Controlled formation of heterotypic hepatic micro-organoids in anisotropic hydrogel microfibers for long-term preservation of liver-specific functions
Controlled formation of heterotypic hepatic micro-organoids in anisotropic hydrogel microfibers for long-term preservation of liver-specific functions
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DOI：
10.1016/j.biomaterials.2012.07.068
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发表时间：
2012-11-01
期刊：
影响因子：
14
通讯作者：
Seki, Minoru
中科院分区：
文献类型：
--
作者：
Yamada, Masumi;Utoh, Rie;Seki, Minoru
我们已经开发了一种基于水凝胶的细胞培养平台，用于形成由原代大鼠肝细胞和饲养细胞（Swiss 3 T3细胞）组成的3D网状肝脏微类器官。将含有肝细胞/3 T3细胞的海藻酸钠溶液连续引入微流体通道中以产生掺入细胞的各向异性Ba-海藻酸盐水凝胶微纤维，其中中心的肝细胞被3 T3细胞紧密夹在中间。在高氧张力下基于水凝胶纤维的培养能够形成异型微型类器官，其长度可达1 mm，直径类似于50 μ m，模仿肝脏中发现的肝索结构，同时在30天内保持高肝细胞活力（类似于80%）。长达90天的长期观察显示，由于异型和同型细胞的相互作用，包括白蛋白分泌和尿素合成以及肝细胞特异性基因的表达，与传统的单层培养和水凝胶纤维中的单一培养相比，肝功能显著增强。通过使用藻酸盐裂解酶酶促消化水凝胶基质，将包封的肝构建体回收为无支架的微型类器官。这种用于创建具有精确有序的多种细胞类型的异型微型类器官的技术将有助于开发新的肝脏组织工程方法，并可能适用于体外生物人工肝（BAL）装置的制造和药物开发和测试的评估工具。(C)2012爱思唯尔有限公司保留所有权利。
We have developed a hydrogel-based cell cultivation platform for forming 3D restiform hepatic micro-organoids consisting of primary rat hepatocytes and feeder cells (Swiss 3T3 cells). Sodium alginate solutions containing hepatocytes/3T3 cells were continuously introduced into a microfluidic channel to produce cell-incorporating anisotropic Ba-alginate hydrogel microfibers, where hepatocytes at the center were closely sandwiched by 3T3 cells. Hydrogel fiber-based cultivation under high oxygen tension enabled the formation of heterotypic micro-organoids with a length of up to 1 mm and a diameter of similar to 50 mu m, mimicking the hepatic cord structures found in the liver, while maintaining a high hepatocyte viability (similar to 80%) over 30 days. Long-term observation of up to 90 days revealed a significant enhancement of hepatic functions because of heterotypic and homotypic cell cell interactions, including albumin secretion and urea synthesis as well as expression of hepatocyte-specific genes, compared with conventional monolayer culture and single cultivation in the hydrogel fibers. The encapsulated hepatic constructs were recovered as scaffold-free micro-organoids by enzymatically digesting the hydrogel matrices using alginate lyase. This technique for creating heterotypic micro-organoids with precisely ordered multiple cell types will be useful for the development of a new liver tissue engineering approach and may be applicable to the fabrication of extracorporeal bioartificial liver (BAL) devices and assessment tools for drug development and testing. (C) 2012 Elsevier Ltd. All rights reserved.