UCLA

Background:Myocardial infarctions (MI) are a leading cause of death worldwide. Unfortunately, the adult heart cannot generate new vessels after MI leading to impaired myocardial perfusion and function. This study aims to identify reprogramming factors during embryonic angiogenesis, which may enhance neovascularization in the adult heart after MI. Our lab discovered 56 novel secreted ligand genes expressed by epicardial cells during development. One factor, collagen, type XVIII, alpha 1 (Col18a1), an angiogenesis inhibitor, has not been thoroughly investigated in the heart.

Methods:Wilm’s tumor gene 1 (Wt1) transgenic and C57BL/6 mouse strains were used to investigate Col18a1 expression at embryonic (E) and early post-natal (P) time points. Overexpression of COL18A1 in the epicardium was performed at E12.5 to investigate alterations in coronary angiogenesis using gene expression and immunohistochemical analyses.

Results:Col18a1 increased in Wt1+ epicardial cells during coronary vasculature formation, peaking at E16.5, and stabilized during the early postnatal periods determined by fluorescence in situ hybridization assays. Gene expression data was corroborated following immunohistochemical analysis of endostatin, a proteolytically produced C-terminal fragment of COL18A1 and potent antiangiogenic protein. COL18A1 overexpression in the developing epicardium caused dysregulation of endothelial cell (EC) positioning within the epicardium, observed after immunolabeling for ERG+ cells. Gene expression data confirmed dysregulation of EC differentiation with a downregulation in arterial Nrp1 and upregulation of venous Nr2f2.

Conclusions:Our data suggest that mRNA and protein levels of Col18a1 and endostatin increased during embryogenesis to limit coronary angiogenesis. Overexpression of COL18A1 inhibited migration and preserved a venous and coronary plexus phenotype in ECs. Furthermore, overexpression of COL18A1 produced immature and proliferative epicardial cells, confirmed by an upregulation in epithelial-to-mesenchymal genes. We theorize that inhibition of epicardial cell differentiation by COL18A1 overexpression drives the dysregulation in EC placement and coronary development.