Mixed inhibition of adenosine deaminase activity by 1,3-dinitrobenzene: a model for understanding cell-selective neurotoxicity in chemically-induced energy deprivation syndromes in brain.
Mixed inhibition of adenosine deaminase activity by 1,3-dinitrobenzene: a model for understanding cell-selective neurotoxicity in chemically-induced energy deprivation syndromes in brain.
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1,3-二硝基苯对腺苷脱氨酶活性的混合抑制：了解化学诱导的大脑能量剥夺综合征中细胞选择性神经毒性的模型。
DOI：
10.1093/toxsci/kfr317
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发表时间：
2012
期刊：
影响因子：
--
通讯作者：
Philbert,MartinA
中科院分区：
文献类型：
--
作者：
Wang,Yipei;Liu,Xin;Schneider,Brandon;Zverina,ElainaA;Russ,Kristen;Wijeyesakere,SanjeevaJ;Fierke,CarolA;Richardson,RudyJ;Philbert,MartinA
星形胶质细胞对1，3-二硝基苯（1，3-DNB）非常敏感，而邻近的神经元相对不受影响，这与其他化学诱导的能量剥夺综合征一致。以前的研究已经调查了星形胶质细胞通过腺苷释放在保护神经元免受缺氧和化学损伤中的作用。腺苷被认为是神经保护性的，但它被细胞外脱氨酶如腺苷脱氨酶（ADA）迅速清除。本研究验证了ADA被1，3-DNB作为底物模拟物抑制，从而防止腺苷catalysis的假设。1，3-DNB抑制ADA，IC_（50）为284μM，Hill斜率，n= 4.8 ± 0.4。天然凝胶电泳显示1，3-DNB不使ADA变性。此外，加入Triton X-100（0.01- 0.05%，wt/vol）、Nonidet P-40（0.0015- 0.0036%，wt/vol）或牛血清白蛋白（0.05 mg/ml）或改变[ADA]（0.2和2 nM）基本上不会改变1，3-DNB的IC 50值。同样，动态光散射显示在149-1043μM的（1，3-DNB）范围内未形成颗粒。动力学显示1，3-DNB与ADA（KI= 520 ± 100μM，n= 1 ± 0.6）和ADA-腺苷复合物（KIS= 262 ± 7μM，n= 6 ± 0.6，表明正协同性）结合的混合抑制。与动力学雅阁，对接预测结合的1，3-DNB的活性位点和三个周边网站。此外，DI TNC-1星形胶质细胞暴露于10-500μM 1，3-DNB后，24 h细胞外腺苷浓度依赖性增加。总之，结果表明，1，3-DNB是ADA的混合抑制剂，因此可能导致细胞外腺苷增加。这一发现可能为指导未来化学诱导能量剥夺的工作提供见解。
Astrocytes are acutely sensitive to 1,3-dinitrobenzene (1,3-DNB) while adjacent neurons are relatively unaffected, consistent with other chemically-induced energy deprivation syndromes. Previous studies have investigated the role of astrocytes in protecting neurons from hypoxia and chemical injury via adenosine release. Adenosine is considered neuroprotective, but it is rapidly removed by extracellular deaminases such as adenosine deaminase (ADA). The present study tested the hypothesis that ADA is inhibited by 1,3-DNB as a substrate mimic, thereby preventing adenosine catabolism. ADA was inhibited by 1,3-DNB with an IC50of 284μM, Hill slope,n= 4.8 ± 0.4. Native gel electrophoresis showed that 1,3-DNB did not denature ADA. Furthermore, adding Triton X-100 (0.01–0.05%, wt/vol), Nonidet P-40 (0.0015–0.0036%, wt/vol), or bovine serum albumin (0.05 mg/ml or changing [ADA] (0.2 and 2nM) did not substantially alter the 1,3-DNB IC50value. Likewise, dynamic light scattering showed no particle formation over a (1,3-DNB) range of 149–1043μM. Kinetics revealed mixed inhibition with 1,3-DNB binding to ADA (KI= 520 ± 100μM,n= 1 ± 0.6) and the ADA-adenosine complex (KIS= 262 ± 7μM,n= 6 ± 0.6, indicating positive cooperativity). In accord with the kinetics, docking predicted binding of 1,3-DNB to the active site and three peripheral sites. In addition, exposure of DI TNC-1 astrocytes to 10–500μM 1,3-DNB produced concentration-dependent increases in extracellular adenosine at 24 h. Overall, the results demonstrate that 1,3-DNB is a mixed inhibitor of ADA and may thus lead to increases in extracellular adenosine. The finding may provide insights to guide future work on chemically-induced energy deprivation.
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