Gene expression of human mesenchymal stem cells in 3D cell culture
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE207210 | GEO | 2022/06/30
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.
2012-11-01 | E-GEOD-41962 | biostudies-arrayexpress
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 3D hepatocyte cell culture that enables cellular maintenance in a polycarbonate scaffold structure. Our data indicate that an actively perfused three dimensional cell culture displays a more pronounced metabolic genotype than statically cultivated hepatocytes. Human hepatocytes of three donors were cultivated for five days under 2D and 3D statical and perfused conditions. Cultivation was started with 0.25 x106 in 2D and with 1x 106 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 culture and BR denotes perfused 3D culture of hepatocytes. The perfusable bioreactor system was operated using a peristaltic pump. It houses the MatriGrid, a polycarbonate based microporous cellular support. For 3Dstatic cultivation, cell- inoculated MatriGrids were placed in wells of a 24 wells plate. Microarray experiments of three 2D (i.e. control), three 3D statically and three actively perfused 3D cultivations, respectively, were performed at SIRS-Lab GmbH (SIRS-Lab GmbH, Jena, Germany) according to the manufacturerM-bM-^@M-^Ys instructions (Illumina, San Diego, CA). Altogether, 8 RNA samples of hepatocyte cultures and an internal control RNA were hybridized on two HumanHT-12 v4 Expression BeadChips.
2013-03-31 | E-GEOD-44580 | biostudies-arrayexpress
Project description:The traditional method for studying cancer in vitro is to grow immortalized cancer cells in two-dimensional (2D) monolayers on plastic. However, many cellular features are impaired in these unnatural conditions and big alterations in gene expression in comparison to tumors have been reported. Three-dimensional (3D) cell culture models have become increasingly popular and are suggested to be better models than 2D monolayers due to improved cell-to-cell contacts and structures that resemble in vivo architecture. The aim of this study was to develop a simple high-throughput 3D drug screening method and to compare drug responses in JIMT1 breast cancer cells when grown in 2D, in polyHEMA coated anchorage independent 3D models and in Matrigel on-top 3D cell culture models. We screened 102 compounds with multiple concentrations and biological replicates for their effects on cell proliferation. The cells were either treated immediately upon plating or they were allowed to grow in 3D for four days prior to the drug treatment. Big variations in drug responses were observed between the models indicating that comparisons of culture model influenced drug sensitivities cannot be made based on effects of a single drug. However, we show with the 63 most prominent drugs that, in general, JIMT1 cells grown on Matrigel were significantly more sensitive to drugs than cells grown in 2D cultures, while responses of cells grown in polyHEMA resembled those of 2D. Furthermore, comparison of gene expression profiles of the cell culture models to xenograft tumors indicated that cells cultured in Matrigel and as xenografts most closely resembled each other. In this study we also suggest that 3D cultures can provide a platform for systematic experimentation of larger compound collections in a high-throughput mode and be used as alternatives for traditional 2D screens towards better comparability to in vivo state. Gene expression analysis of JIMT1 breast cancer cells cultured as xenografts for 43 days, in two dimensional cultures for seven days (2D7d), in polyHEMA three dimensional cell culture models for four and seven days (PH7d and PH7d), and in Matrigel three dimensional cultures for four and seven days (MG4d and MG7d). Two biological replicates was included for each sample.
