Real-time monitoring of 3T3-L1 preadipocyte differentiation using a commercially available electric cell-substrate impedance sensor system

Kramer, Adam H.; Joos-Vandewalle, Julia; Edkins, Adrienne L.; Frost, Carminita; Prinsloo, Earl

doi:10.1016/j.bbrc.2013.12.123

Cited by 32 publications

(24 citation statements)

References 19 publications

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“…30 In the present study we aimed to characterize the full differentiation process of both 3T3L1 fibroblasts and human hASCs on RTCA system, including cell growth and/or survival (i.e. cell number), morphological changes (i.e., cell size and cell force adhesion) in real-time assays on xCELLigence system to give a global image and point out specific events further characterized by complimentary approaches such as cell size analyses, lipid uptake measurements, cell growth and/or survival assays and cell imaging.…”

Section: Resultsmentioning

confidence: 99%

Pathways commonly dysregulated in mouse and human obese adipose tissue: FAT/CD36 modulates differentiation and lipogenesis

et al. 2015

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Obesity is linked to adipose tissue hypertrophy (increased adipocyte cell size) and hyperplasia (increased cell number). Comparative analyses of gene datasets allowed us to identify 1426 genes which may represent common adipose phenotype in humans and mice. Among them we identified several adipocyte-specific genes dysregulated in obese adipose tissue, involved in either fatty acid storage (acyl CoA synthase ACSL1, hormone-sensitive lipase LIPE, aquaporin 7 AQP7, perilipin PLIN) or cell adhesion (fibronectin FN1, collagens COL1A1, COL1A3, metalloprotein MMP9, or both (scavenger receptor FAT/CD36). Using real-time analysis of cell surface occupancy on xCELLigence system we developed a new method to study lipid uptake and differentiation of mouse 3T3L1 fibroblasts and human adipose stem cells. Both processes are regulated by insulin and fatty acids such as oleic acid. We showed that fatty acid addition to culture media increased the differentiation rate and was required for full differentiation into unilocular adipocytes. Significant activation of lipogenesis, i.e. lipid accumulation, by either insulin or oleic acid was monitored in times ranging from 1 to 24 h, depending on differentiation state, whereas significant effects on adipogenesis, i.e., surperimposed lipid accumulation and gene transcriptional regulations were measured after 3 to 4 d. Combination of selected times for analysis of lipid contents, cell counts, size fractionations, and gene transcriptional regulations showed that FAT/CD36 specific inhibitor AP5258 significantly increased cell survival of oleic acid-treated mouse and human adipocytes, and partially restored the transcriptional response to oleic acid in the presence of insulin through JNK pathway. Taken together, these data open new perspectives to study the molecular mechanisms commonly dysregulated in mouse and human obesity at the level of lipogenesis linked to hypertrophy and adipogenesis linked to hyperplasia

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Section: Resultsmentioning

confidence: 99%

Pathways commonly dysregulated in mouse and human obese adipose tissue: FAT/CD36 modulates differentiation and lipogenesis

et al. 2015

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“…The continuous monitoring of cell viability by the xCELLigence system makes it possible to distinguish between different perturbations, such as proliferation, migration and invasion [28]. Recently, this platform has proved very informative in monitoring the toxicity of compounds [29], biomaterials [30], inhibitors [31] and the cell differentiation process [32,33]. …”

Section: Resultsmentioning

confidence: 99%

“…The xCELLigence RTCA platform is proving to be a highly accurate platform to monitor cell behaviour [32,33,36–38] and it correlates very well with conventional adhesion, migration and invasion assays [39]. As well as this, the non-invasive and label-free platform is being used as a tool for high-throughput drug screening and as a robust system to measure the toxicological response to nanoparticles and novel compounds [40–42].…”

Section: Discussionmentioning

confidence: 99%

Using real-time impedance-based assays to monitor the effects of fibroblast-derived media on the adhesion, proliferation, migration and invasion of colon cancer cells

