Specialized pro-resolving mediators enhance the clearance of dead cells.

The failure to resolve inflammation underpins to several prevalent diseases, like atherosclerosis, and so identifying ways to boost resolution is unmet clinical needs. The resolution of inflammation is governed by several factors such as specialized pro-resolving mediators (SPMs) that counter-regulate pro-inflammatory pathways and promote tissue repair without compromising host defense. A major function of nearly all SPMs is to enhance the clearance of dead cells or efferocytosis. As such, phagocytes, such as macrophages, are essential cellular players in the resolution of inflammation because of their ability to rapidly and efficiently clear dead cells. This review highlights the role of SPMs in the clearance of apoptotic and necroptotic cells and offers insights into how targeting efferocytosis may provide new treatments for non-resolving diseases, like atherosclerosis.

Resolvin D2 limits atherosclerosis progression via myeloid cell-GPR18.

Dysregulated inflammation-resolution programs are associated with atherosclerosis progression. Resolvins, in part, mediate inflammation-resolution programs. Indeed, Resolvin D2 (RvD2) activates GPR18, a G-protein-coupled receptor, and limits plaque progression, though the cellular targets of RvD2 remain unknown. Here, we developed a humanized GPR18 floxed ("fl/fl") and a myeloid (Lysozyme M Cre) GPR18 knockout (mKO) mouse. We functionally validated this model by assessing efferocytosis in bone marrow-derived macrophages (BMDMs) and found that RvD2 enhanced efferocytosis in the fl/fl, but not in the mKO BMDMs. To understand the functions of RvD2-GPR18 in atherosclerosis, we performed a bone marrow transfer of fl/fl or mKO bone marrow into Ldlr-/- recipients. For these experiments, we treated each genotype with either Vehicle/PBS or RvD2 (25 ng/mouse, 3 times/week for 3 weeks). Myeloid loss of GPR18 resulted in significantly more necrosis, increased cleaved caspase-3+ cells and decreased percentage of Arginase-1+ -Mac2+ cells without a change in overall Mac2+ plaque macrophages, compared with fl/fl➔Ldlr-/- transplanted mice. RvD2 treatment decreased plaque necrosis, the percent of cleaved caspase-3+ cells and increased the percent of Arginase-1+ -Mac2+ cells in fl/fl➔Ldlr-/- mice, but not in the mKO➔Ldlr-/- transplanted mice. These results suggest that GPR18 plays a causal role in limiting atherosclerosis progression and that RvD2's ability to limit plaque necrosis is in part dependent on myeloid GRP18.

Institutional Support for the Career Advancement of Women Faculty in Science and Academic Medicine: Successes, Challenges, and Future Directions.

Institutional support is crucial for the successful career advancement of all faculty but in particular those who are women. Evolving from the past, in which gender disparities were prevalent in many institutions, recent decades have witnessed significant progress in supporting the career advancement of women faculty in science and academic medicine. However, continued advancement is necessary as previously unrecognized needs and new opportunities for improvement emerge. To identify the needs, opportunities, and potential challenges encountered by women faculty, the Women's Leadership Committee of the Arteriosclerosis, Thrombosis, and Vascular Biology Council developed an initiative termed GROWTH (Generating Resources and Opportunities for Women in Technology and Health). The committee designed a survey questionnaire and interviewed 19 leaders with roles and responsibilities in faculty development from a total of 12 institutions across various regions of the United States. The results were compiled, analyzed, and discussed. Based on our interviews and analyses, we present the current status of these representative institutions in supporting faculty development, highlighting efforts specific to women faculty. Through the experiences, insights, and vision of these leaders, we identified success stories, challenges, and future priorities. Our article provides a primer and a snapshot of institutional efforts to support the advancement of women faculty. Importantly, this article can serve as a reference and resource for academic entities seeking ideas to gauge their commitment level to women faculty and to implement new initiatives. Additionally, this article can provide guidance and strategies for women faculty as they seek support and resources from their current or prospective institutions when pursuing new career opportunities.

