Department of Surgery 1
Fukushima Medical University
Development of Novel Cell Sheet Technology; Hybrid islet cell sheets Laminated with Mesenchymal Stem Cells
Hirofumi Shimizu1, Hiroyuki Hanayama1, Rie Utoh2, Takahiro Saito1, Teruo Okano2, Mitsukazu Gotoh1.
1Department of Regenerative Surgery, Fukushima Medical University, Fukushima, Japan; 2 Institute of Advanced Biomedical Engineering and Science, Tokyo Women’s Medical University, Tokyo, Japan
Background: We have shown that monolayer islet cell sheets obtained by temperature-responsive culture dishes restore normoglycemia indefinitely when grafted in the subcutaneous space of STZ-induced diabetic SCID mice. It was contrasted with the observation that the same number of dispersed islet single cells (10.0 x 10^6: islets obtained from 5 rats) failed to do so. Mesenchymal stem cells (MSCs) have been demonstrated to behave as feeder cells supporting culture cells and also to provide comfortable environments to composite graft in vivo. In this study, we examine a novel approach of the islet cell sheets laminated with MSCs sheet in order to advance the therapeutic effects of islet cell sheet technology.
Methods: 0.5 x 10^5 MSCs, obtained from bone marrow of Fisher 344 rats, were seeded onto the temperature-responsive culture dishes (35mm diameter), and reached confluency after 5 days of culture. After then, dissociated islet cells, 2.0 x 10^6 (n=4), 1.0 x 10^6 (n=4) and 0.5 x 10^6 (n=4) cells, obtained from Wistar rats were seeded onto MSCs. After 2 days of culture, islet cells and MSCs were found to be laminated and harvested easily as a composite sheet, and were subsequently transplanted into subcutaneous space of diabetic SCID mice. For control, relative large number of 5.0 x 10^6 islet single cells were infused into the subcutaneous space of diabetic SCID mice (n=6).
Results: The islet cells and MSCs in laminated sheet kept their cell-to-cell connections even after detachment from the temperature-responsive culture dishes (Fig1). It took only a few minutes to transplant composite sheet under the skin without losing any islet cells during the procedure. It was easy to handle and contrasted with the previous method using a fragile single islet sheet. All animals given 2.0 and 1.0 x 10^6 cells with MSCs enjoyed restoration of normo-glycemia within 5 days postgrafting (Fig 2). Two of 4 mice given 0.5 x 10^6 islet cells with MSCs finally restored normoglycemia. Meanwhile none of control animals restored normoglycemia. Histological analyses showed well granulated islet tissue mixed with MSCs in the subcutaneous space at day 60 postgrafting.
Conclusions: The present bioengineering techniques using MSCs as laminated sheet for islet sheet technology may offer a novel modality for quite efficient glycemic control of islet cells grafted in the subcutaneous space of recipient mice.
15:30 - 16:30
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