Dendritic cells (DCs) have been identified as effective antigen‐presenting cells (APCs). We demonstrate that extracellular matrix (ECM), hyaluronic acid (HA) and chondroitin sulphate A (CSA), in the presence of granulocyte‐macrophage colony‐stimulating factor (GM‐CSF), can rapidly promote the differentiation of monocyte‐derived immature DCs, as characterized by the remarkable upregulation of human leucocyte antigen (HLA‐DR), CD40, CD54, CD80 and CD86 expression to levels higher than those in the DCs generated by culturing with GM‐CSF and interleukin (IL)‐4 for 7 days and aggregation of the cells within 48 h. The upregulation of expression of HLA‐DR, CD40, CD54, CD80 and CD86 was dose‐dependent. Further studies showed that HA and CSA were able to augment nuclear factor (NF)‐κB activity, as determined by gel mobility shift assay and promote protein phosphorylation. Inhibition of NF‐κB by pyrolidine dithiocarbamate and sodium salicylate, and serine‐threonine and tyrosine kinase by starosporine as well as phosphatidylinositide‐3‐kinase (PI‐3‐K) by wortmannin could prevent the effects of HA and CSA on the expression of HLA‐DR, CD40, CD80 and CD86 in various degrees. Thus, our data demonstrate that HA or CSA can effectively and rapidly promote the differentiation of immature DC, suggesting that HA and CSA may possess a potential capacity in regulating immune responses.