In Vitro Assessment of Bone and Cartilage Stimulating Peptides

Abstract

Spinal fusion is an effective and one of the most commonly used surgeries to treat intervertebral disc degeneration (IDD) related low back pain. However, non-fusion still occurs in about 10-15% of the clinical cases. The global objective of this project is to develop a synthetic bone graft for the spinal fusion, combined with bone growth enhancing factors named bone and cartilage stimulating peptides (BCSP®). The focus of this present study is to assess BCSPs’ performance in various concentrations with respect to their effects on osteoprogenitor proliferation, osteogenic differentiation, cell culture mineralization and cell migration. BCSPs appeared to enhance the bone growth in the previous study using an animal model; however, the cellular and molecular mechanisms of BCSPs performance remain unclear. In this study, we examined three BCSPs in vitro with respect to their ability to enhance proliferation, to induce early osteogenic differentiation and mineralization, and to simulate the migration of osteoprogenitor cells. The proliferation, early differentiation, mineralization and migration assessments were carried out with a double-stranded DNA quantification, an alkaline phosphatase (ALP, early osteogenic differentiation marker) expression assay, an Alizarin Red S staining assay and a Boyden Chamber assay, respectively. Osteoprogenitor cells examined included MC3T3-E1 preosteoblasts, human bone marrow-derived mesenchymal stem cells (hBM-MSCs), human periosteum-derived stem cells (hPDSCs), and rabbit periosteum-derived stem cells (rPDSCs). Both hBM-MSCs and MC3T3-E1 preosteoblasts exhibited a significant increase of ALP expression after the exposure to BCSP1 and BCSP7, and MC3T3-E1 preosteoblasts also exhibited a significant increase of ALP expression after the exposure to BCSP11. However, no significant difference was observed in proliferation, migration or mineralization assessments for any of the examined osteoprogenitor cells after exposure to BCSPs. The present work showed that BCSPs have a stimulating effect on the early differentiation of MC3T3-E1 preosoteblast and human BM-MSCs. However, BCSPs did not show any stimulating effects on the proliferation, mineralization and migration of osteoprogenitors listed, indicating that promoting early differentiation of osteoprogenitors might be one of the mechanisms of BCSPs’ in vivo bone growth stimulating effect.