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Abstract
BackgroundMeniscal cartilage displays a poor repair capacity, especially when injury is located in the avascular region of the tissue. Cell-based tissue engineering strategies to generate functional meniscus substitutes is a promising approach to treat meniscus injuries. Meniscus fibrochondrocytes (MFC) can be used in this approach. However, MFC are unable to retain their phenotype when expanded in culture. In this study, we explored the effect of oxygen tension on MFC expansion and on their matrix-forming phenotype.Methodology/Principal FindingsMFC were isolated from human menisci followed by basic fibroblast growth factor (FGF-2) mediated cell expansion in monolayer culture under normoxia (21%O2) or hypoxia (3%O2). Normoxia and hypoxia expanded MFC were seeded on to a collagen scaffold. The MFC seeded scaffolds (constructs) were cultured in a serum free chondrogenic medium for 3 weeks under normoxia and hypoxia. Constructs containing normoxia-expanded MFC were subsequently cultured under normoxia while those formed from hypoxia-expanded MFC were subsequently cultured under hypoxia. After 3 weeks of in vitro culture, the constructs were assessed biochemically, histologically and for gene expression via real-time reverse transcription-PCR assays. The results showed that constructs under normoxia produced a matrix with enhanced mRNA ratio (3.5-fold higher; p<0.001) of collagen type II to I. This was confirmed by enhanced deposition of collagen II using immuno-histochemistry. Furthermore, the constructs under hypoxia produced a matrix with higher mRNA ratio of aggrecan to versican (3.5-fold, p<0.05). However, both constructs had the same capacity to produce a glycosaminoglycan (GAG) –specific extracellular matrix.ConclusionsOur data provide evidence that oxygen tension is a key player in determining the matrix phenotype of cultured MFC. These findings suggest that the use of normal and low oxygen tension during MFC expansion and subsequent neo-tissue formation cultures may be important in engineering different regions of the meniscus.
Highlights
The meniscus is a specialized fibrocartilaginous tissue located in the knee joint where it functions to aid joint stability, protect articular cartilage, absorb shock and transmit load [1]
These findings suggest that the use of normal and low oxygen tension during meniscus fibrochondrocytes (MFC) expansion and subsequent neo-tissue formation cultures may be important in engineering different regions of the meniscus
Cell culture and biochemical analysis MFCs were isolated from human knee menisci and cultured in monolayer in the presence of FGF-2 under normal (21% O2) or low (3% O2) oxygen tension
Summary
The meniscus is a specialized fibrocartilaginous tissue located in the knee joint where it functions to aid joint stability, protect articular cartilage, absorb shock and transmit load [1]. Meniscus allograft suffers from donor shortage and risk of transmission of infectious diseases, while implantation of current biomaterialbased substitutes is only compatible with partial meniscectomy To address these limitations, cell-based tissue engineering strategies to repair or to produce functional meniscus substitutes are of interest [22,23,24,25,26,27]. Cell-based tissue engineering strategies to repair or to produce functional meniscus substitutes are of interest [22,23,24,25,26,27] These approaches require an ideal biomaterial (scaffold) to support sufficient number of cells that exhibit the phenotypic characteristics of MFC. We explored the effect of oxygen tension on MFC expansion and on their matrix-forming phenotype
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