Thickness limitation and cell viability of multi-layered cell sheets and overcoming the diffusion limit by a porous-membrane culture insert
Thickness limitation and cell viability of multi-layered cell sheets and overcoming the diffusion limit by a porous-membrane culture insert
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多层细胞片的厚度限制和细胞活力以及通过多孔膜培养插入物克服扩散限制
DOI：
10.4172/2153-0777.s2-001
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发表时间：
2011
影响因子：
4.8
通讯作者：
T. Okano
中科院分区：
文献类型：
--
作者：
Waki Sekine;Y. Haraguchi;Tatsuya Shimizu;A. Umezawa;T. Okano
组织工程的一个主要挑战是创建三维（3D）和功能性组织。然而，组织缺血环境使得较厚的3D组织的生产变得困难。为了评估组织的厚度限制，研究了由多层细胞片组成的新型体外3D组织模型的细胞活力和代谢。人骨髓间充质细胞（EMC）培养在温度响应培养皿。仅通过降低温度将汇合培养的EMCs收获为完整的连续细胞片层。将获得的细胞片成功地分层成3D细胞致密组织，并在体外培养7天。培养基中的葡萄糖消耗和乳酸盐产生根据层数（单层至三层）增加。组织学分析和细胞活力测定表明，在单层至三层细胞片中发现了活组织，在超过四层的细胞片中发现了受损组织。结果得出结论，层叠电池片的厚度极限为约40 mm，这是三层电池片的厚度。重要的是，当在多孔膜上培养多层细胞片时，细胞片之间的粘附和细胞活力得到改善，从而成功地制造出比在正常培养皿上更厚的组织（约100 mm）。代谢分析表明，多层细胞片依赖于它们的厌氧代谢，这表明通过上部和底部组织表面提供营养物质而不是氧气提高了3D组织中的细胞活力。
A major challenge of tissue engineering is the creation of three-dimensional (3D) and functional tissues. However, the tissue ischemic environments make the production of thicker 3D tissues difficult. For evaluating the thickness limitation of tissue, the cell viability and metabolism of a novel in vitro 3D tissue model, which is composed of multi- layered cell sheets, were investigated. Human endometrial-derived mesenchymal cells (EMC) were cultured on temperature-responsive culture dishes. Confluently cultured EMCs were harvested as an intact contiguous cell sheet only by lowering temperature. The obtained cell sheets were successfully layered into 3D cell-dense tissues and cultured in vitro for 7 days. Glucose consumption and lactate production in the culture media increased in accordance with the number of layers (single to triple). Histological analyses and cell viability assays showed that viable tissues were found in single- to triple-layered cell sheets and damaged tissues in over quadruple layers. The results concluded that the thickness limitation of layered cell sheets was approximately 40 mm, which was the thickness of triple-layered cell sheets. Importantly, when multi-layered cell sheets were cultured on porous membranes, the adhesion among cell sheets and cell viability were improved, resulting in successful fabricating thicker tissues (~100 mm) than that on normal culture dishes. The metabolic analyses showed that multi-layered cell sheets rely on their anaerobic metabolism, indicating that supplying nutrients rather than oxygen through both upper and bottom tissue surfaces improved the cell viability in 3D tissues.
影响因子：
4.3
作者：
Berrios, Raul;Totterdell, Peter;Kellett, Stephen
通讯作者：
Kellett, Stephen