Transforming growth factor-beta stimulates articular chondrocyte cell growth through p44/42 MAP kinase (ERK) activation.
Transforming growth factor-beta1 (TGF-beta1) stimulates articular chondrocyte cell proliferation and extracellular matrix formation. We reported previously that immediate and transient expression of c-fos mRNA through protein kinase C activation is required for the mitogenic effect of TGF-beta1 on cultured rat articular chondrocytes (CRAC). In gel kinase assays using myelin basic protein (MBP) showed that total cell lysates from cells treated with TGF-beta1 caused rapid phosphorylation of MBP, which suggests the involvement of mitogen-activated protein kinase (MAPK) activation. To identify specific MAPK pathways activated by TGF-beta1, we performed in vitro kinase assays using specific substrates. TGF-beta1 induced a rapid activation of extracellular signal regulated kinase (ERK) with a peak at 5 min, which decreased to basal levels within 240 min after TGF-beta1 stimulation. In contrast, the c-jun N-terminal kinase activity increased only about 2.5-fold after 240 min of stimulation and p38 MAPK activity did not change significantly. ERK activation by TGF-beta1 was also confirmed by in vivo phosphorylation assays of Elk1. However, a specific MEK1 inhibitor, PD98059, significantly decreased TGF-beta1 induced Elk1 phosphorylation in a dose-dependent manner. Furthermore, PD98059 reduced the TGF-beta1-induced cell growth by 40%. These results indicate that TGF-beta1 specifically activates MEK1 and subsequent ERK pathways in CRAC, and that the activation of this MAPK pathway plays a role in the mitogenic response to TGF-beta1.
Yonekura
A
A
Department of Orthopedic Surgery, Nagasaki University School of Medicine, Japan.
Osaki
M
M
Hirota
Y
Y
Tsukazaki
T
T
Miyazaki
Y
Y
Matsumoto
T
T
Ohtsuru
A
A
Namba
H
H
Shindo
H
H
Yamashita
S
S
D000818:Animals; D002455:Cell Division / Q000187:drug effects; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004789:Enzyme Activation; D008297:Male; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D017207:Rats, Sprague-Dawley; D012997:Solvents / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D012997:Solvents; D016212:Transforming Growth Factor beta; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D004121:Dimethyl Sulfoxide