- Email: [email protected]
In vivo biomarkers of organophosphates and carbamates induced neurotoxicity
S6
Abstracts / Toxicology Letters 196S (2010) S1–S36
chemicals could sum up, resulting in an acceleration of colon carcinogenesis. Data supporting the different roles of inflammation in chemically induced colon carcinogenesis will be presented. doi:10.1016/j.toxlet.2010.03.040
S05 Biomarkers and Molecular Mechanisms in Pesticide Toxicity S05-1 In vivo biomarkers of organophosphates and carbamates induced neurotoxicity R. Gupta 1 , D. Milatovic 2 Murray State University, United States, 2 Vanderbilt University, United States
1
Currently, organophosphates (OPs) and carbamates (CMs) are the most widely used pesticides. Commonly referred to as “Anticholinesterase Pesticides” they illicit poisoning by acetylcholinesterase (AChE) inhibition at the synapses in brain and neuromuscular junctions. Inhibition of other serine-containing esterases (butyrylcholinesterase and carboxylesterase) also occurs in target and nontarget tissues; though not involved in mechanism of toxicity they do impact overall toxicity. The mechanisms involved in OP-/CM-induced neuronal damage/neurotoxicity appear to be linked with excitotoxicity (cholinergic and glutamatergic systems) and free radical mediated injury. Rats acutely exposed to an OP (DFP) or CM (carbofuran) at 1.5 mg/kg, sc, exhibited onset of signs within 5–10 min and maximal severity with hyper cholinergic preponderance within 30–60 min. At the time of maximal severity, brain analysis revealed profound inhibition of AChE activity (7590%), significant increases in markers of reactive oxygen species (ROS: F2-isoprostanes and F4-neuroprostanes; 2-6-times), reactive nitrogen species (RNS: citrulline; 4–8-times), neuroinflammation (PGE2, 2–3-times), and declines in high-energy phosphates (HEP: ATP, 36–60% and phosphocreatine, 28–52%). At the same time, quantitative morphometric analysis of pyramidal neurons of the hippocampal CA1 region revealed significant reductions in dendritic lengths and spine density. Brain regional heterogeneity exists to a great extent for some of these biomarkers. Pretreatment of rats with a NMDA receptor antagonist memantine (18 mg/kg, sc), spin-trapping antioxidant N-tert-butyl-phenylnitrone (200 mg/kg, ip) or chain-breaking antioxidant vitamin E (100 mg/kg, ip for 3 days) significantly attenuated DFP-/carbofuran-induced increases in markers of ROS and RNS, declines in HEP, dendritic length and spine density, by multiple mechanisms. doi:10.1016/j.toxlet.2010.03.043
S05-2 Plasma beta-glucuronidase as an alternative biomarker to cholinesterase for organophosphate exposure T. Satoh HAB Research Institute, United States Acetylcholinesterase and butyrylcholinesterase (BChE) activities in blood are traditionally used as the biomarkers for organophosphorus insecticide (OP) exposure. However, cholinesterase inhibition has several disadvantages such as individual variation. Recently, we proposed more sensitive biomarker of OP exposure than cholinesterase inhibition. Egasyn as one of carboxylesterase
isozymes is tightly bound to beta-glucuronidase(BG) to form egasyn-beta-glucuronidase(EG) complex at the liver microsomal membrane. When OPs are incorporated into the liver microsomes, they are tightly bound to egasyn, and subsequently, BG is dissociated and released into blood. In animal study, a single administration of OPs increased plasma BG activity in approximately 100-fold the control level in rats. In human study, we conducted human studies to evaluate plasma BG level, BChE activity and urinary OP metabolites in Malaysia and Japan. In Malaysian case, plasma BG activity among the farmers who were chronically exposed to OPs was significantly increased. Thus, inhibition of plasma cholinesterase activity was negatively correlated with the increase in plasma BG activity. In Japan, we assessed the relation among plasma BG level, plasma BChE and urinary OP metabolite level in three groups. It consisted of 32 controls (control), 21 pest control operators and their co-workers who had not sprayed OPs within 3 days prior to sample. Plasma BG activity was increased according to their OP exposure level. However, there was no change in BChE activity under the same condition. In conclusion, increase in plasma BG activity and measurement of urinary OP metabolites are much more useful biomarker than BChE inhibition for low-level exposure in humans. doi:10.1016/j.toxlet.2010.03.044
