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Spatial transcriptional profile of the chick and mouse endocardial cushions
identify novel regulators of endocardial EMT in vitro
#MMPMID23557753
DeLaughter DM
; Christodoulou DC
; Robinson JY
; Seidman CE
; Baldwin HS
; Seidman JG
; Barnett JV
J Mol Cell Cardiol
2013[Jun]; 59
(?): 196-204
PMID23557753
show ga
Valvular Interstitial Cells (VICs) are a common substrate for congenital and
adult heart disease yet the signaling mechanisms governing their formation during
early valvulogenesis are incompletely understood. We developed an unbiased
strategy to identify genes important in endocardial epithelial-to-mesenchymal
transformation (EMT) using a spatial transcriptional profile. Endocardial cells
overlaying the cushions of the atrioventricular canal (AVC) and outflow tract
(OFT) undergo an EMT to yield VICs. RNA sequencing (RNA-seq) analysis of gene
expression between AVC, OFT, and ventricles (VEN) isolated from chick and mouse
embryos at comparable stages of development (chick HH18; mouse E11.0) was
performed. EMT occurs in the AVC and OFT cushions, but not VEN at this time. 198
genes in the chick (n=1) and 105 genes in the mouse (n=2) were enriched 2-fold in
the cushions. Gene regulatory networks (GRN) generated from cushion-enriched gene
lists confirmed TGF? as a nodal point and identified NF-?B as a potential node.
To reveal previously unrecognized regulators of EMT four candidate genes, Hapln1,
Id1, Foxp2, and Meis2, and a candidate pathway, NF-?B, were selected. In vivo
spatial expression of each gene was confirmed by in situ hybridization and a
functional role for each in endocardial EMT was determined by siRNA knockdown in
a collagen gel assay. Our spatial-transcriptional profiling strategy yielded gene
lists which reflected the known biology of the system. Further analysis
accurately identified and validated previously unrecognized novel candidate genes
and the NF-?B pathway as regulators of endocardial cell EMT in vitro.
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