Author Report for: Moretto AContact information:Moretto AngeloInstituto di Medicina del Lavoro, Universita degli Studi di Padova, via Giustiniani 2, 35128 PadovaItalyPhone : (39-49) 8212548/8212549 Fax : (39-49) 8212550 The 2nd International Meeting on Esterases Reacting with Organophosphorus Compounds (Salsomaggiore, Italy) The 3rd International Meeting on Esterases Reacting with Organophosphorus Compounds (Dubrovnik, Croatia)
Moretto A, Nicolli A, Lotti M Ref: Toxicol Appl Pharmacol, 219:196, 2007 : PubMed ESTHER: Moretto_2007_Toxicol.Appl.Pharmacol_219_196 PubMedSearch: Moretto 2007 Toxicol.Appl.Pharmacol 219 196 PubMedID: 17207828 Abstract Certain esterase inhibitors, such as carbamates, phosphinates and sulfonyl halides, do not cause neuropathy as some organophosphates, but they may exacerbate chemical or traumatic insults to axons. This phenomenon is called promotion of axonopathies. Given the biochemical and toxicological characteristics of these compounds, the hypothesis was made that the target of promotion is a phenyl valerate (PV) esterase similar to neuropathy target esterase (NTE), the target of organophosphate induced delayed polyneuropathy. However, attempts to identify a PV esterase in hen peripheral nerve have been, so far, unsuccessful. We tested several esters, other than PV, as substrates of esterases from crude homogenate of the hen peripheral nerve. The ideal substrate should be poorly hydrolysed by NTE but extensively by enzyme(s) that are insensitive to non-promoters, such as mipafox, and sensitive to promoters, such as phenyl methane sulfonyl fluoride (PMSF). When phenyl benzoate (PB) was used as substrate, about 65% of total activity was resistant to the non-promoter mipafox (up to 0.5 mM, 20 min, pH 8.0), that inhibits NTE and other esterases. More than 90% of this resistant activity was sensitive to the classical promoter PMSF (1 mM, 20 min, pH 8.0) with an IC(50) of about 0.08 mM (20 min, pH 8.0). On the contrary, the non-promoter p-toluene sulfonyl fluoride caused only about 10% inhibition at 0.5 mM. Several esterase inhibitors including, paraoxon, phenyl benzyl carbamate, di-n-butyl dichlorovinyl phosphate and di-isopropyl fluorophosphate, were tested both in vitro and in vivo for inhibition of this PB activity. Mipafox-resistant PMSF-sensitive PB esterase activity(ies) was inhibited by promoters but not by non promoters and neuropathic compounds. Title: A tiered approach to systemic toxicity testing for agricultural chemical safety assessment Doe JE, Boobis AR, Blacker A, Dellarco V, Doerrer NG, Franklin C, Goodman JI, Kronenberg JM, Lewis R and Wolf DC <9 more author(s)> Doe JE, Boobis AR, Blacker A, Dellarco V, Doerrer NG, Franklin C, Goodman JI, Kronenberg JM, Lewis R, McConnell EE, Mercier T, Moretto A, Nolan C, Padilla S, Phang W, Solecki R, Tilbury L, van Ravenzwaay B, Wolf DC (- 9) Ref: Crit Rev Toxicol, 36:37, 2006 : PubMed ESTHER: Doe_2006_Crit.Rev.Toxicol_36_37 PubMedSearch: Doe 2006 Crit.Rev.Toxicol 36 37 PubMedID: 16708694 Abstract A proposal has been developed by the Agricultural Chemical Safety Assessment (ACSA) Technical Committee of the ILSI Health and Environmental Sciences Institute (HESI) for an improved approach to assessing the safety of crop protection chemicals. The goal is to ensure that studies are scientifically appropriate and necessary without being redundant, and that tests emphasize toxicological endpoints and exposure durations that are relevant for risk assessment. The ACSA Systemic Toxicity Task Force proposes an approach to systemic toxicity testing as one part of the overall assessment of a compound's potential to cause adverse effects on health. The approach is designed to provide more relevant data for deriving reference doses for shorter time periods of human exposure, and includes fewer studies for deriving longer term reference doses-that is, neither a 12-month dog study nor a mouse carcinogenicity study is recommended. All available data, including toxicokinetics and metabolism data and life stages information, are taken into account. The proposed tiered testing approach has the potential to provide new risk assessment information for shorter human exposure durations while reducing the number of animals used and without compromising the sensitivity of the determination of longer term reference doses. Title: Do carbamates cause polyneuropathy? Lotti M, Moretto A Ref: Muscle & Nerve, 34:499, 2006 : PubMed ESTHER: Lotti_2006_Muscle.Nerve_34_499 PubMedSearch: Lotti 2006 Muscle.Nerve 34 499 PubMedID: 16897762 Abstract Carbamates are reversible inhibitors of acetylcholinesterase, and some also inhibit neuropathy target esterase (NTE), the target in organophosphate-induced delayed polyneuropathy. However, based on mechanistic considerations, these carbamates were thought to be unable to initiate polyneuropathy. Consequently, clinical reports of polyneuropathy associated with carbamate exposures have been disregarded. We discuss three cases of polyneuropathy that occurred after severe poisoning by methylcarbamates. In addition, high repeated doses of phenyl N-methyl N-benzylcarbamate caused nearly 100% NTE inhibition and polyneuropathy in the hen model. These data suggest the need to reconsider the long-standing tenet that carbamates cannot cause polyneuropathy. Alternatively, a preexisting subclinical neuropathy in these individuals may have been amplified by carbamates, as observed in animal models. We suggest that individuals with underlying neuropathy (e.g., diabetics) who are poisoned by carbamates should be followed closely. In addition, procedures for the current risk assessment of carbamate pesticides may need to be reconsidered. Title: Organophosphate-induced delayed polyneuropathy Lotti M, Moretto A Ref: Toxicol Rev, 24:37, 2005 : PubMed ESTHER: Lotti_2005_Toxicol.Rev_24_37 PubMedSearch: Lotti 2005 Toxicol.Rev 24 37 PubMedID: 16042503 Abstract Organophosphate-induced delayed polyneuropathy (OPIDP) is a rare toxicity resulting from exposure to certain organophosphorus (OP) esters. It is characterised by distal degeneration of some axons of both the peripheral and central nervous systems occurring 1-4 weeks after single or short-term exposures. Cramping muscle pain in the lower limbs, distal numbness and paraesthesiae occur, followed by progressive weakness, depression of deep tendon reflexes in the lower limbs and, in severe cases, in the upper limbs. Signs include high-stepping gait associated with bilateral foot drop and, in severe cases, quadriplegia with foot and wrist drop as well as pyramidal signs. In time, there might be significant recovery of the peripheral nerve function but, depending on the degree of pyramidal involvement, spastic ataxia may be a permanent outcome of severe OPIDP. Human and experimental data indicate that recovery is usually complete in the young. At onset, the electrophysiological changes include reduced amplitude of the compound muscle potential, increased distal latencies and normal or slightly reduced nerve conduction velocities. The progression of the disease, usually over a few days, may lead to non-excitability of the nerve with electromyographical signs of denervation. Nerve biopsies have been performed in a few cases and showed axonal degeneration with secondary demyelination. Neuropathy target esterase (NTE) is thought to be the target of OPIDP initiation. The ratio of inhibitory powers for acetylcholinesterase and NTE represents the crucial guideline for the aetiological attribution of OP-induced peripheral neuropathy. In fact, pre-marketing toxicity testing in animals selects OP insecticides with cholinergic toxicity potential much higher than that to result in OPIDP. Therefore, OPIDP may develop only after very large exposures to insecticides, causing severe cholinergic toxicity. However, this was not the case with certain triaryl phosphates that were not used as insecticides but as hydraulic fluids, lubricants and plasticisers and do not result in cholinergic toxicity. Several thousand cases of OPIDP as a result of exposure to tri-ortho-cresyl phosphate have been reported, whereas the number of cases of OPIDP as a result of OP insecticide poisoning is much lower. In this article, we mainly discuss OP pesticide poisoning, particularly when caused by chlorpyrifos, dichlorvos, isofenphos, methamidophos, mipafox, trichlorfon, trichlornat, phosphamidon/mevinphos and by certain carbamates. We also discuss case reports where neuropathies were not convincingly attributed to fenthion, malathion, omethoate/dimethoate, parathion and merphos. Finally, several observational studies on long-term, low-level exposures to OPs that sometimes reported mild, inconsistent and unexplained changes of unclear significance in peripheral nerves are briefly discussed. Title: Peripheral nerve esterases and the promotion of organophosphate-induced neuropathy in hens Moretto A, Nicolli A, Lotti M Ref: Chemico-Biological Interactions, 157-158:285, 2005 : PubMed ESTHER: Moretto_2005_Chem.Biol.Interact_157-158_285 PubMedSearch: Moretto 2005 Chem.Biol.Interact 157-158 285 PubMedID: 16243301 Abstract Several esterase