Abstract
The knee meniscus contains a mixed population of cells that exhibit fibroblastic as well as chondrocytic characteristics. Tissue engineering studies and future therapies for the meniscus require a large population of cells that are seeded on scaffolds. To achieve this, monolayer expansion is often used as a technique to increase cell number. However, the phenotype of these cells may be significantly different from that of the primary population. The objective of this study was to investigate changes in meniscal fibrochondrocytes at the gene expression level over four passages using quantitative real-time reverse transcriptase polymerase chain reaction. Cells from the inner two-thirds of bovine medial menisci were used. Four extracellular matrix (ECM) molecules, commonly found in the meniscus, were investigated, namely collagen I, collagen II, aggrecan and cartilage oligomeric matrix protein (COMP). In addition, primary and passaged meniscus fibrochondrocytes were placed on surfaces coated with collagen I or aggrecan protein to investigate whether any gene expression changes resulting from passage could be reversed. Collagen I expression was found to increase with the number of passages, whereas collagen II and COMP expression decreased. Collagen I and aggrecan surface coatings were shown to downregulate and upregulate collagen I and COMP expression levels, respectively, in passaged cells. However, decreases in collagen II expression could not be reversed by either protein coating. These results indicate that although monolayer expansion results in significant changes in gene expression in meniscal fibrochondrocytes, protein coatings may be used to regain the primary cell expression of several ECM molecules.
Highlights
The meniscus is a wedge-shaped fibrocartilaginous tissue located in the knee joint
Primary and passaged meniscus fibrochondrocytes were placed on surfaces coated with collagen I or aggrecan protein to investigate whether any gene expression changes resulting from passage could be reversed
Decreases in collagen II expression could not be reversed by either protein coating. These results indicate that monolayer expansion results in significant changes in gene expression in meniscal fibrochondrocytes, protein coatings may be used to regain the primary cell expression of several extracellular matrix (ECM) molecules
Summary
The meniscus is a wedge-shaped fibrocartilaginous tissue located in the knee joint. As reviewed elsewhere, it serves several mechanical functions including shock absorption, load transmission, joint stability and joint lubrication [1,2]. Fibrochondrocytes, as defined by the authors, are cells that are localized in the middle and inner meniscus and express both collagen I and collagen II. They can be identified by their round or oval shape and by the presence of a pericellular matrix. Cells of the superficial zone are located below the surface of ANOVA = analysis of variance; COMP = cartilage oligomeric matrix protein; DEPC = diethyl pyrocarbonate; ECM = extracellular matrix; FBS = fetal bovine serum; GAPDH = glyceraldehyde-3-phosphate dehydrogenase; IGF-I = insulin-like growth factor-I; PBS = phosphate-buffered saline; PSF = penicillin–streptomycin–Fungizone; RT-PCR = reverse transcriptase polymerase chain reaction; TMJ = temporomandibular joint
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