The size‐dependent apoptotic effect of titanium dioxide nanoparticles on endothelial cells by the intracellular pathway

Abstract

Concerns over the health risk of the widely distributed, commonly used titanium dioxide nanoparticles (nano-TiO ) are increasing worldwide. Yet, up-to-now, our understanding in their potential effects on the cardiovascular system is very limited and the toxicological mechanisms are still unclear. In the present study, the CCK-8 assay was performed to determine the cytotoxicity of four sizes (10, 30, 50, and 100 nm) of anatase nano-TiO on human umbilical vein endothelial cells (HUVECs) in culture, and the flow cytometry was employed to investigate the potential of these nano-TiO to induce the apoptosis of HUVECs. The apoptotic pathway was also probed through the determination of the protein expression and activation of p53, Bax, Bcl-2, caspases-9, -7, -3, and PARP by western blot. The results showed that at the administrative levels (1, 5, 25 μg/mL), all the four sizes of nano-TiO could significantly inhibit the viability of HUVECs and elicit significant apoptosis in them, compared with the negative control (P < .05, P < .01). Moreover, the apoptotic rates of HUVECs were increased respectively with the elevating levels and decreasing sizes of the administrative nano-TiO , showing a clear dose- and size-dependent effect relationships. Interestingly, the increasing phosphorylation of p53, decreasing ratio of Bcl-2/Bax, and enhancing activation of the downstream proteins caspase-9, -7, -3, and PARP, were also observed with the decreasing sizes of the administrative nano-TiO in the western blot, indicating that the intracellular approach of apoptosis, the p53-caspase pathway, is the major way of the nano-TiO -mediated apoptosis in HUVECs in culture and that the size is an important parameter that may determine the potential of nano-TiO to induce cellular response. In conclusion, these results suggested that high levels of nano-TiO exposure may pose potential risks to human cardiovascular health by inducing cardiovascular EC apoptosis.

The size‐dependent genotoxic potentials of titanium dioxide nanoparticles to endothelial cells

“…Four sizes [T100 (100 ± 14.3 nm), T50 (50 ± 7.6 nm), T30 (30 ± 5.1 nm), and T10 (10 ± 2.3 nm)] of anatase TiO 2 ‐NPs were provided by the National Center for Nanoscience and Technology, Chinese Academy of Sciences (China) . The size and morphology of the used NPs were described in our previous study .…”

“…Four sizes [T100 (100 ± 14.3 nm), T50 (50 ± 7.6 nm), T30 (30 ± 5.1 nm), and T10 (10 ± 2.3 nm)] of anatase TiO 2 ‐NPs were provided by the National Center for Nanoscience and Technology, Chinese Academy of Sciences (China) . The size and morphology of the used NPs were described in our previous study . Chemicals for cell culture, including MEM with Earle's salts, fetal bovine serum (FBS), l‐glutamin, penicillin/streptomycin, 4‐(2‐hydroxyethyl)‐1‐piperazineethanesulfonic acid (HEPES), and phosphate buffered solution (PBS), were bought from Hyclone (GE Healthcare Life Science, Beijing), and other chemicals were from Sigma (Sigma‐Aldrich Co., Beijing).…”

“…The culture conditions for HUVECs were described in our previous work . A brief description is presented here.…”

“…This highlights the need to study the genotoxicity of ENPs in various types of cells on a case‐by‐case basis. Our previous work has found that TiO 2 ‐NPs could induce a significant apoptosis in human umbilical vein endothelial cells (HUVECs) in culture . But, in this cellular pathological process, it is still unclear whether these NPs have posed a genotoxicity to HUVECs or not.…”

“…In the present study, the genotoxic potential of four sizes of anatase TiO 2 ‐NPs to HUVECs was determined with the alkaline comet assay and cytokinesis‐block micronucleus (CBMN) test. The intracellular redox statuses of HUVECs exposed to these NPs were also explored through the measurement of the levels of reactive oxygen species (ROS) production and reduced glutathione (GSH), respectively.…”