inhibitors, not capable of causing peripheral neuropathy by themselves, exacerbate organophosphate-induced delayed polyneuropathy (OPIDP) and other axonopathies. This effect was called promotion of axonopathies and it was found not to be associated with inhibition of neuropathy target esterase (NTE), the molecular target of OPIDP. The search for an esterase as the target of promotion has started long ago, when an eterogeneous group of esterases-hydrolysing phenyl valerate (PV) was identified in hen's sciatic nerve by means of selective inhibitors. Correlation studies in vivo indicated that the target of promotion may have been among the proteins present in the soluble fraction. When this soluble PV-esterase activity was separated on a Sephacryl-S-300 column, correlation was found between promotion and its inhibition in vivo. The electrophoretic analysis of this fraction indicated the presence of several proteins. Subsequent ion-exchange chromatography identified a protein of about 80 kDa molecular weight that was associated with PV-esterase activity. The inhibition of this activity did also correlate with promotion. The sequence of this protein identified it as ovotransferrin, but commercial preparations of ovotransferrin were found to lack PV-esterase activity. Binding experiments on this purified PV-activity and on commercial ovotransferrin using radiolabelled promoters were inconclusive. Titration of this PV-activity showed that about 20-30% of it is resistant to high concentrations of several inhibitors, suggesting heterogeneity of the fraction. In fact, bi-dimensional electrophoresis indicated the presence of several proteins. Finally, in vivo correlation experiments with p-toluensulfonyl fluoride showed that whereas this chemical does not promote OPIDP induced by dibutyl dichlorovinyl phosphate, it does inhibit about 80% of this PV-activity. In conclusion, available data indicate that the target of promotion is unlikely to be ovotransferrin. However, all promoters identified so far are esterase inhibitors suggesting that the target of promotion might be, indeed, a protein with esteratic activity. Title: The relationship between isofenphos cholinergic toxicity and the development of polyneuropathy in hens and humans Moretto A, Lotti M Ref: Archives of Toxicology, 76:367, 2002 : PubMed ESTHER: Moretto_2002_Arch.Toxicol_76_367 PubMedSearch: Moretto 2002 Arch.Toxicol 76 367 PubMedID: 12107655 Abstract Species differences have been observed between hen and human clinical manifestations of isofenphos toxicities. Hens treated with the insecticide isofenphos (90 mg/kg p.o.) developed severe cholinergic toxicity followed by mild organophosphate-induced delayed polyneuropathy (OPIDP). However, a patient developed severe OPIDP, which was preceded by very mild cholinergic signs, after an attempted suicide with a commercial formulation containing isofenphos and phoxim, an insecticide not causing OPIDP (estimated doses were 500 and 125 mg/kg, respectively). To explain this difference the following hypotheses were tested: (1) phoxim is a promoter of isofenphos-induced OPIDP; (2) whereas neuropathy target esterase (NTE) is thought to be the target of OPIDP, activation of isofenphos by liver microsomes causes the formation of more potent NTE inhibitor(s) in humans than in hens; (3) in contrast to hen NTE, the sensitivity of the human enzyme to such inhibitor(s) is higher than that of acetylcholinesterase (AChE), the target of cholinergic toxicity. Results showed that phoxim (22.5 mg/kg p.o.) was not a promoter of OPIDP in hens and that the ratio AChE inhibition:NTE inhibition by microsome-activated isofenphos was similar for both hen and human enzymes. The schedule of antidotal treatment in hens is the likely explanation for the observed difference from the patient. Peak AChE inhibition was maintained in hen brain up to 6 days after a single dose of isofenphos, suggesting prolonged pharmacokinetics. However, the AChE reactivator pyridine-2-aldoxime (2-PAM) was given to hens before isofenphos and then every 8 h, whereas continuous 2-PAM infusion was provided to the patient. When 2-PAM was given to hens every hour after isofenphos (90 mg/kg p.o.), the birds remained asymptomatic. Since other organophosphates may have a prolonged pharmacokinetics, testing procedures for the potential of these insecticides to cause OPIDP may underestimate the risk for humans. Title: Effects of S-ethyl hexahydro-1H-azepine-1-carbothioate (molinate) on di-n-butyl dichlorovinyl phosphate (DBDCVP) neuropathy Moretto A, Gardiman G, Panfilo S, Colle MA, Lock EA, Lotti M Ref: Toxicol Sci, 62:274, 2001 : PubMed ESTHER: Moretto_2001_Toxicol.Sci_62_274 PubMedSearch: Moretto 2001 Toxicol.Sci 62 274 PubMedID: 11452140 Inhibitor(s) related to this paper: Molinate, Dibutyl-dichlorovinyl-phosphate~DBVP Abstract Certain esterase inhibitors protect from organophosphate-induced delayed polyneuropathy (OPIDP) when given before a neuropathic organophosphate by inhibiting neuropathy target esterase (NTE). In contrast, they can exaggerate OPIDP when given afterwards and this effect (promotion) is associated with inhibition of another esterase (M200). In vitro sensitivities of hen, rat, and human NTE and M200 to the active metabolites of molinate, sulfone, and sulfoxide, were similar. NTE and M200 were irreversibly inhibited (> 78%) in brain and peripheral nerve of hens and rats given molinate (100-180 mg/kg, sc). No clinical or morphological signs of neuropathy developed in these animals. Hens and rats were protected from di-n-butyl dichlorovinyl phosphate neuropathy (DBDCVP, 1 and 5 mg/kg, sc, respectively) by molinate (180 or 100 mg/kg, sc, 24 h earlier, respectively) whereas 45 mg/kg, sc molinate causing about 34% NTE inhibition offered partial protection to hens. Hens treated with DBDCVP (0.4 mg/kg, sc) developed a mild OPIDP; molinate (180 mg/kg, 24 h later) increased the severity of clinical effects and of histopathology in spinal cord and in peripheral nerves. Lower doses of molinate (45 mg/kg, sc), causing about 47% M200 inhibition, did not promote OPIDP whereas the effect of 90 mg/kg, sc (corresponding to about 50-60% inhibition) was mild and not statistically significant. OPIDP induced by DBDCVP (5 mg/kg, sc) in rats was promoted by molinate (100 mg/kg, sc). In conclusion, protection from DBDCVP neuropathy by molinate is correlated with inhibition of NTE whereas promotion of DBDCVP neuropathy is associated with > 50% M200 inhibition Title: Use of human data for the derivation of a reference dose for chlorpyrifos van Gemert M, Dourson M, Moretto A, Watson M Ref: Regul Toxicol Pharmacol, 33:110, 2001 : PubMed ESTHER: van Gemert_2001_Regul.Toxicol.Pharmacol_33_110 PubMedSearch: van Gemert 2001 Regul.Toxicol.Pharmacol 33 110 PubMedID: 11350194 Abstract In 1998 a panel of experts met to discuss the data available on chlorpyrifos, both human and animal, and to determine the most appropriate endpoints to be used for the derivation of the reference dose (RfD). Since that time, additional data have become available on chlorpyrifos from an experimental study involving humans. Moreover, Food Quality Protection Act (FQPA) considerations need to be addressed, and the appropriate cholinesterase endpoint, whether plasma, red blood cell, peripheral nerve, or brain, has become highly debated. Therefore, Dow AgroSciences, one of the manufacturers of chlorpyrifos, convened a second panel of toxicology and medical experts on June 21, 1999, to consider the presently available scientific literature both published and unpublished on chlorpyrifos and to determine the acute and chronic toxicological RfDs for chlorpyrifos. Four questions were posed to this second panel of experts concerning the available data on chlorpyrifos. (1) Should the RfD for chlorpyrifos be based on acetylcholinesterase (AChE) inhibition or butyrylcholinesterase (BuChE) inhibition as an endpoint for adverse effect? (2) Should the RfDs for chlorpyrifos be based on the data set from three human studies, which are supported by animal data? (3) Should the FQPA safety factor be reduced to 1xbased on animal studies of pre- or postnatal toxicity? (4) If an RfD for chlorpyrifos were to be based on animal data, then is a 10-fold interspecies uncertainty factor necessary? The panel of experts concluded that: (1) inhibition of BuChE is not an adverse effect, and the RfD for chlorpyrifos should be based on AChE inhibition; (2) the RfD for chlorpyrifos should be based on the three available human studies, which are also supported by animal data; (3) the extra FQPA safety factor should be reduced to 1x, because chlorpyrifos shows no pre- or postnatal toxicity of concern at relevant human exposure conditions; and (4) the extra 10-fold safety factor for interspecies variation appears overly conservative because no differences in species sensitivity to chlorpyrifos is evident. Title: The relevance of inhibitor-substrate interactions when measuring neuropathy target esterase inhibition Moretto A, Jokanovic M, Lotti M Ref: Archives of Toxicology, 73:655, 2000 : PubMed ESTHER: Moretto_2000_Arch.Toxicol_73_655 PubMedSearch: Moretto 