Collagen vitrigel promotes hepatocytic differentiation of induced pluripotent stem cells into functional hepatocyte-like cells
Shun Nakai,
Ima Shibata,
Takahiro Shitamichi,
Hiroyuki Yamaguchi,
Nobuyuki Takagi,
Tomoaki Inoue,
Toshito Nakagawa,
Jumpei Kiyokawa,
Satoshi Wakabayashi,
Tomoya Miyoshi,
Eriko Higashi,
Seiichi Ishida,
Nobuaki Shiraki,
Shoen Kume
Affiliations
Shun Nakai
School of Life Science and Technology, Tokyo Institute of Technology, 4259-B-25 Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8501, Japan
Ima Shibata
School of Life Science and Technology, Tokyo Institute of Technology, 4259-B-25 Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8501, Japan
Takahiro Shitamichi
School of Life Science and Technology, Tokyo Institute of Technology, 4259-B-25 Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8501, Japan
Hiroyuki Yamaguchi
Isehara Research Laboratory, Technology and Development Division, Kanto Chemical Co., Inc., 21 Suzukawa, Isehara, Kanagawa 259-1146, Japan
Nobuyuki Takagi
Technology and Development Division, Kanto Chemical Co., Inc., 2-1, Nihonbashi Muromachi 2-chome, Chuo-ku, Tokyo 103-0022, Japan
Tomoaki Inoue
Research Division, Chugai Pharmaceutical Co. Ltd, 1-135 Komakado, Gotemba, Shizuoka 412-8513, Japan
Toshito Nakagawa
Research Division, Chugai Pharmaceutical Co. Ltd, 1-135 Komakado, Gotemba, Shizuoka 412-8513, Japan
Jumpei Kiyokawa
Research Division, Chugai Pharmaceutical Co. Ltd, 1-135 Komakado, Gotemba, Shizuoka 412-8513, Japan
Satoshi Wakabayashi
Pharmacokinetics and Metabolism, Drug Safety and Pharmacokinetics Laboratories, Taisho Pharmaceutical Co., Ltd, 1-403 Yoshino-cho, Saitama-shi, Saitama 330-8530, Japan
Tomoya Miyoshi
Toxicology and Pharmacokinetics Laboratories, Pharmaceutical Research Laboratories, Toray Industries, Inc., 6-10-1 Tebiro, Kamakura, Kanagawa 248-8555, Japan
Eriko Higashi
Toxicology and Pharmacokinetics Laboratories, Pharmaceutical Research Laboratories, Toray Industries, Inc., 6-10-1 Tebiro, Kamakura, Kanagawa 248-8555, Japan
Seiichi Ishida
Division of Pharmacology, National Institute of Health Science, 3-25-26 Tonomati, Kawasaki 210-9501, Japan
Nobuaki Shiraki
School of Life Science and Technology, Tokyo Institute of Technology, 4259-B-25 Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8501, Japan
Shoen Kume
School of Life Science and Technology, Tokyo Institute of Technology, 4259-B-25 Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8501, Japan
Differentiation of stem cells to hepatocytes provides an unlimited supply of human hepatocytes and therefore has been vigorously studied. However, to date, the stem cell-derived hepatocytes were suggested to be of immature features. To obtain matured hepatocytes from stem cells, we tested the effect of culturing human-induced pluripotent stem (hiPS) cell-derived endoderm cells on collagen vitrigel membrane and compared with our previous reported nanofiber matrix. We cultured hiPS cell-derived endoderm cells on a collagen vitrigel membrane and examined the expression profiles, and tested the activity of metabolic enzymes. Gene expression profile analysis of hepatocytic differentiation markers revealed that upon culture on collagen vitrigel membrane, immature markers of AFP decreased, with a concomitant increase in the expression of mature hepatocyte transcription factors and mature hepatocyte markers such as ALB, ASGR1. Mature markers involved in liver functions, such as transporters, cytochrome P450 enzymes and phase II metabolic enzymes were also upregulated. We observed the upregulation of the liver markers for at least 2 weeks. Gene array profiling analysis revealed that hiPS cell-derived hepatocyte-like cells (hiPS-hep) resemble those of the primary hepatocytes. Functions of the CYP enzyme activities were tested in multi-institution and all revealed high CYP1A, CYP2C19, CYP2D6, CYP3A activity, which could be maintained for at least 2 weeks in culture. Taken together, the present approach identified that collagen vitrigel membrane provides a suitable environment for the generation of hepatocytes from hiPS cells that resemble many characteristics of primary human hepatocytes.
