Differential expression of multiple genes during articular chondrocyte redifferentiation

Articular chondrocytes undergo a rapid change in phenotype and gene express ion, termed dedifferentiation, when isolated from cartilage tissue and cult ured on tissue culture plastic. On the other hand, "redifferentiation" of a rticular chondrocytes in suspension culture is characterized by decreased c ellular proliferation and the reinitiation of synthesis of hyaline articula r cartilage extracellular matrix molecules. The molecular triggers for thes e events have yet to be defined. Subtracted cDNA libraries representing gen es involved in the early events of adult human articular chondrocyte rediff erentiation were generated from human articular chondrocytes that were firs t cultured in monolayer, and subsequently transferred to suspension culture at 10(6) cells/ml for redifferentiation. Differential regulation of genes involved in cellular organization, nuclear structure, cellular growth regul ation, and extracellular matrix deposition and remodeling were observed wit hin 48 hr of this transfer. Many of these genes had not been previously ide ntified in the chondrocyte differentiation pathway and a number of the isol ated cDNAs did not have homologies to sequences in the public data banks. G enes involved in IL-6 signal transduction including acute phase response fa ctor (APRF), Mn superoxide dismutase, and IL-6 itself were up-regulated in suspension culture. Membrane glycoprotein gp130, a component of the IL-6 re ceptor, was down-regulated. Other genes involved in cell polarity, cell adh erence, apoptosis, and possibly TGF-beta signaling were differentially regu lated. The differential regulation of the cytokine connective tissue growth factor (CTGF) during the early stages of articular chondrocyte redifferent iation, decreasing within 48 hours of transfer to suspension culture, was p articularly interesting given its reported role in the stimulation of cellu lar proliferation. CTGF was highly expressed in proliferative monolayer cul ture, and then greatly reduced by redifferentiation in standard high-densit y suspension culture. When articular chondrocytes were seeded in suspension at low-density (10(4) cells/ml), however, high levels of CTGF were observe d along with increased levels of mature articular cartilage extracellular m atrix protein RNAs, such as type II collagen and aggrecan. Although the rol e of CTGF in articular cartilage biology remains to be elucidated, the resu lts described here demonstrate the potential utility of subtractive hybridi zation in understanding the process of articular chondrocyte redifferentiat ion. Anat Rec 263:91-98, 2001. (C) 2001 Wiley-Liss, Inc.