2000 Arch.Toxicol 73 655 PubMedID: 10741477 Inhibitor(s) related to this paper: O-hexyl-O-2,5-dichlorophenyl-phosphoramidate Abstract Neuropathy target esterase (NTE), thought to be the target for organophosphate polyneuropathy, is operationally defined as that neural phenyl valerate esterase resistant to paraoxon (40 microM) and sensitive to mipafox (50 microM; 20 min, pH 8.0, 37 degrees C). The time course of inhibition of particulate paraoxon pretreated esterases by mipafox showed that the lines indicating the rate of inhibition did not pass through the log 100% activity when extrapolated at zero time. Slopes of inhibition of NTE were not linearly related to the concentration of mipafox. Kinetic parameters derived from Wilkinson type plots were: Ka = 49-199 microM, k(+2) = 0.24-0.64 min(-1) and k(a) = 3.1-5.0 mM(-1) m(-1). When mipafox was removed (either by dilution or centrifugation) before the addition of phenyl valerate intercepts below 100% disappeared. We confirm that the formation of Michaelis complex between NTE and mipafox is not prevented by phenyl valerate and that inhibition proceeds after addition of phenyl valerate. We compared inhibitions obtained with experiments by using the traditional method (sequential incubation with inhibitors and phenyl valerate) to those obtained with a method where mipafox is removed before the addition of substrate. When calculating fixed-time 50% inhibitory concentrations (IC50s) of some inhibitors for NTE, the longer the hydrolysis time, the lower were the IC50s. Therefore, the inhibitory potency of certain NTE inhibitors, is accurately assessed only when calculating second-order rate constants (k(a)). Title: Promotion of organophosphate induced delayed polyneuropathy by certain esterase inhibitors Lotti M, Moretto A Ref: Chemico-Biological Interactions, 119-120:519, 1999 : PubMed ESTHER: Lotti_1999_Chem.Biol.Interact_119-120_519 PubMedSearch: Lotti 1999 Chem.Biol.Interact 119-120 519 PubMedID: 10421491 Abstract Certain esterase inhibitors elicit or intensify the clinical expression of various insults to axons. This phenomenon was called promotion of axonopathies because these chemicals are not additive neurotoxicants nor do they interfere with the pharmacokinetics. Characterization of promotion was carried out by using organophosphate induced delayed polyneuropathy (OPIDP) as a model. The search for a physiological explanation of promotion has the following background: (1) Promotion expresses clinically the biochemical lesions which are otherwise well compensated (such as 30/40% neuropathy target esterase (NTE) inhibition by neuropathic organophosphates). (2) Promotion is not specific because axonopathies of different origin are affected. (3) Promoters are effective when given several days before the neuropathic insult. (4) Promotion is less effective in young animals as compared with adults. (5) Promotion occurs when axons, but not necessarily the cell body, are targeted by promoters. (6) Repeated dosing with a promoter failed to produce axonopathy. Based on this evidence it is suggested that promotion might interfere with a mechanism(s) of compensation and/or repair of long axons. The target of promotion of axonopathies is thought to be similar or linked to NTE which is defined as the phenyl valerate esterase activity (PVE) in nervous tissues resistant to paraoxon and sensitive to mipafox (40 and 50 microM, pH 8.0, 20 min, respectively). Mipafox (50 microM) resistant PVEs include some activity sensitive to the promoter phenylmethane sulfonylfluoride (PMSF) but no correlation was found between its inhibition and promotion. A complete titration curve of paraoxon-resistant PVEs by mipafox (0-1 mM) dissected, besides NTE (I50 about 10 microM), another PVE with an I50 of approximately 200 microM. This enzyme was present in hen brain, spinal cord and peripheral nerve, corresponding to about 10, 20 and 30% of NTE activity, respectively, and was sensitive both in vitro and in vivo to promoters and much less so to neuropathic NTE inhibitors. By means of chromatography, other workers have identified in soluble extracts of peripheral nerves two forms of mipafox-sensitive PVEs with different molecular weights and different sensitivity to mipafox. These might correspond to NTE and to the other enzyme. Inhibition in vivo of the latter also correlated with promotion. Title: Repeated low doses of O-(2-chloro-2,3,3 trifluorocyclobutyl) O-ethyl S- propyl phosphorothioate (KBR-2822) do not cause neuropathy in hens Jokanovic M, Moretto A, Lotti M Ref: Archives of Toxicology, 72:93, 1998 : PubMed ESTHER: Jokanovic_1998_Arch.Toxicol_72_93 PubMedSearch: Jokanovic 1998 Arch.Toxicol 72 93 PubMedID: 9456080 Inhibitor(s) related to this paper: KBR-2822 Abstract Certain esterase inhibitors such as O-(2-chloro-2,3,3-trifluorocyclobutyl) O-ethyl S-propyl phosphorothioate (KBR-2822) and phenylmethanesulfonyl fluoride (PMSF) cause exacerbation (promotion) of toxic and traumatic axonopathies. Although these chemicals are capable of inhibiting neuropathy target esterase (NTE), which is the target for organophosphate induced delayed neuropathy, the target for promotion is unlikely to be NTE. Experiments were aimed to ascertain if neuropathy is caused by repeated dosing with a promoter not causing NTE inhibition and in the absence of deliberate injury to axons. Hens were treated with KBR-2822 (0.2 or 0.4 mg/kg per day) by gavage for 90 days and observed for clinical signs up to 21-23 days after treatment when histopathological examination was carried out. NTE and acetylcholinesterase (AChE) were measured at intervals and mean percentages of inhibition at steady state of inhibition/resynthesis (on day 20) were as follows: mean inhibition NTE was < or = 8% in the 0.2 mg/kg group and between 15 and 18% in the 0.4 mg/kg group in brain, spinal cord and peripheral nerve; mean AChE inhibition in brain was 31 and 57% in the two experimental groups, respectively. Controls treated with paraoxon (not neuropathic or a promoter and given at 0.05 mg/kg per day by gavage) showed 45% mean AChE inhibition and no NTE inhibition. Neither clinical nor morphological signs of neuropathy were observed in any group. To ascertain whether subclinical lesions were produced by the repeated treatment with KBR-2822, hens were given KBR-2822 (0.2 mg/kg per day) for 21 days by gavage followed by PMSF (120 mg/kg s.c. 24 h after the last dose of KBR-2822). A control group of hens was treated with the neuropathic DFP (0.03 mg/kg s.c. daily for 21 days causing 40-50% NTE inhibition) followed by PMSF (120 mg/kg s.c.). After PMSF, the KBR-2822 treated hens did not develop neuropathy whereas DFP treated hens did. Lack of neuropathy after repeated treatment with KBR-2822 indicates that a continuous promoting 'pressure' on hen axons is harmless in the absence of a concurrent biochemical or neurotoxic injury. Title: Experimental and clinical toxicology of anticholinesterase agents Moretto A Ref: Toxicol Lett, 103:509, 1998 : PubMed ESTHER: Moretto_1998_Toxicol.Lett_103_509 PubMedSearch: Moretto 1998 Toxicol.Lett 103 509 PubMedID: 10022304 Abstract Several organophosphorus compounds (OP) and carbamates (CA) are used as insecticides or warfare agents (OPs only). Their acute toxic effect in the central and peripheral nervous system is due to inhibition of acetylcholinesterase (AChE) at nerve endings which causes accumulation of acetylcholine and consequently overstimulation of the nicotinic and muscarinic receptors. The cholinergic syndrome appears at approximately 50% AChe inhibition whereas death is believed to occur at > 90%. Inhibition of AChE (phosphorylation) by most OPs is irreversible whereas CAs reversibly inhibit AChE (spontaneous reactivation with a t(1/2) of minutes); dimethylphosphorylated AChE partially and slowly (t(1/2) = 1-2 h) reactivates. Although long-term, mild neurobehavioural changes of questionable significance have been reported in some instances, recovery from the cholinergic syndrome appears to be complete, unless lesions develop in the central nervous system as a consequence of either convulsions or anoxia. Certain OPs and CAs have been reported to interact with cholinergic receptors in vitro. The toxicological relevance of these interactions is still not clear. Certain OPs cause OP-induced delayed polyneuropathy (OPIDP) which develops 2-5 weeks after an acute poisoning. The molecular target is believed to be neuropathy target esterase (NTE). OP insecticides are more potent AChE inhibitors rather than NTE inhibitors and therefore, the dose required to cause OPIDP is much higher than that causing the cholinergic syndrome. In the experimental animal, OPIDP is associated with > 70% NTE inhibition after single or repeated exposures. The threshold in man is not known, although there are indications that it is similar. Some non-neuropathic esterase inhibitors (OPs, CAs, sulfonyl fluorides) exacerbate the clinical outcome of OPIDP and other chemical axonopathies, and of nerve crush. The phenomenon has been called promotion and has so far been observed in experimental animals only. Title: Poisoning by organophosphorus