Title: Comparative effectiveness of organophosphorus protoxicant activating systems in neuroblastoma cells and brain homogenates Barber D, Correll L, Ehrich M Ref: J Toxicol Environ Health, 57:63, 1999 : PubMed
The ability of bromine and rat liver microsomes (RLM) to convert organophosphorus (OP) protoxicants to esterase inhibitors was determined by measuring acetylcholinesterase (AChE) and neuropathy target esterase (NTE) inhibition. Species specific differences in susceptibility to esterase inhibition were determined by comparing the extent of esterase inhibition observed in human neuroblastoma cells and hen, bovine, and rodent brain homogenates. OP protoxicants examined included tri-o-tolyl phosphate (TOTP), O-ethyl O-p-nitrophenyl phenylphosphonothioate (EPN), leptophos, fenitrothion, fenthion, and malathion. Bromine activation resulted in greater AChE inhibition than that produced by RLM activation for equivalent concentrations of fenitrothion, malathion, and EPN. For EPN and leptophos, bromine activation resulted in greater inhibition of NTE than RLM. Only preincubation with RLM activated TOTP; resultant inhibition of AChE was less in hen brain (13 +/- 3%) than in neuroblastoma cells (73 +/- 1%) at 10(-6) M. In contrast, 10(-6) M RLM-activated TOTP produced more inhibition of hen brain NTE (89 +/- 6%) than NTE of human neuroblastoma cells (72 +/- 7%). Human neuroblastoma cells and brain homogenates from hens, the accepted animal model for study of OP-induced neurotoxicity, were relatively similar in sensitivity to esterase inhibition. Homogenates from hens were more sensitive to NTE inhibition induced by phenyl saligenin phosphate (PSP), an active congener of TOTP, than were homogenates from less susceptible species (mouse, rat, bovine). AChE of hen brain homogenates was also more sensitive than homogenates from other species to malaoxon, the active form of malathion.
Title: Inhibition of carboxylesterases in SH-SY5Y human and NB41A3 mouse neuroblastoma cells by organophosphorus esters Ehrich M, Correll L Ref: J Toxicol Environ Health, 53:385, 1998 : PubMed
Carboxylesterases (CbxE) can be inhibited by organophosphorus esters (OPs) without causing clinical evidence of toxicity. CbxE are thought to protect the critical enzyme acetylcholinesterase (AChE) from OP inhibition in animals. CbxE and AChE are both present in neuroblastoma cells, but, even though these cells have potential to be an in vitro model of OP toxicity, the effect of OPs on CbxE and the relationship of CbxE inhibition and AChE inhibition have not yet been examined in these cells. Therefore, this study examined concentration-related OP-induced inhibition of CbxE in human SH-SY5Y and mouse NB41A3 neuroblastoma cells with 11 active esterase inhibitors: paraoxon, malaoxon, chlorpyrifos-oxon, tolyl saligenin phosphate (TSP), phenyl saligenin phosphate (PSP), diisopropyl phosphorofluoridate (DFP), mipafox, dichlorvos, trichlorfon, dibutyryl dichlorovinyl phosphate (DBVP), and dioctyl dichlorovinyl phosphate (DOVP). All could inhibit CbxE, although the enzyme was less likely to be inhibited than AChE following exposure to 9 of the test compounds in the human cell line and to all 11 of the test compounds in the murine cell line. Species differences in concentration-related inhibitions of CbxE were evident. When cells were exposed first to an OP with a low IC50 toward CbxE (PSP), followed by an OP with high affinity for AChE (paraoxon or malaoxon), inhibitions of CbxE and AChE were additive. This indicated that CbxE did not protect AChE from OP-induced inhibition in this cell culture model.
