Expression of a stable articular cartilage phenotype without evidence of hypertrophy by adult human articular chondrocytes in vitro

Abstract

Chondrocytes that were isolated from adult human articular cartilage changed phenotype during monolayer tissue culture, as characterized by a fibroblastic morphology and cellular proliferation. Increased proliferation was accompanied by downregulation of the cartilage-specific extracellular matrix proteoglycan, aggrecan, by cessation of type-II collagen expression, and by upregulation of type-I collagen and versican. This phenomenon observed in monolayer was reversible after the transfer of cells to a suspension culture system. The transfer of chondrocytes to suspension culture in alginate beads resulted in the rapid upregulation of aggrecan and type-II collagen and the downregulation of expression of versican and type-I collagen. Type-X collagen and osteopontin, markers of chondrocyte hypertrophy and commitment to endochondral ossification, were not expressed by adult articular chondrocytes cultured in alginate, even after 5 months. In contrast, type-X collagen was expressed within 2 weeks in a population of cells derived from a fetal growth plate. The inability of adult articular chondrocytes to express markers of chondrocyte hypertrophy has underscored the fundamental distinction between the differentiation pathways that lead to articular cartilage or to bone. Adult articular chondrocytes expressed only hyaline articular cartilage markers without evidence of hypertrophy.

Corticosteroids alter the differentiated phenotype of articular chondrocytes

“…The higher baseline of type I1 collagen expression in monolayer chondrocyte cultures may magnify methylprednisolone-induced suppression. Maintaining a rounded morphology [5,7,24] and integrity of the cartilage extracellular matrix [9,22,44,49] supports stability of the chondrocytic phenotype in culture and may lessen the effects of corticosteroid exposure. Finally, although the articular cartilage was selected only from weight-bearing locations, neither the explant nor isolated chondrocyte culture systems can address the cellular heterogeneity of articular cartilage.…”

“…Using an in vivo model of synovitis, we showed that one dose of intraarticular MPA significantly decreased steady-state type I1 procollagen mRNA in articular cartilage [34]. Protein analyses in cartilage explants from these same animals, however, demonstrated increased total collagen synthesis in MPA-treated joints [5 11. An earlier study, using cartilage explants and metabolic labeling observed concentration-dependent effects of methylprednisolone.…”

Differential expression of multiple genes during articular chondrocyte redifferentiation

“…Adult human articular chondrocytes were isolated from healthy cartilage tissue [National Disease Research Interchange Tissue Bank (NDRI), Philadelphia, PA] from three donors aged 24, 31, and 52 years, as described previously (Binette et al, 1998). Briefly, tissue samples were minced and digested in 0.1% collagenase (Sigma, St. Louis, MO) for 18 hr at 37°C.…”

“…The morphological changes caused by suspension culture are accompanied by a simultaneous down-regulation of genes that are characteristic of proliferating fibroblasts and an up-regulation of cartilage specific genes, a process termed redifferentiation, which occurs over a period of several weeks (Binette et al, 1998;Yaeger et al, 1997). Because the molecular mechanisms involved in the modulation of the articular chondrocyte phenotype were not well established, we employed a subtractive hybridization strategy to generate cDNA libraries of genes that are differentially regulated between monolayer and suspension cultures of articular chondrocytes Gurskaya et al, 1996).…”

Engineering custom-designed osteochondral tissue grafts

“…hMSCs have advantages because of their availability and multipotency, but the effects of donor age and health are still not well understood [57], and these cells have less capacity to form cartilage than chondrocytes [58]. By contrast, harvesting of adult autologous articular chondrocytes involves joint biopsy, and the cells tend to dedifferentiate during expansion [35,59]. Approaches currently being studied include growth factor supplementation during cell expansion and culture [60,61], use of additional cell sources (such as nasal, ear and rib cartilage) [62], adipose-derived hMSCs [24] and cell co-culture [27,63].…”