Monolayer and Spheroid Culture of Human Liver Hepatocellular Carcinoma Cell Line Cells Demonstrate Distinct Global Gene Expression Patterns and Functional Phenotypes
Monolayer and Spheroid Culture of Human Liver Hepatocellular Carcinoma Cell Line Cells Demonstrate Distinct Global Gene Expression Patterns and Functional Phenotypes
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DOI：
10.1089/ten.tea.2007.0434
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发表时间：
2009-03-01
影响因子：
4.1
通讯作者：
Hughes-Fulford, Millie
中科院分区：
文献类型：
--
作者：
Chang, Tammy T.;Hughes-Fulford, Millie
了解三维（3D）生物结构的细胞生物学对于更完整地了解体内组织功能和推进离体器官工程工作非常重要。为了阐明3D结构如何影响肝细胞的细胞反应，我们比较了在组织培养皿（TCD）上单层培养或在旋转壁容器（RWV）生物反应器内球体培养的人肝细胞癌细胞系（HepG2）细胞的整体基因表达。在RWVs中生长的HepG2细胞在72小时内形成直径达100 mm的球状体，长期培养形成直径达1 mm的球状体。单层细胞的肌动蛋白细胞骨架显示应力纤维形成，而球状体具有皮质肌动蛋白组织。全球基因表达分析表明，上调的结构基因，如细胞外基质，细胞骨架，粘附分子在单层，而RWV球状体显示上调的代谢和合成基因，表明功能的差异。事实上，肝细胞色素P450活性和白蛋白生产的特定功能在球状体中更高。增强的肝功能需要维持3D结构和环境，因为球状体转移到TCD导致球状体解体和随后的功能丧失。这些发现说明了物理环境对细胞组织的重要性及其对肝细胞过程的影响。
Understanding cell biology of three-dimensional (3D) biological structures is important for more complete appreciation of in vivo tissue function and advancing ex vivo organ engineering efforts. To elucidate how 3D structure may affect hepatocyte cellular responses, we compared global gene expression of human liver hepatocellular carcinoma cell line (HepG2) cells cultured as monolayers on tissue culture dishes (TCDs) or as spheroids within rotating wall vessel (RWV) bioreactors. HepG2 cells grown in RWVs form spheroids up to 100 mm in diameter within 72 h and up to 1 mm with long-term culture. The actin cytoskeleton in monolayer cells show stress fiber formation while spheroids have cortical actin organization. Global gene expression analysis demonstrates upregulation of structural genes such as extracellular matrix, cytoskeletal, and adhesion molecules in monolayers, whereas RWV spheroids show upregulation of metabolic and synthetic genes, suggesting functional differences. Indeed, liver-specific functions of cytochrome P450 activity and albumin production are higher in the spheroids. Enhanced liver functions require maintenance of 3D structure and environment, because transfer of spheroids to a TCD results in spheroid disintegration and subsequent loss of function. These findings illustrate the importance of physical environment on cellular organization and its effects on hepatocyte processes.