Title: Acetylcholinesterase and neuropathy target esterase inhibitions in neuroblastoma cells to distinguish organophosphorus compounds causing acute and delayed neurotoxicity Ehrich M, Correll L, Veronesi B Ref: Fundamental & Applied Toxicology, 38:55, 1997 : PubMed
The differential inhibition of the target esterases acetylcholinesterase (AChE) and neuropathy target esterase (NTE, neurotoxic esterase) by organophosphorus compounds (OPs) is followed by distinct neurological consequences in exposed subjects. The present study demonstrates that neuroblastoma cell lines (human SH-SY5Y and murine NB41A3) can be used to differentiate between neuropathic OPs (i.e., those inhibiting NTE and causing organophosphorus-induced delayed neuropathy) and acutely neurotoxic OPs (i.e., those highly capable of inhibiting AChE). In these experiments, concentration-response data indicated that the capability to inhibit AChE was over 100x greater than the capability to inhibit NTE for acutely toxic, nonneuropathic OPs (e.g., paraoxon and malaoxon) in both cell lines. Inhibition of AChE was greater than inhibition of NTE, without overlap of the concentration-response curves, for OPs which are more likely to cause acute, rather than delayed, neurotoxic effects in vivo (e.g., chlorpyrifos-oxon, dichlorvos, and trichlorfon). In contrast, concentrations inhibiting AChE and NTE overlapped for neuropathy-causing OPs. For example, apparent IC50 values for NTE inhibition were less than 9.6-fold the apparent IC50 values for AChE inhibition when cells were exposed to the neuropathy-inducing OPs diisopropyl phosphorofluoridate, cyclic tolyl saligenin phosphate, phenyl saligenin phosphate, mipafox, dibutyl dichlorovinyl phosphate, and di-octyl-dichlorovinyl phosphate. In all cases, esterase inhibition occurred at lower concentrations than those needed for cytoxicity. These results suggest that either mouse or human neuroblastoma cell lines can be considered useful in vitro models to distinguish esterase-inhibiting OP neurotoxicants.
Title: Effect of laundering on ability of glove fabrics to decrease the penetration of organophosphate insecticides through in vitro epidermal systems Keeble VB, Correll L, Ehrich M Ref: Journal of Applied Toxicology, 16:401, 1996 : PubMed
Two knit glove fabrics, one of 100% cotton and one of 100% polypropylene, were examined for their capability to decrease the penetration of the organophosphate insecticides (OPs), azinphos-methyl and paraoxon after 4 h at field concentrations (3000 and 15 ppm, respectively) through an in vitro epidermal system (Skin2, Advanced Tissue Systems, LaJolla, CA). The glove fabrics were examined under three different conditions of use: new, after they had been abraded and after they had been abraded and then laundered. New and laundered cotton fabric was also examined for its capability to decrease the penetration of azinphos-methyl through another in vitro epidermal system (Epiderm, MatTek Corp., Ashland, MA), after 4 and 24 h of exposure. Capability of the media under the in vitro epidermal systems to inhibit brain acetylcholinesterase (AChE) was used as the indicator of penetration. Results were compared to OP-caused inhibitions seen in media under the fabric alone and in media under the in vitro epidermal systems alone. Incubations of azinphos-methyl suspensions and the in vitro epidermal systems covered with fabric indicated that both the epidermal cells and fabric provided protection against AChE inhibition caused by this OP and that the protective effects were additive, whether measured after 4 or 24 h of exposure. Therefore, neither laundering nor abrasion followed by laundering altered the capability of the in vitro epidermal systems to absorb azinphos-methyl suspension. For paraoxon solution, however, new cotton glove fabric prevented absorption, and this protective effect, noted after 4 h of exposure, was lost when the fabric was laundered. Abrading the fabric did not cause a greater effect than laundering alone. These results suggest that the pesticide as well as its formulation may be factors of consideration when protective fabrics are chosen, and that, for cotton glove fabric, the protection against some OPs may best be provided before the fabric is laundered.
