Vuong Ngoc Q, Goegan Patrick, Mohottalage Susantha, Breznan Dalibor, Ariganello Marianne, Williams Andrew, Elisma Fred, Karthikeyan Subramanian, Vincent Renaud, Kumarathasan Premkumari
Inhalation Toxicology Laboratory, Environmental Health Science and Research Bureau, Health Canada, Ottawa, ON K1A 0K9, Canada; Department of Biochemistry, Faculty of Science, University of Ottawa, Ottawa, ON K1H 8M5, Canada.
Inhalation Toxicology Laboratory, Environmental Health Science and Research Bureau, Health Canada, Ottawa, ON K1A 0K9, Canada.
J Proteomics. 2016 Oct 21;149:53-63. doi: 10.1016/j.jprot.2016.03.046. Epub 2016 Apr 12.
This study combined cytotoxicity assays with proteomic analysis to characterize the unique biological responses of the A549 human lung epithelial cell line to two physicochemically distinct respirable particles titanium dioxide (TiO) and carbon black (CB). Cellular LDH, ATP, BrdU incorporation and resazurin reduction indicated that CB was more potent than TiO. Proteomic analysis was done using 2D-GE and MALDI-TOF-TOF-MS. Proteomic changes reflected common and particle-specific responses. Particle-specific proteomic responses were associated with cell death (necrosis and apoptosis), viability and proliferation pathways. Our results suggested that these pathways were consistent with the cytotoxicity data. For instance, increased expressions of anti-proliferative proteins LMNA and PA2G4 were in agreement with the decreased BrdU incorporation in A549 cells after exposure to CB. Similarly, increased expression of HSPA5 that is associated with ATPase activity was consistent with decreased cellular ATP levels in these cells. These findings reveal that proteomic changes can explain the cellular cytotoxicity characteristics of the particles. In essence, our results demonstrate that the in vitro toxicoproteomic approach is a promising tool to gain insight into molecular mechanisms underlying particle exposure-specific cytotoxicity.
In this study we have shown that toxicoproteomics is a sensitive and informative method to resolve the toxicity characteristics of particles with different physicochemical properties. This approach can be useful in the investigation of molecular mechanisms underpinning cellular cytotoxic responses elicited by particle exposures. Thus, the toxicoproteomic approach can be valuable in assessing the risk associated with particle exposures in vitro.
本研究将细胞毒性检测与蛋白质组学分析相结合，以表征A549人肺上皮细胞系对两种物理化学性质不同的可吸入颗粒二氧化钛（TiO）和炭黑（CB）的独特生物学反应。细胞乳酸脱氢酶（LDH）、三磷酸腺苷（ATP）、溴脱氧尿苷（BrdU）掺入和刃天青还原表明，炭黑比二氧化钛的毒性更强。使用二维凝胶电泳（2D-GE）和基质辅助激光解吸电离飞行时间质谱（MALDI-TOF-TOF-MS）进行蛋白质组学分析。蛋白质组学变化反映了共同的和颗粒特异性的反应。颗粒特异性蛋白质组学反应与细胞死亡（坏死和凋亡）、活力和增殖途径相关。我们的结果表明，这些途径与细胞毒性数据一致。例如，抗增殖蛋白核纤层蛋白A/C（LMNA）和前列腺凋亡反应蛋白4（PA2G4）的表达增加与暴露于炭黑后A549细胞中BrdU掺入减少一致。同样，与ATP酶活性相关的热休克蛋白A5（HSPA5）表达增加与这些细胞中细胞ATP水平降低一致。这些发现表明，蛋白质组学变化可以解释颗粒的细胞毒性特征。本质上，我们的结果表明，体外毒理蛋白质组学方法是深入了解颗粒暴露特异性细胞毒性潜在分子机制的一种有前途的工具。
在本研究中，我们表明毒理蛋白质组学是一种灵敏且信息丰富的方法，可解析具有不同物理化学性质颗粒 的毒性特征。这种方法可用于研究颗粒暴露引发的细胞毒性反应的潜在分子机制。因此，毒理蛋白质组学方法在评估体外颗粒暴露相关风险方面可能具有重要价值。