Resolvin D2-G-Protein Coupled Receptor 18 Enhances Bone Marrow Function and Limits Steatosis and Hepatic Collagen Accumulation in Aging.

Aging is associated with nonresolving inflammation and tissue dysfunction. Resolvin D2 (RvD2) is a proresolving ligand that acts through the G-protein-coupled receptor called GPR18. Unbiased RNA sequencing revealed increased Gpr18 expression in macrophages from old mice, and in livers from elderly humans, which was associated with increased steatosis and fibrosis in middle-aged (MA) and old mice. MA mice that lacked GPR18 on myeloid cells had exacerbated steatosis and hepatic fibrosis, which was associated with a decline in Mac2+ macrophages. Treatment of MA mice with RvD2 reduced steatosis and decreased hepatic fibrosis, correlating with increased Mac2+ macrophages, increased monocyte-derived macrophages, and elevated numbers of monocytes in the liver, blood, and bone marrow. RvD2 acted directly on the bone marrow to increase monocyte-macrophage progenitors. A transplantation assay further demonstrated that bone marrow from old mice facilitated hepatic collagen accumulation in young mice. Transient RvD2 treatment to mice transplanted with bone marrow from old mice prevented hepatic collagen accumulation. Together, this study demonstrates that RvD2-GPR18 signaling controls steatosis and fibrosis and provides a mechanistic-based therapy for promoting liver repair in aging.

Radiation-Induced Macrophage Senescence Impairs Resolution Programs and Drives Cardiovascular Inflammation.

Radiation is associated with tissue damage and increased risk of atherosclerosis, but there are currently no treatments and a very limited mechanistic understanding of how radiation impacts tissue repair mechanisms. We uncovered that radiation significantly delayed temporal resolution programs that were associated with decreased efferocytosis in vivo. Resolvin D1 (RvD1), a known proresolving ligand, promoted swift resolution and restored efferocytosis in sublethally irradiated mice. Irradiated macrophages exhibited several features of senescence, including increased expression of p16INK4A and p21, heightened levels of SA-ß-gal, COX-2, several proinflammatory cytokines/chemokines, and oxidative stress (OS) in vitro, and when transferred to mice, they exacerbated inflammation in vivo. Mechanistically, heightened OS in senescent macrophages led to impairment in their ability to carry out efficient efferocytosis, and treatment with RvD1 reduced OS and improved efferocytosis. Sublethally irradiated Ldlr -/- mice exhibited increased plaque necrosis, p16INK4A cells, and decreased lesional collagen compared with nonirradiated controls, and treatment with RvD1 significantly reduced necrosis and increased lesional collagen. Removal of p16INK4A hematopoietic cells during advanced atherosclerosis with p16-3MR mice reduced plaque necrosis and increased production of key intraplaque-resolving mediators. Our results demonstrate that sublethal radiation drives macrophage senescence and efferocytosis defects and suggest that RvD1 may be a new therapeutic strategy to limit radiation-induced tissue damage.

What Makes a Great Mentor: Interviews With Recipients of the ATVB Mentor of Women Award.

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Resolvin D1 Enhances Necroptotic Cell Clearance Through Promoting Macrophage Fatty Acid Oxidation and Oxidative Phosphorylation.

