| Abstract |
Chondroitin sulfate (CS) is a principal extracellular matrix component in cartilaginous tissues, well known for its role in cartilage elasticity owing to its physicochemical properties. However, its intrinsic role in chondrogenesis remains unelucidated. This study aimed to investigate the expression pattern and corresponding effects of 4-sulfated CS chains throughout the differentiation processes in chondrogenic ATDC5 cells. Herein, chondroitin 4-O-sulfotransferase-1 (C4ST-1) knockdown (KD) subclones, a potential late-limiting enzyme in 4-sulfated CS biosynthesis, were constructed. Notably, C4ST-1 KD did not affect the initial cellular condensation of ATDC5 cells, albeit it suppressed subsequent chondrogenic differentiation, markedly upregulating the production of cartilage matrix macromolecules, including proteoglycans bearing 4-sulfated CS chains. Additionally, bone morphogenetic protein (BMP)-4-mediated chondrogenic signaling events, including small mothers against decapentaplegic homolog 1/5/9 phosphorylation and N-cadherin cleavage, showed marked downregulation in C4ST-1 KD cells. Bath application of exogenous BMP-4 partially restored N-cadherin cleavage rate and upregulated the expression of chondrocyte markers in C4ST-1 KD cell cultures. Altogether, the findings of this study strongly suggest the function of 4-sulfated CS chains as an extracellular regulator, in addition to their role as a structural support for cartilage tissues, which effectively controls the progression of chondrocyte differentiation.
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