Controlled cell morphology and liver-specific function of engineered primary hepatocytes by fibroblast layer cell densities

Yusuke Sakai, Makiko Koike, Daisuke Kawahara, Hideko Hasegawa, Tomomi Murai, Kosho Yamanouchi, Akihiko Soyama, Masaaki Hidaka, Mitsuhisa Takatsuki, Fumihiko Fujita, Tamotsu Kuroki, Susumu Eguchi

研究成果: Contribution to journalArticle査読

7 被引用数 (Scopus)

抄録

Engineered primary hepatocytes, including co-cultured hepatocyte sheets, are an attractive to basic scientific and clinical researchers because they maintain liver-specific functions, have reconstructed cell polarity, and have high transplantation efficiency. However, co-culture conditions regarding engineered primary hepatocytes were suboptimal in promoting these advantages. Here we report that the hepatocyte morphology and liver-specific function levels are controlled by the normal human diploid fibroblast (TIG-118 cell) layer cell density. Primary rat hepatocytes were plated onto TIG-118 cells, previously plated 3 days before at 1.04, 5.21, and 26.1 x 10(3) cells/cm(2). Hepatocytes plated onto lower TIG-118 cell densities expanded better during the early culture period. The hepatocytes gathered as colonies and only exhibited small adhesion areas because of the pushing force from proliferating TIG-118 cells. The smaller areas of each hepatocyte result in the development of bile canaliculi. The highest density of TIG-118 cells downregulated albumin synthesis activity of hepatocytes. The hepatocytes may have undergone apoptosis associated with high TGF-beta 1 concentration and necrosis due to a lack of oxygen. These occurrences were supported by apoptotic chromatin condensation and high expression of both proteins HIF-la and HIF-lb. Three types of engineered hepatocyte/fibroblast sheets comprising different TIG-118 cell densities were harvested after 4 days of hepatocyte culture and showed a complete cell sheet format without any holes. Hepatocyte morphology and liver-specific function levels are controlled by TIG-118 cell density, which helps to design better engineered hepatocytes for future applications such as in vitro cell-based assays and transplantable hepatocyte tissues. (C) 2018, The Society for Biotechnology, Japan. All rights reserved.
本文言語英語
ページ(範囲)249-257
ページ数9
ジャーナルJournal of Bioscience and Bioengineering
126
2
DOI
出版ステータス出版済み - 8 2018

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