High-Throughput Aggregate Culture System to Assess the Chondrogenic Potential of Mesenchymal Stem Cells

Penick, Kitsie; Solchaga, Luis A.; Welter, Jean F.

doi:10.2144/000112009

Cited by 108 publications

(125 citation statements)

References 13 publications

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“…that described by Penick (Penick et al, 2005)]. and subjecting the cells to gentle centrifugation (800 g for 5 min) in a 15-ml conical polypropylene tube, after which the cap was loosened, and the tube was placed in the incubator, where the cells adhered to one another and consolidated into a cell pellet within 24 h. After 3 to 4 days, the hMSCs had formed a 1-mm ball in the bottom of the tube.…”

Section: Assay For Chondrogenesis Of Hmscsmentioning

confidence: 99%

ATF6a, a Runx2-activable transcription factor, is a novel regulator of chondrocyte hypertrophy

Guo

Han

et al. 2015

Journal of Cell Science

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Our previous research has shown that the spliced isoform of XBP1 (XBP1s) is an important downstream mediator of BMP2 and is involved in BMP2-stimulated chondrocyte differentiation. Herein, we report that ATF6 and its cleaved N-terminal cytoplasmic domain (known as ATF6a) are expressed in growth plate chondrocytes. We find that these proteins are differentially induced during BMP2-triggered chondrocyte differentiation. This differential expression probably results from the activation of the ATF6 gene by Runx2 and its repression by the Sox6 transcription factor. Runx2 and Sox6 act through their respective binding elements on the ATF6 gene. When overexpressed, ATF6 and ATF6a intensify chondrogenesis; our studies demonstrate that under the stimulation of ATF6 and ATF6a, chondrocytes tend to be hypertrophied and mineralized, a process leading to bone formation. By contrast, lowering expression of ATF6a by use of its specific siRNA suppresses chondrocyte differentiation. Moreover, ATF6a interacts with Runx2 and augments the Runx2-mediated hypertrophication of chondrocytes. Importantly, overexpression and knockdown of ATF6a during the chondrocyte hypertrophy process also led to altered expressions of IHH and PTHrP (also known as PTHLH). Taken together, these findings indicate that ATF6a favorably controls chondrogenesis and bone formation (1) by acting as a co-factor of Runx2 and enhancing Runx2-incited hypertrophic chondrocyte differentiation, and (2) by affecting IHH and PTHrP signaling.

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Section: Assay For Chondrogenesis Of Hmscsmentioning

confidence: 99%

ATF6a, a Runx2-activable transcription factor, is a novel regulator of chondrocyte hypertrophy

Guo

Han

et al. 2015

Journal of Cell Science

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“…[26][27][28][29] For each sample, the entire construct was digested in 1 mL of protease (2.5 mg/mL; Type XIV Bacterial from Streptomyces griseus; Sigma) in trisbuffered saline in a 558C water bath overnight. A 100 mL aliquot of the digest was assayed for total GAG content by addition of 2 mL of 1,9-dimethyl methylene blue dye solution (Polyscience, Northampton, United Kingdom).…”

Section: Biochemical Analysis Of Glycosaminoglycanmentioning

confidence: 99%

Magnetization Transfer Imaging Provides a Quantitative Measure of Chondrogenic Differentiation and Tissue Development

Hong

et al. 2010

Tissue Engineering Part C: Methods

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The goal of the present investigation was to test whether quantitative magnetization transfer imaging can be used as a noninvasive evaluation method for engineered cartilage. In this work, we used magnetic resonance imaging (MRI) to monitor the chondrogenesis of stem-cell-based engineered tissue over a 3-week period by measuring on a pixel-by-pixel basis the relaxation times (T 1 and T 2 ), the apparent diffusion coefficient, and the magnetization transfer parameters: bound proton fraction and cross-relaxation rate (k). Tissue-engineered constructs for generating cartilage were created by seeding mesenchymal stem cells in a gelatin sponge. Every 7 days, tissue samples were analyzed using MRI, histological, and biochemical methods. The MRI measurements were verified by histological analysis, and the imaging data were correlated with biochemical analysis of the developing cartilage matrix for glycosaminoglycan content. The MRI analysis for bound proton fraction and k showed a statistically significant increase that was correlated with the increase of glycosaminoglycan (R ¼ 0.96 and 0.87, respectively, p < 0.05), whereas T 1 , T 2 , and apparent diffusion coefficient results did not show any significant changes over the 3-week measurement period.

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“…Three-dimensional (3D) cell culture systems eliminate all these drawbacks of 2D cell culture systems, provide a high surface area for growth and migration, which can be tuned to support other cell behaviors, such as morphology, viability, proliferation, response to stimuli, differentiation or maturation and mimic human tissue micro-environment, pathological conditions and biological mechanisms much more closely [1,2]. 3D culture systems are used in an extensive area of cell-based studies [3], including cell adhesion/migration, tumour biology [4][5][6], stem cell research [7], regenerative medicine, tissue engineering and preclinical testing in drug discovery [8][9][10][11].…”

Section: Introductionmentioning

confidence: 99%

Comparison of Microtissue Forming Capacity of Sh-Sy5y and Sk-N-as Cell Lines

Taşdemir¹,

Ürkmez²,

Dogan³

2016

deufmd

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Two-dimensional (2D) cell culture systems are important tools for basic in vitro research. However, they form a thin monolayer structure and poorly mimic the complex in vivo conditions in terms of biochemical signals, cell-cell and cell-extracellular matrix (ECM) interactions. In this study, we performed a comparative study on SH-SY5Y and SK-N-AS neuroblastoma cell lines in terms of their microtissue forming capacity. Both cell lines are commonly used to model neurodegenerative diseases in vitro. Cells were cultured using 3D Petri Dish ® technique. The cells' microtissue forming capacity was observed morphologically and microtissues' size was analized. Results indicate that microtissue forming capacity of SH-SY5Ycell line was better than that of SK-N-AS cell line. SH-SY5Y microtissues can be used as an alternative, scaffold-free in vitro 3D model for neurodegenerative diseases and neuroblastoma research. Keywords: Cell culture, Three-dimensional (3D) Cell culture, Microtissue, SH-SY5Y, SK-N-A ÖZ

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High-Throughput Aggregate Culture System to Assess the Chondrogenic Potential of Mesenchymal Stem Cells

Cited by 108 publications

References 13 publications

ATF6a, a Runx2-activable transcription factor, is a novel regulator of chondrocyte hypertrophy

ATF6a, a Runx2-activable transcription factor, is a novel regulator of chondrocyte hypertrophy

Magnetization Transfer Imaging Provides a Quantitative Measure of Chondrogenic Differentiation and Tissue Development

Comparison of Microtissue Forming Capacity of Sh-Sy5y and Sk-N-as Cell Lines

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