insecticides and sensory neuropathy Moretto A, Lotti M Ref: Journal of Neurology Neurosurg Psychiatry, 64:463, 1998 : PubMed ESTHER: Moretto_1998_J.Neurol.Neurosurg.Psychiatry_64_463 PubMedSearch: Moretto 1998 J.Neurol.Neurosurg.Psychiatry 64 463 PubMedID: 9576536 Abstract OBJECTIVES: Poisoning by organophosphate insecticides causes cholinergic toxicity. Organophosphate induced delayed polyneuropathy (OPIDP) is a sensory-motor distal axonopathy which usually occurs after ingestion of large doses of certain organophosphate insecticides and has so far only been reported in patients with preceding cholinergic toxicity. Surprisingly, it was recently reported by other authors that an exclusively sensory neuropathy developed in eight patients after repeated unquantified exposures to chlorpyrifos, which did not cause clear-cut cholinergic toxicity. The objective was to assess whether an exclusively sensory neuropathy develops in patients severely poisoned by various OPs. Title: Phenyl valerate esterases other than neuropathy target esterase and the promotion of organophosphate polyneuropathy Milatovic D, Moretto A, Osman KA, Lotti M Ref: Chemical Research in Toxicology, 10:1045, 1997 : PubMed ESTHER: Milatovic_1997_Chem.Res.Toxicol_10_1045 PubMedSearch: Milatovic 1997 Chem.Res.Toxicol 10 1045 PubMedID: 9305588 Abstract Certain esterase inhibitors (such as phenylmethanesulfonyl fluoride, PMSF) enhance the clinical and morphological signs of organophosphate-induced delayed polyneuropathy (OPIDP) in hens. This is called promotion of OPIDP. The target of promotion is unknown, but it is likely to be different from neuropathy target esterase (NTE), the target of OPIDP, NTE is a neural phenyl valerate (PV) esterase, operationally defined by selective inhibition with organophosphates. This study was aimed to ascertain whether the target for promotion is a PV esterase other than NTE. Brain and sciatic nerve PV esterases of hens were incubated with diisopropylphosphorofluoridate (DFP; 5 microM) or N,N-diisopropyl phosphorodiamidofluoridate (mipafox; 50 microM) to inhibit NTE and other esterases thought not to be relevant to promotion. Remaining activities, quantitatively similar after either inhibition, were titrated with PMSF (up to 500 microM) and analysis of time course of inhibition showed first-order kinetics. Mipafox (50 microM)-resistant PMSF (500 microM)-sensitive activity (about 80% of mipafox-resistant ones) was tested both in vitro and in vivo with several inhibitors. No correlation was found between inhibition of mipafox-resistant PMSF-sensitive activity and the capability of several inhibitors to promote OPIDP. We conclude that the target of promotion is unlikely to be a PV esterase resistant to mipafox (50 microM). Title: Sulfonyl fluorides and the promotion of diisopropyl fluorophosphate neuropathy Osman KA, Moretto A, Lotti M Ref: Fundamental & Applied Toxicology, 33:294, 1996 : PubMed ESTHER: Osman_1996_Fundam.Appl.Toxicol_33_294 PubMedSearch: Osman 1996 Fundam.Appl.Toxicol 33 294 PubMedID: 8921348 Abstract Phenylmethanesulfonyl fluoride (PMSF) enhances the neuropathic response when given to hens after organophosphates causing delayed polyneuropathy. This study was undertaken to ascertain whether other sulfonyl fluorides promote diisopropyl fluorophosphate (DFP) neuropathy in hens and if they inhibit neuropathy target esterase (NTE), the target for organophosphate-induced delayed polyneuropathy. Among seven sulfonyl fluoride analogs of PMSF (alkyl-, and phenylsulfonyl fluorides), only n-butanesulfonyl fluoride was found to be an NTE inhibitor in vitro at a concentration (I50 = 60 microM) similar to that of PMSF, n-Butanesulfonyl fluoride (0.2 mmol.kg-1 sc to hens) caused both NTE inhibition in nervous tissues (> 80%) and promotion of neuropathy after DFP (0.003 mmol.kg-1 sc) similar to those observed after the same molar dose of PMSF. These results confirm that, so far, all known promoters of organophosphate polyneuropathy are also NTE inhibitors. Title: Cholinergic symptoms and Gulf War syndrome Lotti M, Moretto A Ref: Nat Med, 1:1225, 1995 : PubMed Title: Organophosphate polyneuropathy and neuropathy target esterase: studies with methamidophos and its resolved optical isomers Lotti M, Moretto A, Bertolazzi M, Peraica M, Fioroni F Ref: Archives of Toxicology, 69:330, 1995 : PubMed ESTHER: Lotti_1995_Arch.Toxicol_69_330 PubMedSearch: Lotti 1995 Arch.Toxicol 69 330 PubMedID: 7654138 Abstract Methamidophos (O,S-dimethyl phosphorothioamidate) causes polyneuropathy in man and hens. However, experiments in the hen show that lower doses of methamidophos either protect from or promote the neuropathy caused by certain organophosphates. The initiation of neuropathy as well as protection from neuropathy are thought to be related to neuropathy target esterase (NTE), whereas promotion is likely to be due to interactions with another unknown target. Methamidophos is a racemate and we report studies with its resolved optical isomers, aimed at elucidating which isomer is responsible for the described effects. The time-course of acetylcholinesterase (AChE) and NTE activity in nervous tissues of hens after inhibition by single doses of either isomer showed that after D-(+) methamidophos (25 mg/kg PO) peak inhibition of both enzymes was achieved within 24 h (80-90%). However, after L-(-) methamidophos (15 mg/kg PO), peak inhibition (80-90%) was obtained within 24 h for AChE, whereas similar NTE inhibition (120 mg/kg PO) was observed only 4 days after dosing. The minimal neuropathic doses of D-(+) and L-(-) methamidophos were 60 and 120 mg/kg PO, respectively, and correlated with > 80% NTE inhibition in nervous tissues. OPIDP initiation by either isomer was slightly promoted by phenylmethanesulfonyl fluoride (120 mg/kg SC). D-(+) Methamidophos (25 mg/kg PO) partially protected from dibutyl dichlorovinyl-phosphate (DBDCVP) neuropathy (up to 0.8 mg/kg SC). This effect correlated with about 70% NTE inhibition. L-(-) Methamidophos (15 or 60 mg/kg PO) did not protect from DBDCVP neuropathy (0.2-0.8 mg/kg SC). Title: A novel probe for characterisation of neuropathy target esterase Moretto A Ref: Hum Exp Toxicol, 14:930, 1995 : PubMed ESTHER: Moretto_1995_Hum.Exp.Toxicol_14_930 PubMedSearch: Moretto 1995 Hum.Exp.Toxicol 14 930 PubMedID: 8588957 Inhibitor(s) related to this paper: OBDPO Title: Selective promotion by phenylmethanesulfonyl fluoride of peripheral and spinal cord neuropathies initiated by diisopropyl phosphorofluoridate in the hen Peraica M, Moretto A, Lotti M Ref: Toxicol Lett, 80:115, 1995 : PubMed ESTHER: Peraica_1995_Toxicol.Lett_80_115 PubMedSearch: Peraica 1995 Toxicol.Lett 80 115 PubMedID: 7482578 Abstract This paper reports studies in hens showing that diisopropyl phosphorofluoridate (DFP) neuropathy is promoted by PMSF when initiated either in central (spinal cord) or peripheral nervous system. Moreover, the critical site for promotion is in peripheral nerve axons rather than in their cell bodies. Selective promotion in peripheral nerves was achieved by giving PMSF into sciatic artery monolaterally (7 mg/kg) to birds where neuropathy was initiated by DFP, either systematically (0.3 mg/kg s.c.) or intra-arterially (0.04 mg/kg in the same artery). Birds developed monolateral neuropathy in the leg where PMSF was delivered. Promotion of spinal cord neuropathy was achieved by giving PMSF (120 mg/kg s.c.) to birds where neuropathy was initiated selectively in spinal cord. This was obtained by protecting peripheral axons with intra-arterial bilateral injections of PMSF (0.55 x 2 mg/kg) followed by DFP (0.3, 0.4 or 0.7 mg/kg s.c.). The resulting syndrome was characterized by spastic ataxia. Title: The phosphorothioic acid O-(2-chloro-2,3,3-trifluorocyclobutyl) O-ethyl S-propyl ester exacerbates organophosphate polyneuropathy without inhibition of neuropathy target esterase Moretto A, Bertolazzi M, Lotti M Ref: Toxicol Appl Pharmacol, 129:133, 1994 : PubMed ESTHER: Moretto_1994_Toxicol.Appl.Pharmacol_129_133 PubMedSearch: Moretto 1994 Toxicol.Appl.Pharmacol 129 133 PubMedID: 7974486 Inhibitor(s) related to this paper: KBR-2822 Abstract Organophosphate-induced delayed polyneuropathy (OPIDP) is thought to be initiated by a variety of neuropathy target esterase (NTE) inhibitors. However, certain inhibitors such as phenylmethanesulfonyl fluoride, phenyl N-methyl N-benzyl carbamate, and phenyl di-n-pentyl phosphinate protect from OPIDP when given to hens before organophosphorus esters. They protect from neuropathy by preventing the binding of neuropathic inhibitors to NTE catalytic site. In contrast, when such NTE inhibitors are given afterward, the resulting clinical effect is more severe. This phenomenon was called promotion of OPIDP. Promotion has been tentatively explained by the interaction of promoters with a target other than the catalytic center of NTE. However, the doses of promoters which cause the effect have, so far, been found to always be inhibitory of NTE. We report that the phosphorothioic acid O-(2-chloro-2,3,3-trifluorocyclobutyl) O-ethyl S propyl ester (KBR-2822) given to hens at doses which did not