Functional behavior of primary rat liver cells in a three-dimensional perfused microarray bioreactor
Functional behavior of primary rat liver cells in a three-dimensional perfused microarray bioreactor
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DOI：
10.1089/107632702760184745
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发表时间：
2002-06-01
影响因子：
--
通讯作者：
Griffith, LG
中科院分区：
文献类型：
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作者：
Powers, MJ;Janigian, DM;Griffith, LG
我们之前已经描述了一种微制造生物反应器的设计和操作，该反应器支持肝细胞的灌注3D培养，并促进组织样形态结构的进化。在这里，我们描述了在这个微阵列生物反应器中维持的细胞的功能活力，并与静态培养相比，研究了不同的播种方案对结构和功能进化的影响。原代大鼠肝细胞在分离后立即作为单细胞悬浮液或在2至3天内形成球形聚集体植入灌注反应器。在最初的研究中，细胞在放置后培养7天，结果显示，与单细胞直接播种相比，在反应器中播种前预聚集3天的细胞具有更大的功能活性和形态稳定性。在单细胞反应器培养中，总白蛋白分泌和尿素生成率在初始培养期间显著下降，而在预聚集反应器培养中保持不变。长期研究表明，白蛋白分泌率和尿素生成率在15天内保持在恒定水平。这些代谢率比在可比培养基比例和交换条件下静态培养的相同预聚集结构的代谢率高一个数量级。光镜图像显示存活的组织结构，电镜图像显示紧密连接、糖原储存和胆管支持代谢功能数据。
We have previously described the design and operation of a microfabricated bioreactor that supports perfused 3D culture of liver cells and facilitates evolution of tissue-like morphological structures. Here, we describe the functional viability of cells maintained in this microarray bioreactor and examine the influence of different seeding protocols on the evolution of structure and function in comparison with static culture. Primary rat hepatocytes were seeded into the perfusion reactors either as single-cell suspensions immediately after isolation or as spheroidal aggregates formed over a 2- to 3-day period. Initial studies in which cells were cultured for 7 days postisolation revealed significantly greater functional activity and morphological stability of cells that were preaggregated for up to 3 days before seeding in the reactor, compared with direct seeding of single cells. Total albumin secretion and urea genesis rates in single-cell reactor cultures declined significantly during this initial culture period while remaining constant in preaggregated reactor cultures. Longer term studies indicate that rates of albumin secretion and urea genesis are maintained at constant levels through 15 days postisolation. These metabolic rates are an order of magnitude higher than observed for the same preaggregated structures cultured statically with comparable medium ratio and exchange conditions. The metabolic function data are supported by light microscopy images showing viable tissue structures, and electron microscopy images that reveal tight junctions, glycogen storage, and bile canaliculi.