Effects of CTGF/Hcs24, a product of a hypertrophic chondrocyte-specific gene, on the proliferation and differentiation of chondrocytes in culture

Recently, we cloned a messenger RNA (mRNA) predominantly expressed in chond rocytes from a human chondrosarcoma-derived chondrocytic cell line, HCS-2/8 , by differential display PCR and found that its gene, named hcs24, was ide ntical with that of connective tissue growth factor (CTGF). Here we investi gated CTGF/Hcs24 func- tion in the chondrocytic cell line HCS-2/8 and rabbi t growth cartilage (RGC) cells. HCS-2/8 cells transfected with recombinant adenoviruses that generate CTGF/Hcs24 sense RNA (mRNA) proliferated more ra pidly than HCS-2/8 cells transfected with control adenoviruses. HCS-2/8 cel ls transfected with recombinant adenoviruses that generate CTGF/Hcs24 sense RNA expressed more mRNA of aggrecan and type X collagen than the control c ells. To elucidate the direct action of CTGF/Hcs24 on the cells, we transfe cted HeLa cells with CTGF/Hcs24 expression vectors, obtained stable transfe ctants, and purified recombinant CTGF/Hcs24 protein from conditioned medium of the transfectants. The recombinant CTGF/Hcs24 effectively promoted the proliferation of HCS-2/8 cells and RGC cells in a dose-dependent manner and also dose dependently increased proteoglycan synthesis in these cells. In addition, these stimulatory effects of CTGF/Hcs24 were neutralized by the a ddition of anti-CTGF antibodies. Furthermore, the recombinant CTGF/Hcs24 ef fectively increased alkaline phosphatase activity in RGC cells in culture. Moreover, RT-PCR analysis revealed that the recombinant CTGF/Hcs24 stimulat ed gene expression of aggrecan and collagen types II and X in RGC cells in culture. These results indicate that CTGF/Hcs24 directly promotes the proli feration and differentiation of chondrocytes.