Focal Adhesion Kinase Is Important for Fluid Shear Stress-Induced Mechanotransduction in Osteoblasts

Abstract

Mechanical loading of bone is important for maintenance of bone mass and structural stability of the skeleton. When bone is mechanically loaded, movement of fluid within the spaces surrounding bone cells generates fluid shear stress (FSS) that stimulates osteoblasts, resulting in enhanced anabolic activity. The mechanisms by which osteoblasts convert the external stimulation of FSS into biochemical changes, a process known as mechanotransduction, remain poorly understood. Focal adhesions are prime candidates for transducing external stimuli. Focal adhesion kinase (FAK), a nonreceptor tyrosine kinase found in focal adhesions, may play a key role in mechanotransduction, although its function has not been directly examined in osteoblasts. We examined the role of FAK in osteoblast mechanotransduction using short interfering RNA (siRNA), overexpression of a dominant negative FAK, and FAK 2/2 osteoblasts to disrupt FAK function in calvarial osteoblasts. Osteoblasts were subjected to varying periods oscillatory fluid flow (OFF) from 5 min to 4 h, and several physiologically important readouts of mechanotransduction were analyzed including: extracellular signal-related kinase 1/2 phosphorylation, upregulation of c-fos, cyclooxygenase-2, and osteopontin, and release of prostaglandin E 2 . Osteoblasts with disrupted FAK signaling exhibited severely impaired mechanical responses in all endpoints examined. These data indicate the importance of FAK for both short and long periods of FSS-induced mechanotransduction in osteoblasts.

Protein Kinase G and Focal Adhesion Kinase Converge on Src/Akt/β-Catenin Signaling Module in Osteoblast Mechanotransduction

“…We found a similar decrease in FSS-induced Erk phosphorylation (supplemental Fig. 3, A and D), confirming previous reports in FAKdeficient primary rodent osteoblasts exposed to FSS (20,37). In stark contrast, cGMP-induced Akt and Erk phosphorylation were unaffected by FAK knockdown (Fig.…”

“…Previous work suggests that focal adhesion kinase (FAK) is upstream of Akt, because a dominant negative FAK blocks FSSinduced Akt and Erk activation (19,20). In many cell types, FAK and Src function within a network of integrin-stimulated signaling pathways leading to activation of the Ras/Raf/MEK/Erk and phosphatidylinositol 3-kinase (PI3K)/Akt signaling cascades (21)(22)(23).…”

“…In many cell types, FAK and Src function within a network of integrin-stimulated signaling pathways leading to activation of the Ras/Raf/MEK/Erk and phosphatidylinositol 3-kinase (PI3K)/Akt signaling cascades (21)(22)(23). However, whether FAK and Src interact to regulate Akt in shear-stressed osteoblasts is unknown, and whether FAK is activated by FSS is controversial (19,20,24). Akt activation by growth factors requires PI3K-mediated synthesis of phosphatidylinositol 3,4,5-triphosphate, which recruits Akt and its activating kinases to the plasma membrane (25).…”

“…Erk is blocked by expression of a dominant negative FAK, but the latter may disrupt scaffolding functions of FAK (19,24,38); osteoblasts from FAK-deficient mice show reduced FSSinduced Erk phosphorylation, but Akt was not examined (20,37). We sought to determine the roles of FAK and Src in FSSand cGMP-induced Akt phosphorylation using an siRNA approach, which allowed specific depletion of each protein (Fig.…”

MicroRNA-138 regulates osteogenic differentiation of human stromal (mesenchymal) stem cells in vivo

“…The focal adhesion kinase (FAK) signaling pathway is suggested to provide a link between activation of ERK1/2 by ECM proteins (7), thereby stimulating subsequent phosphorylation of Runx2/Cbfa-1, a key transcription factor controlling osteogenic gene expression (7). Furthermore, disruption of FAK signaling in osteoblasts indicates that FAK is a critical factor in the mechanotransduction pathway for osteoblast differentiation (8).…”