Titanium dioxide nanoparticles induce mitochondria-associated apoptosis in HepG2 cells

Widespread applications of nanosized materials over the past decade have prompted investigations of desirable properties and potential hazards to humans and the environment. Titanium dioxide (TiO 2 ) nanoparticles are one of the most widely used nanoparticles. To investigate the effect of biological functions induced by TiO 2 nanoparticles (10 nm: TiO 2 NPs) on human liver cell lines, normal liver cell line L02 and hepatoma cell line HepG2 were co-cultured with exogenous TiO 2 NPs. Cell growth and proliferation, cell cycle, and the apoptosis rate were analyzed. The effects of TiO 2 NPs on the expression levels of apoptosis-associated protein caspase-3 and the membrane channel protein αENaC and caspase-3/7 activity were determined. Moreover, the influence of TiO 2 NPs on the expression levels of the mitochondria-related proteins SIRT3, VDAC1, and ACSS1, the mitochondrial membrane potential and the ADP/ATP ratio were also examined. Our results revealed that TiO 2 NPs inhibited the growth and proliferation of HepG2 cells, suppressed the S phase of cell cycling, and induced apoptosis of HepG2 cells. Following an increase in concentration, the inhibitory effect induced by TiO 2 NPs on proliferation and cell cycle was more evident, and the apoptosis rate increased in a significant concentration-dependent manner, whereas there was no significant effect on the growth, proliferation, apoptosis, and cell cycle of L02 cells. In addition, the results of western blot showed that in HepG2 cells, TiO 2 NPs upregulated the expressions of the apoptosis-related protein caspase-3 and the membrane channel protein αENaC in a concentration-dependent manner. However, in L02 cells, there was no significant difference in the expression levels of caspase-3 or αENaC. Furthermore, TiO 2 NPs induced depolarization of the mitochondrial membrane, upregulated the expression levels of the mitochondria-related proteins SIRT3 and VDAC1, and downregulated the expression level of the key respiratory chain protein ACSS1 in HepG2 cells. However, in L02 cells, the expressions of SIRT3, VDAC1, and ACSS1 exhibited no clear change. The apoptosis of HepG2 cells induced by TiO 2 NPs may be achieved by regulating intracellular osmotic pressure; moreover, upregulating the expression of the channel protein αENaC or the mitochondrial porin VDAC1 and depolarizing the mitochondrial membrane of HepG2 cells resulted in the loss of Cyt- c and ATP and further activated caspase-3. To further confirm the above r