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Spatial control of oxygen delivery to 3D cultures alters cancer cell growth and gene expression

ABSTRACT: Commonly used monolayer cell cultures lack the capacity to provide a physiologically relevant environment for cell culture in terms of cell-cell architecture, extracellular matrix composition, and spatiotemporal delivery of key growth factors and small molecules, such as oxygen. Here, we describe a three-dimensional (3D) approach to cell culture in vitro, utilizing a bioreactor system designed to control oxygenation of 3D cancer cell cultures, in order to better mimic tumor microenvironments observed in vivo. We found transcriptomic differences in breast and ovarian cancer cell cultures grown in traditional monolayer cultures as compared to cultures grown in a Matrigel three-dimensional matrix. We also investigated the transcriptomes of 3D cultures grown in 21% O2, 3% O2, and a gradient of 3% O2 to 0% O2 using our bioreactor system. By controlling oxygen delivery, we observed differences in cell growth morphology and transcriptome regulation under the three conditions.

ORGANISM(S): Homo sapiens

PROVIDER: GSE129111 | GEO | 2019/04/02

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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Spatial control of oxygen delivery to 3D cultures alters cancer cell growth and gene expression

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