PURIFICATION OF GUINEA-PIG YKL40 AND MODULATION OF ITS SECRETION BY CULTURED ARTICULAR CHONDROCYTES

The aim of this study was to purify, characterize, and study the regul ation at the chondrocyte level of the guinea pig (gp) homologue of hum an (R) YKL40, a putative marker of arthritic disorders. Studying YKL40 in guinea pigs is of particular interest, as age-related osteoarthrit is develops in this species spontaneously. Both N-terminal sequencing and total amino acid composition of gpYKL40 purified from the secretio n medium of cultured articular chondrocytes indicate a high degree of identity with hYKL40. gpYKL40 was found to contain complex N-linked ca rbohydrate, as demonstrated by N-glycosidase F and endoglycosidase F d igestion. Isoelectric focusing demonstrated the presence of a major ba nd at pi 6.7. The secretion of gpYKL40 by confluent articular chondroc ytes in the extracellular medium was studied by immunoblotting. gpYKL4 0 was released by chondrocytes continuously over a 7 day period and di d not appeal to be degraded by proteinases, as its signal intensity in cell-free medium at 37 degrees C did not decrease with time. Thus, gp YKL40 displays high stability and accumulates in extracellular medium without reaching a steady-state level. Among the main factors known to regulate cartilage metabolism IL-1 beta, TNF-alpha, bFGF, or 1,25(OH) (2)D-3 did not alter the basal level of gpYKL40, and retinoic acid had a slight inhibitory effect; TGF-beta and IGF-I and -II dose-dependent ly and inversely modulated this basal level. TGF-beta at 5 ng/ml decre ased extracellular gpYKL40 2.9-fold, whereas IGF-I and IGF-II at 50 ng /ml increased extracellular gpYKL40 3.6- and 3.4-fold, respectively. T he present biochemical and biological Findings give new insights for s tudying the function of YKL40 in cartilage. (C) 1998 Wiley-Liss, Inc.