Homeostasis of barrier integrity in cerebral microvessels is maintained by active regulation of endothelial cell-cell adhesive proteins. During aberrant inflammatory responses, a host of factors act upon the endothelium to disrupt these adhesive proteins and diminish barrier integrity. The pleiotropic cytokine oncostatin M (OSM) is one such factor, which has been shown to be elevated in several inflammatory conditions associated with increased microvascular leakage, such as multiple sclerosis. Germaine to this proposal, some evidence suggests that OSM may directly induce cerebral microvascular endothelial cell (CMVEC) barrier dysfunction, but the mechanisms remain unclear. We recently discovered that OSM-induced barrier dysfunction in human CMVECs is associated with a decrease in the tight-junction-associated MARVEL proteins (TAMPs), occludin and tricellulin. Here we are proposing two novel approaches to determine the endothelial-specific signaling mechanisms leading to downregulation of TAMPs during inflammation. In the first aim, we will utilize both pharmacologic (OSM receptor antagonists) and biologic (OSM receptor knockout mice) approaches to modulate OSM receptor (OSMR) signaling in endothelial cells. The goal is to determine the relative role of OSMR signaling in OSM-induced TAMP repression and cerebral microvascular barrier dysfunction. Snail is a transcriptional repressor, which has previously been identified to downregulate epithelial cell-cell adhesive proteins. Therefore, we evaluated a potential role of Snail in barrier dysfunction of CMVECs. At baseline, there was minimal detection of Snail protein; however, OSM induced a robust upregulation of Snail protein preceding the onset of barrier dysfunction. Moreover, siRNA-mediated knockdown of SNAI1 increased baseline levels of occludin, reversed OSM-induced occludin downregulation and attenuated OSM-mediated CMVEC barrier dysfunction. In the second aim, we are proposing to characterize the novel role of Snail-dependent repression of TAMPs in cerebral endothelial cells. We will use a Snail-specific inhibitor and inducible endothelial-specific deletion of Snail to determine its functional role in regulating endothelial barrier function during inflammation. The overarching hypothesis to be tested in this proposal is that OSM promotes OSMR/JAK/STAT3 signaling and triggers cerebral endothelial barrier dysfunction via Snail-dependent repression of TAMPs.
|Program type||Scientist Development Grant|
|Effective start/end date||07/01/2017 → 06/30/2020|