Abstract 17304: Stem Cell Model for Down Syndrome-Related Heart Defects
About 60% of newborns with Down syndrome (DS) are born with heart defects. The most common heart defect is the failure of formation of the part of the heart that arises from an embryonic structure called endocardial cushions, which is responsible for separating the central parts of the heart. Endocardial cushions are thought to arise from a subset of endothelial cells that undergo epithelial to mesenchymal transformation and migrate into the cardiac jelly to populate endocardial cushions with mesenchymal cells. We employed directed differentiation of iPS (induced pluripotent stem) cells generated from individuals with and without DS to define the phenotypic features distinguishing the development of normal and DS mesodermal precursors that give rise to endothelial and mesechymal cells, the most important structural components of endocardial cushion. The iPS lines were generated by forced expression of transcription factors in the fibroblasts of the patients using retroviral transduction. The established iPS lines were shown to express a set of characteristic pluripotency markers, to be capable of differentiation in vitro to all three germ layers and form teratomas. Mesodermal and hematoendothelial iPS differentiation was performed either in chemically defined conditions or in the co-culture with OP9 stromal cells. The endothelial potential of CD31+VE-Cadherin+ isolated cells was evaluated by cell tube formation assay, immunostaining with von Willebrand Factor (vWF) and the ability to internalize Acetylated-low density lipoprotein (Ac-LDL). The progenitor cells were collected at days 2 and 3 of iPS differentiation on OP9 and assessed by flow cytometry analysis as well as frequencies of mesenchymal and blast colonies in semisolid media (MS-CFU and BL-CFU). We identified the earliest stages in mesodermal differentiation that demonstrate the difference in colony forming potential, thus showing a distinct pattern of differentiation in DS vs. euploid iPS cells. In particular the colony forming and hematoendothelial potential of DS iPS cells is significantly reduced. These results of in vitro differentiation will be supplemented with molecular analysis in an effort to decipher the complex developmental pattern of mesodermal precursors in DS.
Author Disclosures: Y. Galat: None. I. Elcheva: None. M. Perepitchka: None. P. Iannaccone: None. V. Galat: None.
- © 2014 by American Heart Association, Inc.