inhibit NTE (2.5 mg/kg p.o.) promoted the neuropathies initiated by either dibutyl-2,2-dichlorovinyl phosphate (DBDCVP, 0.4 mg/kg s.c., 24 hr earlier) or diisopropyl phosphorofluoridate (DFP, 0.3 mg/kg sc or 0.5 mg/kg s.c., 24 hr earlier). When given alone, DBDCVP and DFP (0.5 mg/kg) caused mild OPIDP, whereas the lower dose of DFP did not cause clinical effects. Dose-response relationships with KBR-2822 indicated that clinical effects of the combined treatments are unlikely to be additive because the compound did not cause OPIDP up to the maximum tolerated dose (10 mg/kg p.o.). Promotion also occurred when KBR-2822 (2.5 mg/kg p.o.) was given before either DBDCVP (0.4 mg/kg s.c.) or DFP (0.3 mg/kg s.c.). NTE inhibitions in the nervous tissues caused by DBDCVP or DFP were not affected by pretreatment with KBR-2822, suggesting that the delivery of neuropathic. NTE inhibitors was not modified. We conclude that KBR-2822 promotes OPIDP initiated by either DBDCVP or DFP by affecting a target other than NTE catalytic site. Title: Interactions between neuropathy target esterase and its inhibitors and the development of polyneuropathy Lotti M, Moretto A, Capodicasa E, Bertolazzi M, Peraica M, Scapellato ML Ref: Toxicol Appl Pharmacol, 122:165, 1993 : PubMed ESTHER: Lotti_1993_Toxicol.Appl.Pharmacol_122_165 PubMedSearch: Lotti 1993 Toxicol.Appl.Pharmacol 122 165 PubMedID: 8211998 Abstract This paper combines new and old data in order to offer a modified perspective of the mechanism of organophosphate-induced delayed polyneuropathy. Neuropathy target esterase (NTE) is though to be the molecular target and neuropathy to be initiated with a two-step mechanism: progressive inhibition of NTE and aging of the phosphorylated enzyme. When neuropathic organophosphates modify more than 70% of NTE in this way, neuropathy develops 2 weeks later. Other chemicals producing an inhibited NTE, which is incapable of aging, were thought to be not neuropathic. When given before a challenging dose of a neuropathic organophosphate they protect animals from neuropathy. However, recent evidence indicates that aging may not always be essential in causing neuropathy. In fact, mipafox and methamidophos as well as certain classic protective inhibitors such as carbamate and sulfonyl fluoride form an inhibited NTE which apparently does not age and yet produces neuropathy. We propose that all NTE inhibitors may have the potential to cause neuropathy. In analogy with pharmacological models of drug-receptor interactions, NTE inhibitors might have variable intrinsic activities to initiate neuropathy once attached to the protein. Strong neuropathic chemicals require about 70% inhibition of NTE, others 80-90%, and the least potent almost 100%. These differences have been amplified by means of promotion. Different levels of NTE inhibition as caused by different compounds were promoted by the same dose of phenylmethanesulfonyl fluoride to similar degrees of ataxia. Conversely nearly complete NTE inhibitions obtained in chicks with different chemicals were promoted to varying severities of ataxia. Protection from delayed polyneuropathy by the least neuropathic inhibitors can be explained by their weak intrinsic activity: occupying NTE, they prevent the binding of more neuropathic compounds. Methamidophos represents a particular example because it is protective at lower doses and neuropathic at high doses. Moreover, the levels of NTE inhibited by methamidophos which can be promoted to neuropathy are lower than those required for classic protective chemicals and higher than those of classic neuropathic OPs. This suggests that methamidophos has an intermediate position between the most and the least neuropathic NTE inhibitors. Title: The search for the physiological functions of NTE; is NTE a receptor? Lotti M, Moretto A Ref: Chemico-Biological Interactions, 87:407, 1993 : PubMed ESTHER: Lotti_1993_Chem.Biol.Interact_87_407 PubMedSearch: Lotti 1993 Chem.Biol.Interact 87 407 PubMedID: 8393746 Abstract Neuropathy target esterase (NTE) was identified as the molecular target for organophosphate-induced delayed polyneuropathy several years ago but its physiological functions are still unknown. The mechanism which initiates neuropathy was thought to be a two step process: inhibition (phosphorylation) of NTE and aging of phosphorylated NTE. Depending on the occurrence of the second reaction (aging), inhibitors were ranked as neuropathic (forming an ageable NTE) and non-neuropathic (forming a non-ageable NTE). Non-neuropathic inhibitors protect from neuropathy if given before the neuropathic ones, because they occupy the catalytic centre of NTE. Thus the catalytic function of NTE seems irrelevant in maintaining the health of neurons. This paper reviews some new information concerning the interaction of NTE with its inhibitors as well as on a phenomenon called promotion of neuropathy. Some inhibitors which apparently form a non-ageable inhibited NTE were found to cause neuropathy, even though some of them must be given at very high doses. Moreover some 'non-neuropathic-protective' NTE inhibitors were found to exacerbate (promote) neuropathy when given after a neuropathic one. It is likely that the target for promotion is other than NTE. The hypothesis that NTE has some unknown receptorial functions where inhibitors act with different efficacy is discussed. NTE inhibitors have been ranked as full agonists (classic neuropathic inhibitors such as diisopropylfluorophosphate), partial agonists (protective or neuropathic, depending on the dose, such as methamidophos) and antagonists (protective, and neuropathic at the highest doses, such as phenylmethanesulfonyl fluoride). Age-related differences in the 'receptor' NTE might be responsible for the different sensitivities of juvenile and adult animals. Title: Promotion of peripheral axonopathies by certain esterase inhibitors Moretto A, Lotti M Ref: Toxicol Ind Health, 9:1037, 1993 : PubMed ESTHER: Moretto_1993_Toxicol.Ind.Health_9_1037 PubMedSearch: Moretto 1993 Toxicol.Ind.Health 9 1037 PubMedID: 8191502 Abstract Certain esterase inhibitors were found to exacerbate the clinical signs of polyneuropathy caused by various neurotoxic compounds and to delay the recovery from nerve crush. This phenomenon is referred to as promotion of axonopathies. The molecular target of promotion has not yet been identified. However, all known promoters are also inhibitors of neuropathy target esterase (NTE), the putative target of organophosphate neuropathy, but it has been shown that the target of promotion is unlikely to be NTE. Available data suggest that promoters might affect a target and a mechanism present in the nervous system that is not activated by axonal lesions. Promotion may be important to understand the physiological mechanism of nerve damage and repair. This finding also implies a changing perspective for the risk assessment of exposures to esterase inhibitors, some of which are used as pesticides and might be promoters. Title: Phenylmethanesulfonyl fluoride delays the recovery from crush of peripheral nerves in hens Moretto A, Capodicasa E, Peraica M, Lotti M Ref: Chemico-Biological Interactions, 87:457, 1993 : PubMed ESTHER: Moretto_1993_Chem.Biol.Interact_87_457 PubMedSearch: Moretto 1993 Chem.Biol.Interact 87 457 PubMedID: 8344003 Abstract Several esterase inhibitors (carbamates, phosphinates and sulfonyl halides) have been shown to promote organophosphate-induced delayed polyneuropathy (OPIDP). The mechanism of promotion is not understood, but indirect evidence suggests impairments of peripheral nerve repair. Also, other toxic neuropathies, such as those caused by 2,5-hexanedione in hens and bromophenylacetylurea in rats, have been reported to be promoted by phenylmethanesulfonyl fluoride (PMSF). Hen sciatic nerve was crushed at the bifurcation. Either mild or heavy pressure was applied by forceps obtaining a mild and rapidly recovering lesion (possibly myelinic) or a more severe, long-lasting lesion (possibly axonal), respectively. Hens were then treated with PMSF (120 mg/kg s.c. or 200 mg/kg s.c. x 2, 24 h apart) either before (5-48 h) crush or afterwards (5-48 h). Controls received vehicle only. Animals were observed for reappearance of digit movements, and standing and walking ability. PMSF treatment did not change the clinical outcome when animals received a mild crush. In hens receiving the more severe crush the reappearance of digit movements and the complete clinical recovery were observed after 43 +/- 14 and 63 +/- 9 days, respectively. In animals treated with PMSF there was a significant delay in both reappearance of digit movements (56 +/- 11 days when PMSF was given 24 and 48 h before crush, and 55 +/- 10 days, when given 24 and 48 h after crush) and in clinical recovery (75 +/- 15 and 80 +/- 18 days, respectively). It is concluded that traumatic axonopathy as well as toxic neuropathies can be promoted by PMSF. Moreover, it appears that PMSF promotion involves a target and a mechanism which are present in healthy axons and do not need to be activated by the insult to the axon. Title: Organophosphate polyneuropathy