE and NTE could be used to assess neuropathic potential using mouse brain preparations in vitro or employing mouse brain assays following dosing of OP compounds in vivo. Excellent correlations were obtained for inhibition kinetics in vitro of mouse brain enzymes vs. hen brain and human recombinant enzymes. Furthermore, inhibition of mouse brain AChE and NTE after dosing with OP compounds afforded ED50 ratios that agreed with relative inhibitory potencies assessed in vitro. Taken together, results with mouse brain enzymes demonstrated consistent correspondence between in vitro and in vivo predictors of neuropathic potential, thus adding to previous studies supporting the validity of a mouse model for biochemical assessment of the ability of OP compounds to produce OPIDN. Copyright © 2014 John Wiley & Sons, Ltd.
... PrDChVP is a known delayed neurotoxicant which was shown to induce ataxia in hens. It has rather high neuropathic potential: its relative inhibitor potencyRIP = IC 50 (AChE)/ IC 50 ( NTE) = 2.6 [13], and high acute toxicity: LD 50 = 10 mg/kg, hens [14].PrDChVP was intensively studied in our previous experiments in vitro and in vivo in hens and rats [9,15,16,17]. This OPC was used as a known neuropathic OPC to develop a mouse model for biochemical assessment of neuropathic potential [9]. ...
... NTE activity was assayed according to the differential inhibition method of Johnson [22] with an electrochemical endpoint as described previously [16,23]. A new sensitive, stable and reproducible planar tyrosinase biosensor was used for phenol detection. ...
... NTE) is a specifi c target and biomarker of OP compound-induced delayed neuropathy (OPIDN). A good correlation between the inhibition/aging of brain/lymphocyte/ blood NTE within hours of exposure and the subsequent induction of OPIDN [7,[12][13][14][15] enables application of blood NTE activity determination for earlier diagnosis of OPIDN. However, because of the low level of NTE activity in blood and interference from hemoglobin absorbance, the colorimetric assay cannot be used to determine NTE activity in whole blood [16]. ...
A tyrosinase-based phenol biosensors were formed via layer-by-layer (LBL) approach to deposit poly(dimethyldiallylammonium chloride) (PDDA) and tyrosinase onto graphite-based substrates, followed by optional glutaraldehyde (GA) crosslinking. The good analytical parameters of phenol detection thogether with low detection limit enabled phenol measurements in highly diluted blood samples, which also minimized interference from extraneous (non-analyte) substances in blood (e.g., adrenaline, ascorbate, glucose, L-tyrosine, and urate). Applicability of the biosensor to analysis of carboxylesterase (CaE), paraofonase 1 (PON1) and neuropathy target esterase (NTE) activities in mouse and human blood was demonstrated. Parallel biosensor and spectrophotometric CaE analyses were carried out 1 h after intraperitoneal injection of mice with the model dialkylphosphate, (C2H5O)2P(O)OCH(CF3)2 (DEHFPP). Dose-related inhibition of CaE activity was observed, yielding ED50 values [mean (95% CI) (n)] of 25.5 (23.2, 28.0) mg/kg (6) and 21.1 mg/kg (18.9, 23.5) (4) for spectrophotometric and electrochemical assays, respectively. Although the ED50 values were significantly different from each other (p < 0.007), excellent correlation between biosensor and spectrophotometric measurements was found (r = 0.99), indicating that the differences between the two methods are systematic and providing validation of the biosensor assay.
... The discovery of NTE in circulating lymphocytes and platelets [13][14][15][16] enabled it to be used as a biomarker of animal and human exposure to neuropathic OPCs [16][17][18][19][20][21]. The development by our team an electrochemical method for NTE assay using tyrosinase-based biosensors enabled measuring NTE activity in whole blood [22][23][24][25] [23]. These studies indicate that NTE in whole blood can be assayed and used as a biomarker of exposure to delayed neuropathic OPCs. ...
... The discovery of NTE in circulating lymphocytes and platelets [13][14][15][16] enabled it to be used as a biomarker of animal and human exposure to neuropathic OPCs [16][17][18][19][20][21]. The development by our team an electrochemical method for NTE assay using tyrosinase-based biosensors enabled measuring NTE activity in whole blood [22][23][24][25] [23]. These studies indicate that NTE in whole blood can be assayed and used as a biomarker of exposure to delayed neuropathic OPCs. ...
... Blood NTE assay NTE activity was assayed according to the differential inhibition method of Johnson [47] with an electrochemical endpoint as described previously [22,23]. A new sensitive, stable and reproducible planar tyrosinase biosensor of construction SPE/PDDA/Tyr was used for phenol detection. ...
Development of biomarkers of human exposures to OPCs and their quantifi cation is a vital component of a system of prediction and early diagnosis of induced diseases. Our study was focused on investigation of esterase status as a complex biomarker of exposure to OPCs and an aid in accurate diagnosis. We suggest that this complex biomarker should be more effective and informative than standard assays of plasma BChE, RBC AChE, and lymphocyte neuropathy target esterase ( NTE). It will allow us: 1) to assess an exposure as such and to confi rm the nonexposure of individuals suspected to have been exposed; 2) to determine if the exposure was to
agents expected to produce acute and/or delayed neurotoxicity; 3) to perform dosimetry of the exposure, which provides valuable information for medical treatment. To confirm this hypothesis, we examined the changes in activity of blood AChE, NTE, BChE and carboxylesterase ( CaE) 1 h after i.p. administration of increasing doses of three OPCs with different esterase profi les: (C2H5O)2P(O)OCH(CF3)2, (C4H9O)2P(O)OCH(CF3)2 and (C3H7O)2P(O)OCH=CCl2. The esterases assay was performed in hemolysed blood by biosensor and spectrophotometric methods. Analysis of the obtained dose-dependences for blood esterases inhibition showed that blood BChE and CaE are the most sensitive biomarkers, allowing detection of low doses. Inhibition of blood NTE and AChE can be used to assess the likelihood that an exposure to OPC would produce cholinergic and/or delayed neuropathic effects. Thus, determination of esterase status allows one to improve the possibilities of diagnostics of exposure to OPCs.