2014

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Increasing our knowledge of the mechanisms regulating cell proliferation, migration and invasion are central to understanding tumour progression and metastasis. The local tumour microenvironment contributes to the transformed phenotype in cancer by providing specific environmental cues that alter the cells behaviour and promotes metastasis. Fibroblasts have a strong association with cancer and in recent times there has been some emphasis in designing novel therapeutic strategies that alter fibroblast behaviour in the tumour microenvironment. Fibroblasts produce growth factors, chemokines and many of the proteins laid down in the ECM (extracellular matrix) that promote angiogenesis, inflammation and tumour progression. In this study, we use a label-free RTCA (real-time cell analysis) platform (xCELLigence) to investigate how media derived from human fibroblasts alters cancer cell behaviour. We used a series of complimentary and novel experimental approaches to show HCT116 cells adhere, proliferate and migrate significantly faster in the presence of media from human fibroblasts. As well as this, we used the xCELLigence CIM-plates system to show that HCT116 cells invade matrigel layers aggressively when migrating towards media derived from human fibroblasts. These data strongly suggest that fibroblasts have the ability to increase the migratory and invasive properties of HCT116 cells. This is the first study that provides real-time data on fibroblast-mediated migration and invasion kinetics of colon cancer cells.

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“…Impedance measurements have been used in a number of different studies to determine morphological changes in real time;27 cisplatin-induced cell death;28 differentiation of 3T3 cells into adipocytes;29 effects of curcumin on physicochemical characterization and effects on MCF7 cancer cell proliferation;30 monitoring of dynamic interactions of tumor cells with tissues and immune cells on a lab-on-a-chip;31 cell senescence measurements of adipose tissue-derived stem cells;32 and real-time evaluation of nanoparticle-induced cytotoxic effects 33. By this method, Moodley et al34 conducted real-time profiling of NK cell killing of human astrocytes.…”

Section: Discussionmentioning

confidence: 99%

A simple in vitro method for evaluating dendritic cell-based vaccinations

Pham¹,

Nguyen²,

Nguyen³

et al. 2014

OTT

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BackgroundDendritic cell (DC) therapy is a promising therapy for cancer-targeting treatments. Recently, DCs have been used for treatment of some cancers. We aimed to develop an in vitro assay to evaluate DC therapy in cancer treatment using a breast cancer model.MethodsDCs were induced from murine bone marrow mononuclear cells in Roswell Park Memorial Institute (RPMI) 1640 medium supplemented with GM-CSF (20 ng/mL) and IL-4 (20 ng/mL). Immature DCs were primed with breast cancer stem cell (BCSC)-derived antigens. BCSCs were sorted from 4T1 cell lines based on aldehyde dehydrogenase expression. A mixture of DCs and cytotoxic T lymphocytes (CTLs) were used to evaluate the inhibitory effect of antigen-primed DCs on BCSCs. BCSC proliferation and doubling time were recorded based on impedance-based cell analysis using the xCELLigence system. The specification of inhibitory effects of DCs and CTLs was also evaluated using the same system.ResultsThe results showed that impedance-based analysis of BCSCs reflected cytotoxicity and inhibitory effects of DCs and CTLs at 72 hours. Differences in ratios of DC:CTL changed the cytotoxicity of DCs and CTLs.ConclusionThis study successfully used impedance-based cell analysis as a new in vitro assay to evaluate DC efficacy in cancer immunotherapy. We hope this technique will contribute to the development and improvement of immunotherapies in the near future.

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Real-time monitoring of 3T3-L1 preadipocyte differentiation using a commercially available electric cell-substrate impedance sensor system

Cited by 32 publications

References 19 publications

Pathways commonly dysregulated in mouse and human obese adipose tissue: FAT/CD36 modulates differentiation and lipogenesis

Pathways commonly dysregulated in mouse and human obese adipose tissue: FAT/CD36 modulates differentiation and lipogenesis

Using real-time impedance-based assays to monitor the effects of fibroblast-derived media on the adhesion, proliferation, migration and invasion of colon cancer cells

A simple in vitro method for evaluating dendritic cell-based vaccinations

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