Abstract
Titanate nanosheet (TiNS) is 2D material with thickness of about 1 nm, and it is expected to be applied for various kinds of industrial use. TiNS is composed of titanium and oxygen as well as titanium dioxide (TiO2) particles, but it has a unique feature of lepidocrocite-type crystal structure, which suggests different toxic effects. Therefore, we proceeded with the analyses for toxicity by cell culture experiments. TiNS caused caspase-dependent apoptosis of human monocytes with giant vacuoles, in which titanium was identified. TiNS induced the expression of autophagy-related genes, but the results suggested a toxic mechanism through abnormally increased lysosomal function. Recently, the significance of lysosome-derived Ca2+ signals has been pointed out. Therefore, we focused on the lysosomal Ca2+ release channel of transient receptor potential mucolipin (TRPML). It was found that the monocytes exposed to TiNS showed increases in intracellular Ca2+ concentration and mRNA levels of v-ATPase and TRPML1 genes following incorporation of TiNS. Additionally, it was also confirmed that Ca2+ binds on TiNS. ML-SA1, which works on release of Ca2+ from lysosomes as TRPML agonist, increased apoptosis of monocytes in the culture with TiNS. The intracellular Ca2+ chelating agent BAPTA-AM suppressed the increases in the mRNA levels of lysosomal genes. It is known that lysosomal cell death is dependent on catepsin (CTS), but CTS inhibitor exacerbated apoptosis caused by TiNS exposure. Those findings indicate that the increase in TRPML1 channel augmented lysosome-derived Ca2+ signals, which induced the expression of the lysosomal genes, leading to a positive feedback loop that amplify the Ca2+ signal and then causes apoptosis in monocytes. The toxic effects of TiNS due to its unique crystalline structure and three-dimensional feature illustrate a new mechanism of immunotoxicity by metal nanomaterials.
