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Fluid shear stress (FSS) is an important biomechanical factor regulating the osteogenic differentiation of human mesenchymal stem cells (hMSCs) and is therefore widely used in bone tissue engineering. However, the mechanotransduction of FSS in hMSCs remains largely unknown. As β1 integrins are considered to be important mechanoreceptors in other cells, we suspect that β1 integrins should also be important for hMSCs to sense the stimulation of FSS. We used a perfusion culture system to produce FSS loading on hMSCs seeded in PLGA three-dimensional (3D) scaffolds and investigated the roles of β1 integrins, FAK and ERK1/2 in FSS-induced osteogenic differentiation of hMSCs. Our results showed that FSS not only markedly increased ALP activity and the expression of ALP, OCN, Runx2 and COLIα genes but also significantly enhanced the phosphorylation of ERK1/2, Runx2 and FAK. FSS-induced activation of ERK1/2 and FAK was inhibited by blockade of the connection between β1 integrins and ECM with RGDS peptide and integrins β1 monoclonal antibody. Our study also found that FSS could upregulate the expression level of β1 integrins and that this upregulation could be abolished by PD98059. Further investigation indicated that FSS-activated ERK1/2 led to the phosphorylation of IκBα and NFκB p65. The activation of NFκB p65 resulted in the upregulation of β1 integrin expression. Therefore, it could be inferred that β1 integrins should sense the stimulation of FSS and thus activate ERK1/2 through activating of FAK, and FSS-activated ERK1/2 feedback to upregulate the expression of β1 integrins through activating NFκB., (Copyright © 2012 John Wiley & Sons, Ltd.)

A reasonable mechanical microenvironment similar to the bone microenvironment in vivo is critical to the formation of engineering bone tissues. As fluid shear stress (FSS) produced by perfusion culture system can lead to the osteogenic differentiation of human mesenchymal stem cells (hMSCs), it is widely used in studies of bone tissue engineering. However, effects of FSS on the differentiation of hMSCs largely depend on the FSS application manner. It is interesting how different FSS application manners influence the differentiation of hMSCs. In this study, we examined the effects of intermittent FSS and continuous FSS on the osteogenic differentiation of hMSCs. The phosphorylation level of ERK1/2 and FAK is measured to investigate the effects of different FSS application manners on the activation of signaling molecules. The results showed that intermittent FSS could promote the osteogenic differentiation of hMSCs. The expression level of osteogenic genes and the alkaline phosphatase (ALP) activity in cells under intermittent FSS application were significantly higher than those in cells under continuous FSS application. Moreover, intermittent FSS up-regulated the activity of ERK1/2 and FAK. Our study demonstrated that intermittent FSS is more effective to induce the osteogenic differentiation of hMSCs than continuous FSS.

It is a classical signaling pathway that the activation of extracellular signal-regulated kinase1/2 (ERK1/2) results in the phosphorylation of runt-related transcription factor 2 (Runx2) and thereby initiates the transcription of osteogenic genes. Recently, it is found that the activation of ERK1/2 resulted from fluid shear stress (FSS) also increased the expression of Runx2 and β1 integrins, and finally enhanced osteogenic differentiation. However, it has been remained largely unknown how ERK1/2 regulates the expression of Runx2 and β1 integrins. We use the perfusion culture system to produce FSS exerting on human bone marrow-derived mesenchymal stem cells (hMSCs) and thus activate ERK1/2. Our study demonstrated that FSS-activated ERK1/2 mediated the expression of osteogenic genes via two novel signaling pathways except for the classical signaling pathway: feedback up-regulation of β1 integrins expression via activating nuclear factor kappa B (NF-κB), and activation of bone morphogenesis proteins (BMPs)/mothers against decapentaplegic (Smad) pathway via activating NF-κB and thereby regulating Runx2 expression. These signaling pathways combined with the classical signaling pathway, with ERK1/2 as a hub node molecule, form a molecular signaling cross-talking network to induce the osteogenic differentiation of hMSCs. The understanding on the mechanism of FSS inducing the osteogenic differentiation of hMSCs will not only be helpful to develop the bone tissue engineering but also provide new targets for drug discovery for treatment of osteoporosis and other related bone-wasting diseases., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Mechanical stimuli can improve bone function by promoting the proliferation and differentiation of bone cells and osteoblasts. As precursors of osteoblasts, human mesenchymal stem cells (hMSCs) are sensitive to mechanical stimuli. In recent years, fluid shear stress (FSS) has been widely used as a method of mechanical stimulation in bone tissue engineering to induce the osteogenic differentiation of hMSCs. However, the mechanism of this differentiation is not completely clear. Several signaling pathways are involved in the mechanotransduction of hMSCs responding to FSS, such as MAPK, NO/cGMP/PKG and Ca(2+) signaling pathway. Here, we briefly review how hMSCs respond to fluid flow stimuli and focus on the signal molecules involved in this mechanotransduction.

Microgravity (MG) leads to a decrease in osteogenic potential of human bone marrow-derived mesenchymal stem cells (hMSCs). In the present study, we used large gradient high magnetic field (LGHMF) produced by a superconducting magnet to model MG (LGHMF-MG) and analyzed the effects of LGHMF-MG on survival, cytoskeleton and osteogenic potential of hMSCs. Results showed that the LGHMF-MG treatment for 6 h disrupted the cytoskeleton of hMSCs, and the LGHMF-MG treatment for 24 h led to cell death. LGHMF-MG treatments for 6 h in early stages of osteogenic induction (the pre-treatment before osteogenic induction, the beginning-treatment in the beginning-stage of osteogenic induction and the middle-treatment in the middle-stage of osteogenic induction) resulted in suppression on osteogenesis of hMSCs. The suppression intensity was reduced gradually as the treatment stage of LGHMF-MG was postponed. The LGHMF-MG treatment for 6 h in the ending-stage of osteogenic induction (the ending-treatment) had no obvious effect on osteogenesis of hMSCs. These results indicated that LGHMF-MG should affect the initiation of osteogenesis. Finally, the possible mechanism for the inhibition effect of LGHMF-MG on osteogenesis of hMSCs is discussed.