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Biofabrication. 2011 Sep;3(3):034111. doi: 10.1088/1758-5082/3/3/034111. Epub 2011 Aug 11.

Avidin-biotin-based approach to forming heterotypic cell clusters and cell sheets on a gas-permeable membrane.

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Institute of Industrial Science, University of Tokyo, 4-6-1 Komaba, Meguro-ku, Tokyo 153-8505, Japan.


Implantation of sheet-like liver tissues is a promising method in hepatocyte-based therapies, because angiogenesis is expected to occur upon implantation from the surrounding tissues. In this context, we introduce here a new methodology for the formation of a functional thick hepatic tissue usable for cell sheet technology. First, we report the formation of composite tissue elements in suspension culture. Composite elements were composed of human hepatoma Hep G2 cells and mouse NIH/3T3 fibroblasts which are important modulators for thick-tissue formation. To overcome the very low attachment and organization capability between different cells in suspension, we synthesized a new cell-to-cell binding molecule based on the avidin-biotin binding system that we previously applied to attach hepatocytes on artificial substrata. This newly synthesized biotin-conjugated biocompatible anchoring molecule was inserted in the plasma membrane of both cell types. NIH/3T3 cells were further conjugated with avidin and incubated with biotin-presenting Hep G2 cells to form highly composite tissue elements. Then, we seeded those elements on highly gas-permeable membranes at their closest packing density to induce the formation of a thick, composite, functional hepatic tissue without any perfusion. This methodology could open a new way to engineer implantable thick liver tissue sheets where different cell types are spatially organized and well supplied with oxygen.

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