Authorship note: KJL and DK are co-senior authors. Conflict of interest: DK serves on The Scientific Advisory Board of and receives research support from Compass Therapeutics. DK and KJL have a pending patent entitled "Compositions and methods for detecting CCR2 receptors" (US patent application no. 15/611,577).

IntroductionCigarette smoke is a profound pro-inflammatory stimulus that contributes to acute lung injuries and to chronic lung disease including COPD (emphysema and chronic bronchitis). Until recently, it was assumed that resolution of inflammation was a passive process that occurred once the inflammatory stimulus was removed. It is now recognized that resolution of inflammation is a bioactive process, mediated by specialized lipid mediators, and that normal homeostasis is maintained by a balance between pro-inflammatory and pro-resolving pathways. These novel small lipid mediators, including the resolvins, protectins and maresins, are bioactive products mainly derived from dietary omega-3 and omega-6 polyunsaturated fatty acids (PUFA). We hypothesize that resolvin D1 (RvD1) has potent anti-inflammatory and pro-resolving effects in a model of cigarette smoke-induced lung inflammation.MethodsPrimary human lung fibroblasts, small airway epithelial cells and blood monocytes were treated with IL-1β or cigarette smoke extract in combination with RvD1 in vitro, production of pro-inflammatory mediators was measured. Mice were exposed to dilute mainstream cigarette smoke and treated with RvD1 either concurrently with smoke or after smoking cessation. The effects on lung inflammation and lung macrophage populations were assessed.ResultsRvD1 suppressed production of pro-inflammatory mediators by primary human cells in a dose-dependent manner. Treatment of mice with RvD1 concurrently with cigarette smoke exposure significantly reduced neutrophilic lung inflammation and production of pro-inflammatory cytokines, while upregulating the anti-inflammatory cytokine IL-10. RvD1 promoted differentiation of alternatively activated (M2) macrophages and neutrophil efferocytosis. RvD1 also accelerated the resolution of lung inflammation when given after the final smoke exposure.ConclusionsRvD1 has potent anti-inflammatory and pro-resolving effects in cells and mice exposed to cigarette smoke. Resolvins have strong potential as a novel therapeutic approach to resolve lung injury caused by smoke and pulmonary toxicants.

IntroductionSeptic shock syndrome resulting from excessive host immune responses induced by infectious organisms is a leading cause of death in hospitalized patients. [1][2][3] Pathophysiologic changes in sepsis involve the pathogen-induced uncontrolled release from immune cells, particularly monocytes and macrophages, of proinflammatory mediators. 4 Gram-negative bacterial infection is one of the major causes of systemic bacterial sepsis. 5 Lipopolysaccharide (LPS), a constituent of the Gram-negative outer membrane, is the leading cause of sepsis. LPS induces a rapid increase of proinflammatory mediators, leading to lethal systemic tissue damage and multiple organ failure, which mimics the inflammatory responses of septic syndrome. 6 In mammals, membrane-bound CD14 and toll-like receptor 4 (TLR4)-MD-2 participate in cellular recognition of LPS. 7 Binding of LPS to TLR4 triggers the activation of members of the mitogen-activated protein kinase (MAPK) pathway including p38, p42/p44 extracellular signal-regulated kinase (ERK1/2), and c-Jun N-terminal kinase (JNK). 8 In resting unstimulated cells, nuclear factor-B (NF-B), a heterodimeric complex composed of 50-and 65-kDa (p50/p65) protein subunits, 9 retains as an inactive complex bound to inhibitory B␣ (IB␣) in the cytoplasm. While the cells are under proinflammatory stimulation by LPS, phosphorylation and degradation of IB␣ permit NF-B nuclear translocation and promote the expression of inflammatory genes including inducible nitric oxide synthase (iNOS), tumor necrosis factor-␣ (TNF-␣), and others. 9Thrombomodulin (TM) is a 557 amino acid type I glycosylated transmembrane protein 10 with an NH 2 -terminal lectinlike region (domain 1; D1) followed by 6 epidermal growth factor (EGF)-like structures (domain 2; D2), an O-glycosylation site-rich domain (domain 3; D3), a transmembrane domain (domain 4; D4), and a cytoplasmic tail domain (domain 5; D5). TM domain 2 (TMD2) EGF-like structures are responsible for the anticoagulant activity of TM via the alteration of thrombin substrate specificity. TMD2-thrombin complex sequentially activates anticoagulant protein C inactivating procoagulant cofactors Va and VIIIa. 11 TM expression also occurs in keratinocytes, 12 polymorphonuclear neutrophils (PMNs), 13 monocytes, 14 and endothelial cells, 15 indicating additional functions of TM besides anticoagulation. 16 Indeed TM domains function as an adhesion molecule, 17 an angiogenic factor, 18 and an anti-inflammatory agent through protein C-dependent and -independent mechanisms. 16,19 Recently, anti-inflammatory activity of TM domain 1 (TMD1) was implied by observing that mice with a deleted TM lectinlike domain (TM LeD/LeD ) become more sensitive to LPS challenge through the suppressed expression of adhesion molecules via NFB and MAPK signaling pathways. 20 Moreover, mice with a mutation in the TM gene (TM pro/pro ) strongly reduce the capacity to generate activated protein C, an anti-inflammatory agent in treatment of sepsis. 21 Mice harboring the latter mutation display an u...

