Grant: Discovery Grants Program - Individual

Normal tissue growth and development is dependent on the coordinated ability of cells to communicate with one another and develop specific cell fates and polarity. These processes are critical for angiogenesis, the growth of new blood vessels from pre-existing vessels. We are interested in determining the molecular regulation of processes important to organized cell behavior using angiogenesis as a model system. These processes include those that regulate cell growth, migration and polarity ultimately ending in vessel sprouting and maturation. We have recently discovered that the angiogenic factor VEGF differentially regulates micro (mi)RNA, small 22 nucleotide RNAs, in endothelial cells (EC). miRNA regulate cellular message (m)RNA expression via their ability to bind and either sequester or cause degradation of their bound target mRNA, ultimately regulating protein production. Importantly, it appears that various angiogenic factors regulate the expression of different miRNA, suggesting multiple regulatory mechanisms may exist. In addition to miRNA changes directly in VEGF-stimulated cells, we speculate that miRNA may have paracrine effects on distant EC. We believe this is in part due to delivery to and uptake of exosomes (EX) by cells. EX are small vesicles that are released by cells which have recently been shown to modulate the angiogenic phenotype of EC, and it is well known that they encapsulate and deliver miRNA to target cells. As such, we wish to further investigate whether miRNA contained in EX are a means by which EC communicate with other EC to modulate angiogenic phenotypes in a paracrine fashion. We propose 3 broad areas of analysis: 1) Assess the miRNA content of exosomes following angiogenic stimulation of endothelial cells, 2) Investigate a role for paracrine effects of miRNA via exosome mediated delivery to distal EC, and 3) Identify mRNA targets of exosome-delivered miRNA to elucidate important regulatory networks. We will evaluate whether angiogenic factor stimulation of EC alters miRNA content of the EX they produce, and whether these EX can modulate the phenotype of untreated `distant' EC. We will use a variety of in vitro and in vivo angiogenesis assays as model systems of cell polarity, organization and fate to assess how specific exosomal-miRNA regulated by angiogenic factors can influence important angiogenic phenotypes. This work will further our basic understanding of the role of miRNA in coordinated cell regulation and communication, cell polarity and cell fate and their contributions to vessel formation.