Title: Catecholamine concentrations and contractile responses of isolated vessels from hens treated with cyclic phenyl saligenin phosphate or paraoxon in the presence or absence of verapamil McCain WC, Flaherty DM, Correll L, Jortner B, Ehrich M Ref: Journal of Toxicology & Environmental Health, 48:397, 1996 : PubMed
Blood samples and vascular segments from the ischiadic artery of hens treated with either cyclic phenyl saligenin phosphate (PSP; 2.5 micrograms/kg, im) or paraoxon (PXN; 0.1 micrograms/kg, im) in the presence or absence of verapamil, a calcium channel antagonist (7 micrograms/kg, im, given 4 consecutive days beginning the day before PSP or PXN administration), were examined 1, 3, 7, and 21 d after PSP or PXN administration in order to determine the contribution of catecholamines and peripheral blood vessel physiology and morphology to organophosphorus-induced delayed neuropathy (OPIDN). The levels of plasma catecholamines were measured by high-performance liquid chromatograpy (HPLC) and indicated a different effect with PSP, which causes OPIDN, and PXN, which does not. PSP treatment elevated the levels of norepinephrine and epinephrine throughout the study, while PXN treatment depressed the levels of these catecholamines. Verapamil treatment attenuated the OP response by approximately 50% for both compounds. Ischiadic vessel segments were isolated from OP-treated hens and perfused at a constant flow rate of 12 ml/min, then examined for their response to potassium chloride (KCl, 3 x 10(-3) M), acetylcholine (ACh), phenylephrine (PE), an alpha 1 adrenergic agonist, and salbutamol (SAL), a beta 2 adrenergic agonist. Agents were delivered in concentrations of 10(-8) to 10(-3) M. Vascular segments did not respond to ACh or SAL at any concentration used. Vessels displayed a significant reduction in contractile response to both KCl (3 x 10(-3) M) and PE (10(-8) to 10(-3) M) 3 and 21 d after exposure to either PSP or PXN. This reduced response was not altered by the presence of verapamil. Innervation of the peripheral vasculature was unchanged after OP treatment. This study indicates that plasma catecholamine levels could be differentially altered by treatment with OPs that do and do not cause OPIDN and suggests that the alterations involve intracellular calcium. In contrast, vascular response of the ischiadic artery was altered following OP treatment, but the effect was not specific for the neuropathy-inducing OP, PSP, and response was not mediated by Ca 2+, nor was it the result of autonomic nerve deterioration.
Title: Effect of beta-naphthoflavone on o-tolyl saligenin phosphate-induced delayed neuropathy in two lines of chickens Bursian SJ, Lehning EJ, Correll L, Ehrich M Ref: J Toxicol Environ Health, 28:461, 1989 : PubMed
The effect of the microsomal enzyme inducer beta-naphthoflavone (beta NF) on the development of organophosphorus-induced delayed neuropathy (OPIDN) was examined in two laboratories (VPI and MSU), utilizing two strains of White Leghorn hens. A single intraperitoneal injection of beta NF at 80 mg/kg body weight 48 h prior to administration of o-tolyl saligenin phosphate (TSP), the neuroactive metabolite of tri-o-tolyl phosphate (TOTP), caused a significant increase in hepatic microsomal cytochrome P-450 concentrations and aniline hydroxylase activities after 72 h in both strains. Hepatic carboxylesterase and cholinesterase activities were not affected by beta NF treatment in either strain. Administration of TSP in single subcutaneous doses of 20 and 25 mg/kg body weight (VPI) or 30 and 60 mg/kg body weight (MSU) caused significant inhibition of whole-brain neuropathy target esterase (NTE) activity 24 h postdosing, and hens subsequently developed clinical signs characteristics of OPIDN. beta NF had no significant effect on NTE inhibition or on initiation or severity of OPIDN clinical signs. However, OPIDN clinical signs were less severe in the strain of bird (MSU) with the higher intrinsic hepatic carboxylesterase activity and the higher beta NF-induced cytochrome P-450 concentration. The study indicates that microsomal enzyme induction, which has been shown to alleviate TOTP-induced delayed neuropathy, could not alleviate OPIDN resulting from exposure to TSP. This study also suggests that strain may affect susceptibility to TSP-induced delayed neuropathy.
