Dose-dependent autophagic effect of titanium dioxide nanoparticles in human HaCaT cells at non-cytotoxic levels

Abstract

BackgroundInteractions between nanoparticles and cells are now the focus of a fast-growing area of research. Though many nanoparticles interact with cells without any acute toxic responses, metal oxide nanoparticles including those composed of titanium dioxide (TiO2-NPs) may disrupt the intracellular process of macroautophagy. Autophagy plays a key role in human health and disease, particularly in cancer and neurodegenerative diseases. We herein investigated the in vitro biological effects of TiO2-NPs (18 nm) on autophagy in human keratinocytes (HaCaT) cells at non-cytotoxic levels.ResultsTiO2-NPs were characterized by transmission electron microscopy (TEM) and dynamic light scattering techniques. Cellular uptake, as evaluated by TEM and NanoSIMS revealed that NPs internalization led to the formation of autophagosomes. TiO2-NPs treatment did not reduce cell viability of HaCaT cells nor increased oxidative stress. Cellular autophagy was additionally evaluated by confocal microscopy using eGFP-LC3 keratinocytes, western blotting of autophagy marker LC3I/II, immunodetection of p62 and NBR1 proteins, and gene expression of LC3II, p62, NBR1, beclin1 and ATG5 by RT-qPCR. We also confirmed the formation and accumulation of autophagosomes in NPs treated cells with LC3-II upregulation. Based on the lack of degradation of p62 and NBR1 proteins, autophagosomes accumulation at a high dose (25.0 μg/ml) is due to blockage while a low dose (0.16 μg/ml) promoted autophagy. Cellular viability was not affected in either case.ConclusionsThe uptake of TiO2-NPs led to a dose-dependent increase in autophagic effect under non-cytotoxic conditions. Our results suggest dose-dependent autophagic effect over time as a cellular response to TiO2-NPs. Most importantly, these findings suggest that simple toxicity data are not enough to understand the full impact of TiO2-NPs and their effects on cellular pathways or function.

The Role of Cell Autophagy in Toxicity Caused by Dental Nanomaterials

“…TiO 2 NPs mediated autophagy induction at low dose can be interpreted as a standard mechanism by which cells attempt to increase the clearance of NPs. While the halt in autophagy at high dose may indicate that the cellular response cannot cope with a NPs overload and thus show a deficient degradative capacity [40].Other studies showed that TiO 2 NPs could induce autophagy in normal lung cells and in primary human keratinocytes [65]. Based on the studies illustrated above, we hypothesize that TiO 2 NPs can induce autophagy dysfunctions in brain tissues and cells.…”

“…The mechanism is similar to carbon nano as we said above. The uptake of TiO 2 NPs led to a dose dependent increase in autophagic effect under non cytotoxic conditions, which has been demonstrated in Lopes's research [40]. Leaching of Cu ions, reactive oxygen species generation and autophagy appear to be the underlying mechanisms of Cu NP toxicity in lung cells [41].…”

“…TiO 2 NPs mediated autophagy induction at low dose can be interpreted as a standard mechanism by which cells attempt to increase the clearance of NPs. While the halt in autophagy at high dose may indicate that the cellular response cannot cope with a NPs overload and thus show a deficient degradative capacity [40].…”

Nanofeatured Titanium Surfaces for Dental Implantology: Biological Effects, Biocompatibility, and Safety

“…This process may cause peri-implant inflammatory reactions and cytotoxicity [232,233]. It has been suggested that a combined action of oxidative stress and "foreign-body" phenomena associated with the release of titanium nanoparticles from surface coatings may induce host immune responses, autophagic changes, and lysosomal dysfunction [234,235]. Furthermore, high concentrations of titanium dioxide nanoparticles have been shown to cause dose-dependent proinflammatory effects on human gingival fibroblasts [236].…”

Versatile Multicolor Nanodiamond Probes for Intracellular Imaging and Targeted Labeling

“…It is a known phenomenon that some types of nanoparticles induce a process of autophagy. [27][28][29] Typically, in autophagy process inside stressed cells is induced to degrade unnecessary or dysfunctional cellular components that can provide energy. This process is suppressed under normal physiological conditions.…”