OBJECTIVE: Plaque necrosis is a key feature of defective resolution in atherosclerosis. Recent evidence suggests that necroptosis promotes plaque necrosis; therefore, we sought to determine how necroptotic cells (NCs) impact resolution programs in plaques. Approach and Results: To investigate the role(s) of necroptosis in advanced atherosclerosis, we used mice deficient of Mlkl, an effector of necroptosis. Mlkl-/- mice that were injected with a gain-of-function mutant PCSK9 (AAV8-gof-PCSK9) and fed a Western diet for 16 weeks, showed significantly less plaque necrosis, increased fibrous caps and improved efferocytosis compared with AAV8-gof-PCSK9 injected wt controls. Additionally, hypercholesterolemic Mlkl-/- mice had a significant increase in proresolving mediators including resolvin D1 (RvD1) and a decrease in prostanoids including thromboxane in plaques and in vitro. We found that exuberant thromboxane released by NCs impaired the clearance of both apoptotic cells and NCs through disruption of oxidative phosphorylation in macrophages. Moreover, we found that NCs did not readily synthesize RvD1 and that exogenous administration of RvD1 to macrophages rescued NC-induced defective efferocytosis. RvD1 also enhanced the uptake of NCs via the activation of p-AMPK (AMP-activated protein kinase), increased fatty acid oxidation, and enhanced oxidative phosphorylation in macrophages. CONCLUSIONS: These results suggest that NCs derange resolution by limiting key SPMs and impairing the efferocytic repertoire of macrophages. Moreover, these findings provide a molecular mechanism for RvD1 in directing proresolving metabolic programs in macrophages and further suggests RvD1 as a potential therapeutic strategy to limit NCs in tissues. Graphic Abstract: A graphic abstract is available for this article.

Resolvin D1 promotes efferocytosis in aging by limiting senescent cell-induced MerTK cleavage.

Inflammation-resolution is mediated by the balance between specialized pro-resolving mediators (SPMs) like resolvin D1 (RvD1) and pro-inflammatory factors, like leukotriene B4 (LTB4). A key cellular process of inflammation-resolution is efferocytosis. Aging is associated with defective inflammation-resolution and the accumulation of pro-inflammatory senescent cells (SCs). Therefore, understanding mechanism(s) that underpin this impairment is a critical gap. Here, using a model of hind limb ischemia-reperfusion (I/R) remote lung injury, we present evidence that aging is associated with heightened inflammation, impaired SPM:LT ratio, defective efferocytosis, and a decrease in MerTK levels in injured lungs. Treatment with RvD1 mitigated I/R lung injury in aging, promoted efferocytosis, and prevented the decrease of MerTK in injured lungs from old mice. Old MerTK cleavage-resistant mice (MerTKCR) exhibited less neutrophils or polymorpho nuclear cells infiltration and had improved efferocytosis compared with old WT controls. Mechanistically, macrophages that were treated with conditioned media (CM) from senescent cells had increased MerTK cleavage, impaired efferocytosis, and a defective RvD1:LTB4 ratio. Macrophages from MerTKCR mice were resistant to CM-induced efferocytosis defects and had an improved RvD1:LTB4 ratio. RvD1-stimulated macrophages prevented CM-induced MerTK cleavage and promoted efferocytosis. Together, these data suggest a new mechanism and a potential therapy to promote inflammation-resolution and efferocytosis in aging.

MerTK cleavage limits proresolving mediator biosynthesis and exacerbates tissue inflammation.

The acute inflammatory response requires a coordinated resolution program to prevent excessive inflammation, repair collateral damage, and restore tissue homeostasis, and failure of this response contributes to the pathology of numerous chronic inflammatory diseases. Resolution is mediated in part by long-chain fatty acid-derived lipid mediators called specialized proresolving mediators (SPMs). However, how SPMs are regulated during the inflammatory response, and how this process goes awry in inflammatory diseases, are poorly understood. We now show that signaling through the Mer proto-oncogene tyrosine kinase (MerTK) receptor in cultured macrophages and in sterile inflammation in vivo promotes SPM biosynthesis by a mechanism involving an increase in the cytoplasmic:nuclear ratio of a key SPM biosynthetic enzyme, 5-lipoxygenase. This action of MerTK is linked to the resolution of sterile peritonitis and, after ischemia-reperfusion (I/R) injury, to increased circulating SPMs and decreased remote organ inflammation. MerTK is susceptible to ADAM metallopeptidase domain 17 (ADAM17)-mediated cell-surface cleavage under inflammatory conditions, but the functional significance is not known. We show here that SPM biosynthesis is increased and inflammation resolution is improved in a new mouse model in which endogenous MerTK was replaced with a genetically engineered variant that is cleavage-resistant (Mertk(CR)). Mertk(CR) mice also have increased circulating levels of SPMs and less lung injury after I/R. Thus, MerTK cleavage during inflammation limits SPM biosynthesis and the resolution response. These findings contribute to our understanding of how SPM synthesis is regulated during the inflammatory response and suggest new therapeutic avenues to boost resolution in settings where defective resolution promotes disease progression.

