Carbon black and titanium dioxide nanoparticles induce pro-inflammatory responses in bronchial epithelial cells: need for multiparametric evaluation due to adsorption artifacts.

The initiation of an inflammatory process is the main adverse effect observed following the exposure of the airway epithelium to nanoparticles (NPs). This study was designed to explore the pro-inflammatory potential of two different NPs of similar size but of different compositions (CB 13 nm and TiO(2) 15 nm) on a human bronchial epithelial cell line (16HBE14o-). The expression of granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin (IL-6), and tumor necrosis factor alpha (TNFalpha) was evaluated in terms of mRNA, intracellular proteins, and released cytokines. Exposure to NPs induced a dose-dependent expression of all these cytokines, depending upon the chemical composition of NPs. The released cytokines appeared to be an inaccurate methodology to evaluate the pro-inflammatory response. Indeed, NPs adsorbed cytokines, and the binding was dependent on the nature of both the cytokine and NPs. Furthermore, addition of fetal calf serum or bovine serum albumin improved the detection of cytokines but also reduced cellular responses. Use of different detergents (Tween, Triton, and NP40) demonstrated limited efficiency to desorb cytokines from NPs. Thus, this study demonstrated the pro-inflammatory potential for CB and TiO(2) NP but underlines the methodological artifacts faced during the in vitro evaluation of cytokine release that necessitates a multiparametric evaluation.