**7. Relation between TiNSs-caused vacuoles and endosomal pathway**

Previous studies have reported formation of vacuoles originating from endocytic organelles, where the vacuolization of late endosomes was induced by the inhibition of kinases for regulation of vesicular transport and sorting [45–47], and several bacterial toxins induced vacuoles having an endosomal/lysosomal origin [48–50]. To examine the relation between TiNS-induced vacuoles and the endosomal pathway, monocyte-derived adherent cells were incubated with fluorescence-dye-labeled dextran to visualize endosome structures. The adherent cells were prepared using a pre-culture of PBMCs. TiNSs were then added to the culture of endosome-visualized adherent cells at 10 μg/ml, and observations were then made of vacuoles using the fluorescence derived from dextran. In the control culture without TiNSs, a diffuse fluorescence was observed. However, exposure to TiNSs induced vacuoles in the adherent cells, where most of the vacuoles showed co-localization with the fluorescence derived from endocytosed dextran. These observations indicate that TiNS-induced formation of vacuoles is related with some part of the endosomal pathway.

> that this type of cell death does not require caspases [54, 55]. Some of the machineries mentioned above might be related to the cell death caused by TiNSs. TiNSs in the endosomal pathway might affect immune functions executed by cell surface receptors through interference of endosomal

> **Figure 3.** Illustration of a comparison of volume and surface area between nanosphere and nanosheet. A nanosphere with

30-nm diagonals and depth of 1 nm has the same density as a nanosphere. Fourteen nanosheets have almost the same

and surface area 1257 nm2

suggests possible alteration of immune responses in a direct manner caused by TiNSs. Further investigation concerning these issues will contribute to a clarification of the toxic machinery of

The authors thank Ms. Tamayo Hatayama, Shoko Yamamoto and Miho Ikeda for their technical help. The authors declare that there is no conflict of interests regarding the publication of

and Takemi Otsuki1

, Naoko Kumagai-Takei1

, Suni Lee1

. A diamond-shaped nanosheet with 20- and

as the sum of both sides, which are 6.68 times as large

Toxicity of Titanate Nanosheets on Human Immune Cells

http://dx.doi.org/10.5772/intechopen.72234

171

,

lymphocytes were also damaged by exposure to TiNSs, which

trafficking. Additionally, CD4<sup>+</sup>

as the surface area of the nanosphere.

a diameter of 20 nm has a volume of 4189 nm3

**Acknowledgements**

this chapter.

**Author details**

Yasumitsu Nishimura1

Hidenori Matsuzaki1

TiNSs and the immunotoxicological effects of TiNSs.

volume as a nanosphere but possess a total surface area of 8400 nm2

\*, Daisuke Yoshioka2

1 Department of Hygiene, Kawasaki Medical School, Kurashiki, Japan

2 Department of Natural Sciences, Kawasaki Medical School, Kurashiki, Japan

, Kei Yoshitome1

\*Address all correspondence to: yas@med.kawasaki-m.ac.jp
