Proteoglycan production by immortalized human chondrocyte cell lines cultured under conditions that promote expression of the differentiated phenotype.
Large and small proteoglycans are essential components of articular cartilage. How to induce chondrocytes to repair damaged cartilage with normal ratios of matrix components after their loss due to degenerative joint disease has been a major research focus. We have developed immortalized human chondrocyte cell lines for examining the regulation of cartilage-specific matrix gene expression. However, the decreased synthesis and deposition of cartilage matrix associated with a rapid rate of proliferation has presented difficulties for further examination at the protein level. In these studies, proteoglycan synthesis was characterized in two chondrocyte cell lines, T/C-28a2 and tsT/AC62, derived, respectively, from juvenile costal and adult articular cartilage, under culture conditions that either promoted or decreased cell proliferation. Analysis of proteo[36S]glycans by Sepharose CL-4B chromatography and SDS-PAGE showed that the large proteoglycan aggrecan and the small, leucine-rich proteoglycans, decorin and biglycan, were produced under every culture condition studied. In monolayer cultures, a high initial cell density and conditions that promoted proliferation (presence of serum for T/C-28a2 cells or permissive temperature for the temperature-sensitive tsT/AC62 cells) favored cell survival and ratios of proteoglycans expected for differentiated chondrocytes. However, the tsT/AC62 cells produced more proteoglycans at the nonpermissive temperature. Culture of cells suspended in alginate resulted in a significant decrease in proteoglycan production in all culture conditions. While the tsT/AC62 cells continued to produce a larger amount of aggrecan than small proteoglycans, the T/C-28a2 cells lost the ability to produce significant amounts of aggrecan in alginate culture. In addition, our data indicate that immortalized chondrocytes may alter their ability to retain pericellular matrix under changing culture conditions, although the production of the individual matrix components does not change. These findings provide critical information that will assist in the development of a reproducible chondrocyte culture model for the study of regulation of proteoglycan biosynthesis in cartilage.
Kokenyesi
R
R
Department of Obstetrics, Gynecology and Women's Health, Saint Louis University, Missouri 63117, USA.
Tan
L
L
Robbins
J R
JR
Goldring
M B
MB
D000328:Adult; D002356:Cartilage / Q000378:metabolism; D002452:Cell Count; D002455:Cell Division; D002461:Cell Line, Transformed; D019902:Chondrocytes / Q000378:metabolism* / Q000473:pathology; D005109:Extracellular Matrix / Q000378:metabolism; D006801:Humans; D011509:Proteoglycans / Q000378:metabolism*; D013696:Temperature
D011509:Proteoglycans