Interference of engineered nanoparticles with in vitro toxicity assays
Interference of engineered nanoparticles with in vitro toxicity assays
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DOI：
10.1007/s00204-012-0837-z
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发表时间：
2012-07-01
影响因子：
6.1
通讯作者：
Schnekenburger, Juergen
中科院分区：
文献类型：
--
作者：
Kroll, Alexandra;Pillukat, Mike Hendrik;Schnekenburger, Juergen
准确的纳米颗粒细胞毒性体外评估需要仔细选择测试系统。由于高吸附能力和光学活性，工程纳米颗粒在影响经典的细胞毒性检测方面具有很大的潜力。在这里，四种常见的体外氧化应激、细胞活力、细胞死亡和炎性细胞因子产生的检测方法(DCF、四甲基偶氮唑盐、乳酸脱氢酶和IL-8酶联免疫吸附试验)在24个特征良好的工程纳米颗粒中进行了有效性评估。对所有纳米粒子可能对光学检测方法的干扰、底物转化能力、对酶活性的影响以及与促炎细胞因子结合的可能性进行了详细的分析。根据所使用的检测系统，结果会有很大的差异。当进行DCF、四甲基偶氮唑蓝和乳酸脱氢酶检测时，所有被测试的纳米颗粒都在50微克厘米(-2)或以上的浓度下干扰光学测量。除碳黑外，可以通过改变检测程序和将颗粒浓度降低到10微克厘米(-2)来防止颗粒干扰。研究还发现，在无细胞体系中，炭黑能够氧化H_2DCF-DA，而只有纳米氧化锌显著降低LDH活性。在测试的三种二氧化钛颗粒中，只有一种观察到免疫反应IL-8的显著丧失。我们的结果表明，工程纳米颗粒以高度浓度、颗粒和检测特异性的方式干扰经典的细胞毒性检测。这些发现强烈表明，每个体外测试系统都必须针对每一种单一的纳米颗粒类型进行评估，以准确地评估纳米颗粒的毒性。
Accurate in vitro assessment of nanoparticle cytotoxicity requires a careful selection of the test systems. Due to high adsorption capacity and optical activity, engineered nanoparticles are highly potential in influencing classical cytotoxicity assays. Here, four common in vitro assays for oxidative stress, cell viability, cell death and inflammatory cytokine production (DCF, MTT, LDH and IL-8 ELISA) were assessed for validity using 24 well-characterized engineered nanoparticles. For all nanoparticles, the possible interference with the optical detection methods, the ability to convert the substrates, the influence on enzymatic activity and the potential to bind proinflammatory cytokines were analyzed in detail. Results varied considerably depending on the assay system used. All nanoparticles tested were found to interfere with the optical measurement at concentrations of 50 mu g cm(-2) and above when DCF, MTT and LDH assays were performed. Except for Carbon Black, particle interference could be prevented by altering assay protocols and lowering particle concentrations to 10 mu g cm(-2). Carbon Black was also found to oxidize H2DCF-DA in a cell-free system, whereas only ZnO nanoparticles significantly decreased LDH activity. A dramatic loss of immunoreactive IL-8 was observed for only one of the three TiO2 particle types tested. Our results demonstrate that engineered nanoparticles interfere with classic cytotoxicity assays in a highly concentration-, particle- and assay-specific manner. These findings strongly suggest that each in vitro test system has to be evaluated for each single nanoparticle type to accurately assess the nanoparticle toxicity.