Aldosterone stimulates collagen gene expression and synthesis via activation of ERK1/2 in rat renal fibroblasts.
Recently, we demonstrated that in rats treated chronically with aldosterone and salt, severe tubulointerstitial fibrosis is associated with the activation of mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinases (ERK1/2). Here, we investigated whether aldosterone stimulates collagen synthesis via ERK1/2-dependent pathways in cultured rat renal fibroblasts. Gene expression of mineralocorticoid receptor (MR) and types I, II, III, and IV collagen was measured by real-time polymerase chain reaction (PCR). MR protein expression and ERK1/2 activity were evaluated by Western blotting analysis with anti-MR and anti-phospho-ERK1/2 antibodies, respectively. Collagen synthesis was determined by [3H]-proline incorporation. Significant levels of MR mRNA and protein expression were observed in rat renal fibroblasts. Treatment with aldosterone (0.1 to 10 nmol/L) increased ERK1/2 phosphorylation in a concentration-dependent manner with a peak at 5 minutes. Aldosterone (10 nmol/L) also increased the mRNA levels of types I, III, and IV collagen at 36 hours but had no effect on the type II collagen mRNA level. [3H]-proline incorporation was significantly increased by aldosterone in both the medium and cell layer at 48 hours. Aldosterone-induced ERK1/2 phosphorylation was markedly attenuated by pretreatment with eplerenone (10 micromol/L), a selective MR antagonist, or PD98059 (10 micromol/L), a specific inhibitor of MAPK kinase/ERK kinase, which is the upstream activator of ERK1/2. In addition, both eplerenone and PD98059 prevented the aldosterone-induced increases in types I, III, and IV collagen mRNA and [3H]-proline incorporation. These results suggest that aldosterone stimulates collagen gene expression and synthesis via MR-mediated ERK1/2 activation in renal fibroblasts, which may contribute to the progression of aldosterone-induced tubulointerstitial fibrosis.
Nagai
Yukiko
Y
Research Equipment Center, Kagawa Medical University, Kagawa, Japan.
Miyata
Kayoko
K
Sun
Guang-Ping
GP
Rahman
Matlubur
M
Kimura
Shoji
S
Miyatake
Akira
A
Kiyomoto
Hideyasu
H
Kohno
Masakazu
M
Abe
Youichi
Y
Yoshizumi
Masanori
M
Nishiyama
Akira
A
D000450:Aldosterone / Q000494:pharmacology*; D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis* / Q000235:genetics*; D003199:Computer Systems; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects*; D007668:Kidney / Q000166:cytology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D018161:Receptors, Mineralocorticoid / Q000378:metabolism
D012333:RNA, Messenger; D018161:Receptors, Mineralocorticoid; D000450:Aldosterone; D003094:Collagen; D048049:Extracellular Signal-Regulated MAP Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3