Effect of low-frequency pulsatile flow on expression of osteoblastic genes by bone marrow stromal cells

Perfusion culture of osteoprogenitor cells is a promising means to form a bone-like extracellular matrix for tissue engineering applications, but the mechanism by which hydrodynamic shear stimulates expression of bone extracellular matrix (ECM) proteins is not understood. Osteoblasts are mechanosensitive and respond differently to steady and pulsatile flow. Therefore, to probe the effect of flow, bone marrow stromal cells (BMSCs)--cultured under osteogenic conditions--were exposed to steady or pulsatile flow at frequencies of 0.015, 0.044, or 0.074 Hz. Following 24 h of stimulus, cells were cultured statically for an additional 13 days and then analyzed for the expression of bone ECM proteins collagen 1alpha1 (Col1alpha1), osteopontin, osteocalcin (OC), and bone sialoprotein (BSP). All mRNA levels were elevated by flow, but OC and BSP were enhanced modestly with pulsatile flow. To determine if these effects were related to gene induction during flow, BMSCs were again exposed to steady or pulsatile flow for 24 h, but then analyzed immediately for expression of growth and differentiation factors bone morphogenetic proteins (BMP)-2, -4, and -7, transforming growth factor (TGF)-beta1, and vascular endothelial growth factor-A. All growth and differentiation factors were significantly elevated by flow, except BMP-4 which was suppressed. In addition, expression of BMP-2 and -7 were enhanced and TGF-beta1 suppressed by pulsatile flow relative to steady flow. These results demonstrate that pulsatile flow modulates expression of BMP-2, -7, and TGF-beta1 and suggest that enhanced expression of bone ECM proteins by pulsatile flow may be mediated through the induction of BMP-2 and -7.