Abstract
The proteolytic activation by thrombin of the proteinase-activated receptor 1 unveils the tethered peptide ligand and cleaves a 41-amino acid peptide. In this report, we show that this peptide, which we have designated as “parstatin,” is a potent inhibitor of angiogenesis. Synthesized parstatin suppressed both the basic angiogenesis and that stimulated by basic fibroblast growth factor and vascular endothelial growth factor in the chick embryo model in vivo and in the rat aortic ring assay. Parstatin also abrogated endothelial cell migration and capillary-like network formation on the Matrigel and fibrin angiogenesis models in vitro. Treatment of endothelial cells with parstatin resulted in inhibition of cell growth by inhibiting the phosphorylation of extracellular signal-regulated kinases in a specific and reversible fashion and by promoting cell cycle arrest and apoptosis through a mechanism involving activation of caspases. We have shown that parstatin acts as a cell-penetrating peptide, exerting its biological effects intracellularly. The uptake into cells and the inhibitory activity were dependent on parstatin hydrophobic region. These results support the notion that parstatin may represent an important negative regulator of angiogenesis with possible therapeutic applications.
Footnotes
-
This work was supported by the University of Patras Research Committee [Program “K. Karatheodoris 2005”].
-
This work was previously presented in part: Gourni D, Zania P, Tsilimidou A, Kalavrizioti D, Flordellis SC, Maragoudakis ME, and Tsopanoglou NE (2007). Parstatin. The cleaved peptide of protease-activated peceptor-1, as a negative regulator of angiogenesis. 5th General Meeting of International Proteolysis Society; 20–24 October 2007; Patras, Greece.
-
N.E.T and M.E.M declare competing financial interest because of submission of patent application covering parstatin.
-
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
-
doi:10.1124/jpet.108.145664.
-
ABBREVIATIONS: PAR, protease-activated receptor; MMP, matrix metalloprotease; VEGF, vascular endothelial growth factor; FITC, fluorescein isothiocyanate; HUVEC, human umbilical vein endothelial cell; CAM, chorioallantoic membrane; bFGF, basic fibroblast growth factor; FBS, fetal bovine serum; BSA, bovine serum albumin; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium; HB, heparin binding; EGF, epidermal growth factor; ERK, extracellular signal-regulated kinase; PARP, poly(ADP-ribose) polymerase; PI, propidium iodide; DAPI, 4,6-diamidino-2-phenylindole; MAPK, mitogen-activated protein kinase; parst, parstatin; Z, N-benzyloxycarbonyl; FMK, fluoromethyl ketone.
- Received September 3, 2008.
- Accepted November 5, 2008.
- U.S. Government work not protected by U.S. copyright
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.
|
Log in using your username and password
Purchase access
In this issue
Parstatin, the Cleaved Peptide on Proteinase-Activated Receptor 1 Activation, Is a Potent Inhibitor of Angiogenesis
