Abstract
Vascular smooth muscle (VSM) proliferation and migration are key components in vessel remodeling. Cyclic nucleotide signaling is protective and has long-served as a therapeutic target against undesired VSM growth. The present work analyzed the effects of the soluble guanylate cyclase (sGC) stimulator 3-(4-amino-5-cyclopropylpyrimidine-2-yl)-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine [BAY 41-2272 (BAY)] on VSM growth, and we hypothesize that BAY has the capacity to reduce proliferation and migration via cyclic nucleotide-driven kinase signaling. Perivascular BAY postballoon injury reduced neointimal growth by ∼40% compared with vehicle controls after 2 weeks. In VSM cells, BAY (10 μM) reduced proliferation by ∼40% after 72 h and migration by ∼40% after 6 h and ∼60% after 18 h without deleterious effects on cell viability. cGMP content peaked (248×) 20 min after BAY treatment and remained elevated (140×) through 60 min; however, BAY did not affect cAMP levels compared with controls. Conventional and In-Cell Western analyses showed increases in vasodilator-stimulated phosphoprotein (VASP) phosphorylation (pVASP) at serines 239 (3×) and 157 (2×), respective markers of cGMP- and cAMP-directed protein kinases (PKG and PKA, respectively). The PKG inhibitor YGRKKRRQRRRPPLRKKKKKH peptide (DT-2) completely reversed BAY-mediated increases in pVASPSer239 and BAY-mediated inhibition of migration. In comparison, the PKA inhibitor peptide PKI further potentiated BAY-stimulated pVASPSer157 and pVASPSer239 and partially reversed the antiproliferative effects of BAY. This is the first report demonstrating the effectiveness of BAY in reducing neointimal growth with direct evidence for PKG-specific antimigratory and PKA-specific antiproliferative mechanisms. Conclusively, the sGC stimulator BAY reduces VSM growth through cGMP-dependent PKG and PKA processes, providing support for continued evaluation of its clinical utility.
Footnotes
This study was supported by the National Institutes of Health National Heart, Lung, and Blood Institute [Grants R01-HL081720, R01-HL081720-03S1, R01-HL081720-03S2].
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
doi:10.1124/jpet.111.183400.
-
ABBREVIATIONS:
- VSM
- vascular smooth muscle
- VSMC
- VSM cell
- ANOVA
- analysis of variance
- BAY
- BAY 41-2272 [3-(4-amino-5-cyclopropylpyrimidine-2-yl)-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine]
- CA
- carotid artery
- Cdk
- cyclin-dependent kinase
- CO
- carbon monoxide
- DMEM
- Dulbecco's modified Eagle's medium
- DMSO
- dimethyl sulfoxide
- DT-2
- YGRKKRRQRRRPPLRKKKKKH peptide
- FBS
- fetal bovine serum
- HO
- heme oxygenase
- IBMX
- 3-isobutyl-1-methylxanthine
- ICW
- In-Cell Western
- MW
- medial wall
- NI
- neointima
- NO
- nitric oxide
- NOS
- NO synthase
- PBS
- phosphate-buffered saline
- PDE
- phosphodiesterase
- PDGF
- platelet-derived growth factor
- PKA
- cAMP-dependent protein kinase
- PKI
- PKA inhibitor
- PKG
- cGMP-dependent protein kinase
- sGC
- soluble guanylate cyclase
- VASP
- vasodilator-stimulated phosphoprotein
- pVASP
- VASP phosphorylation
- pPKASer
- phosphorylation of PKA at serine
- VASPTot
- total VASP
- YC-1
- 3-(5′-hydroxymethyl-2′-furyl)-1-benzyl indazole.
- Received April 27, 2011.
- Accepted August 5, 2011.
- Copyright © 2011 by The American Society for Pharmacology and Experimental Therapeutics
JPET articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|