Phenotypic characterization of osteoblasts from the sclerotic zones of osteoarthritic subchondral bone

Abstract

Objective: To determine the phenotype of osteoblasts from the sclerotic zones of human osteoarthritic (OA) subchondral bone.

Methods: Human osteoblasts were isolated from sclerotic or nonsclerotic areas of subchondral bone and cultured for 14 days in monolayer. The expression of 14 genes was investigated by real-time reverse transcription-polymerase chain reaction. The activities of alkaline phosphatase (AP) and transglutaminases (TGases) were quantified by enzymatic assays. C-terminal type I procollagen propeptide (CPI), interleukin-1beta (IL-1beta), IL-6, IL-8, transforming growth factor beta1 (TGFbeta1), osteocalcin (OC), and osteopontin (OPN) were assayed in the culture medium by immunoassay.

Results: Gene expression levels of matrix metalloproteinase 13, COL1A1 and COL1A2, OPN, tissue-nonspecific AP, OC, vascular endothelial growth factor, ANKH, TGase 2, factor XIIIA, and dentin matrix protein 1 were significantly up-regulated in sclerotic osteoblasts compared with nonsclerotic osteoblasts. In contrast, parathyroid hormone receptor gene expression was depressed in sclerotic osteoblasts, but bone sialoprotein levels were unchanged. The activities of AP and TGases were increased in sclerotic osteoblasts, while matrix mineralization, revealed by alizarin red staining, was decreased. In parallel, protein synthesis of CPI, OC, OPN, IL-6, IL-8, and TGFbeta1 was significantly higher in sclerotic osteoblasts than in nonsclerotic osteoblasts, while IL-1beta production was similar in both groups.

Conclusion: These findings contribute to a better understanding of the mechanisms involved in subchondral bone sclerosis and identify osteoblasts with an altered phenotype as a potential target for future OA therapies.