Deep zone articular chondrocytes in vitro express genes that show specificchanges with mineralization

We have developed a method to form reconstituted mineralized articular cart ilagenous tissue in vitro from isolated deep zone chondrocytes. The aim of this study was to characterize further these cultures Drier to and during m ineralization. Histologic examination of the cells up to 8 days in culture showed that the chondrocytes had formed cartilagenous tissue. Similar to th e in vivo cartilage, the chondrocytes expressed aggrecan, types II, I, and X collagens, osteopontin, and alkaline phosphatase (ALP). No osteocalcin mR NA expression was detected in either the in vivo cartilage or In vitro-gene rated tissue. Addition of beta-glycerophosphate (beta-GP) to the medium on day 5 induced mineralization and changes in gene expression. Expression of ape X collagen, type II collagen, aggrecan core protein, and ALP were inhib ited significantly between 2 h and 24 h after the addition of beta-GP. At 7 2 h, expression of these genes were still significantly depressed. These ch anges correlated with a decrease in collagen and proteoglycan synthesis, an d ALP activity. Osteopontin expression increased within 8 h but returned to constitutive levels by 72 h. No change in type I collagen expression was d etected. The changes in gene expression were not due to a direct effect of beta-GP itself, because similar gene changes occurred in the presence of ph osphoethanolamine, another agent which induces mineralization. No changes i n gene expression were seen in nonmineralizing cultures. In summary, articu lar chondrocytes grown on filter culture show expression of similar genes t o the chondrocytes in the deep zone of articular cartilage and that changes in expression of specific genes were observed during tissue mineralization , suggesting that it is a suitable model to use to study the mechanism(s) r egulating the localized mineralization of articular cartilage.