二氧化钛纳米颗粒与乳酸脱氢酶的结合。
Binding of titanium dioxide nanoparticles to lactate dehydrogenase.
机构信息
Department of Occupational Toxicology, Institute of Industrial Ecological Sciences, University of Occupational and Environmental Health, Yahatanishi-ku, Kitakyushu 807-8555, Japan.
出版信息
Environ Health Prev Med. 2012 Jul;17(4):341-5. doi: 10.1007/s12199-011-0245-7. Epub 2011 Oct 13.
OBJECTIVE
Measurement of released lactate dehydrogenase (LDH) activity, a commonly used marker of lethal cell injury in both in vitro and in vivo screenings, has been used to assess the cytotoxicity of nanoparticles (NPs), chemical compounds, and environmental factors. We have recently demonstrated that titanium dioxide (TiO₂) particles bind to several serum proteins. In the present study we investigated the binding of TiO₂ NPs to LDH.
METHODS
Purified LDH was incubated with TiO₂ NPs at 37°C for 1 h. The particles were then sedimented by centrifugation, and the activity and quantity of LDH in the supernatant and precipitated fraction were analyzed.
RESULTS
Incubation with TiO₂ reduced the LDH activity in the supernatant in a dose-dependent manner, while LDH activity in the precipitated fraction increased in a dose-dependent manner. Moreover, sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed a TiO₂ dose-dependent reduction in the quantity of LDH protein in the supernatant and an increase of LDH in particulate re-suspensions.
CONCLUSIONS
These findings, although based on a purified form of LDH, suggest that TiO₂ NPs bind to LDH, and consequently, TiO₂ NP-induced toxicity could be underestimated by the LDH activity assay.
目的
在体外和体内筛选中，释放的乳酸脱氢酶（LDH）活性的测量常被用作衡量致命细胞损伤的指标，用于评估纳米颗粒（NPs）、化学化合物和环境因素的细胞毒性。我们最近证明了二氧化钛（TiO₂）颗粒会与几种血清蛋白结合。在本研究中，我们研究了 TiO₂ NPs 与 LDH 的结合。
方法
将纯化的 LDH 与 TiO₂ NPs 在 37°C 下孵育 1 小时。然后通过离心沉淀颗粒，并分析上清液和沉淀部分中 LDH 的活性和数量。
结果
与 TiO₂ 孵育以剂量依赖性方式降低上清液中的 LDH 活性，而沉淀部分中的 LDH 活性则以剂量依赖性方式增加。此外，十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分析显示上清液中 LDH 蛋白的量随 TiO₂ 剂量的降低而减少，而在重新悬浮的颗粒中 LDH 则增加。
结论
这些发现虽然基于纯化的 LDH，但表明 TiO₂ NPs 与 LDH 结合，因此，LDH 活性测定可能低估了 TiO₂ NP 诱导的毒性。
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