Prolyl oligopeptidase induces angiogenesis both in vitro and in vivo in a novel regulatory manner.
BACKGROUND AND PURPOSE A serine protease, prolyl oligopeptidase (POP) has been reported to be involved in the release of the pro-angiogenic tetrapeptide acetyl-N-Ser-Asp-Lys-Pro (Ac-SDKP) from its precursor, 43-mer thymosin β4 (Tβ4). Recently, it was shown that both POP activity and the levels of Ac-SDKP are increased in malignant tumours. The aim of this study was to clarify the release of Ac-SDKP, and test if POP and a POP inhibitor, 4-phenyl-butanoyl-L-prolyl-2(S)-cyanopyrrolidine (KYP-2047), can affect angiogenesis. EXPERIMENTAL APPROACH We used HPLC for bioanalytical and an enzyme immunoassay for pharmacological analysis. Angiogenesis of human umbilical vein endothelial cells was assessed in vitro using a 'tube formation' assay and in vivo using a Matrigel plug assay (BD Biosciences, San Jose, CA, USA) in adult male rats. Moreover, co-localization of POP and blood vessels was studied. KEY RESULTS We showed the sequential hydrolysis of Tβ4: the first-step hydrolysis by proteases to <30-mer peptides is followed by an action of POP. Unexpectedly, POP inhibited the first hydrolysis step, revealing a novel regulation system. POP with Tβ4 significantly induced, while KYP-2047 effectively prevented, angiogenesis in both models compared with Tβ4 addition itself. POP and endothelial cells were abundantly co-localized in vivo. CONCLUSIONS AND IMPLICATIONS We have now revealed that POP is a second-step enzyme in the release of Ac-SDKP from Tβ4, and it has novel autoregulatory effect in the first step. Our results also advocate a role for Ac-SDKP in angiogenesis, and suggest that POP has a pro-angiogenic role via the release of Ac-SDKP from its precursor Tβ4 and POP inhibitors can block this action.
Division of Pharmacology and Toxicology, Faculty of Pharmacy, University of Helsinki, Viikinkaari 5E, Helsinki, Finland. Timo.Myöhänen@helsinki.fi
T T
TT
Myöhänen
J
J
Tenorio-Laranga
B
B
Jokinen
R
R
Vázquez-Sánchez
M J
MJ
Moreno-Baylach
J A
JA
García-Horsman
P T
PT
Männistö
D020533:Angiogenesis Inhibitors / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001672:Biocompatible Materials / Q000378:metabolism; D003094:Collagen / Q000378:metabolism; D004338:Drug Combinations; D042783:Endothelial Cells / Q000187:drug effects*; D006131:Growth Inhibitors / Q000032:analysis / Q000096:biosynthesis; D006801:Humans; D007668:Kidney / Q000187:drug effects*; D007797:Laminin / Q000378:metabolism; D008297:Male; D009389:Neovascularization, Pathologic / Q000378:metabolism; D009842:Oligopeptides / Q000032:analysis / Q000096:biosynthesis; D011392:Proline / Q000031:analogs & derivatives* / Q000494:pharmacology; D000085822:Prolyl Oligopeptidases; D011509:Proteoglycans / Q000378:metabolism; D051381:Rats; D011920:Rats, Inbred WF; D012697:Serine Endopeptidases / Q000502:physiology*; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology*; D013947:Thymosin / Q000378:metabolism; D014471:Umbilical Veins / Q000187:drug effects*; D000099201:Thymosin beta(4)
D020533:Angiogenesis Inhibitors; D001672:Biocompatible Materials; D003094:Collagen; D004338:Drug Combinations; D006131:Growth Inhibitors; D007797:Laminin; D009842:Oligopeptides; D011392:Proline; D000085822:Prolyl Oligopeptidases; D011509:Proteoglycans; D012697:Serine Endopeptidases; D015842:Serine Proteinase Inhibitors; D013947:Thymosin; D000099201:Thymosin beta(4); C551540:4-phenylbutanoyl-prolylcyanopyrrolidine; C059485:matrigel; C000705287:PREPL protein, human; C058504:goralatide