Garcia-Pérez Adolfo G, Barril José, Estévez Jorge, Vilanova Eugenio
Division de Toxicologia, Instituto de Bioingeniería, Universidad Miguel Hernández de Elche, Avenida del Ferrocarril s/n. E-03202 Alicante, Spain.
Toxicol Lett. 2003 Apr 30;142(1-2):1-10. doi: 10.1016/s0378-4274(02)00469-1.
Chicken serum, the usual in vivo animal for testing organophosphorus delayed neuropathy, has long been reported not to contain a homologous activity of the neuronal neuropathy target esterase (NTE) activity when it is assayed according to standard methods as the phenyl valerate esterase (PVase) activity, which is resistant to paraoxon and sensitive to mipafox. However, a PVase activity (1000-1500 nmol/min/ml) can be measured in serum that is extremely sensitive to both paraoxon, a non-neuropathic organophosphorus compound and mipafox, a model neuropathy inducer. The inhibition was time progressive in both cases, suggesting a covalent phosphorilating reaction. The fixed time inhibition curves suggest at least two sensitive components. The IC50 for 30 min, at 37 degrees C are 6 and 51 nM for paraoxon and 4 and 110 nM for mipafox, for every sensitive component. When paraoxon was removed from a serum sample pretreated with the inhibitor, the paraoxon sensitive PVase activity was recovered, in spite of showing a time progressive inhibition suggesting that hydrolytic dephosphorylating reaction recovered at a significant rate. The reactivation of the phosphorylated enzyme could explain that the time progressive inhibitions curves for long time with paraoxon tend to reach a plateau depending on the inhibition concentration. However, with mipafox, the curve approached the same maximal inhibitions at all concentrations as expected for a permanent covalent irreversible phosphorylation, which is coherent with the observations that the activity remained inhibited after removing the inhibitor. Data of serum esterases described in this paper showed similar properties to those previously reported for peripheral nerve soluble phenylvalerate esterase: (1) extremely high sensitivity to paraoxon and mipafox; (2) time progressive kinetic with two sensitive components; (3) recovery of activity after removal of paraoxon; and (4) permanent inhibition with mipafox. These properties of serum esterases are very similar to those of soluble fraction of peripheral nerves. So, serum PVases could be considered as appropriate biomarkers, as a mirror for the neural soluble paraoxon and mipafox sensitive soluble esterases that could be used for biomonitoring purpose.
鸡血清是用于测试有机磷迟发性神经病的常用体内动物，长期以来的报告显示，按照标准方法将其作为对氧磷抗性且对丙胺氟磷敏感的苯基戊酸酯酶（PVase）活性进行测定时，鸡血清中不含有神经元神经病靶酯酶（NTE）活性的同源活性。然而，在血清中可以检测到一种PVase活性（1000 - 1500 nmol/分钟/毫升），它对非神经性有机磷化合物对氧磷和模型神经病诱导剂丙胺氟磷都极为敏感。在这两种情况下，抑制作用都是随时间进展的，表明存在共价磷酸化反应。固定时间抑制曲线表明至少有两个敏感成分。在37℃下30分钟的IC50，对氧磷和丙胺氟磷的每个敏感成分分别为6 nM和51 nM以及4 nM和110 nM。当从用抑制剂预处理的血清样品中去除对氧磷时，尽管显示出随时间进展的抑制作用，但对氧磷敏感的PVase活性得以恢复，这表明水解去磷酸化反应以显著速率恢复。磷酸化酶的再活化可以解释为什么对氧磷长时间的随时间进展的抑制曲线会根据抑制浓度趋于达到一个平台期。然而，对于丙胺氟磷，曲线在所有浓度下都接近相同的最大抑制，这是永久性共价不可逆磷酸化所预期的，这与去除抑制剂后活性仍被抑制的观察结果一致。本文中描述的血清酯酶数据显示出与先前报道的外周神经可溶性苯基戊酸酯酶相似的性质：（1）对对氧磷和丙胺氟磷极高的敏感性；（2）具有两个敏感成分的随时间进展的动力学；（3）去除对氧磷后活性恢复；（4）丙胺氟磷导致永久性抑制。血清酯酶的这些性质与外周神经可溶性部分的性质非常相似。因此，血清PVases可被视为合适的生物标志物，作为神经可溶性对氧磷和丙胺氟磷敏感的可溶性酯酶的一种反映，可用于生物监测目的。