Boosting Inflammation Resolution in Atherosclerosis: The Next Frontier for Therapy.

Defective inflammation resolution is the underlying cause of prevalent chronic inflammatory diseases, such as arthritis, asthma, cancer, and neurodegenerative and cardiovascular diseases. Inflammation resolution is governed by several endogenous factors, including fatty acid-derived specialized proresolving mediators and proteins, such as annexin A1. Specifically, specialized proresolving mediators comprise a family of mediators that include arachidonic acid-derived lipoxins, omega-3 fatty acid eicosapentaenoic acid-derived resolvins, docosahexaenoic acid-derived resolvins, protectins, and maresins. Emerging evidence indicates that imbalances between specialized proresolving mediators and proinflammatory mediators are associated with several prevalent human diseases, including atherosclerosis. Mechanisms that drive this imbalance remain largely unknown and will be discussed in this review. Furthermore, the concept of dysregulated inflammation resolution in atherosclerosis has been known for several decades. Recently, there has been an explosion of new work with regard to the therapeutic application of proresolving ligands in experimental atherosclerosis. Therefore, this review will highlight recent advances in our understanding of how inflammation resolution may become defective in atherosclerosis and the potential for proresolving therapeutics in atherosclerosis. Last, we offer insight for future implications of the field.

Infection regulates pro-resolving mediators that lower antibiotic requirements.

Underlying mechanisms for how bacterial infections contribute to active resolution of acute inflammation are unknown. Here, we performed exudate leukocyte trafficking and mediator-metabololipidomics of murine peritoneal Escherichia coli infections with temporal identification of pro-inflammatory (prostaglandins and leukotrienes) and specialized pro-resolving mediators (SPMs). In self-resolving E. coli exudates (10(5) colony forming units, c.f.u.), the dominant SPMs identified were resolvin (Rv) D5 and protectin D1 (PD1), which at 12 h were at significantly greater levels than in exudates from higher titre E. coli (10(7) c.f.u.)-challenged mice. Germ-free mice had endogenous RvD1 and PD1 levels higher than in conventional mice. RvD1 and RvD5 (nanograms per mouse) each reduced bacterial titres in blood and exudates, E. coli-induced hypothermia and increased survival, demonstrating the first actions of RvD5. With human polymorphonuclear neutrophils and macrophages, RvD1, RvD5 and PD1 each directly enhanced phagocytosis of E. coli, and RvD5 counter-regulated a panel of pro-inflammatory genes, including NF-κB and TNF-α. RvD5 activated the RvD1 receptor, GPR32, to enhance phagocytosis. With self-limited E. coli infections, RvD1 and the antibiotic ciprofloxacin accelerated resolution, each shortening resolution intervals (R(i)). Host-directed RvD1 actions enhanced ciprofloxacin's therapeutic actions. In 10(7) c.f.u. E. coli infections, SPMs (RvD1, RvD5, PD1) together with ciprofloxacin also heightened host antimicrobial responses. In skin infections, SPMs enhanced vancomycin clearance of Staphylococcus aureus. These results demonstrate that specific SPMs are temporally and differentially regulated during infections and that they are anti-phlogistic, enhance containment and lower antibiotic requirements for bacterial clearance.

