FrzA/sFRP-1, a secreted antagonist of the Wnt-Frizzled pathway, controls vascular cell proliferation in vitro and in vivo
Cardiovascular Research. 2004-09-01; 63(4): 731-738
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Ezan J(1), Leroux L, Barandon L, Dufourcq P, Jaspard B, Moreau C, Allières C, Daret D, Couffinhal T, Duplàa C.
(1)Inserm U441, Avenue du Haut Lévêque, 33 600 Pessac, France, and Université Victor Segalen Bordeaux 2, rue léo Saignat, 33 000 Bordeaux, France.
OBJECTIVE: FrzA, a member of the group of secreted frizzled related proteins
(sFRP) that is expressed in the cardiovascular system, has been shown to
antagonize the Wnt/frizzled signaling pathway. We have recently demonstrated its
role in vascular cell growth control in vitro. In this study, we aimed to examine
the mechanisms by which FrzA exerts its antiproliferative effect on vascular
cells in vitro and its potential effect in vivo.
METHODS AND RESULTS: On synchronized, growth-arrested endothelial cells (EC) and
smooth muscle cells (SMC) treated with the recombinant purified FrzA protein,
flow cytometry analysis showed that the recombinant FrzA protein delayed G1 phase
and entry into S-phase. Western blot experiments demonstrated that the treatment
of EC or SMC with FrzA was associated with a decrease in the level of the cyclins
and cyclin-dependent kinases and an increase in cytosolic phospho-beta-catenin
levels. The FrzA-induced cell cycle delay was resolved by 24 h. C57BL/6J mice
underwent surgery to produce unilateral hindlimb ischemia and empty adenoviruses
(AdE) or adenoviruses coding for FrzA (AdFrzA) were injected at the time of the
surgery. In AdFrzA-treated mice in the 7 days following surgery, we showed a
decrease in cell proliferation, capillary density, and blood flow recovery and a
reduced expression of cyclin and cdk activity in the ischemic muscle compared to
that in the AdE-treated ischemic muscle. To gain insight into the pathway
activated by FrzA overexpression, we showed an increase in the level of cytosolic
phospho-beta-catenin, a marker of beta-catenin degradation, in AdFrzA-treated
ischemic muscle compared to that in control AdE-treated ischemic muscle.
CONCLUSION: We provided the first evidence that an impairment of the Wnt-Frizzled
pathway, via FrzA overexpression, controlled proliferation and neovascularization
after muscle ischemia.