Establishment of a novel chondrocytic cell line N1511 derived from p53-null mice

We established a clonal chondrocytic cell line N1511 derived from rib cartilage of a p53-null mouse. N1511 cells proliferated in polygonal shape and elicited differentiation at confluence when treated with combination of bone morphogenetic protein (BMP) 2 and insulin or parathyroid hormone (PTH) and dexamethasone. BMP-2/insulin-treated cells became refractile without forming cartilaginous nodules and reached terminal differentiation, became positive for alizarin red staining, and developed considerable ALP activity. In contrast, PT]PTH/dexamethasone-treated cells formed Alcian blue-positive nodules but remained negative for alizarin red staining and ALP activity. Northern blot analysis revealed that BMP-2/insulin-treated cells sequentially expressed type II, IX, and X collagens, whereas PTH/dexamethasone-treated cells slowly expressed type II collagen and then type IX, and they did not exhibit type X collagen expression. These results show that BMP-2/insulin treatment induces full differentiation toward hypertrophy, whereas treatment with PTII/ dexamethasone slows and limits differentiation. Recovery of p53 expression in N1511 cells by transient transfection inhibited cell proliferation, suggesting that cell proliferation could be regulated with p53 in this cell line. These results indicate that N1511 is the only cell line with known genetic mutation, which undergoes multiple steps of chondrocyte differentiation toward hypertrophy, and because proliferation could be regulated by expression of p53, N1511 could be an excellent model for studies of chondrogenesis, the function of p53, and genetic engineering of cartilage tissue.