Fine-tuning inflammation-resolution programs: focus on atherosclerosis.

PURPOSE OF REVIEW: Nonresolving inflammation is now considered the underpinning of several prevalent human diseases, including atherosclerosis. The resolution of inflammation is a highly coordinated program to counterbalance proinflammatory signals for a swift return to tissue homeostasis. This process is controlled in part by endogenous specialized proresolving lipid mediators (SPMs). Emerging evidence has revealed that the balance of SPMs and proinflammatory mediators during acute inflammation regulates the duration of the inflammatory response and the timing of tissue resolution. Moreover, an imbalance between SPMs and proinflammatory mediators has been linked to several prevalent chronic inflammatory diseases in humans, including atherosclerosis. RECENT FINDINGS: Lipid mediator imbalances have recently been linked to atherosclerotic plaque instability. Administration of key SPMs restored this imbalance and led to plaque stability. SPMs have also recently been shown to be protective in other cardiovascular disease models including myocardial infarction, stroke and neointimal hyperplasia. SUMMARY: The current review highlights recent work that supports the concept of dysregulated inflammation-resolution in atherosclerosis with a particular focus on mechanisms and therapeutic opportunities associated with SPM receptors and lipid mediator imbalances. This article is based on experimental studies.

Pro-resolution therapeutics for cardiovascular diseases.

Studies over the last couple of decades suggest that failed resolution of a chronic inflammatory response is an important driving force in the progression of atherosclerosis. Resolution of inflammation is mediated in part by lipid-derived specialized pro-resolving mediators (SPMs) such as lipoxins, resolvins, protectins and maresins. The major functions of SPMs are to quell inflammation and repair tissue damage in a manner that does not compromise host defense. An imbalance between SPMs and pro-inflammatory mediators like leukotriene B4 (LTB4) are associated with several prevalent human diseases, including atherosclerosis. Because atherosclerosis is marked by persistent, unresolved inflammation and arterial tissue injury, SPMs have garnered immense interest as a potential treatment strategy. This mini review will highlight recent advances in the application of SPMs in atherosclerosis as well as the ability of SPMs to control several of the risk factors associated with cardiovascular diseases.

Resolvin D1 limits 5-lipoxygenase nuclear localization and leukotriene B4 synthesis by inhibiting a calcium-activated kinase pathway.

Imbalances between proinflammatory and proresolving mediators can lead to chronic inflammatory diseases. The balance of arachidonic acid-derived mediators in leukocytes is thought to be achieved through intracellular localization of 5-lipoxygenase (5-LOX): nuclear 5-LOX favors the biosynthesis of proinflammatory leukotriene B4 (LTB4), whereas, in theory, cytoplasmic 5-LOX could favor the biosynthesis of proresolving lipoxin A4 (LXA4). This balance is shifted in favor of LXA4 by resolvin D1 (RvD1), a specialized proresolving mediator derived from docosahexaenoic acid, but the mechanism is not known. Here we report a new pathway through which RvD1 promotes nuclear exclusion of 5-LOX and thereby suppresses LTB4 and enhances LXA4 in macrophages. RvD1, by activating its receptor formyl peptide receptor2/lipoxin A4 receptor, suppresses cytosolic calcium and decreases activation of the calcium-sensitive kinase calcium-calmodulin-dependent protein kinase II (CaMKII). CaMKII inhibition suppresses activation P38 and mitogen-activated protein kinase-activated protein kinase 2 kinases, which reduces Ser271 phosphorylation of 5-LOX and shifts 5-LOX from the nucleus to the cytoplasm. As such, RvD1's ability to decrease nuclear 5-LOX and the LTB4:LXA4 ratio in vitro and in vivo was mimicked by macrophages lacking CaMKII or expressing S271A-5-LOX. These findings provide mechanistic insight into how a specialized proresolving mediator from the docosahexaenoic acid pathway shifts the balance toward resolution in the arachidonic acid pathway. Knowledge of this mechanism may provide new strategies for promoting inflammation resolution in chronic inflammatory diseases.