It is well established that maladaptive innate immune responses to sterile tissue injury represent a fundamental mechanism of disease pathogenesis. In the context of cardiac ischemia reperfusion injury, neutrophils enter inflamed heart tissue, where they play an important role in potentiating tissue damage and contributing to contractile dysfunction. The precise mechanisms that govern how neutrophils are recruited to and enter the injured heart are incompletely understood. Using a model of cardiac transplant–mediated ischemia reperfusion injury and intravital 2-photon imaging of beating mouse hearts, we determined that tissue-resident CCR2+ monocyte–derived macrophages are essential mediators of neutrophil recruitment into ischemic myocardial tissue. Our studies revealed that neutrophil extravasation is mediated by a TLR9/MyD88/CXCL5 pathway. Intravital 2-photon imaging demonstrated that CXCL2 and CXCL5 play critical and nonredundant roles in guiding neutrophil adhesion and crawling, respectively. Together, these findings uncover a specific role for a tissue-resident monocyte-derived macrophage subset in sterile tissue inflammation and support the evolving concept that macrophage ontogeny is an important determinant of function. Furthermore, our results provide the framework for targeting of cell-specific signaling pathways in myocardial ischemia reperfusion injury.

Ischemia-reperfusion injury, a form of sterile inflammation, is the leading risk factor for both short-term mortality following pulmonary transplantation and chronic lung allograft dysfunction. While it is well recognized that neutrophils are critical mediators of acute lung injury, processes that guide their entry into pulmonary tissue are not well understood. Here, we found that CCR2+ classical monocytes are necessary and sufficient for mediating extravasation of neutrophils into pulmonary tissue during ischemia-reperfusion injury following hilar clamping or lung transplantation. The classical monocytes were mobilized from the host spleen, and splenectomy attenuated the recruitment of classical monocytes as well as the entry of neutrophils into injured lung tissue, which was associated with improved graft function. Neutrophil extravasation was mediated by MyD88-dependent IL-1β production by graft-infiltrating classical monocytes, which downregulated the expression of the tight junction-associated protein ZO-2 in pulmonary vascular endothelial cells. Thus, we have uncovered a crucial role for classical monocytes, mobilized from the spleen, in mediating neutrophil extravasation, with potential implications for targeting of recipient classical monocytes to ameliorate pulmonary ischemia-reperfusion injury in the clinic.

Chronic obstructive pulmonary disease is characterized, in part, by chronic inflammation that persists even after smoking cessation, suggesting that a failure to resolve inflammation plays an important role in the pathogenesis of the disease. It is widely recognized that the resolution of inflammation is an active process, governed by specialized proresolving lipid mediators, including lipoxins, resolvins, maresins, and protectins. Here, we report that proresolving signaling and metabolic pathways are disrupted in lung tissue from patients with chronic obstructive pulmonary disease, suggesting that supplementation with proresolving lipid mediators might reduce the development of emphysema by controlling chronic inflammation. Groups of mice were exposed long-term to cigarette smoke and treated with the proresolving mediator resolvin D1. Resolvin D1 was associated with a reduced development of cigarette smokeeinduced emphysema and airspace enlargement, with concurrent reductions in inflammation, oxidative stress, and cell death. Interestingly, resolvin D1 did not promote the differentiation of M2 macrophages and did not promote tissue fibrosis. Taken together, our results suggest that cigarette smoking disrupts endogenous proresolving pathways and that supplementation with specialized proresolving lipid mediators is an important therapeutic strategy in chronic lung disease, especially if endogenous specialized proresolving lipid mediator signaling is impaired. (Am J Pathol 2015, 185: 3189e3201; http:// dx.doi.org/10.1016/j.ajpath.2015 Chronic obstructive pulmonary disease (COPD) is a major global health problem and a leading cause of death and disability. Tobacco smoking remains the major causative factor in the development of COPD; however, new evidence suggests that household air pollution from fuel used for indoor cooking and heating is also a significant cause.