S05-3 Application of biomarkers of exposure, effect and susceptibility to subjects long-term exposed to pesticides A.F. Hernández University of Granada School of Medicine, Spain Pesticides represent a large and important class of chemicals showing acute and long-term toxicity in non target organisms, such as humans. The major results of our studies performed in plastic greenhouses workers from Southeast Spain on different categories of biomarkers (exposure, of effect and of susceptibility) will be addressed. Exposure was assessed by measuring serum cholinesterase (BChE) and red blood cell cholinesterase (AChE). Application of pesticides can result in exposure by either the dermal or respiratory route, resulting in low-grade depressions in both cholinesterases. Lifetime exposure to pesticides significantly decreased AChE in the exposed population. Significant lower levels of -aminolevullinic acid dehydratase (ALA-D) and AChE were observed in pesticide applicators as compared to controls (41.3 and 14.5%, respectively). A number of well-established measurements were assessed for early biological effects after pesticide exposure. New serum esterases, such as -glucuronidase and paraoxonase (PON1) as well as certain serum enzymes (aspartate aminotransferase, lactate dehydrogenase, amine oxidase) underwent significant changes in pesticide-exposed workers, which supports a subtle biochemical dysfunction resulting in cytotoxicity. Pesticide-induced oxidative stress may result from their biotransformation or toxic action in the body. We found that greenhouse workers were exposed to more oxidative stress as evidenced by changes in their antioxidant status (decreased levels of superoxide dismutase, catalase and glutathione reductase). The individual susceptibility to pesticides is caused by polymorphisms in biotransformation enzymes such as esterases, transferases and CYP450 s. Human serum PON1 hydrolyses organophosphates entering the blood circulation and tissues thus limiting toxicity. PON1-192R allele was associated with both a higher risk of BChE inhibition and a lesser risk of having a previous episode of acute pesticide poisoning. PON1-192R and null GSTT1 were associated
Abstracts / Toxicology Letters 196S (2010) S1–S36
chemicals could sum up, resulting in an acceleration of colon carcinogenesis. Data supporting the different roles of inflammation in chemically induced colon carcinogenesis will be presented. doi:10.1016/j.toxlet.2010.03.040
S05 Biomarkers and Molecular Mechanisms in Pesticide Toxicity S05-1 In vivo biomarkers of organophosphates and carbamates induced neurotoxicity R. Gupta 1 , D. Milatovic 2 Murray State University, United States, 2 Vanderbilt University, United States
1
Currently, organophosphates (OPs) and carbamates (CMs) are the most widely used pesticides. Commonly referred to as “Anticholinesterase Pesticides” they illicit poisoning by acetylcholinesterase (AChE) inhibition at the synapses in brain and neuromuscular junctions. Inhibition of other serine-containing esterases (butyrylcholinesterase and carboxylesterase) also occurs in target and nontarget tissues; though not involved in mechanism of toxicity they do impact overall toxicity. The mechanisms involved in OP-/CM-induced neuronal damage/neurotoxicity appear to be linked with excitotoxicity (cholinergic and glutamatergic systems) and free radical mediated injury. Rats acutely exposed to an OP (DFP) or CM (carbofuran) at 1.5 mg/kg, sc, exhibited onset of signs within 5–10 min and maximal severity with hyper cholinergic preponderance within 30–60 min. At the time of maximal severity, brain analysis revealed profound inhibition of AChE activity (7590%), significant increases in markers of reactive oxygen species (ROS: F2-isoprostanes and F4-neuroprostanes; 2-6-times), reactive nitrogen species (RNS: citrulline; 4–8-times), neuroinflammation (PGE2, 2–3-times), and declines in high-energy phosphates (HEP: ATP, 36–60% and phosphocreatine, 28–52%). At the same time, quantitative morphometric analysis of pyramidal neurons of the hippocampal CA1 region revealed significant reductions in dendritic lengths and spine density. Brain regional heterogeneity exists to a great extent for some of these biomarkers. Pretreatment of rats with a NMDA receptor antagonist memantine (18 mg/kg, sc), spin-trapping antioxidant N-tert-butyl-phenylnitrone (200 mg/kg, ip) or chain-breaking antioxidant vitamin E (100 mg/kg, ip for 3 days) significantly attenuated DFP-/carbofuran-induced increases in markers of ROS and RNS, declines in HEP, dendritic length and spine density, by multiple mechanisms. doi:10.1016/j.toxlet.2010.03.043
