Comparison of bovine undifferentiated MSCs, chondrogenic MSCs and chondrocytes in 3D culture
Ontology highlight
ABSTRACT:
SUBMITTER: 
Ashley Stein Robert L Mauck Alice H Huang
PROVIDER: E-GEOD-18394 | ArrayExpress | 2010-10-02
SECONDARY ACCESSION(S): GSE18394PRJNA118081
REPOSITORIES: GEO, ArrayExpress
Dataset's files
primary
| Action | DRS | |||
|---|---|---|---|---|
| E-GEOD-18394.README.txt | Txt | |||
| E-GEOD-18394.eSet.r | Other | |||
| E-GEOD-18394.idf.txt | Idf | |||
| E-GEOD-18394.processed.1.zip | Processed | |||
| E-GEOD-18394.raw.1.zip | Raw |
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Project description:Bovine chondrocyte-seeded and mesenchymal stem cell (MSC)-seeded agarose were cultured for 28 days in chemically defined media containing 10 ng/mL TGF-beta3. Chondrogenic differentiated MSCs were compared to chondrocytes at this timepoint and to undifferentiated MSCs harvested at day 0. Overall design: Donor-matched sets of chondrocytes and MSCs were used, with three donors total.
2010-10-02 | GSE18394 | GEO
Project description:Mesenchymal stem cells (MSCs) are an attractive cell source for cartilage tissue engineering and regenerative medicine. However, the use of these cells has been limited by their reduced ability to form functional tissue compared to chondrocytes when placed in three-dimensional culture systems. To optimize MSC functional chondrogenesis, we examined the effects of increasing seeding density and transient application of transforming growth factor beta 3 (TGF-beta3), two factors previously shown to improve growth of chondrocyte-based constructs. Chondrocytes seeded in agarose at 20 million cells/mL and MSCs seeded at 20 or 60 million cells/mL agarose were cultured for 7 weeks under continuous or transient application of TGF-beta3. In the transient group, cell-laden constructs were exposed to TGF-beta3 for the initial 3 weeks, followed by 4 weeks of culture in medium without TGF-beta3. Compressive properties, biochemical content, and gene expression were assessed at 3, 5, and 7 weeks. Matrix distribution and collagen type was determined using histology and immunohistochemistry, and chondrogenic and osteogenic markers were assessed using real-time polymerase chain reaction. When maintained continuously with TGF-beta3, chondrocyte-seeded constructs achieved a higher equilibrium compressive modulus than MSCs similarly maintained. Although properties of both groups increased with respect to starting values, there was no difference in bulk mechanical or biochemical properties with higher seeding density when MSCs were cultured with constant TGF-beta3. Findings also showed that while transient application of TGF-beta3 elicited robust growth from chondrocyte-laden gels, MSCs seeded at the same density failed to respond, although constructs maintained their previously accrued properties and continued to express cartilaginous genes after TGF-beta3 removal. Conversely, MSCs seeded at 60 million cells/mL exhibited a strong anabolic response with transient TGF-beta3 exposure, achieving an equilibrium modulus of approximately 200 kPa. Although this represents the highest modulus we have been able to achieve with MSC-seeded constructs using our culture system, further work remains to optimize MSC chondrogenesis for cartilage tissue engineering, particularly in terms of collagen content and dynamic mechanical properties.
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Project description:Long-term dynamic compression enhanced the mechanical properties of MSC-seeded constructs only when loading was initiated after 21 days of chondrogenic differentiation. This study examined the molecular differences of chondrogenic MSCs compared to undifferentiated MSCs (TGF-beta vs no TGF-beta) and the effects of dynamic loading on MSC chondrogenesis (loading vs free-swelling). Free-swelling MSC-seeded constructs were cultured for 21 days in chemically defined media. Chondrogenesis was induced with TGF-beta3. Undifferentiated controls were maintained in parallel. After 21 days of chondrogenic differentiation, a subset of constructs were subjected to 21 days of dynamic compressive loading. On days 21 and 42, construct mechanical properties and biochemical content were assessed. Microarray analysis was carried out on day 3, day 21 and day 42 constructs. 6 arrays.
