Anti-inflammatory M2 Phenotype Research Articles

Hidden Treasures: Macrophage Long Non-Coding RNAs in Lung Cancer Progression.

Simple SummaryCancer immunotherapy mainly targets immune system components, such as immune-suppressive networks generated by cancer cells in the tumor microenvironment (TME). Programmed cell death ligand 1, which is a secretory immune-suppressive factor, is released by tumor-associated macrophages (TAMs). The TME also disrupts production of tumor-specific T cells and generates immunosuppressive leukocytes, regulatory T cells, and myeloid-derived suppressor cells. Immune checkpoint inhibitors are effective in various cancers but only in a subset of patients. Non-coding RNAs, such as microRNAs (miRNAs), long non-coding RNAs (lncRNAs), and circular RNAs (circRNAs), are dysregulated in cancer through transcriptional, post-transcriptional, and epigenetic changes and have significant roles in cancer initiation and progression, which depends on deregulation of lncRNA expression. TAM function can be influenced by lncRNAs in various ways. However, our understanding of lncRNA dysregulation and function in cancer remains in the early stage.Ever since RNA sequencing of whole genomes and transcriptomes became available, numerous RNA transcripts without having the classic function of encoding proteins have been discovered. Long non-coding RNAs (lncRNAs) with a length greater than 200 nucleotides were considered as “junk” in the beginning, but it has increasingly become clear that lncRNAs have crucial roles in regulating a variety of cellular mechanisms and are often deregulated in several diseases, such as cancer. Lung cancer is the leading cause of cancer-related deaths and has a survival rate of less than 10%. Immune cells infiltrating the tumor microenvironment (TME) have been shown to have a great effect on tumor development with macrophages being the major cell type within the TME. Macrophages can inherit an inflammatory M1 or an anti-inflammatory M2 phenotype. Tumor-associated macrophages, which are predominantly polarized to M2, favor tumor growth, angiogenesis, and metastasis. In this review, we aimed to describe the complex roles and functions of lncRNAs in macrophages and their influence on lung cancer development and progression through the TME.

Macrophages: The Good, the Bad, and the Gluttony.

Macrophages are dynamic cells that play critical roles in the induction and resolution of sterile inflammation. In this review, we will compile and interpret recent findings on the plasticity of macrophages and how these cells contribute to the development of non-infectious inflammatory diseases, with a particular focus on allergic and autoimmune disorders. The critical roles of macrophages in the resolution of inflammation will then be examined, emphasizing the ability of macrophages to clear apoptotic immune cells. Rheumatoid arthritis (RA) is a chronic autoimmune-driven spectrum of diseases where persistent inflammation results in synovial hyperplasia and excessive immune cell accumulation, leading to remodeling and reduced function in affected joints. Macrophages are central to the pathophysiology of RA, driving episodic cycles of chronic inflammation and tissue destruction. RA patients have increased numbers of active M1 polarized pro-inflammatory macrophages and few or inactive M2 type cells. This imbalance in macrophage homeostasis is a main contributor to pro-inflammatory mediators in RA, resulting in continual activation of immune and stromal populations and accelerated tissue remodeling. Modulation of macrophage phenotype and function remains a key therapeutic goal for the treatment of this disease. Intriguingly, therapeutic intervention with glucocorticoids or other DMARDs promotes the re-polarization of M1 macrophages to an anti-inflammatory M2 phenotype; this reprogramming is dependent on metabolic changes to promote phenotypic switching. Allergic asthma is associated with Th2-polarised airway inflammation, structural remodeling of the large airways, and airway hyperresponsiveness. Macrophage polarization has a profound impact on asthma pathogenesis, as the response to allergen exposure is regulated by an intricate interplay between local immune factors including cytokines, chemokines and danger signals from neighboring cells. In the Th2-polarized environment characteristic of allergic asthma, high levels of IL-4 produced by locally infiltrating innate lymphoid cells and helper T cells promote the acquisition of an alternatively activated M2a phenotype in macrophages, with myriad effects on the local immune response and airway structure. Targeting regulators of macrophage plasticity is currently being pursued in the treatment of allergic asthma and other allergic diseases. Macrophages promote the re-balancing of pro-inflammatory responses towards pro-resolution responses and are thus central to the success of an inflammatory response. It has long been established that apoptosis supports monocyte and macrophage recruitment to sites of inflammation, facilitating subsequent corpse clearance. This drives resolution responses and mediates a phenotypic switch in the polarity of macrophages. However, the role of apoptotic cell-derived extracellular vesicles (ACdEV) in the recruitment and control of macrophage phenotype has received remarkably little attention. ACdEV are powerful mediators of intercellular communication, carrying a wealth of lipid and protein mediators that may modulate macrophage phenotype, including a cargo of active immune-modulating enzymes. The impact of such interactions may result in repair or disease in different contexts. In this review, we will discuss the origin, characterization, and activity of macrophages in sterile inflammatory diseases and the underlying mechanisms of macrophage polarization via ACdEV and apoptotic cell clearance, in order to provide new insights into therapeutic strategies that could exploit the capabilities of these agile and responsive cells.

