Mechanical Loading Regulates NFATc1 and -Catenin Signaling through a GSK3 Control Node*

Mechanical stimulation can prevent adipogenic and improve osteogenic lineage allocation of mesenchymal stem cells (MSC), an effect associated with the preservation of -catenin levels. We asked whether mechanical up-regulation of -catenin was critical to reduction in adipogenesis as well as other mechanical events inducing alternate MSC lineage selection. In MSC cultured under strong adipogenic conditions, mechanical load (3600 cycles/day, 2% strain) inactivated GSK3 in a Wnt-independent fashion. Small interfering RNA targeting GSK3 prevented both strain-induced induction of -catenin and an increase in COX2, a factor associated with increased osteoprogenitor phenotype. Small interfering RNA knockdown of -catenin blocked mechanical reduction of peroxisome proliferator-activated receptor and adiponectin, implicating -catenin in strain inhibition of adipogenesis. In contrast, the effect of both mechanical and pharmacologic inhibition of GSK3 on the putative -catenin target, COX2, was unaffected by -catenin knockdown. GSK3 inhibition caused accumulation of nuclear NFATc1; mechanical strain increased nuclear NFATc1, independent of -catenin. NFATc1 knockdown prevented mechanical stimulation of COX2, implicating NFATc1 signaling. Finally, inhibition of GSK3 caused association of RNApolymerase IIwith theCOX2 gene, suggesting transcription initiation. These results demonstrate that mechanical inhibition of GSK3 induces activation of both -catenin and NFATc1 signaling, limiting adipogenesis via the former and promoting osteoblastic differentiation via NFATc1/COX2. Our novel findings suggest thatmechanical loading regulates mesenchymal stem cell differentiation through inhibition of GSK3 , which in turn regulates multiple downstream effectors.