Scaffold-free three-dimensional cell culture utilizing micromolded nonadhesive hydrogels
Scaffold-free three-dimensional cell culture utilizing micromolded nonadhesive hydrogels
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DOI：
10.2144/000112591
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发表时间：
2007-10-01
期刊：
影响因子：
2.7
通讯作者：
Morgan, Jeffrey R.
中科院分区：
文献类型：
--
作者：
Napolitano, Anthony P.;Dean, Dylan M.;Morgan, Jeffrey R.
使细胞自组装成三维（3-D）球体微组织的技术提供了强大的体外模型，这些模型正变得越来越受欢迎，特别是在干细胞研究、组织工程和癌症生物学等领域。不幸的是，涉及规模，费用，几何形状和实用性的警告阻碍了这些技术的广泛采用。我们提出了一种易于使用，廉价，可扩展的技术，用于生产复杂形状的3-D微组织。利用各种原代细胞和永生细胞系来证明该技术适用于人类细胞类型，并突出它们自组装现象的差异。当接种到微成型的非粘附性琼脂糖凝胶上时，细胞会沉降到凹槽中，其结构优化了自发自组装所需的细胞间相互作用。通过一个移液步骤，我们能够创建数百个均匀的球状体，其大小由接种密度决定。多细胞肿瘤球状体（MCTS）由单细胞组装或生长，并且使用改良的4-[3-（4-碘苯基）-2-（4-硝基苯基）-2H-5-四唑]-1.3-苯二磺酸盐（WST-1）测定来定量它们的增殖。复杂形状（例如，蜂窝状）的同质或混合细胞群的微组织可以很容易地产生，为3D组织培养开辟了新的可能性。
Techniques that allow cells to self-assemble into three-dimensional (3-D) spheroid microtissues provide powerful in vitro models that are becoming increasingly popular-especially in fields such as stem cell research, tissue engineering, and cancer biology. Unfortunately, caveats involving scale, expense, geometry, and practicality have hindered the widespread adoption of these techniques. We present an easy-to-use, inexpensive, and scalable technology for production of complex-shaped, 3-D microtissues. Various primary cells and immortal cell lines were utilized to demonstrate that this technique is applicable to man), cell types and highlight differences in their self-assembly phenomena. When seeded onto micromolded, nonadhesive agarose gels, cells settle into recesses, the architectures of which optimize the requisite cell-to-cell interactions for spontaneous self-assembly With one pipeting step, we were able to create hundreds of uniform spheroids whose size was determined by seeding density. Multicellular tumor spheroids (MCTS) were assembled or grown from single cells, and their proliferation was quantified using a modified 4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1.3-benzene disulfonate (WST-1) assay. Complex-shaped (e.g., honeycomb) microtissues of homogeneous or mixed cell populations can be easily produced, opening new possibilities for 3-D tissue culture.