Alternative splicing of type II procollagen pre‐mRNA in chondrocytes is oppositely regulated by BMP‐2 and TGF‐β1

Valcourt, Ulrich; Gouttenoire, Jérôme; Aubert-Foucher, E.; Herbage, D.; Malléin-Gerin, Frédéric

doi:10.1016/s0014-5793(03)00510-6

Cited by 49 publications

(39 citation statements)

References 42 publications

(45 reference statements)

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“…As a first attempt to compare the capabilities of BMP-2 and TGF-b1 to direct redifferentiation of chondrocytes, we used embryonic mouse chondrocytes, a cell model routinely used in our laboratory. [3][4][5][6][7][8] We found that BMP-2 was more effective than TGF-b1 to restore the chondrogenic character of chondrocytes, as judged by the re-expression of specific chondrocytic markers such as Sox9, a10 integrin subunit, and type IIB procollagen isoform. 5 However, this redifferentiation occurred to a certain extent only, since type I collagen expression persisted indifferently after the addition of BMP-2 or TGF-b1.…”

Section: Introductionmentioning

confidence: 85%

Control of Collagen Production in Mouse Chondrocytes by Using a Combination of Bone Morphogenetic Protein-2 and Small Interfering RNA TargetingCol1a1for Hydrogel-Based Tissue-Engineered Cartilage

Perrier‐Groult

Pasdeloup

Malbouyres

et al. 2013

Tissue Engineering Part C: Methods

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Because articular cartilage does not self-repair, tissue-engineering strategies should be considered to regenerate this tissue. Autologous chondrocyte implantation is already used for treatment of focal damage of articular cartilage. Unfortunately, this technique includes a step of cell amplification, which results in dedifferentiation of chondrocytes, with expression of type I collagen, a protein characteristic of fibrotic tissues. Therefore, the risk of producing a fibrocartilage exists. The aim of this study was to propose a new strategy for authorizing the recovery of the differentiated status of the chondrocytes after their amplification on plastic. Because the bone morphogenetic protein (BMP)-2 and the transforming growth factor (TGF)-b1 are cytokines both proposed as stimulants for cartilage repair, we undertook a detailed comparative analysis of their biological effects on chondrocytes. As a cellular model, we used mouse chondrocytes after their expansion on plastic and we tested the capability of BMP-2 or TGF-b1 to drive their redifferentiation, with special attention given to the nature of the proteins synthesized by the cells. To prevent any fibrotic character of the newly synthesized extracellular matrix, we silenced type I collagen by transfecting small interfering RNA (siRNA) into the chondrocytes, before their exposure to BMP-2 or TGF-b1. Our results showed that addition of siRNA targeting the mRNA encoded by the Col1a1 gene (Col1a1 siRNA) and BMP-2 represents the most efficient combination to control the production of cartilage-characteristic collagen proteins. To go one step further toward scaffold-based cartilage engineering, Col1a1 siRNA-transfected chondrocytes were encapsulated in agarose hydrogel and cultured in vitro for 1 week. The analysis of the chondrocyte-agarose constructs by using real-time polymerase chain reaction, Western-blotting, immunohistochemistry, and electron microscopy techniques demonstrated that the BMP-2/Col1a1 siRNA combination is effective in reinitializing correct production and assembly of the cartilage-characteristic matrix in agarose hydrogel, without production of type I collagen. Because agarose is known to favor long-term expression of the chondrocyte phenotype and agarose-based hydrogels are approved for clinical trials, this strategy appears very promising to repair hyaline cartilage.

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Section: Introductionmentioning

confidence: 85%

Control of Collagen Production in Mouse Chondrocytes by Using a Combination of Bone Morphogenetic Protein-2 and Small Interfering RNA TargetingCol1a1for Hydrogel-Based Tissue-Engineered Cartilage

Perrier‐Groult

Pasdeloup

Malbouyres

et al. 2013

Tissue Engineering Part C: Methods

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“…It is well known that cultured chondrocytes readily dedifferentiate in monolayer culture with increasing expression levels of COL1A1 across passage and decreases in COL2A1 across passage (Darling and Athanasiou 2005a;Darling and Athanasiou 2005b;Valcourt et al 2003). Cultured chondrocytes will attach and maintain their native rounded morphology when plated on tissue culture plastic but will then slowly flatten and occupy larger amounts of surface area in monolayer culture.…”

Section: Discussionmentioning

confidence: 99%

Monolayer cell expansion conditions affect the chondrogenic potential of adipose‐derived stem cells

Estes

Diekman

Guilak

2008

Biotech & Bioengineering

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Adipose-derived stem cells (ASCs) are an abundant, readily available population of multipotent progenitor cells that reside in adipose tissue. Isolated ASCs are typically expanded in monolayer on standard tissue culture plastic with a basal medium containing 10% fetal bovine serum. However, recent data suggests that altering the monolayer expansion conditions by using suspension culture plastic, adding growth factors to the medium, or adjusting the seeding density may affect the self-renewal rate, multipotency, and lineage-specific differentiation potential of the ASCs. We hypothesized that variation in any of these expansion conditions would influence the chondrogenic potential of ASCs. ASCs were isolated from human liposuction waste tissue and expanded through two passages with different tissue culture plastic, feed medium, and cell seeding densities. Once expanded, the cells were cast in an agarose gel and subjected to identical chondrogenic culture conditions for 7 days, at which point cell viability, radiolabel incorporation, and gene expression were measured. High rates of matrix synthesis upon chondrogenic induction were mostly associated with smaller cells, as indicated by cell width and area on tissue culture plastic, and it appears that expansion in a growth factor supplemented medium is important in maintaining this morphology. All end-point measures were highly dependent on the specific monolayer culture conditions. These results support the hypothesis that monolayer culture conditions may "prime" the cells or predispose them towards a specific phenotype and thus underscore the importance of early culture conditions in determining the growth and differentiation potential of ASCs.

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“…Collagen IIA is broadly expressed in embryonic or nonchondrogenic tissues as well as prechondrocytes and mesenchymal cells. The expression of collagen IIB, an isoform lacking a 69 amino acid cysteine-rich domain encoded by exon 2 is specifically localized in differentiating and adult chondrocytes, indicating the differentiation into functional chondrocytes (Ng et al, 1993;Sandell et al, 1994;Valcourt et al, 2003). Moreover, the high expression level of total collagen II and an isoform IIB in a primary culture of chondrocytes dramatically decreased within a few days followed by the first passage, whereas the expression level of collagen IIA relatively increased (Valcout et al, 2003).…”

Section: Discussionmentioning

confidence: 99%

Overexpression of SOX9 in mouse embryonic stem cells directs the immediate chondrogenic commitment

Kim

Do²,

Yang³

et al. 2005

Exp Mol Med

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Alternative splicing of type II procollagen pre‐mRNA in chondrocytes is oppositely regulated by BMP‐2 and TGF‐β1

Cited by 49 publications

References 42 publications

Control of Collagen Production in Mouse Chondrocytes by Using a Combination of Bone Morphogenetic Protein-2 and Small Interfering RNA TargetingCol1a1for Hydrogel-Based Tissue-Engineered Cartilage

Control of Collagen Production in Mouse Chondrocytes by Using a Combination of Bone Morphogenetic Protein-2 and Small Interfering RNA TargetingCol1a1for Hydrogel-Based Tissue-Engineered Cartilage

Monolayer cell expansion conditions affect the chondrogenic potential of adipose‐derived stem cells

Overexpression of SOX9 in mouse embryonic stem cells directs the immediate chondrogenic commitment

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