A static, closed and scaffold-free bioreactor system that permits chondrogenesis in vitro

Grogan, Shawn P.; Rieser, Franz; Winkelmann, Verena; Berardi, Simona; Mainil‐Varlet, Pierre

doi:10.1016/s1063-4584(03)00053-0

Cited by 61 publications

(53 citation statements)

References 44 publications

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“…This assessment takes into consideration (i) the uniformity and intensity of Safranin-O stain, (ii) the distance between cells and amount of matrix produced, and (iii) the cell morphology found within sections. Sections were also processed for immunohistochemistry as previously described (Grogan et al, 2003). Briefly, after pre-treatment with 1 mg/ml chondroitinase, sections were incubated with monoclonal primary antibodies for type II collagen (II-II6B3, Hybridoma Bank, University of Iowa, USA) for 1 h. The slides were then incubated with a biotinylated goat antimouse secondary antibody for 35 min followed by StrABC/AP for 45 min (DAKO, Denmark).…”

Section: Methodsmentioning

confidence: 99%

A low percentage of autologous serum can replace bovine serum to engineer human nasal cartilage

Wolf¹,

Haug²,

Farhadi³

et al. 2008

eCM

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For the generation of cell-based therapeutic products, it would be preferable to avoid the use of animal-derived components. Our study thus aimed at investigating the possibility to replace foetal bovine serum (FBS) with autologous serum (AS) for the engineering of cartilage grafts using expanded human nasal chondrocytes (HNC). HNC isolated from 7 donors were expanded in medium containing 10% FBS or AS at different concentrations (2%, 5% and 10%) and cultured in pellets using serum-free medium or in Hyaff ® -11 meshes using medium containing FBS or AS. Tissue forming capacity was assessed histologically (Safranin O), immunohistochemically (type II collagen) and biochemically (glycosaminoglycans -GAG-and DNA). Differences among experimental groups were assessed by Mann Whitney tests. HNC expanded under the different serum conditions proliferated at comparable rates and generated cartilaginous pellets with similar histological appearance and amounts of GAG. Tissues generated by HNC from different donors cultured in Hyaff ® -11 had variable quality, but the accumulated GAG amounts were comparable among the different serum conditions. Staining intensity for collagen type II was consistent with GAG deposition. Among the different serum conditions tested, the use of 2% AS resulted in the lowest variability in the GAG contents of generated tissues. In conclusion, a low percentage of AS can replace FBS both during the expansion and differentiation of HNC and reduce the variability in the quality of the resulting engineered cartilage tissues.

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Section: Methodsmentioning

confidence: 99%

A low percentage of autologous serum can replace bovine serum to engineer human nasal cartilage

Wolf¹,

Haug²,

Farhadi³

et al. 2008

eCM

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“…Sections were also processed for immunohistochemistry to visualize collagen type II (II-II6B3, Hybridoma Bank, University of Iowa, Iowa City, IA), as previously described. 16 Biochemical analysis. Pellets were digested with protease K (0.5 mL of 1 mg/mL protease K in 50 mM Tris with 1mM EDTA, 1 mM iodoacetamide, and 10 mg/mL pepstatin-A) for 15 h at 568C, as previously described.…”

Section: Methodsmentioning

confidence: 99%

Growth Factors for Clinical-Scale Expansion of Human Articular Chondrocytes: Relevance for Automated Bioreactor Systems

Francioli

Martin

Sie³

et al. 2007

Tissue Engineering

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The expansion of chondrocytes in automated bioreactors for clinical use requires that a relevant number of cells be generated, starting from variable initial seeding densities in one passage and using autologous serum. We investigated whether the growth factor combination transforming growth factor beta 1/fibroblast growth factor 2/platelet-derived growth factor BB (TFP), recently shown to enhance the proliferation capacity of human articular chondrocytes (HACs), allows the efficiency of chondrocyte use to be increased at different seeding densities and percentages of human serum (HS). HACs were seeded at 1,000, 5,000, and 10,000 cells/cm 2 in medium containing 10% fetal bovine serum or 10,000 cells/cm 2 with 1%, 5%, or 10%HS. The chondrogenic capacity of post-expanded HACs was then assessed in pellet cultures. Expansion with TFP allowed a sufficient number of HACs to be obtained in one passage even at the lowest seeding density and HS percentage and variability in cartilage-forming capacity of HACs expanded under the different conditions to be reduced. Instead, larger variations and insufficient yields were found in the absence of TFP. By allowing large numbers of cells to be obtained, starting from a wide range of initial seeding densities and HS percentages, the use of TFP may represent a viable solution for the efficient expansion of HACs and addresses constraints of automated clinical bioreactor systems.

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“…Sections were also processed for immunohistochemistry to visualize type II collagen (II-II6B3, Hybridoma Bank, University of Iowa, Iowa City, IA, USA), as previously described (Grogan et al, 2003).…”

Section: Histological and Immunohistochemical Analysismentioning

confidence: 99%

An RGD-restricted substrate interface is sufficient for the adhesion, growth and cartilage forming capacity of human chondrocytes

Vonwil¹,

Schüler²,

Barbero³

et al. 2010

eCM

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This study aimed at testing whether an RGD-restricted substrate interface is sufficient for adhesion and growth of human articular chondrocytes (HAC), and whether it enhances their post expansion chondrogenic capacity. HAC/ substrate interaction was restricted to RGD by modifying tissue culture polystyrene (TCPS) with a poly(ethylene glycol) (PEG) based copolymer system that renders the surface resistant to protein adsorption while at the same time presenting the bioactive RGD-containing peptide GCRGYGRGDSPG (RGD). As compared to TCPS, HAC cultured on RGD spread faster (1.9-fold), maintained higher type II collagen mRNA expression (4.9-fold) and displayed a 19% lower spreading area. On RGD, HAC attachment efficiency (66±10%) and proliferation rate (0.56±0.04 doublings/day), as well as type II collagen mRNA expression in the subsequent chondrogenic differentiation phase, were similar to those of cells cultured on TCPS. In contrast, cartilaginous matrix deposition by HAC expanded on RGD was slightly but consistently higher (15% higher glycosaminoglycan-to-DNA ratio). RDG (bioinactive peptide) and PEG (no peptide ligand) controls yielded drastically reduced attachment efficiency (lower than 11%) and proliferation (lower than 0.20 doublings/day). Collectively, these data indicate that restriction of HAC interaction with a substrate through RGD peptides is sufficient to support their adhesion, growth and maintenance of cartilage forming capacity. The concept could thus be implemented in materials for cartilage repair, whereby in situ recruited/infiltrated chondroprogenitor cells would proliferate while maintaining their ability to differentiate and generate cartilage tissue.

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A static, closed and scaffold-free bioreactor system that permits chondrogenesis in vitro

Abstract: In this study, we provide evidence of a scaffold-free system that can produce immature hyaline-like cartilaginous constructs suitable for in vivo implantation, or that may be useful for in vitro studies of events related to the process of chondrogenesis.

Cited by 61 publications

References 44 publications

A low percentage of autologous serum can replace bovine serum to engineer human nasal cartilage

A low percentage of autologous serum can replace bovine serum to engineer human nasal cartilage

Growth Factors for Clinical-Scale Expansion of Human Articular Chondrocytes: Relevance for Automated Bioreactor Systems

An RGD-restricted substrate interface is sufficient for the adhesion, growth and cartilage forming capacity of human chondrocytes

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