Abstract 16723: Complementation of Mutant Embryos Demonstrates a Cell Autonomous Requirement of Etv2
Our previous studies have demonstrated that the disruption of Etv2, a gene encoding an Ets related transcription factor, resulted in the absence of the endothelial/endocardial lineages. Etv2 is unique in that it has a limited window of expression from E7 to E9. It is unknown whether all endothelial/endocardial progenitors require Etv2 expression or whether this is only a requirement for the initial vasculogenic progenitor/stem cells. We hypothesized that complementation of Etv2 mutant embryoid bodies (EBs) or embryos with wildtype embryonic stem cells (ESCs) would rescue the endothelial defect in a cell autonomous manner and mutant cells would not contribute to vessel development. To address this hypothesis, we co-differentiated mouse wildtype and mutant ESCs with EYFP-labeled wildtype ESCs. We observed that differentiation of wildtype EYFP-ESCs resulted in contributions to the endothelial lineage. Further, co-differentiation of Etv2 mutant and EYFP-labeled ESCs resulted in endothelial cells derived solely from the wildtype EYFP-labeled cells, indicating that the rescue event is cell autonomous. We further addressed this hypothesis using blastocyst complementation in vivo. We interbred mice hemizygous for two distinct Etv2 gene disruptions and harvested blastocysts at E3.5. EYFP-labeled ESCs were injected into the blastocoele cavity and transferred into pseudopregnant mice. At E10.5, a day after the death of Etv2 mutant embryos, the developing embryos were analyzed for chimerism and the contribution of EYFP-labeled ESCs to various cell lineages. We observed that the EYFP-labeled ESCs contributed randomly to all lineages in the wildtype or heterozygous Etv2 embryo. However, when the recipient was an Etv2 mutant blastocyst, the EYFP-labeled cells preferentially gave rise to endothelial lineages. These wildtype ES cells rescued the phenotype of mutant embryos in direct correlation with the degree of chimerism. Furthermore, this was a cell autonomous event as unlabeled mutant cells were unable to give rise to endothelium. We conclude that all endothelial progenitors during embryogenesis require Etv2 expression.
Author Disclosures: T.L. Rasmussen: None. C.A. Walter: None. J.E. Rosenberg: None. X. Shi: None. M.G. Garry: None. N. Koyano-Nakagawa: None. D.J. Garry: None.
This research has received full or partial funding support from the American Heart Association
- © 2014 by American Heart Association, Inc.