Aldosterone Stimulates Collagen Gene Expression and Synthesis Via Activation of ERK1/2 in Rat Renal Fibroblasts

Abstract

Abstract-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 [ 3 H]-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.[3 H]-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 mol/L), a selective MR antagonist, or PD98059 (10 mol/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 [ Key Words: aldosterone Ⅲ collagen Ⅲ fibroblasts Ⅲ mineralocorticoids R ecent studies have indicated the usefulness of mineralocorticoid receptor (MR) antagonists in ameliorating renal injury. [1][2][3][4][5][6][7][8][9][10][11][12][13] In stroke-prone spontaneously hypertensive rats 4 and rats treated with angiotensin II and a nitric oxide synthase inhibitor, 5 cyclosporine A, 6 or radiation, 7 MR antagonists had no effect on systemic blood pressure but markedly ameliorated glomerular and tubulointerstitial fibrosis. In clinical studies, the addition of a nonselective MR antagonist, spironolactone, to angiotensin-converting enzyme inhibitors had no hemodynamic effects but markedly reduced proteinuria in patients with chronic renal failure 8 and early diabetic nephropathy. 9 It has also been shown that monotherapy with spironolactone 10 or eplerenone, 11 a selective MR antagonist, is more effective than angiotensin-converting enzyme inhibitors in reducing proteinuria in hypertensive patients. Furthermore, White et al 12 showed that in hypertensive patients, eplerenone had a similar blood pressurelowering effect to that of a calcium antagonist, amlodipine, but reduced the urinary albumin-to-creatinine ratio to a greater extent. These observations suggest that MR blockade has renoprotective effects through mechanisms that cannot be simply explained by blood pressure and hemodynamic changes.R...

Involvements of Rho-Kinase and TGF-β Pathways in Aldosterone-Induced Renal Injury

“…We recently found that aldosterone stimulates reactive oxygen species production (15) and induces cell proliferation (16) or deformation (16) in cultured glomerular mesangial cells. In addition, aldosterone stimulated collagen synthesis in cultured renal fibroblasts (17). Collectively, these data suggest that aldosterone has deleterious effects on the kidney that cannot be explained simply by BP changes.…”

“…␤ 1 , CTGF, and monocyte chemoattractant protein-1 (MCP-1) were analyzed by real-time PCR using an ABI Prism 7000 Sequence Detection System (Applied Biosystems, Foster City, CA) (17,33,34). The oligonucleotide primer sequences of GAPDH and types I and III collagen were reported previously (17,33).…”

“…here is increasing evidence to support potential roles of aldosterone in the pathogenesis of renal injury (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). Severe glomerular injury and interstitial fibrosis were observed in rats that were treated long term with aldosterone/ salt (3,4).…”

Aldosterone-induced osteopontin gene transcription in vascular smooth muscle cells involves glucocorticoid response element

“…Promoter activity was determined by a GloMax 96 Microplate Luminometer (Promega) using a Dual-Glo Luciferase Assay System (Promega). As for the concentration of Aldo 10 À7 M, in the promoter assay, we referred other investigators' studies 23 …”

Emodin inhibits extracellular matrix synthesis by suppressing p38 and ERK1/2 pathways in TGF-β1-stimulated NRK-49F cells

“…The most important downstream factors mediating the ECM synthesis of TGF-β1 are mitogen-activated protein kinase (MAPK) signals, including extracellular signal-regulated kinase (ERK), p38 MAP kinase and c-jun N-terminal kinase (JNK). ERK mediates TGF-β1-induced synthesis of plasminogen activator inhibitor type I (6), CTGF (7) and aldsterone-stimulated collagen synthesis (8). ERK is also involved in the ECM synthesis in diabetic rats (9).…”