Neutrophil (PMN) infiltration of the intestinal mucosa is a hallmark of inflammatory bowel diseases (IBD). Although PMNs are critical players in host defense, their accumulation and activity in tissues can result in exacerbated inflammation and damage. PMN mobilization to sites of inflammation requires crossing of the vascular wall and the endothelial barrier. While sequential PMN adhesive interactions with EC during TEM have been well-defined, mechanisms underlying the initiation and termination of this process and particularly in gut inflammation are not well-understood. Perivascular macrophages (PVMs) in the brain and the skin were recently implicated in regulating vascular barrier function and via chemokine release to promote PMN recruitment. Our data uniquely suggest that unlike in the brain, gut interstitial macrophages are actively recruited by vascular ECs to the vessel wall, where they can engage in homophylic interactions with vascular PECAM to promote PMN TEM specific Ca2+ responses. Our findings further suggest that macrophages via the release of extracellular vesicles (EVs) can transfer regulatory microRNAs to modulate EC function. Specifically, a targeted microRNA screen of macrophages-EVs revealed miR-125b to be one of the highest expressed microRNAs in EVs. miR-125b targets NFkB inhibitor (intracellular ubiquitin-editing protein A20 or TNFAIP3) to activate NFkB signaling, which in ECs induces PMN TEM-regulatory adhesion molecules including ICAM-1 and VCAM-1. Thus, we will test the hypothesis that in inflamed mucosa, interstitial macrophages are actively recruited to the vessel wall, where they via binding interactions and microRNA activity can locally prime EC responses to guide PMN TEM. Using models of acute inflammation/mucosal injury, state of the art imaging approaches and complementary ex vivo transwell migration assays we will 1. Define specific chemotactic cues generated by inflamed EC that guide recruitment of interstitial macrophages to the vessel wall. 2. Establish the novel role of macrophages-expressed ICAM-1 in macrophages mobilization towards inflamed vascular wall (Aim 1). We will further use specific miRNA mimics/inhibition, novel EC-specific Ca2+ reporter mice and macrophages depletion approaches to 1. Determine whether macrophages via miR-125b activity trigger localized increases in EC adhesion molecule expression. 2. Determine whether macrophages engage EC-PECAM to trigger PMN TEM-specific Ca2+ responses and cytoskeletal reorganization. 3. Determine whether targeting miR-125b or TEM-specific Ca2+ responses to inhibit PMN TEM will improve mucosal healing (Aim 2).
|Effective start/end date||7/1/22 → 6/30/25|
- Crohn's and Colitis Foundation of America (937347)
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