in chicks Peraica M, Capodicasa E, Moretto A, Lotti M Ref: Biochemical Pharmacology, 45:131, 1993 : PubMed ESTHER: Peraica_1993_Biochem.Pharmacol_45_131 PubMedSearch: Peraica 1993 Biochem.Pharmacol 45 131 PubMedID: 8381002 Inhibitor(s) related to this paper: Dibutyl-dichlorovinyl-phosphate~DBVP Abstract Young animals are resistant to organophosphate-induced delayed neuropathy (OPIDP), although biochemical changes on Neuropathy Target Esterase (NTE) caused by neuropathic organophosphorus esters (OP) are similar to those observed in the sensitive hen. We report here that the resistance of chicks to single doses of neuropathic OPs is not absolute because ataxia was produced in 40-day-old chicks by 2,2-dichlorovinyl dibutyl phosphate (DBDCVP, 5.0 or 10.0 mg/kg s.c.) and by diisopropyl phosphorofluoridate (DFP, 2.0 mg/kg s.c.). However, the clinical picture was different from that usually seen in hens; spasticity and complete recovery being the main features. alpha-Tolyl sulphonyl fluoride (PMSF, 300 mg/kg s.c.) promoted both DBDCVP neuropathy (5.0 or 10.0 mg/kg s.c.) and non-neuropathic doses of DFP (1.5 mg/kg s.c.) or DBDCVP (1.0 mg/kg s.c.). The lowest promoting dose of PMSF given 24 hr after 1.5 mg/kg of DFP was 30 mg/kg. Higher doses had a more severe effect but no further increase of OPIDP severity was obtained with doses ranging from 90 to 300 mg/kg. PMSF (30 mg/kg) protected 40-day-old chicks from subsequent doses of neuropathic OPs even when a promoting dose of PMSF followed. At 60 days of age, chicks' resistance to OPIDP decreased because lower doses of neuropathic OPs became effective and, similarly to hens, PMSF did not fully protect from subsequent promotion. In 40-day-old chicks the threshold of NTE inhibition for OPIDP development was 95-97% (DBDCVP 5.0 mg/kg). When promotion followed initiation, the minimal effective inhibition of NTE for initiation by neuropathic OPs was about 90%. In 36-day-old chicks, PMSF (300 mg/kg) promoted OPIDP when given up to 5 days after DFP (1.5 mg/kg) when residual NTE inhibition in brain and sciatic nerve was about 40%. We conclude that chicks' resistance to OPIDP might reflect either a less effective initiation by phosphorylated NTE or a more efficient repair mechanism or both, and also that promotion is likely to involve a target other than NTE. Title: Phenylmethanesulfonyl fluoride elicits and intensifies the clinical expression of neuropathic insults Moretto A, Bertolazzi M, Capodicasa E, Peraica M, Richardson RJ, Scapellato ML, Lotti M Ref: Archives of Toxicology, 66:67, 1992 : PubMed ESTHER: Moretto_1992_Arch.Toxicol_66_67 PubMedSearch: Moretto 1992 Arch.Toxicol 66 67 PubMedID: 1316117 Abstract It has been recently reported that phenylmethanesulfonyl fluoride (PMSF) when given to hens after a neuropathic organophosphate (OP) promotes organophosphate-induced delayed polyneuropathy (OPIDP). Chicks are resistant to OPIDP despite high inhibition/aging of neuropathy target esterase (NTE), the putative target of OPIDP initiation. However, when PMSF (300 mg/kg s.c.) is given to chicks after di-butyl 2,2-dichlorovinyl phosphate (DBDCVP, 1 or 5 mg/kg s.c.), OPIDP is promoted. Inhibition/aging of at least 30% of NTE was thought to be an essential prerequisite for promotion to be elicited in adult hens. However, we observed in hens that when NTE is maximally affected (greater than 90%) by phenyl N-methyl N-benzyl carbamate (40 mg/kg i.v.), a non-ageable inhibitor of NTE, and then PMSF is given (120 mg/kg/day s.c. x 3 days) clinical signs of neuropathy become evident. Methamidophos (50 mg/kg p.o. to hens), which produces in vivo a reactivatable form of inhibited NTE, was shown either to protect from or promote OPIDP caused by DBDCVP (0.45 mg/kg s.c.), depending on the sequence of dosing. Because very high doses of methamidophos cause OPIDP, we considered this effect to be a "self-promoted" OPIDP. We concluded that NTE inhibitors might have different intrinsic activities for producing OPIDP once NTE is affected. Aging might differentiate highly neuropathic OPs, like DBDCVP, from less neuropathic OPs, like methamidophos, or from the least neuropathic carbamates, which require promotion in order for neuropathy to be expressed.(ABSTRACT TRUNCATED AT 250 WORDS) Title: Neuropathy target esterase is not reduced in neural tissues of diabetic rats Moretto A, Lotti M, Triban C, Di Gregorio F, Fiori MG Ref: Diabete Metab, 18:218, 1992 : PubMed ESTHER: Moretto_1992_Diabete.Metab_18_218 PubMedSearch: Moretto 1992 Diabete.Metab 18 218 PubMedID: 1397476 Abstract Marked (greater than 70%) reduction of the neuropathy target enzyme (NTE) shortly after exposure to organophosphorus compounds heralds the onset of delayed neuropathic damage in animals and humans. One previous study reported that lymphocyte NTE from diabetic patients was depressed by greater than 70%; such a reduction was considered to be a biological marker of diabetic polyneuropathy. To ascertain whether NTE from target tissues might be involved in diabetic neuropathy, we measured NTE activity in the brain, spinal cord and peripheral nerves of streptozotocin-diabetic rats. No reduction in NTE activity was detected in these neural specimens. Therefore, it is concluded that NTE is not involved in diabetic nerve damage and that the meaning of low NTE activity in peripheral lymphocytes of diabetic patients remains unclear. Title: Clinical expression of organophosphate-induced delayed polyneuropathy in rats [published erratum appears in Toxicol Lett 1992 Dec;63(3):355] Moretto A, Capodicasa E, Lotti M Ref: Toxicol Lett, 63:97, 1992 : PubMed ESTHER: Moretto_1992_Toxicol.Lett_63_97 PubMedSearch: Moretto 1992 Toxicol.Lett 63 97 PubMedID: 1412529 Inhibitor(s) related to this paper: Dibutyl-dichlorovinyl-phosphate~DBVP Abstract Single doses of certain organophosphates (OP), such as dibutyl-2,2-dichlorovinyl phosphate (DBDCVP) cause organophosphate-induced delayed polyneuropathy (OPIDP) in hens. Clinical effects correlate with inhibition of neuropathy target esterase (NTE) which is considered the target for this toxicity. Pre-treatment with non-neuropathic NTE inhibitors, such as phenylmethanesulfonyl fluoride (PMSF), protects from OPIDP. However, when given after OPs, these compounds promote OPIDP. Chicks are relatively resistant to OPIDP despite high NTE inhibition. It has also always been reported that rats represent a species which is resistant to OPIDP and that they might develop morphological but not clinical signs of OPIDP. We report here that clinical OPIDP can be produced in 3.5- and 6-month-old rats by DBDCVP (5 mg/kg s.c.) and that it correlates with high (> 90%) NTE inhibition. When PMSF (120 mg/kg s.c. x 2) was given after DBDCVP, OPIDP was promoted. Pretreatment with PMSF protected from OPIDP. We conclude that resistance to OPIDP in the rat is age-related, as it is in the hen. Title: Interaction of methamidophos with hen and human acetylcholinesterase and neuropathy target esterase Bertolazzi M, Caroldi S, Moretto A, Lotti M Ref: Archives of Toxicology, 65:580, 1991 : PubMed ESTHER: Bertolazzi_1991_Arch.Toxicol_65_580 PubMedSearch: Bertolazzi 1991 Arch.Toxicol 65 580 PubMedID: 1781739 Abstract Methamidophos causes acute cholinergic toxicity in several species, including man, and organophosphate-induced delayed polyneuropathy which has been reported in man but not in the hen. Acetylcholinesterase (AChE) and neuropathy target esterase (NTE) are thought to be the molecular targets of acute and delayed toxicity, respectively. The rate constants of inhibition (ka) and reactivation (k + 3) of human and hen brain AChE and NTE by methamidophos resolved optical isomers are here reported. NTE inhibition was progressive and irreversible. Human and hen NTE ka (M-1.m-1) for D-(+) methamidophos was 88 and 59, respectively, and for L-(-) methamidophos 3.2 and 3.0, respectively. AChE spontaneously reactivates after inhibition. D-(+) methamidophos 10(-3).ka (M-1.m-1) for human and hen AChE was 0.24 and 0.13; 10(3).k+3 (m-1) was 0.83 and 0.69, respectively. L-(-) Methamidophos 10(-3).ka (M-1.m-1) for human and hen AChE was 5.7 and 2.8, whereas 10(3).k+3 (m-1) was 6.50 and 1.52, respectively. L-(-)-Inhibited AChE reactivated to about 60% for human and 30% for hen enzymes, respectively. D-(+)-Inhibited AChE reactivated to about 10-20% for both species. Maximal reactivation occurred within 4-6 h when a plateau was reached. The larger and faster reactivation of human AChE inhibited in vitro by L-(-) methamidophos suggests that a corresponding effect might be possible in vivo and therefore explain, in part, the relatively higher susceptibility of man to delayed polyneuropathy induced by racemic methamidophos which occurs, however, with doses always causing severe cholinergic toxicity. Title: Chlorpyrifos-induced delayed polyneuropathy Capodicasa E, Scapellato ML, Moretto A, Caroldi S, Lotti M Ref: Archives of Toxicology, 65:150, 1991 : PubMed ESTHER: Capodicasa_1991_Arch.Toxicol_65_150 PubMedSearch: Capodicasa 1991 Arch.Toxicol 65 150 PubMedID: 1711837 Abstract Chlorpyrifos [0,0-diethyl 0-(3,5,6-trichloro-pyridyl) phosphorothioate] caused