Project Summary/Abstract The goal of this two PI research proposal is to determine the immunosuppressive and injurious effects of e- cigarettes on human distal lung structural cells (i.e. small airway epithelial cells and lung microvascular endothelial cells) that are critical to the pathogenesis of chronic obstructive pulmonary disease (COPD), typically induced by tobacco cigarette smoke exposure. The use of e-cigarettes is rapidly increasing in the US youth population, but its health effects on distal lungs have not been investigated. We have found that inhaling e- cigarette vapor (ECVap) can reach small airways to cause distal lung injury. Specifically, we have demonstrated that exposures to ECVap constituents, including nicotine, significantly weaken distal lung (e.g. small airway) innate immunity against influenza A virus infection and microvascular endothelial barrier function, and enhance lung inflammation. We will test the hypothesis that e-cigarettes impair the antiviral immunity, causing increased neutrophilic inflammation and distal lung structural cell (small airway and microvascular endothelial cell) injury by proposing three specific aims. In Aim 1, we will identify the mechanisms whereby ECVap impairs the antiviral immunity of human distal lungs by performing RNA sequencing in precision-cut human (15 to 25 years old) lung slices exposed to ECVap and influenza A viruses (IAV), and carrying out human primary small airway epithelial cell air-liquid interface culture in the presence or absence of ECVap and IAV. In Aim 2, we will determine how ECVap enhances neutrophilic inflammatory response to IAV infection of the human distal lung by using the human lung slices and the co-culture system of human microvascular endothelial cells and neutrophils. In Aim 3, we will determine how ECVap increases distal lung injury following IAV infection by testing if ECVap exposure reduces sphingolipid pro-survival signaling and subsequently increases the vulnerability of small airway epithelial and endothelial cells to virus-induced injury. Research findings from our proposed studies will significantly improve our understanding about the distal lung health effects of e-cigarette use in youth, and inform policies of e-cigarette regulation.