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Global gene expression changes including drug metabolism and disposition induced by three-dimensional culture of HepG2

ABSTRACT: We found constitutive upregulation and higher degree induction of drug metabolism and disposition-related genes in a three-dimensional HepG2 culture. The upregulated genes are those believed to be regulated by different regulatory factors. The global gene expression analysis by Affymetrix GeneChip indicated that altered expressions of microtubule-related genes may change expressed levels of drug metabolism and disposition genes. Stabilization of the microtubule molecules with docetaxel, a tubulin stabilizing agent, in the two-dimensional culture showed gene expression patterns similar to those in the three-dimensional culture, indicating that culture environment affects drug metabolism functions in HepG2 cells. Keywords: radial flow bioreactor cell culture system, three dimensional culture, HepG2, GeneChip U133A,

ORGANISM(S): Homo sapiens

PROVIDER: GSE12939 | GEO | 2009/02/24

SECONDARY ACCESSION(S): PRJNA110961

REPOSITORIES: GEO

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Project description:The development of more complex but reliable systems for compound testing in a pharmaceutical context is a challenging task to date. Three-dimensional (3D), organ mimetic cell culture is aiming to become an alternative to common two-dimensional (2D) cell culture or animal testing in that field. We developed a biocompatible 3D cell culture environment for a hepatocellular carcinoma (HCC) model that enables cellular maintenance in a polycarbonate scaffold structure. Albumin, regarded as a differentiation marker, was elevated in statically 3D cultivated HepG2 cells. Expression of HCC tumor marker alpha-fetoprotein (AFP) was reduced compared to immunofluorescence stainings of 2D cultivated cells. Remarkably, expression of cytokeratin and pathophysiologically relevant beta-1 integrin (ITGB1) was found enhanced in nonperfused 3D cell culture. Changes in gene expression induced by the 3D cultivation environment were investigated using Ingenuity Pathway Analysis (IPA). Our findings revealed involvement of the insulin growth factor (IGF) signaling pathway in upregulation of matrix metalloproteinases (MMP) and ITGB1. The experimental data indicate a more differentiated state in 3D cultivated HepG2 cells than in the respective 2D experiments. Hence, scaffold-supported 3D cultivation of HepG2 cells may lead to a gain of information valuable for both drug testing and cancer research. HepG2 cells were cultivated for five days under 2D and 3D statical and perfused conditions. Cultivation was started with 0.25x10^6 cells in 2D and with 1x10^6 vital cells for the 3D experiments. The day of seeding was defined as d0. The groups were classified as follows: 2D, i.e., monolayer cultures, 3D, i.e., statical 3D cultures and BR, which denotes perfused 3D culture of HepG2 cells. The perfusable bioreactor system was operated using a peristaltic pump. It houses the MatriGrid, a polycarbonate-based microporous cellular support. For 3D static cultivation, cell-inoculated MatriGrids were placed in wells of a 24-well plate. Microarray experiments of three 2D (i.e., control), three 3D statically and three actively perfused 3D cultivations were performed at SIRS-Lab GmbH (SIRS-Lab GmbH, Jena, Germany) according to the manufacturer's instructions (Illumina, San Diego, CA). Altogether, 9 RNA samples of hepatocyte cultures and an internal control RNA were hybridized on two HumanHT-12 v4 Expression BeadChips.

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Global gene expression changes including drug metabolism and disposition induced by three-dimensional culture of HepG2

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