No abstract

Human paraoxonase (PON1) is a polymorphic, high-density lipoprotein (HDL)-associated esterase that hydrolyzes the toxic metabolites of several organophosphorus (OP) insecticides and nerve agents. The activity polymorphism is determined by a Gln/Arg (Q/R) substitution at position 192. Injection of purified PON1 protects animals from OP poisoning. In the present study, we investigated the in-vivo function of PON1 for detoxifying organophosphorus insecticides in PON1-knockout mice that were challenged via dermal exposure with diazoxon, diazinon and paraoxon. PON1-knockout mice were extremely sensitive to diazoxon. Doses (2 and 4 mg/kg) that caused no cholinesterase (ChE) inhibition in wild-type mice were lethal to the knockout mice, which also showed slightly increased sensitivity to the parent compound diazinon. Surprisingly, these knockout mice did not show increased sensitivity to paraoxon. In-vitro assays indicated that the PON1R192 isoform hydrolyzed diazoxon less rapidly than did the PON1Q192 isoform. In-vivo analysis, where PON1-knockout mice received the same amount of either PON1(192) isoform via intraperitoneal (i.p.) injection 4 h prior to exposure, showed that both isoforms provided a similar degree of protection against diazoxon, while PON1R192 conferred better protection against chlorpyrifos-oxon than PON1Q192. Injection of purified rabbit PON1 or either human PON1(192) isoform did not protect PONI-knockout mice from paraoxon toxicity, nor did over-expression of the human PON1R192 transgene in wild-type mice. Kinetic analysis of the two human PON1(192) isoforms revealed that the catalytic efficiency (Vmax/Km) determines the in-vivo efficacy of PON1 for organophosphorus detoxication. The results indicate that PON1 plays a major role in the detoxication of diazoxon and chlorpyrifos oxon but not paraoxon.

Abstract-The paraoxonase (PON1) PON1-Q192R and PON1-L55M polymorphisms have been inconsistently associated with vascular disease. Plasma PON1 activity phenotypes vary markedly within genotypes and were, therefore, expected to add to the informativeness of genotype for predicting vascular disease. The case-control sample included 212 ageand race-matched men (mean age 66.4 years). The 106 carotid artery disease (CAAD) cases had Ͼ80% carotid stenosis, and the 106 controls had Ͻ15%. [15][16][17] The cardioprotective role of HDL, the inhibition or reduction of atherogenic LDL oxidation, appears to be, in large part, a function of PON1, which is associated with HDL. 18 -22 PON1 metabolizes mildly oxidized phospholipids, presumably by eliminating hydroperoxy derivatives of unsaturated fatty acids. 20 Thus, the PON1-CVD association is expected to result from the role of PON1 in the metabolism of bioactive lipid molecules and protection against damage due to oxidized LDL.PON1 hydrolyzes a variety of substrates, including the toxic components of the pesticides parathion, chlorpyrifos, and diazinon; aryl esters, such as phenyl acetate; and the nerve agents soman and sarin. There is 10-to 40-fold interindividual variability in rates of paraoxon hydrolysis. 23 The PON1 192Q allele has the higher rate of in vitro hydrolysis of diazoxon, sarin, and soman, 24 whereas the PON1 192R allele has higher activity for the hydrolysis of paraoxon and chlorpyrifos oxon. 24,25 These rates of substrate hydrolysis are quite variable within PON1 genotypes (at least 13-fold) and represent phenotypes that can add information about PON1 status beyond genotyping alone. 24 Paraoxon hydrolysis activity is lost in the plasma of the PON1 knockout mouse, and these mice are more susceptible to atherosclerosis. 26 We compared the PON1 192 and PON1 55 genotypes with PON1 rates of hydrolysis of paraoxon (POase activity) and diazoxon (DZOase activity) for their predictiveness in vascular disease of the carotid arteries. These 2 substrates were chosen because, relative to the other isoform, the PON1 192Q isoform has a higher DZOase activity and the PON1 192R isoform has a higher POase activity in the in vitro assays. The resulting 2D plot (Figure 1) allows an accurate inference of PON1 192 genotype, in addition to providing PON1 phenotype information. 27 Methods Sample