Inflammation and its resolution as determinants of acute coronary syndromes.

Inflammation contributes to many of the characteristics of plaques implicated in the pathogenesis of acute coronary syndromes. Moreover, inflammatory pathways not only regulate the properties of plaques that precipitate acute coronary syndromes but also modulate the clinical consequences of the thrombotic complications of atherosclerosis. This synthesis will provide an update on the fundamental mechanisms of inflammatory responses that govern acute coronary syndromes and also highlight the ongoing balance between proinflammatory mechanisms and endogenous pathways that can promote the resolution of inflammation. An appreciation of the countervailing mechanisms that modulate inflammation in relation to acute coronary syndromes enriches our fundamental understanding of the pathophysiology of this important manifestation of atherosclerosis. In addition, these insights provide glimpses into potential novel therapeutic interventions to forestall this ultimate complication of the disease.

Development and in vivo efficacy of targeted polymeric inflammation-resolving nanoparticles.

Excessive inflammation and failed resolution of the inflammatory response are underlying components of numerous conditions such as arthritis, cardiovascular disease, and cancer. Hence, therapeutics that dampen inflammation and enhance resolution are of considerable interest. In this study, we demonstrate the proresolving activity of sub-100-nm nanoparticles (NPs) containing the anti-inflammatory peptide Ac2-26, an annexin A1/lipocortin 1-mimetic peptide. These NPs were engineered using biodegradable diblock poly(lactic-co-glycolic acid)-b-polyethyleneglycol and poly(lactic-co-glycolic acid)-b-polyethyleneglycol collagen IV-targeted polymers. Using a self-limited zymosan-induced peritonitis model, we show that the Ac2-26 NPs (100 ng per mouse) were significantly more potent than Ac2-26 native peptide at limiting recruitment of polymononuclear neutrophils (56% vs. 30%) and at decreasing the resolution interval up to 4 h. Moreover, systemic administration of collagen IV targeted Ac2-26 NPs (in as low as 1 µg peptide per mouse) was shown to significantly block tissue damage in hind-limb ischemia-reperfusion injury by up to 30% in comparison with controls. Together, these findings demonstrate that Ac2-26 NPs are proresolving in vivo and raise the prospect of their use in chronic inflammatory diseases such as atherosclerosis.

Resolvin E1 and chemokine-like receptor 1 mediate bone preservation.

The polyunsaturated ω-3 fatty acid eicosapentaenoic acid-derived resolvin E1 (RvE1) enhances resolution of inflammation, prevents bone loss, and induces bone regeneration. Although the inflammation-resolving actions of RvE1 are characterized, the molecular mechanism of its bone-protective actions are of interest. To test the hypothesis that receptor-mediated events impact bone changes, we prepared transgenic mice overexpressing the RvE1 receptor chemokine-like receptor 1 (chemR23) on leukocytes. In zymosan-initiated peritonitis, neutrophil polymorphonuclear leukocyte infiltration in response to RvE1 was limited requiring log order lower doses in chemR23tg mice. Ligature-induced alveolar bone loss was diminished in chemR23tg mice. Local RvE1 treatment of uniform craniotomy in the parietal bone significantly accelerated regeneration of the bone defect. In in vitro bone cultures, RvE1 significantly enhanced expression of osteoprotegerin (OPG) without inducing change in receptor activator of NF-κB ligand levels, whereas the osteogenic markers alkaline phosphatase, bone sialoprotein, and Runt-related transcription factor 2 remained unchanged. These results indicate that RvE1 modulates osteoclast differentiation and bone remodeling by direct actions on bone, rescuing OPG production and restoring a favorable receptor activator of NF-κB ligand/OPG ratio, in addition to known anti-inflammatory and proresolving actions.