Abstract 14437: Protein Kinase D1 Signaling is the Key to Arterial Differentiation of Vascular Endothelial Cells
Endothelial cell (EC) differentiation through transcriptional reprogramming plays a fundamental role in functional angiogenesis that is important in recovering blood supply in ischemic cardiovascular diseases. However, it is unclear how terminally differentiated ECs are reprogrammed. Our recent study implicates that protein kinase D1 (PKD-1) regulates reprogramming of microvascular ECs (MVECs) for arterial differentiation. We hypothesized that PKD-1 signaling is critical in the differentiation of terminally differentiated ECs. Different types of ECs, and GS4012, a small chemical molecule that was screened in a Gridlock Zebrafish model and is critical in arterial differentiation, were used to elucidate the role of PKD-1 signaling in EC differentiation. qPCR, lentiviral transduction, western blot assays, immunofluorescence microscopy, Boyden chamber migration assays, in vitro Matrigel assays, and EC-specific pkd-1 null mice were respectively used for the analysis of gene expression, signaling, and change of EC morphology and function. Our results showed that GS4012 treatment stimulated the phosphorylation of PKD-1, and of its downstream signaling molecule Erk1/2, but did not show any synergistic effect with VEGF. Intriguingly, prolonged EC exposure to GS4012 increased the mRNA expression of key arterial markers ephrin B2, neuropilin1, and DLL4, which were attenuated by the PKD inhibitor (p<0.01). Functionally, EC migration increased in response to GS4012 exposure via PKD-1 signaling (p<0.01). Moreover, transduction of constitutively active PKD-1 into primary ECs induced extensive EC branching morphogenesis accompanied by increased mRNA expression of DLL4. GS4012 treatment also increased EC branching morphogenesis and showed a synergistic effect with VEGF in two- and three-dimensional Matrigel assays. Importantly, the vascular endothelium in the heart and lung of the EC-specific pkd-1 deficient mice showed abnormal vascular structures with defective EC coverage. In conclusion, our study indicates an essential role of PKD-1 signaling in promoting arterial differentiation of ECs, and it supports the concept of EC plasticity or terminal differentiated EC can be reprogrammed with certain environmental cues.
Author Disclosures: C. Opansky: None. B. Best: None. R. Yuan: None. Q. Cao: None. B. Ren: None.
- © 2016 by American Heart Association, Inc.