CXCR4+/FLK-1+ biomarkers select a cardiopoietic lineage from embryonic stem cells.
Pluripotent stem cells demonstrate an inherent propensity for unrestricted multi-lineage differentiation. Translation into regenerative applications requires identification and isolation of tissue-specified progenitor cells. From a comprehensive pool of 11,272 quality-filtered genes, profiling embryonic stem cells at discrete stages of cardiopoiesis revealed 736 transcripts encoding membrane-associated proteins, where 306 were specifically upregulated with cardiogenic differentiation. Bioinformatic dissection of exposed surface biomarkers prioritized the chemokine receptor cluster as the most significantly over-represented gene receptor family during pre cardiac induction, with CXCR4 uniquely associated with mesendoderm formation. CXCR4(+) progenitors were sorted from the embryonic stem cell pool into mesoderm-restricted progeny according to co-expression with the early mesoderm marker Flk-1. In contrast to CXCR4(-)/Flk-1(-) cells, the CXCR4(+)/Flk-1(+) subpopulation demonstrated overexpressed cardiac lineage transcription factors (Mef2C, Myocardin, Nkx2.5), whereas pluripotent genes (Oct4, Fgf4, Sox2) as well as neuroectoderm (Sox1) and endoderm alpha-fetoprotein markers were all depleted. In fact, the CXCR4(+)/Flk-1(+) biomarker combination identified embryonic stem cell progeny significantly enriched with Mesp-1, GATA-4, and Tbx5, indicative of pre cardiac mesoderm and the primary heart field. Although the CXCR4(+)/Flk-1(+) transcriptome shared 97% identity with the CXCR4(-)/Flk-1(-) counterpart, the 818 divergent gene set represented predominantly cardiovascular developmental functions and formed a primitive cardiac network. Differentiation of CXCR4(+)/Flk-1(+) progenitors yielded nuclear translocation of myocardial transcription factors and robust sarcomerogenesis with nascent cardiac tissue demonstrating beating activity and calcium transients. Thus, the CXCR4/Flk-1 biomarker pair predicts the emergence of cardiogenic specification within a pluripotent stem cell pool, enabling targeted selection of cardiopoietic lineage. Disclosure of potential conflicts of interest is found at the end of this article.
Nelson
Timothy J
TJ
Department of Medicine, Marriott Heart Disease Research Program, Division of Cardiovascular Diseases, Mayo Clinic, Rochester, Minnesota 55905, USA.
Faustino
Randolph S
RS
Chiriac
Anca
A
Crespo-Diaz
Ruben
R
Behfar
Atta
A
Terzic
Andre
A
D000818:Animals; D015415:Biomarkers / Q000032:analysis; D002118:Calcium / Q000502:physiology; D002454:Cell Differentiation; D018076:DNA, Complementary / Q000235:genetics; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000502:physiology; D004707:Endoderm / Q000166:cytology / Q000502:physiology; D020869:Gene Expression Profiling; D008648:Mesoderm / Q000166:cytology / Q000502:physiology; D051379:Mice; D032383:Myocytes, Cardiac / Q000166:cytology / Q000502:physiology*; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D019718:Receptors, CXCR4 / Q000235:genetics*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics*
D015415:Biomarkers; C514936:CXCR4 protein, mouse; D018076:DNA, Complementary; D019718:Receptors, CXCR4; D012313:RNA; D040301:Vascular Endothelial Growth Factor Receptor-2; D002118:Calcium