Magnesium ion stimulation of bone marrow stromal cells enhances osteogenic activity, simulating the effect of magnesium alloy degradation

Magnesium alloys are being investigated for load-bearing bone fixation devices due to their initial mechanical strength, modulus similar to native bone, biocompatibility and ability to degrade in vivo. Previous studies have found Mg alloys to support bone regeneration in vivo, but the mechanisms have not been investigated in detail. In this study, we analyzed the effects of Mg2+ stimulation on intracellular signaling mechanisms of human bone marrow stromal cells (hBMSCs). hBMSCs were cultured in medium containing 0.8, 5, 10, 20 and 100 mM MgSO4, either with or without osteogenic induction factors. After 3 weeks, mineralization of extracellular matrix (ECM) was analyzed by Alizarin red staining, and gene expression was analyzed by quantitative polymerase chain reaction array. Mineralization of ECM was enhanced at 5 and 10 mM MgSO4, and collagen type X mRNA (COL10A1, an ECM protein deposited during bone healing) expression was increased at 10 mM MgSO4 both with and without osteogenic factors. We also confirmed the increased production of collagen type X protein by Western blotting. Next, we investigated the mechanisms of intracellular signaling by analyzing the protein production of hypoxia-inducible factor (HIF)-1α and 2α (transcription factors of COL10A1), vascular endothelial growth factor (VEGF) (activated by HIF-2α) and peroxisome proliferator-activated receptor gamma coactivator (PGC)-1α (transcription coactivator of VEGF). We observed that 10 mM MgSO4 stimulation enhanced COL10A1 and VEGF expression, possibly via HIF-2α in undifferentiated hBMSCs and via PGC-1α in osteogenic cells. These data suggest possible ECM proteins and transcription factors affected by Mg2+ that are responsible for the enhanced bone regeneration observed around degradable Mg orthopedic/craniofacial devices.