Mechanical loading regulates NFATc1 and beta-catenin signaling through a GSK3beta control node.
Mechanical stimulation can prevent adipogenic and improve osteogenic lineage allocation of mesenchymal stem cells (MSC), an effect associated with the preservation of beta-catenin levels. We asked whether mechanical up-regulation of beta-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 GSK3beta in a Wnt-independent fashion. Small interfering RNA targeting GSK3beta prevented both strain-induced induction of beta-catenin and an increase in COX2, a factor associated with increased osteoprogenitor phenotype. Small interfering RNA knockdown of beta-catenin blocked mechanical reduction of peroxisome proliferator-activated receptor gamma and adiponectin, implicating beta-catenin in strain inhibition of adipogenesis. In contrast, the effect of both mechanical and pharmacologic inhibition of GSK3beta on the putative beta-catenin target, COX2, was unaffected by beta-catenin knockdown. GSK3beta inhibition caused accumulation of nuclear NFATc1; mechanical strain increased nuclear NFATc1, independent of beta-catenin. NFATc1 knockdown prevented mechanical stimulation of COX2, implicating NFATc1 signaling. Finally, inhibition of GSK3beta caused association of RNA polymerase II with the COX2 gene, suggesting transcription initiation. These results demonstrate that mechanical inhibition of GSK3beta induces activation of both beta-catenin and NFATc1 signaling, limiting adipogenesis via the former and promoting osteoblastic differentiation via NFATc1/COX2. Our novel findings suggest that mechanical loading regulates mesenchymal stem cell differentiation through inhibition of GSK3beta, which in turn regulates multiple downstream effectors.
Sen
Buer
B
Department of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599, USA. buer_sen@med.unc.edu
Styner
Maya
M
Xie
Zhihui
Z
Case
Natasha
N
Rubin
Clinton T
CT
Rubin
Janet
J
D050156:Adipogenesis / Q000502:physiology; D052242:Adiponectin / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology / Q000502:physiology; D019703:Calcineurin / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D038362:Glycogen Synthase Kinase 3 / Q000235:genetics / Q000378:metabolism*; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000502:physiology*; D051379:Mice; D050778:NFATC Transcription Factors / Q000235:genetics / Q000378:metabolism*; D047495:PPAR gamma / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D013314:Stress, Mechanical; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*
D052242:Adiponectin; D050778:NFATC Transcription Factors; D047495:PPAR gamma; D034741:RNA, Small Interfering; D051176:beta Catenin; D051546:Cyclooxygenase 2; D038362:Glycogen Synthase Kinase 3; D019703:Calcineurin; D002118:Calcium