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Abstract

The anatomic arrangement of microvascular endothelial cells and cardiomyocytes in vivo enables close interactions among these cells. In our in vitro co-culture system, ANP and BNP mRNA expression in cardiomyocytes and subsequent ANP release were significantly upregulated when co-cultured with endothelial cells or exposed to endothelial cell-conditioned medium. Endothelin-1 (ET-1) activation of endothelial cells remarkably enhanced their paracrine effects on cardiomyocyte gene expression in our study, suggesting for the first time that ET-1 stimulation of endothelial cells affects expression of fetal genes such as ANP and BNP in adult cardiomyocytes through paracrine signalling. Combined with previous results indicating a crucial role for ANP and BNP in cardiac homeostasis, our finding provide further evidence that paracrine signalling by cardiac microvascular endothelial cells modulates cardiomyocyte function. Exposure of HL-1 cells to authentic Angiopoietin-2 (Ang2) caused a concentration-dependent decrease in ANP expression while ET-1-induced ANP expression was augmented by low concentrations of Ang2 (1 ng/ml) but inhibited by high concentrations of Ang2 (10 ng/ml). Mass spectrometry analysis of EC-conditioned medium revealed qualitative differences in the release of proteins from endothelial cells with and without ET-1 stimulation. Gene ontology annotation of the detected proteins points towards a role for exosomes and proteins involved in RNA folding in endothelial cell-cardiomyocyte crosstalk.