Abstract 14770: Secreted Frizzled Related Protein 2 Reduces Resident Cardiac Progenitor Cell Proliferation and Induces Cardiac Marker Expression
Introduction: Endogenous repair via activation of resident cardiac progenitor cells represents an attractive therapeutic approach for limiting cardiac remodeling and enhancing cardiac regeneration post infarction. Secreted Frizzled Related Protein 2 (Sfrp2) has been shown to reduce myocyte apoptosis and to affect infarct size in vivo, but its effect on resident cardiac progenitor cells has yet to be investigated. In this study, we examined the effects of Sfrp2 in the proliferation and differentiation of mouse resident cardiac progenitor cells (mCPCs).
Methods: cKit+/Sca1+ mCPCs were isolated by magnetic beads and single cell clones were derived after plating cells by limiting dilutions. Expression of stem cell and mesenchymal markers was evaluated by FACS analysis and Sfrps by qRT-PCR. To examine the effects of Sfrp2 on mCPC proliferation, cell cycle was assessed by BrdU incorporation and DNA content staining using FACS analysis and confirmed by BrdU ELISA. The effect of Sfrp2 on mCPC differentiation was evaluated by both qRT-PCR and immunofluorescent staining for cardiac markers Nkx2.5, Actinin, and α-MHC.
Results: Isolated mCPC clones maintained expression of Sca1, CD44, CD105, CD29 and CD106 throughout the study. These cells expressed high levels of Sfrp1 and 3. Sfrp2 was detectable but at significantly lower levels. Treatment of mCPCs with 1–100 nM recombinant Sfrp2 protein showed a dose dependent reduction in the percent of cells in S-phase (maximum reduction at 10 nM of 35–40%, P<0.01). Likewise, the percent of cells in the G0/G1 phase increased by similar numbers with no change in G2-M phase. In addition, mCPC clones treated with Sfrp2 upregulated Nkx2.5 by 2 weeks and maintained this upregulation for up to 4 weeks. Expression of Actinin and α-MHC was detectable at three weeks albeit at low levels and in a small number of cells.
Conclusions: Our data suggests that Sfrp2 induces cell cycle arrest of mCPCs by specifically blocking progression of cells from the G0/G1 to S-phase without effecting cell division. This inhibition in proliferation is accompanied by induction of mCPC differentiation as evident by increased expression cardiac specific markers, such as Nkx2.5, but Sfrp2 failed to drive these cells to fully differentiate to mature cardiomyocytes.
- © 2010 by American Heart Association, Inc.