2010-11-01 | E-GEOD-18879 | ArrayExpress
Project description:Mesenchymal stem cells (MSCs) seeded on specific carrier materials are a promising source for the repair of traumatic cartilage injuries. The best supportive carrier material has not yet been determined. As natural components of cartilage's extracellular matrix, hyaluronic acid and collagen are the focus of biomaterial research. In order to optimize chondrogenic support, we investigated three different scaffold compositions of a hyaluronic acid (HA)-gelatin based biomaterial.Human MSCs (hMSCs) were seeded under vacuum on composite scaffolds of three different HA-gelatin ratios and cultured in chondrogenic medium for 21 days. Cell-scaffold constructs were assessed at different time points for cell viability, gene expression patterns, production of cartilage-specific extracellular matrix (ECM) and for (immuno-)histological appearance. The intrinsic transforming growth factor beta (TGF-beta) uptake of empty scaffolds was evaluated by determination of the TGF-beta concentrations in the medium over time.No significant differences were found for cell seeding densities and cell viability. hMSCs seeded on scaffolds with higher ratios of HA showed better cartilage-like differentiation in all evaluated parameters. TGF-beta uptake did not differ between empty scaffolds.Higher ratios of HA support the chondrogenic differentiation of hMSCs seeded on a HA-gelatin composite scaffold.
2016-01-01 | S-EPMC5503045 | BioStudies
Project description:Mesenchymal stem cells (MSCs) are an attractive cell source for cartilage tissue engineering given their ability to undergo chondrogenesis in 3D culture systems. Mechanical forces play an important role in regulating both cartilage development and MSC chondrogenic gene expression, however, mechanical stimulation has yet to enhance the mechanical properties of engineered constructs. In this study, we applied long-term dynamic compression to MSC-seeded constructs and assessed whether varying pre-culture duration, loading regimens and inclusion of TGF-beta3 during loading would influence functional outcomes and these phenotypic transitions. Loading initiated before chondrogenesis decreased functional maturation, although chondrogenic gene expression increased. In contrast, loading initiated after chondrogenesis and matrix elaboration further improved the mechanical properties of MSC-based constructs, but only when TGF-beta3 levels were maintained and under specific loading parameters. Although matrix quantity was not affected by dynamic compression, matrix distribution, assessed histologically and by FT-IRIS analysis, was significantly improved on the micro- (pericellular) and macro- (construct expanse) scales. Further, whole genome expression profiling revealed marked shifts in the molecular topography with dynamic loading. These results demonstrate, for the first time, that dynamic compressive loading initiated after a sufficient period of chondro-induction and with sustained TGF-beta exposure enhances matrix distribution and the mechanical properties of MSC-seeded constructs.
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Project description:OBJECTIVE: This preliminary study aims to determine the differentially expressed proteins from chondrogenic differentiated multipotent stromal cells (cMSCs) in comparison to undifferentiated multipotent stromal cells (MSCs) and adult chondrocytes (ACs). METHODS: ACs and bone marrow-derived MSCs were harvested from New Zealand White rabbits (n = 3). ACs and cMSCs were embedded in alginate and were cultured using a defined chondrogenic medium containing transforming growth factor-beta 3 (TGF-?3). Chondrogenic expression was determined using type-II collagen, Safranin-O staining and glycosaminoglycan analyses. Two-dimensional gel electrophoresis (2-DE) was used to isolate proteins from MSCs, cMSCs and ACs before being identified using liquid chromatography-mass spectrometry (LC-MS). The differentially expressed proteins were then analyzed using image analysis software. RESULTS: Both cMSCs and ACs were positively stained with type-II collagen and safranin-O. The expression of glycosaminoglycan in cMSCs was comparable to AC at which the highest level was observed at day-21 (p>0.05). Six protein spots were found to be most differentially expressed between MSCs, cMSCs and ACs. The protein spots cofilin-1 (CFL1) and glycealdehyde-3-phosphate dehydrogenase (GAPD) from cMSCs had expression levels similar to that of ACs whereas the others (ie. MYL6B, ALDOA, TAGLN2, EF1-alpha), did not match the expression level of ACs. CONCLUSION: Despite having similar phenotypic expressions to ACs, cMSCs expressed proteins which were not typically expected. This may explain the reason for the unexplained lack of improvement in cartilage repair outcomes reported in previous studies.
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