Combined pulmonary inflammation and fibrosis, often referred to as """"""""interstitial lung disease"""""""" (ILD), is debilitat- ing and even deadly. ILD may develop for a variety of reasons, and with no confirmed therapies, ILD is the main cause of death in patients with scleroderma, whereas median survival is only 2 to 3 years in patients with another ILD, idiopathic pulmonary fibrosis (IPF). Military exposures, aging, and autoimmune diseases make ILD prevalent in Veterans. Although much progress has been made in uncovering the mechanisms of ILD, the disease still remains poorly understood. Recent findings implicate nuclear factor from high endothelial venules (NFHEV), also known as interleukin (IL)-33, in the pathogenesis of scleroderma and IPF. NFHEV/IL-33 is des- ignated by these two different names because of its dual biology. As a nuclear factor, it is expressed in a varie- ty of cell types and binds directly to DNA. As a cytokine, proteolytically activated extracellular IL-33 is a power- ful inducer of Th2 responses, such as those in allergies and asthma. The latter activities of the IL-33 cytokine are under active investigation, whereas the former activities of NFHEV as a nuclear factor have been underap- preciated and not studied in sufficient detail or in association with a disease process. Our data suggest that, as a nuclear factor, NFHEV drives ILD. NFHEV is elevated in the lungs of patients with ILD and predominant over the IL-33 form. In these patients, NFHEV is expressed in inflammatory cells and, notably, fibroblasts, which are central to the pathogenesis of ILD. Consistent with this, the expression of NFHEV is elevated in the lungs of mice in the bleomycin model of ILD. Gene delivery of NFHEV in cell culture and in the animal model leads to increased production of MCP-1, IL-6, MMP3, MMP10, and MMP13, all of which are known to contribute to ILD. Gene delivery of NFHEV also induces inflammation and local collagen deposition. Although extracellular IL-33 in patients may also be somewhat elevated, there is an accompanying elevation in soluble T1/ST2, which neu- tralizes IL-33 cytokine activity but has no effect on the function of NFHEV. Gene deficiency of T1/ST2 has min- imal if any attenuating effect on the severity of ILD in the bleomycin model. Based on these data, the Hypothe- sis of this proposal is that NFHEV is a central T1/ST2-independent regulator of ILD, acting through stimulation of proinflammatory and profibrotic gene expression. If successful in validating this role of NFHEV, this study will lay the groundwork for future development of NFHEV-targeting strategies and thus lead to better therapies for ILD. To assess the validity of this Hypothesis, three Specific Aims will be addressed.
Aim 1 is to prove, in the bleomycin model of interstitial lung disease, that NFHEV worsens pulmonary inflammation and fibrosis through T1/ST2-independent regulation of proinflammatory and profibrotic gene expression.
Aim 2 is to define pathophysiologically important NFHEV-expressing cell types by determining whether ubiquitous and/or cell type-specific deficiency of NFHEV protects against pulmonary inflammation and fibrosis in the bleomycin mod- el of interstitial lung disease.
This Aim will utilize mice with inducible cell type-specific NFHEV gene knockout.
Aim 3 will take advantage of the spatial heterogeneity of inflammation and fibrosis in the lungs of human pa- tients with IPF to determine whether increased expression of NFHEV is associated with active fibrotic foci, cel- lular inflammation, and expression of candidate NFHEV-driven proinflammatory and profibrotic factors. This work investigates unknown aspects of NFHEV biology in association with a deadly lung disease prevalent in Veterans. The results will lay the groundwork for future development of NFHEV-targeting therapies in Veterans and in the general population. Thus, this proposal directly addresses the needs of Veterans and contributes to the VA Healthcare Mission.

Public Health Relevance

Interstitial lung disease (ILD) can be severely debilitating and even deadly. This is particularly true in Veterans, who, because of their military exposures and aging, develop ILD more frequently and in more severe forms. Despite progress in understanding ILD, it continues taking toll on Veterans. Our work suggests that a molecule called NFHEV/IL-33 plays a critical role in ILD. This research will determine how this molecule propels ILD and will create the basis for the development of future medicines. As such, this research is directly applicable to the VA patient care mission, because it is aimed at developing better therapies for Veterans and thus at improving Veterans'welfare, quality of life, and survival.

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