2013-10-30 | E-GEOD-42529 | biostudies-arrayexpress
Project description:Background: The main focus of the work was the evaluation of gene expression differences between our established NSCLC 3D cell culture model and the 2D cell culture in regard to the use of our model for drug screening applications. Methods: The non-small cell lung cancer (NSCLC) cell lines Colo699 and A549 were cultivated as monolayer (2D) on cell culture plates for five days or as microtissues (3D) in a hanging-drop system for five and ten days, respectively. Cells and microtissues were harvested and Affymetrix chip analyses were performed with the prior isolated RNA. This was repeated in three independent experiments. Subsequent biostatistical data analyses tested for reproducibility, comparability and significant differences in gene expression profiles between cell lines, experiments and culture methods. Results: The analyses revealed a high interassay correlation within the distinct culture systems, thus proving a high validity of our data. The comparison of 3D versus 2D cell cultures revealed significant differences in RNA expression (979 genes for A549; 1106 genes for Colo699), but the overlap of changes in RNA profiles between the cell lines at the individual gene level was small (149 genes), potentially reflecting overall heterogeneity and their origin, i.e. primary vs pleural effusion. Nevertheless, these RNA expression changes affected most relevant cancer-associated pathways as DNA methylation, cell cycle, rRNA expression and meiosis pathways. Furthermore, the expression differences between 2D and 3D were more evident after longer cultivation time, which supports the hypothesis of cultivation related mechanisms and the usage of long-time cultivation systems. Conclusion: In summary, our data support the need of innovative 3D drug testing systems to close the gap between in-vitro drug screening and in-vivo data. Thus, our 3D NSCLC model might provide a model to address the challenge of microenviroment associated resistance mechanisms, as well as cell-cell interaction related effects.
2018-02-07 | GSE78210 | GEO
Project description:Complex three-dimensional (3D) in vitro model systems that recapitulate human tumor biology are essential to better understand the pathophysiology of the disease and to aid in the discovery of novel anti-cancer therapies. 3D organotypic cultures exhibit intercellula communication, nutrient and oxygen gradients, and cell polarity that is lacking in traditional two-dimensional (2D) monolayer cultures. In the present study, we could demonstrate that 2D and 3D cancer models exhibit different drug sensitivities towards both targeted inhibitors of EGFR signaling and broad acting cytotoxic agents. Changes in the kinase activities of Erb family members and differential expression of apoptosis- and survival-associated genes before and after drug treatment may account for the differential drug sensitivities. Importantly, EGFR oncoprotein addiction was evident only in the 3D cultures mirroring the effect of EGFR inhibition in the clinic. Furthermore, targeted drug efficacy was strongly increased when incorporating cancer-associated fibroblasts into the 3D cultures. Taken together, we could provide conclusive evidence that complex 3D cultures are more predictive of the clinical outcome than their 2D counterparts. In the future, 3D cultures will be instrumental for understanding the mode of action of drugs, identifying genotype-drug response relationships and developing patient-specific and personalized cancer treatments.
2017-11-30 | GSE102722 | GEO
Project description:The traditional method for studying cancer in vitro is to grow immortalized cancer cells in two-dimensional (2D) monolayers on plastic. However, many cellular features are impaired in these unnatural conditions and big alterations in gene expression in comparison to tumors have been reported. Three-dimensional (3D) cell culture models have become increasingly popular and are suggested to be better models than 2D monolayers due to improved cell-to-cell contacts and structures that resemble in vivo architecture. The aim of this study was to develop a simple high-throughput 3D drug screening method and to compare drug responses in JIMT1 breast cancer cells when grown in 2D, in polyHEMA coated anchorage independent 3D models and in Matrigel on-top 3D cell culture models. We screened 102 compounds with multiple concentrations and biological replicates for their effects on cell proliferation. The cells were either treated immediately upon plating or they were allowed to grow in 3D for four days prior to the drug treatment. Big variations in drug responses were observed between the models indicating that comparisons of culture model influenced drug sensitivities cannot be made based on effects of a single drug. However, we show with the 63 most prominent drugs that, in general, JIMT1 cells grown on Matrigel were significantly more sensitive to drugs than cells grown in 2D cultures, while responses of cells grown in polyHEMA resembled those of 2D. Furthermore, comparison of gene expression profiles of the cell culture models to xenograft tumors indicated that cells cultured in Matrigel and as xenografts most closely resembled each other. In this study we also suggest that 3D cultures can provide a platform for systematic experimentation of larger compound collections in a high-throughput mode and be used as alternatives for traditional 2D screens towards better comparability to in vivo state.
