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Open Access Preconditioned Cell Array Optimized for a Three-Dimensional Culture of Hepatocytes

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Three-dimensional culture procedures have attracted attention in various fields of cell biology. A newly developed cell array assisted in the formation of hepatocyte spheroids by two innovations: 1) micropatterning by a hydrophilic polymer, and 2) the use of bovine carotid artery-derived HH cells as feeder cells. The former contributes to the standardization of the spheroid size and the latter to the maintenance of the spheroids. We created a way to provide a ready-to-use cell array by cryopreservation of an HH feeder cell cultured array. After inoculation of HH cells on the cell array, the culture medium was replaced by freezing medium containing dimethyl sulfoxide. Thereafter, the array was frozen and stored in a −80°C deep freezer. At the start of the hepatocyte culture, the cryopreserved HH cell array was thawed by adding warmed (37°C) culture medium. The morphology and biological activities of the cryopreserved HH cells were intact, as confirmed by phase contrast microscopy and functional staining with calcein and formazan. The rat hepatocytes formed perfect spheroids on the cryopreserved HH cell array without any differences from those on the freshly prepared HH cell array. The CYP3A drug metabolism activities of the hepatocytes were well maintained on the cryopreserved and fresh cell arrays. The present protocol greatly shortened the time and labor required to prepare a cell array for culturing hepatocytes.

Keywords: Cell array; Cryopreservation; Heptocytes; Three-dimensional culture

Document Type: Research Article

Affiliations: Department of Innovative Surgery, National Research Institute for Child Health and Development, Tokyo, Japan

Publication date: 2009-05-01

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  • Cell Transplantation publishes original, peer-reviewed research and review articles on the subject of cell transplantation and its application to human diseases. To ensure high-quality contributions from all areas of transplantation, separate section editors and editorial boards have been established. Articles deal with a wide range of topics including physiological, medical, preclinical, tissue engineering, and device-oriented aspects of transplantation of nervous system, endocrine, growth factor-secreting, bone marrow, epithelial, endothelial, and genetically engineered cells, among others. Basic clinical studies and immunological research papers are also featured. To provide complete coverage of this revolutionary field, Cell Transplantation will report on relevant technological advances, and ethical and regulatory considerations of cell transplants. Cell Transplantation is now an Open Access journal starting with volume 18 in 2009, and therefore there will be an inexpensive publication charge, which is dependent on the number of pages, in addition to the charge for color figures. This will allow work to be disseminated to a wider audience and also entitle the corresponding author to a free PDF, as well as prepublication of an unedited version of the manuscript.
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