delayed polyneuropathy in man. Contrary to previous studies, we report here that it also causes delayed polyneuropathy in the hen, the animal model for this toxicity. The minimal neuropathic dose was 60-90 mg/kg p.o., corresponding to 4-6 times the estimated LD50. Consequently, pralidoxime (2-PAM) in conjunction with atropine was necessary to reverse acetylcholinesterase (AChE) inhibition and cholinergic toxicity in hens given high enough doses of chlorpyrifos to cause neuropathy. Chlorpyrifos was slowly absorbed after single oral doses and the threshold of inhibition (greater than 70%) of neuropathy target esterase (NTE), the putative target for delayed neuropathy, was reached within 5-6 days. High AChE inhibition (greater than 90%), however, was measured within hours after dosing because of the higher potency of chlorpyrifos to inhibit this enzyme. In vitro studies showed that chlorpyrifos-oxon, the active metabolite of chlorpyrifos, was 10-20 times more active against AChE than against NTE, confirming the clinical observation. No differences were seen between human and hen enzymes in this respect. Hen and human brain homogenates contain A-esterases which hydrolysed chlorpyrifos to about the same extent in both species. In conclusion, chlorpyrifos causes delayed polyneuropathy in the hen, as was reported in man. The reasons for previous negative data in the hen are probably due to the relatively lower doses which were used. Judging from in vitro studies with hen and human enzymes, there are no differences in the two species as far as their relative sensitivity to delayed polyneuropathy. It is likely that delayed polyneuropathy would develop in both species only after severe cholinergic toxicity requiring aggressive antidotal treatment. Title: Promotion of organophosphate-induced delayed polyneuropathy by phenylmethanesulfonyl fluoride Lotti M, Caroldi S, Capodicasa E, Moretto A Ref: Toxicol Appl Pharmacol, 108:234, 1991 : PubMed ESTHER: Lotti_1991_Toxicol.Appl.Pharmacol_108_234 PubMedSearch: Lotti 1991 Toxicol.Appl.Pharmacol 108 234 PubMedID: 2017753 Abstract Certain sulfonates, like phenylmethanesulfonyl fluoride (PMSF), carbamates, and phosphinates, when given prior to neuropathic doses of organophosphates such as diisopropyl phosphorofluoridate (DFP), protect hens from organophosphate-induced delayed polyneuropathy (OPIDP). Protection was related to inhibition of the putative target of OPIDP, which is called Neuropathy Target Esterase (NTE). NTE inhibition above 70-80% in the nervous system of hens followed by a molecular rearrangement called aging initiates OPIDP. PMSF and other protective chemicals inhibit NTE but OPIDP does not develop because aging cannot occur. DFP (1 mg/kg sc) inhibited NTE above 70-80% in peripheral nerve and caused OPIDP in hens. Lower doses (0.3 and 0.5 mg/kg sc) caused about 40-60% NTE inhibition and no or marginal OPIDP. Chlorpyrifos (90 mg/kg po) also caused OPIDP. When repeated (30 mg/kg sc daily for 9 days) or single (5-120 mg/kg sc) doses of PMSF were given after either DFP or chlorpyrifos, OPIDP developed in birds treated with nonneuropathic doses of DFP and was more severe in birds treated with chlorpyrifos or higher doses of DFP. PMSF increased NTE inhibition to greater than 90%. Promotion of OPIDP with a single dose of PMSF (120 mg/kg sc) was obtained in birds up to 11 days after a marginally neuropathic dose of DFP (0.5 mg/kg sc). Promotion was also obtained with phenyl N-methyl N-benzyl carbamate (40 mg/kg iv) but not with non-NTE inhibitors in vivo such as paraoxon or benzenesulfonyl fluoride when given at maximum tolerated doses. These results indicate that protection from OPIDP is only one effect of PMSF because promotion of OPIDP is also observed depending upon the sequence of dosing. Either effect is always related to the doses of PMSF, which inhibit NTE. Title: Age sensitivity to organophosphate-induced delayed polyneuropathy. Biochemical and toxicological studies in developing chicks Moretto A, Capodicasa E, Peraica M, Lotti M Ref: Biochemical Pharmacology, 41:1497, 1991 : PubMed ESTHER: Moretto_1991_Biochem.Pharmacol_41_1497 PubMedSearch: Moretto 1991 Biochem.Pharmacol 41 1497 PubMedID: 2018554 Inhibitor(s) related to this paper: Dibutyl-dichlorovinyl-phosphate~DBVP Abstract Young animals are resistant to organophosphate-induced delayed polyneuropathy (OPIDP). The putative target protein in the nervous system for initiation of OPIDP in the adult hen is an enzyme called Neuropathy Target Esterase (NTE), which is dissected by selective inhibitors among nervous tissue esterases hydrolysing phenyl valerate (PV). We report here that the pool of PV-esterases sensitive to paraoxon was different in peripheral nerves of chicks as compared to that of hens while that of brain and spinal cord was not. NTE activity decreased with age in brain, spinal cord and peripheral nerve, but its sensitivity to several inhibitors remained unchanged. In the adult hen more than 70% inhibition of peripheral nerve NTE by neuropathic OPs is followed by deficit of retrograde axonal transport, axonal degeneration and paralysis. Similar NTE inhibition in 40-day-old or younger chicks however is not followed by changes in retrograde axonal transport nor by OPIDP. Chicks aged 60 to 80 days are only marginally sensitive to a single dose of DFP otherwise clearly neuropathic to hens. In vitro and in vivo phosphorylation by DFP and subsequent aging of brain NTE is similar both in chicks and in hens. The recovery of NTE activity monitored in vivo after inhibition by DFP is faster (half-life of about 3 days) in chick peripheral nerves as compared to chick brain, hen brain and hen peripheral nerve (half-life of about 5 days). It is concluded that the reduced sensitivity to OPIDP in chicks is not due to differences in OP-NTE interactions. The resistance might be explained by a more efficient repair mechanism, as suggested by the faster recovery of peripheral nerve NTE activity. Title: Distribution and some biochemical properties of rat paraoxonase activity Pellin MC, Moretto A, Lotti M, Vilanova E Ref: Neurotoxicology & Teratology, 12:611, 1990 : PubMed ESTHER: Pellin_1990_Neurotoxicol.Teratol_12_611 PubMedSearch: Pellin 1990 Neurotoxicol.Teratol 12 611 PubMedID: 2175012 Abstract The calcium-dependent enzyme activity which hydrolyzes the p-nitrophenyl-O-P bond of paraoxon (paraoxonase) has been studied in several rat and human tissues. Rat plasma and liver showed the highest activities (1.31 +/- 0.19, 0.82 +/- 0.09 nmol/min mg protein +/- SEM, respectively), while other tissues showed less than 2% plasma activity. The Arrhenius plot showed monophasic patterns in both tissues with activation energy values of Ea = 57 +/- 3 and 69 +/- 4 kcal/mol degree K for rat liver and plasma, respectively. Rat plasma and liver paraoxonase lost about 80% activity after 24-hr storage at 27-30 degrees C and was not restored by calcium addition. There was no loss of activity in human serum after 3 days and only 33% after 5 days. The pH optimum for paraoxonase activities was about 7.4 for both rat tissues. It is concluded that plasma paraoxonase is similar to the liver enzyme and is a good mirror for total body detoxifying activity. Title: In vivo and in vitro regional differential sensitivity of neuropathy target esterase to di-n-butyl-2,2-dichlorovinyl phosphate Moretto A, Lotti M, Spencer PS Ref: Archives of Toxicology, 63:469, 1989 : PubMed ESTHER: Moretto_1989_Arch.Toxicol_63_469 PubMedSearch: Moretto 1989 Arch.Toxicol 63 469 PubMedID: 2619560 Inhibitor(s) related to this paper: Dibutyl-dichlorovinyl-phosphate~DBVP Abstract Organophosphate-induced delayed polyneuropathy (OPIDP) is initiated by inhibition/aging of more than 70-75% of neuropathy target esterase (NTE). Di-n-butyl-2,2-dichlorovinyl phosphate (DBDCVP) (1 mg/kg s.c.) inhibited 96%, 86% and 83% of NTE in brain, spinal cord and peripheral nerve, respectively, and induced a typical central peripheral distal axonopathy in hens. A lower dose (0.45 mg/kg s.c.) caused 90%, 83% and 54% NTE inhibition in the same organs; by contrast, hens developed a spastic ataxia with axonal degeneration in spinal cord but not in peripheral nerve. With a dose of 0.2 mg/kg s.c., a suprathreshold inhibition of NTE was produced in brain (78%) but not in spinal cord (56%) and peripheral nerve (33%) and no morphological or clinical signs of neuropathy developed in hens. With doses up to 4.0 mg/kg s.c., acetylcholinesterase (AChE) inhibition was similar throughout the nervous system. In vitro time-course inhibition studies showed a different sensitivity to DBDCVP of NTE from peripheral nerve (ka = 5.4 x 10(6)) relative to that from spinal cord (ka = 13.9 x 10(6)) or brain (ka = 20.6 x 10(6)). In vitro I50s of DBDCVP for AChE were similar in brain, spinal cord and peripheral nerve (11-17 nM). These data support the hypothesis that the critical target for initiation of OPIDP is located in the nerve fiber, possibly in the axon and also suggest that peripheral nerve NTE has a different sensitivity to DBDCVP than the brain enzyme.