Abstract
The pathological relevance of articular cartilage calcification in osteoarthritis (OA) is becoming increasingly evident. We are only beginning to understand the pathobiological mechanisms that contribute to articular cartilage calcification in OA. How molecular environmental factors interact with calcification mechanisms is poorly explored. In this study, we developed an in vitro phosphate-driven calcification model for human OA articular chondrocytes, in which these cells are cultured in the presence of calcification medium containing adenosine triphosphate (ATP) and β-glycerophosphate (BGP). We employed this model to investigate the role of transforming growth factor β (TGF-β) in chondrocyte calcification. Chondrocyte culture in calcification medium resulted in mineral nodule formation over a time course of 7 days. The presence of calcium and phosphate deposition in these nodules was validated with von Kossa staining, scanning electron microscopy-energy-dispersive X-ray spectroscopy (SEM-EDX), and colorimetric calcium and phosphate assays. Supplementation of calcification medium with TGF-β resulted in enhanced nodule formation with a different morphology and changed the expression of extracellular matrix-related genes such as collagen type I and III. In conclusion, we developed a new in vitro model for human OA articular chondrocyte calcification, in which we demonstrated a pro-calcifying role for TGF-β. This in vitro model may be used as a basis to aid the investigation of the influence of environmental factors on chondrocyte calcification and the development of new anti-calcification disease-modifying osteoarthritis drugs.