Biological Effects of Phosphocitrate on Osteoarthritic Articular Chondrocytes

Abstract

Background: Phosphocitrate (PC) inhibits osteoarthritis (OA) in Hartley guinea pigs. However, the underlying molecular mechanisms remain poorly understood.

Objective: This study sought to examine the biological effect of PC on OA chondrocytes and test the hypothesis that PC may exert its OA disease modifying effect, in part, by inhibiting the expression of genes implicated in OA disease process and stimulating the production of extracellular matrices.

Method: OA chondrocytes were cultured in the absence or presence of PC. Total RNA was extracted and subjected to microarray analyses. The effect of PC on proliferation and chondrocyte-mediated calcification were examined in monolayer culture. The effect of PC on the production of extracellular matrices was examined in micromass culture.

Results: PC downregulated the expression of numerous genes classified in proliferation and apoptosis while upregulating the expression of many genes classified in transforming growth factor-β (TGF-β) receptor signaling pathway and ossification. PC also downregulated the expressions of many genes classified in inflammatory response and Wnt receptor signaling pathways. Consistent with its effect on the expression of genes classified in proliferation, ossification, and skeletal development, PC inhibited the proliferation of OA chondrocytes and chondrocyte-mediated calcification while stimulating the production of extracellular matrices.

Conclusion: PC may exert its OA disease modifying effect, in part, through a crystal-independent mechanism or by inhibiting the expressions of many genes implicated in OA disease process, and at the same time, stimulating the expression of genes implicated in chondroprotection and production of extracellular matrices.

Keywords: Calcification; Chondrocyte; Matrix; Microarray; Osteoarthritis; Phosphocitrate.

Fig. (1)

Effect of PC on chondrocyte proliferation. Left bar group-numbers of foreskin fibroblasts cultured in the absence and presence of PC. No effect was noted. Right bar group - numbers of OA chondrocytes cultured in the absence and presence of PC. PC inhibited the proliferation of OA chondrocytes. * = p < 0.01 versus untreated control.

Fig. (2)

Representative images of alizarin red stained OA chondrocytes. Calcium deposits were detected in monolayer of OA chondrocytes cultured in osteogenetic differentiation medium, but not in the monolayer of OA chondrocytes cultured in osteogenetic differentiation medium containing 1 mm PC. (bar = 10 μm).

Fig. (3)

Representative images of picrosirius red and alcian blue stained sections of micromasses of OA chondrocytes. Arrows – outer layers of the micromasses of OA chondrocytes. (bar = 100 μm). PC stimulated the production of collagens and proteoglycans.