(ABSTRACT TRUNCATED AT 250 WORDS) Title: Human serum A-esterases. Hydrolysis of O,O-dimethyl-2,2-dichlorovinyl phosphate Traverso R, Moretto A, Lotti M Ref: Biochemical Pharmacology, 38:671, 1989 : PubMed ESTHER: Traverso_1989_Biochem.Pharmacol_38_671 PubMedSearch: Traverso 1989 Biochem.Pharmacol 38 671 PubMedID: 2537085 Abstract Some characteristics of the hydrolysis of O,O-dimethyl-2,2 dichlorovinyl phosphate (DDVP) by human serum are reported and compared with the hydrolysis of O,O-diethyl-4-nitrophenyl phosphate (paraoxon) which is a substrate for Paraoxonase, a known "A"-esterase of human serum. When incubated with human serum, DDVP was losing its inhibitory power toward acetylcholinesterase (AChE). The loss of DDVP followed first order kinetics and was proportional to serum dilution. The disappearance of DDVP after incubation with human serum was not due to protein binding. Apparent Km and Vm for the hydrolysis of DDVP were 7.1 mM and 143 nmol.min-1.ml-1. The pH sensitivity, EDTA inhibitory and Ca2+ requirements of DDVP-ase were similar to those of Paraoxonase. DDVP inhibited the Paraoxonase activity and paraoxon inhibited the DDVP-ase activity. Ca2+, Ag+ and Hg2+ were better inhibitors of the Paraoxonase than the DDVP-ase. The rate of heat inactivation was also different; at 55 degrees Paraoxonase inactivated almost completely within 10 min, while DDVP-ase lost only about 10% activity over 1 hr. Consequently, DDVP-ase and Paraoxonase can be differentiated by means of heat sensitivity. The DDVP-ase was normally distributed in a population of 60 individuals, while Paraoxonase is known to show a marked polymorphism. Title: Blood copper in organophosphate-induced delayed polyneuropathy Lotti M, Caroldi S, Moretto A Ref: Toxicol Lett, 41:175, 1988 : PubMed ESTHER: Lotti_1988_Toxicol.Lett_41_175 PubMedSearch: Lotti 1988 Toxicol.Lett 41 175 PubMedID: 2835831 Abstract Some organophosphorous esters cause a polyneuropathy which becomes clinically evident 2 weeks after a single dose. The pathogenesis involves modifications of a target protein, neuropathy target esterase, in the axons and a selective inhibition of retrograde axonal transport. It was suggested that copper metabolism might also be involved because of increased levels of plasma copper and ceruloplasmin in animals developing this polyneuropathy. Our results do not confirm this observation; treatment of hens with highly neuropathic single doses of two organophosphates (dihexyl-2,2-dichlorovinyl phosphate and mono-o-cresyl diphenyl phosphate) does not affect total and plasma free copper when measured several times during the development of polyneuropathy. We concluded that copper homeostasis is not affected and that copper changes are unlikely to be involved in the pathogenesis of this polyneuropathy. Title: Organ distribution of neuropathy target esterase in man Moretto A, Lotti M Ref: Biochemical Pharmacology, 37:3041, 1988 : PubMed ESTHER: Moretto_1988_Biochem.Pharmacol_37_3041 PubMedSearch: Moretto 1988 Biochem.Pharmacol 37 3041 PubMedID: 3395378 Title: Central-peripheral delayed neuropathy caused by diisopropyl phosphorofluoridate (DFP): segregation of peripheral nerve and spinal cord effects using biochemical, clinical, and morphological criteria Lotti M, Caroldi S, Moretto A, Johnson MK, Fish CJ, Gopinath C, Roberts NL Ref: Toxicol Appl Pharmacol, 88:87, 1987 : PubMed ESTHER: Lotti_1987_Toxicol.Appl.Pharmacol_88_87 PubMedSearch: Lotti 1987 Toxicol.Appl.Pharmacol 88 87 PubMedID: 3564033 Abstract Systemic injection of diisopropyl phosphorofluoridate (DFP; 1 mg/kg, sc) causes delayed neuropathy in hens. This effect is associated with a high level of organophosphorylation of neuropathy target esterase (NTE) followed by an intramolecular rearrangement called "aging." Phenylmethanesulfonyl fluoride (PMSF) also attacks the active center of NTE but "aging" cannot occur. This compound does not cause neuropathy and protects against a subsequent challenge systemic dose of DFP. Intraarterial injection of DFP (0.185 mg/kg) into only one leg of hens caused a high NTE inhibition (greater than 80%) in the sciatic nerve of the injected leg, but not in other parts of the nervous system (37% average). A unilateral neuropathy with typical histopathological lesions developed in the injected leg. PMSF (0.55 mg/kg) injected into each sciatic artery caused 47% inhibition of sciatic nerve NTE but only 17-22% inhibition of NTE elsewhere; it did not produce clinical or histopathological lesions. When these hens were challenged with DFP (1 mg/kg, sc), high inhibition of residual-free NTE (greater than 85%) occurred throughout the nervous system and clinical signs of a syndrome different from the classical delayed neuropathy developed: this spinal cord type of ataxia was associated with histopathological lesions in the spinal cord but not in peripheral nerve. PMSF (1 mg/kg) injected into only one sciatic artery caused selective protective inhibition of sciatic nerve NTE of that leg. After systemic challenge by DFP, clinical effects expressed were a combination of spinal cord ataxia plus unilateral peripheral neuropathy. The challenge dose of DFP (1 mg/kg, sc) was insufficient to produce clear histopathological lesions in unprotected peripheral nerves although spinal lesions were found in these hens. Thus clinical evaluation of the peripheral nervous system by means of walking tests and a simple test of "leg retraction" reflexes was more sensitive and specific in diagnosis of peripheral neuropathy than was the histopathology. Title: Progressive deficit of retrograde axonal transport is associated with the pathogenesis of di-n-butyl dichlorvos axonopathy Moretto A, Lotti M, Sabri MI, Spencer PS Ref: Journal of Neurochemistry, 49:1515, 1987 : PubMed ESTHER: Moretto_1987_J.Neurochem_49_1515 PubMedSearch: Moretto 1987 J.Neurochem 49 1515 PubMedID: 2444671 Inhibitor(s) related to this paper: Dibutyl-dichlorovinyl-phosphate~DBVP Abstract The induction of central-peripheral distal axonopathy in hens singly dosed with some organophosphorus (OP) compounds, such as di-n-butyl-2,2-dichlorovinyl phosphate (DBDCVP), requires greater than 80% organophosphorylation and subsequent intramolecular rearrangement ("aging") of a protein [neuropathy target esterase (NTE)] in the axon. Suprathreshold biochemical reaction, 24 h after dosing with DBDCVP (0.75-1.00 mg/kg s.c.), is shown to be associated with progressive decrement of retrograde axonal transport in sensory and motor fibers. The maximum transport deficit (about 70% reduction) is reached 7 days after DBDCVP, prior to the appearance of axonal degeneration and the onset of clinical signs of neuropathy (day 10-11). By contrast, phenylmethylsulfonyl fluoride (30 mg/kg s.c.), an agent that prevents the development of OP neuropathy by inhibiting NTE without the "aging" reaction, had no effect on axon transport, nerve fiber integrity, or clinical status and, when administered prior to a neurotoxic dose of DBDCVP (1.00 mg/kg s.c.), prevented DBDCVP effects. Paraoxon (0.2 mg/kg s.c.) neither inhibited NTE nor caused deficits in retrograde transport or neuropathy. Taken in concert, these studies demonstrate that induced deficits in retrograde transport are associated with the pathogenesis of OP-induced nerve-fiber degeneration and the threshold-initiating mechanism thereof. Title: Inhibition of lymphocytic neuropathy target esterase predicts the development of organophosphate-induced delayed polyneuropathy Lotti M, Moretto A, Zoppellari R, Dainese R, Rizzuto N, Barusco G Ref: Archives of Toxicology, 59:176, 1986 : PubMed ESTHER: Lotti_1986_Arch.Toxicol_59_176 PubMedSearch: Lotti 1986 Arch.Toxicol 59 176 PubMedID: 2434058 Abstract Neuropathy Target Esterase (NTE) is the molecular target in the nervous system for organophosphorus esters (OP) when they cause delayed polyneuropathy. Some NTE activity was recently found also in blood lymphocytes. An unsuccessful suicide attempt with the widely used pesticide chlorpyrifos (0,0-diethyl-0-3,5,6,-trichloro-2-pyridyl phosphorothioate) is reported, where prior inhibition of lymphocytic NTE correlates with the delayed development of polyneuropathy. A 42-year-old man drank approximately 300 mg/kg chlorpyrifos. The subsequent severe cholinergic syndrome lasted for 17 days with varying degrees of severity. Thirty days after intoxication the clinical and electrophysiological examination of the peripheral nervous system was normal but lymphocytic NTE was about 60% inhibited. On day 43 the patient began to complain of paresthesia and leg weakness. Clinical examination, electrophysiology and a nerve biopsy revealed signs of a peripheral polyneuropathy, axonal in type. This case report indicates that measurement of lymphocytic NTE might be used as a clinical test to predict the development of OP-induced delayed polyneuropathy. Title: Poster 89. Neurotoxic esterase in extranervous tissues of man Moretto A, Fassina A, Lotti M Ref: In: Cholinesterases, fundamental and applied aspects : proceedings of the Second International Meeting on Cholinesterases, (Brzin M, Barnard EA, Sket D, Eds) De Gruyter:, 1984 : PubMed | |||||||
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