Subscribe to RSS
DOI: 10.1055/s-0034-1367198
Identification and characterization of a postnatal Nkx2-5 enhancer positive cardiac progenitor cell population after myocardial infarction
Objectives: Recent studies have demonstrated a limited capacity of the postnatal mammalian heart to undergo cardiomyocyte renewal. This process may be mediated by stem-/progenitor cells. However, the exact cellular mechanisms remain poorly understood. By using an Nkx2-5 cardiac enhancer-eGFP transgenic mouse model we sought to investigate cellular origins of postnatal cardiac regeneration after experimental myocardial infarction (MI).
Methods: MI was induced experimentally by LAD ligation in Nkx2-5 cardiac enhancer-eGFP mice of different ages. The number of eGFP+ cells in whole heart tissue samples was quantified by flow cytometry at day 5 and 9 post-MI, respectively. Further characterization of FACS sorted eGFP+ cells after MI was performed using qRT-PCR for gene expression profiling. Selected qRT-PCR results were confirmed by antibody staining and subsequent FACS analysis.
Results: Green fluorescence in the cardiac-specific Nkx2-5 enhancer-eGFP mouse peaks around E8, fades after E11.5 and vanishes in the uninjured adult heart until 3 weeks postnatally. Interestingly, the number of eGFP+ cells increased to about 1% after MI induction in this mouse line (> 6 weeks of age). When compared to the GFP- cell fraction FACS sorted eGFP+ cells revealed high expression levels of several cardiac developmental markers (Tbx20, Baf60c, Gata4, Flk1) and were enriched for embryonic epicardial genes (Tbx18, WT1). eGFP+ cells lack the expression of mature cardiomyocyte marker cTnT. Expression of CD45 (panleukocyte marker) and c-Kit/CD117 (stem cell marker) was absent which was confirmed by antibody staining and subsequent FACS analysis. Interestingly, the stem cell marker Sca1 was upregulated in GFP+ cells compared to the GFP- fraction. This result was also approved by antibody staining. Screening for potential cytokine targets revealed a quite specific expression of receptors for FGF, PDGF, TGFβ and IGF within eGFP+ cells.
Conclusions: By using an Nkx2-5 cardiac enhancer-eGFP transgenic mouse model we identified an eGFP+ population which increases after MI in the adult murine heart. Detailed gene expression profiling of isolated GFP+ cells showed that myocardial injury triggers the expression of cardiac-specific embryonic genes implicating characteristics of cardiac progenitor cells for the GFP+ cells when compared to the GFP- fraction. Moreover, we identified potential cytokine targets to therapeutically boost cardiac regeneration after myocardial infarction.