Inhibitory effects of fasudil on renal interstitial fibrosis induced by unilateral ureteral obstruction.
Renal fibrosis is the major cause of chronic kidney disease, and the Rho/Rho-associated coiled-coil kinase (ROCK) signaling cascade is involved in the renal fibrotic processes. Several studies have reported that ROCK inhibitors attenuate renal fibrosis. However, the mechanism of this process remains to be fully elucidated. The present study assessed the inhibitory effect of fasudil, a ROCK inhibitor using immunohistochemistry, reverse transcription-quantitative polymerase chain reaction and western blot analyses, in vivo and in vitro, to elucidate the mechanisms underlying renal interstitial fibrosis. In mice induced with unilateral ureteral obstruction (UUO), collagen accumulation, the expression of fibrosis‑associated genes and the content of hydroxyproline in the kidney increased 3, 7, and 14 days following UUO. Fasudil attenuated the histological changes, and the production of collagen and extracellular matrix in the UUO kidney. The expression of α‑smooth muscle actin (α‑SMA) and the transforming growth factor‑β (TGFβ)‑Smad signaling pathway, and macrophage infiltration were suppressed by fasudil in the kidneys of the UUO mice. The present study also evaluated the role of intrinsic renal cells and infiltrated macrophages using NRK‑52E, NRK‑49F and RAW264.7 cells. The mRNA and protein expression levels of collagen I and α‑SMA increased in the NRK‑52E and NRK‑49F cells stimulated by TGF‑β1. Hydroxyfasudil, a bioactive metabolite of fasudil, attenuated the increase in the mRNA and protein expression levles of α‑SMA in the two cell types. However, the reduction in the mRNA expression of collagen I was observed in the NRK‑49F cells only. Hydroxyfasudil decreased the mRNA expression of monocyte chemoattractant protein‑1 (MCP‑1) induced by TGF‑β1 in the NRK‑52E cells, but not in the NRK‑49F cells. In the RAW264.7 cells, the mRNA expression levels of MCP‑1, interleukin (IL)‑1β, IL‑6 and tumor necrosis factor α were increased significantly following lipopolysaccharide stimulation, and were not suppressed by hydroxyfasudil. These data suggested that the inhibition of ROCK activity by fasudil suppressed the transformation of renal intrinsic cells into the myofibroblast cells, and attenuated the infiltration of macrophages, without inhibiting the expression or the activation of cytokine/chemokines, in the progression of renal interstitial fibrosis.
Pharmacology Research Laboratories II, Research Division, Mitsubishi Tanabe Pharma Corporation, Toda‑shi, Saitama 335‑8505, Japan.
Itsuko
I
Baba
Pharmacology Research Laboratories II, Research Division, Mitsubishi Tanabe Pharma Corporation, Toda‑shi, Saitama 335‑8505, Japan.
Yasuhiro
Y
Egi
Safety Research Laboratory, Research Division, Mitsubishi Tanabe Pharma Corporation, Toda‑shi, Saitama 335‑8505, Japan.
Hiroyuki
H
Utsumi
Safety Research Laboratory, Research Division, Mitsubishi Tanabe Pharma Corporation, Toda‑shi, Saitama 335‑8505, Japan.
Tetsuhiro
T
Kakimoto
Pharmacology Research Laboratories II, Research Division, Mitsubishi Tanabe Pharma Corporation, Toda‑shi, Saitama 335‑8505, Japan.
Kazuo
K
Suzuki
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000627:therapeutic use; D000199:Actins / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D004195:Disease Models, Animal; D005355:Fibrosis / Q000188:drug therapy / Q000209:etiology*; D007668:Kidney / Q000378:metabolism / Q000473:pathology*; D007674:Kidney Diseases / Q000209:etiology / Q000517:prevention & control; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D047428:Protein Kinase Inhibitors / Q000494:pharmacology* / Q000627:therapeutic use; D015398:Signal Transduction / Q000187:drug effects; D051785:Smad Proteins / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology; D015854:Up-Regulation / Q000187:drug effects; D014517:Ureteral Obstruction / Q000150:complications* / Q000188:drug therapy / Q000473:pathology
D000199:Actins; D024042:Collagen Type I; D047428:Protein Kinase Inhibitors; D051785:Smad Proteins; D053773:Transforming Growth Factor beta1; C544013:alpha-smooth muscle actin, mouse; C431085:hydroxyfasudil; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; C049347:fasudil