S05-2 Plasma beta-glucuronidase as an alternative biomarker to cholinesterase for organophosphate exposure T. Satoh HAB Research Institute, United States Acetylcholinesterase and butyrylcholinesterase (BChE) activities in blood are traditionally used as the biomarkers for organophosphorus insecticide (OP) exposure. However, cholinesterase inhibition has several disadvantages such as individual variation. Recently, we proposed more sensitive biomarker of OP exposure than cholinesterase inhibition. Egasyn as one of carboxylesterase
isozymes is tightly bound to beta-glucuronidase(BG) to form egasyn-beta-glucuronidase(EG) complex at the liver microsomal membrane. When OPs are incorporated into the liver microsomes, they are tightly bound to egasyn, and subsequently, BG is dissociated and released into blood. In animal study, a single administration of OPs increased plasma BG activity in approximately 100-fold the control level in rats. In human study, we conducted human studies to evaluate plasma BG level, BChE activity and urinary OP metabolites in Malaysia and Japan. In Malaysian case, plasma BG activity among the farmers who were chronically exposed to OPs was significantly increased. Thus, inhibition of plasma cholinesterase activity was negatively correlated with the increase in plasma BG activity. In Japan, we assessed the relation among plasma BG level, plasma BChE and urinary OP metabolite level in three groups. It consisted of 32 controls (control), 21 pest control operators and their co-workers who had not sprayed OPs within 3 days prior to sample. Plasma BG activity was increased according to their OP exposure level. However, there was no change in BChE activity under the same condition. In conclusion, increase in plasma BG activity and measurement of urinary OP metabolites are much more useful biomarker than BChE inhibition for low-level exposure in humans. doi:10.1016/j.toxlet.2010.03.044
S05-3 Application of biomarkers of exposure, effect and susceptibility to subjects long-term exposed to pesticides A.F. Hernández University of Granada School of Medicine, Spain Pesticides represent a large and important class of chemicals showing acute and long-term toxicity in non target organisms, such as humans. The major results of our studies performed in plastic greenhouses workers from Southeast Spain on different categories of biomarkers (exposure, of effect and of susceptibility) will be addressed. Exposure was assessed by measuring serum cholinesterase (BChE) and red blood cell cholinesterase (AChE). Application of pesticides can result in exposure by either the dermal or respiratory route, resulting in low-grade depressions in both cholinesterases. Lifetime exposure to pesticides significantly decreased AChE in the exposed population. Significant lower levels of -aminolevullinic acid dehydratase (ALA-D) and AChE were observed in pesticide applicators as compared to controls (41.3 and 14.5%, respectively). A number of well-established measurements were assessed for early biological effects after pesticide exposure. New serum esterases, such as -glucuronidase and paraoxonase (PON1) as well as certain serum enzymes (aspartate aminotransferase, lactate dehydrogenase, amine oxidase) underwent significant changes in pesticide-exposed workers, which supports a subtle biochemical dysfunction resulting in cytotoxicity. Pesticide-induced oxidative stress may result from their biotransformation or toxic action in the body. We found that greenhouse workers were exposed to more oxidative stress as evidenced by changes in their antioxidant status (decreased levels of superoxide dismutase, catalase and glutathione reductase). The individual susceptibility to pesticides is caused by polymorphisms in biotransformation enzymes such as esterases, transferases and CYP450 s. Human serum PON1 hydrolyses organophosphates entering the blood circulation and tissues thus limiting toxicity. PON1-192R allele was associated with both a higher risk of BChE inhibition and a lesser risk of having a previous episode of acute pesticide poisoning. PON1-192R and null GSTT1 were associated