Pulmonary Fibrosis (PF) is a devastating progressive disease in which normal lung structure and function is compromised by scarring. Lung fibrosis can be caused by thoracic radiation, injury from chemotherapy and systemic diseases such as rheumatoid arthritis that involve inflammatory responses. CDDO-Me (Methyl 2-cyano-3,12-dioxooleana-1,9(11)dien-28-oate, Bardoxolone methyl) is a novel triterpenoid with anti-fibrotic and anti-inflammatory properties as shown by our in vitro studies. Based on this evidence, we hypothesized that CDDO-Me would reduce lung inflammation, fibrosis and lung function impairment in a bleomycin model of lung injury and fibrosis. To test this hypothesis, mice received bleomycin via oropharyngeal aspiration (OA) on day zero and CDDO-Me during the inflammatory phase from days -1 to 9 every other day. Bronchoalveolar lavage fluid (BALF) and lung tissue were harvested on day 7 to evaluate inflammation, while fibrosis and lung function were evaluated on day 21. On day 7, CDDO-Me reduced total BALF protein by 50%, alveolar macrophage infiltration by 40%, neutrophil infiltration by 90% (p≤0.01), inhibited production of the inflammatory cytokines KC and IL-6 by over 90% (p≤0.001), and excess production of the pro-fibrotic cytokine TGFβ by 50%. CDDO-Me also inhibited α-smooth muscle actin and fibronectin mRNA by 50% (p≤0.05). On day 21, CDDO-Me treatment reduced histological fibrosis, collagen deposition and αSMA production. Lung function was significantly improved at day 21 by treatment with CDDO-Me, as demonstrated by respiratory rate and dynamic compliance. These new findings reveal that CDDO-Me exhibits potent anti-fibrotic and anti-inflammatory properties in vivo. CDDO-Me is a potential new class of drugs to arrest inflammation and ameliorate fibrosis in patients who are predisposed to lung injury and fibrosis incited by cancer treatments (e.g. chemotherapy and radiation) and by systemic autoimmune diseases.

The respiratory epithelium are lung sentinel cells and are the first to contact inhaled inflammatory insults including air pollutants, smoke and microorganisms. To avoid damaging exuberant or chronic inflammation, the inflammatory process must be tightly controlled and terminated once the insult is mitigated. Inflammation-resolution is now known to be an active process involving a new genus of lipid mediators called “specialized pro-resolving lipid mediators” (SPMs) that includes resolvin D1 (RvD1). We and others have reported that RvD1 counteracts pro-inflammatory signaling and promotes resolution. A knowledge gap is that the specific cellular targets and mechanisms of action for RvD1 remain largely unknown. Here, we identified the mechanism whereby RvD1 disrupts inflammatory mediator production induced by the viral mimic poly(I:C) in primary human lung epithelial cells. RvD1 strongly suppressed the viral mimic poly(I:C)-induced IL-6 and IL-8 production and pro-inflammatory signaling involving MAP kinases and NF-κB. Most importantly, we found that RvD1 inhibited the phosphorylation of TAK1, a key upstream regulatory kinase common to both the MAP kinase and NF-κB pathways, by inhibiting the formation of a poly(I:C)-induced signaling complex composed of TAK1, TAB1 and TRAF6. We confirmed that ALX/FPR2 and GPR32, two RvD1 receptors, were expressed on hSAEC. Furthermore, blocking these receptors abrogated the inhibitory action of RvD1. Herein, we present the idea that RvD1 has the potential to be used as an anti-inflammatory and pro-resolving agent, possibly in the context of exuberant host responses to damaging respirable agents such as viruses.