2013-10-30 | GSE42529 | GEO
Project description:Current preclinical models in tumor biology are limited in their ability to recapitulate relevant (patho-) physiological processes, including autophagy. Three-dimensional (3D) growth cultures have frequently been proposed to overcome the lack of correlation between two-dimensional (2D) monolayer cell cultures and human tumors in preclinical drug testing. Besides 3D growth, it is also advantageous to simulate shear stress, compound flux and removal of metabolites, e.g. via bioreactor systems, through which culture medium is constantly pumped at a flow rate reflecting physiological conditions. Here, we show that both Staticic 3D growth and 3D growth within a bioreactor system modulate key hallmarks of cancer cells, including proliferation and cell death as well as macroautophagy, a recycling pathway often activated by highly proliferative tumors to cope with metabolic stress. The autophagy-related gene expression profiles of 2D- and 3D-grown cells are substantially different, with the 3D-grown cells exhibiting an expression profile closely resembling the (patho-) physiological Statice of a tumor. Underscoring the importance of this pathway, autophagy-controlling transcription factors, such as TFEB and FOXO3, are upregulated in tumors, and 3D-grown cells have increased expression compared with cells grown in 2D conditions. Three-dimensional cultures depleted of the autophagy mediators BECN1, ATG5 or ATG7 or the transcription factor FOXO3, are more sensitive to cytotoxic treatment. Accordingly, combining cytotoxic treatment with compounds affecting late autophagic flux, such as chloroquine, renders the 3D-grown cells more susceptible to therapy and increases intracellular doxorubicin concentration to the level of 2D-grown cells. Altogether, 3D cultures are a valuable tool to study drug response of tumor cells, as these models recapitulate (patho-) physiologically relevant pathways, such as autophagy.
2017-07-04 | GSE100020 | GEO
Project description:The traditional method for studying cancer in vitro is to grow immortalized cancer cells in two-dimensional (2D) monolayers on plastic. However, many cellular features are impaired in these unnatural conditions and big alterations in gene expression in comparison to tumors have been reported. Three-dimensional (3D) cell culture models have become increasingly popular and are suggested to be better models than 2D monolayers due to improved cell-to-cell contacts and structures that resemble in vivo architecture. The aim of this study was to compare gene expression patterns of MCF7 breast cancer cells when grown as xenografts, in 2D, in polyHEMA coated anchorage independent 3D models and in Matrigel on-top 3D cell culture models. Surprisingly small variations in gene expression patterns were observed between the models indicating that 3D and xenograft are not always that different from 2D cell cultures. Gene expression analysis of MCF7 breast cancer cells cultured as xenografts for 43 days, in two dimensional cultures for seven days (2D7d), in polyHEMA three dimensional cell culture models for four and seven days (PH7d and PH7d), and in Matrigel three dimensional cultures for four and seven days (MG4d and MG7d). Two biological replicates was included for each sample.
2013-10-30 | E-GEOD-47583 | biostudies-arrayexpress
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.
2012-11-01 | GSE41962 | GEO
Project description:Tumor cell response to irradiation also depends on their microenvironment. Therefore ongoing investigation of three-dimensional (3D) cell culture models provide researchers with essential data studying and remodeling radiotherapeutic implications in cancer treatment. 3D culture models were shown to mimic in vivo cell microenvironment more accurately than the standard two-dimensional cell monolayer (2D) cultures. Growing evidence suggests that 2D and 3D cultured cell gene expression pattern discrepancies following irradiation is highly dependent on cell-ECM interactions. It has been shown that laminin-rich-extracellular matrix (lr-ECM) used in 3D cultures not only alters cancer cell phenotype and response to external stimuli but also affects their differentiation, migration and survivability. Thus, a change in these fundamental cell properties demands us to reconsider data previously collected using 2D in vitro models. RNA was harvested from two colorectal cancer cell lines cultivated under 3D cell culture conditions, 4h after treatment of single (2 Gy or 10 Gy) or fractionated (5x2 Gy) ionizing radiation dose.
2015-12-12 | E-GEOD-75915 | biostudies-arrayexpress