Mitogen-activated protein kinase signaling in bovine articular chondrocytes in response to fluid flow does not require calcium mobilization.
In the present study, the role of mitogen-activated protein kinases (MAPKs) in chondrocyte mechanotransduction was investigated. We hypothesized that MAPKs participate in fluid flow-induced chondrocyte mechanotransduction. To test our hypothesis, we studied cultured chondrocytes subjected to a well-defined mechanical stimulus generated with a laminar flow chamber. The extracellular signal-regulated kinases 1 and 2 (ERK1/2) were activated 1.6-3-fold after 5-15 min of fluid flow exposure corresponding to a chamber wall shear stress of 1.6 Pa. Activation of ERK1/2 was observed in the presence of both 10% FBS and 0.1% BSA, suggesting that the flow effects do not require serum agonists. Treatment with thapsigargin or EGTA had no significant effect on the ERK1/2 activation response to flow, suggesting that Ca2+ mobilization is not required for this response. To assess downstream effects of the activated MAPKs on transcription, flow studies were performed using chondrocytes transfected with a chimeric luciferase construct containing 2.4 kb of the promoter region along with exon 1 of the human aggrecan gene. Two-hour exposure of transfected chondrocytes to fluid flow significantly decreased aggrecan promoter activity by 40%. This response was blocked by treatment of chondrocytes with the MEK-1 inhibitor PD98059. These findings demonstrate that, under the conditions of the present study, fluid flow-induced signals activate the MEK-1/ERK signaling pathway in articular chondrocytes, leading to down-regulation of expression of the aggrecan gene.
Hung
C T
CT
Cellular Engineering Laboratory, Center for Biomedical Engineering, Columbia University, New York, NY 10027, USA.
Henshaw
D R
DR
Wang
C C
CC
Mauck
R L
RL
Raia
F
F
Palmer
G
G
Chao
P H
PH
Mow
V C
VC
Ratcliffe
A
A
Valhmu
W B
WB
D053674:Aggrecans; D000818:Animals; D001696:Biomechanical Phenomena; D002118:Calcium / Q000378:metabolism*; D002358:Cartilage, Articular / Q000166:cytology / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D004533:Egtazic Acid / Q000494:pharmacology; D016326:Extracellular Matrix Proteins; D005419:Flavonoids / Q000494:pharmacology; D015870:Gene Expression; D006801:Humans; D037181:Lectins, C-Type; D048369:MAP Kinase Kinase 1; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D017346:Protein Serine-Threonine Kinases; D011509:Proteoglycans / Q000235:genetics; D015398:Signal Transduction; D019284:Thapsigargin / Q000494:pharmacology; D014162:Transfection
D053674:Aggrecans; D016326:Extracellular Matrix Proteins; D005419:Flavonoids; D037181:Lectins, C-Type; D011509:Proteoglycans; D004533:Egtazic Acid; D019284:Thapsigargin; D017346:Protein Serine-Threonine Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; C482125:MAP2K1 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; D002118:Calcium