Abstract
It has been shown that the inhibition of the Rho/Rho kinase (ROCK) pathway prevents tubulointerstitial fibrosis and ameliorates renal function in various progressive renal disorders. The present study was to determine whether fasudil, a ROCK inhibitor, has a protective effect on cyclosporine A (CsA)-induced nephropathy. Male Sprague-Dawley rats were treated with CsA (n = 10, 20 mg · kg−1 day−1 s.c.), CsA + fasudil (n = 10, 3 mg · kg−1 day−1 i.p.), or vehicle alone (n = 10) for 28 days. Fasudil cotreatment ameliorated CsA-induced changes and restored renal function. CsA decreased the expression of endothelial nitric-oxide synthase and increased inducible nitric-oxide synthase/3-nitrotyrosine in the kidney. Accordingly, there was infiltration of inflammatory cells and up-regulation of inflammatory cytokines. Fasudil also significantly suppressed the expression of transforming growth factor-β1, Smad signaling, and subsequent epithelial-to-mesenchymal processes. In addition, fasudil augmented p27kip1 expression and decreased the number of proliferating cell nuclear antigen-positive cells. In another series of experiments using HK-2 cells in culture, fasudil also suppressed CsA-induced increases in mitogen-activated protein kinase phosphorylation. CsA induced expression of p53, the degree of which was attenuated by fasudil in association with decreases of proapoptotic markers such as Bad, Bax, and total/cleaved caspase-3. These results suggest that inhibition of the Rho/ROCK pathway attenuates CsA-induced nephropathy through the suppression of the induction of inflammatory, apoptotic, and fibrogenic factors, along with inhibition of Smad, mitogen-activated protein kinases, and nitric oxide signaling pathways.
Footnotes
This work was supported by the Korea Science and Engineering Foundation through the Medical Research Center for Gene Regulation [Grant R13-2002-013-05004-0] at Chonnam National University; and the Chonnam National University Hospital Research Institute of Clinical Medicine [Grant CRI10034-1].
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
doi:10.1124/jpet.111.179457.
↵ The online version of this article (available at http://jpet.aspetjournals.org) contains supplemental material.
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ABBREVIATIONS:
- CsA
- cyclosporine A
- ROCK
- Rho kinase
- MAPK
- mitogen-activated protein kinase
- MYPT1
- myosin light chain phosphatase
- NF-κB
- nuclear factor κB
- TGF-β1
- transforming growth factor-β1
- CDK
- cyclin-dependent kinase
- EMT
- epithelial-to-mesenchymal transition
- eNOS
- endothelial nitric-oxide synthase
- iNOS
- inducible nitric-oxide synthase
- PCNA
- proliferating cell nuclear antigen
- α-SMA
- α-smooth muscle actin
- p-MYPT1
- phosphorylated myosin light chain phosphatase
- p-ERK1/2
- phosphorylated extracellular signal-regulated kinase 1/2
- p-JNK
- phosphorylated c-Jun N-terminal kinase
- P-p38
- phosphorylated p38
- P-p53
- phosphorylated p53
- CTGF
- connective tissue growth factor
- ROS
- reactive oxygen species
- BP
- blood pressure
- OSOM
- outer stripe of outer medulla
- PCR
- polymerase chain reaction
- FBS
- fetal bovine serum
- PBS
- phosphate-buffered saline
- H&E
- hematoxylin and eosin
- TUNEL
- terminal deoxynucleotidyl transferase dUTP nick-end labeling
- IκB-α
- inhibitor of NF-κB α
- P-p65
- phosphorylated NF-κB p65 subunit
- PCr
- plasma creatinine
- CCr
- creatinine clearance
- UOsm
- urine osmolality
- UAE
- urine albumin excretion
- CsA Cmin
- cyclosporine plasma trough level
- Bcl-2
- B-cell lymphoma 2
- N.M.
- not measured.
- Received January 20, 2011.
- Accepted April 6, 2011.
- Copyright © 2011 by The American Society for Pharmacology and Experimental Therapeutics
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