P–238 Impaired oocyte fertilization is associated with a proinflammatory M1 phenotype of macrophages and the activation of the NLRC4 inflammasome

Abstract Study question Are inflammatory signatures in the blood of clinically asymptomatic patients predictive for oocyte fertilization success in ART treatment? Summary answer The proinflammatory M1 phenotype of macrophages combined with increasing numbers of cytotoxic T-cells and an upregulated inflammasome NLRC4 is associated with impaired oocyte fertilization. What is known already Oocyte fertilization is an indispensable step towards embryogenesis and reproductive success. Patients with acute or chronic inflammation are at risk of reduced fertilization rates and failure of assisted reproductive technology (ART) treatment. Inflammasomes are intracellular multiprotein complexes that mediate inflammatory tissue remodeling. They can be activated by endogenous and exogenous stimuli and result in maturation of the cytokines IL–1βand IL–18. Appropriate inflammasome activation serves for pathogen defense and tissue damage repair, while aberrant activation of inflammasomes can enhance uncontrolled tissue damage. Study design, size, duration This a prospective study including 39 patients stratified by fertilization rate (cut off 66%) of mature M II oocytes after ICSI procedure, compares 20 patients with high (mean 84,8%, range 67 –100%) versus 19 patients with low (mean 29.4%, range 0–65%) rates. Participants/materials, setting, methods We performed FACS analysis of immune cell composition (leukocytes, neutrophils, monocytes, macrophage types M1 and M2, cytotoxic T-cells, T-helper cells) and utilized RT-qPCR analysis of three inflammasomes (AIM 2, NLRP3 and NLRC4) and their down-streaming proteins caspase 1 and IL–1ß. We focused on an initial and a late state during controlled ovarian stimulation (COS) procedure. Main results and the role of chance This study reports a cellular and molecular inflammatory signature associated with reduced oocyte fertilization rates after ICSI treatment in clinically asymptomatic patients. On a cellular level proinflammatory M1 macrophages were significantly elevated in the low fertilization group (p < 0.01), additionally the ratio of proinflammatory M1 macrophages to anti-inflammatory M2 phenotype was inversely associated with oocyte fertilization rate (p < 0.001). Cytotoxic T cells (p < 0.05) and the inflammasome NLRC4 (p < 0.01) revealed identical patterns of association with fertilization rates in the group comparison. In line with this pattern, there was a significant upregulation of Caspase 1 with a consecutive increase in IL–1ß activity in women with low fertilization rates (p < 0.05). No significant association was shown for the other tested immune cells (leukocytes, neutrophils, monocytes, T-helper cells) and inflammasomes (NLRC3, AIM2). Limitations, reasons for caution The preliminary results in this small study indicate an activation of the inflammasome NLRC4. This, however, has to be assessed on the protein level in a larger group of patients. Wider implications of the findings: Systemic macrophage augmentation of M1 phenotype and cytotoxic T cell presence in combination with upregulated inflammasome NLRC4 are associated with impaired oocyte fertilization. The predictive information of the identified immunologic signature could indicate targeted treatment to down regulate systemic inflammation and the use of oocyte activators in the ART laboratory. Trial registration number none

Immunometabolic Modulatory Role of Naltrexone in BV-2 Microglia Cells.

Background: Naltrexone is an opioid receptor antagonist commonly used to treat opioid and alcohol dependence. The use of low dose naltrexone (LDN) was found to have anti-inflammatory properties for treatment of diseases such as fibromyalgia, Crohn’s disease, multiple sclerosis and regional pain syndromes. Related to its anti-neuroinflammatory properties, the mechanism of action is possibly mediated via Toll-like receptor 4 antagonism, which is widely expressed on microglial cells. The aim of the present study was to assess the immunometabolic effects of naltrexone on microglia cells in in vitro conditions. Methods: All experiments were performed in the BV-2 microglial cell line. The cells were treated with naltrexone at 100 μM concentrations corresponding to low dose for 24 h. Cell viability was assessed for every drug dose. To induce additional activation, the cells were pretreated with LPS and IFN-γ. Immunofluorescence was used to analyse the classical microglial activation markers iNOS and CD206, while Seahorse was used for real-time cellular metabolic assessments. mTOR activity measured over the expression of a major direct downstream target S6K was assessed using western blot. Results: LDN induced a shift from highly activated pro-inflammatory phenotype (iNOShighCD206low) to quiescent anti-inflammatory M2 phenotype (iNOSlowCD206high) in BV-2 microglia cells. Changes in the inflammatory profile were accompanied by cellular metabolic switching based on the transition from high glycolysis to mitochondrial oxidative phosphorylation (OXPHOS). LDN-treated cells were able to maintain a metabolically suppressive phenotype by supporting OXPHOS with high oxygen consumption, and also maintain a lower energetic state due to lower lactate production. The metabolic shift induced by transition from glycolysis to mitochondrial oxidative metabolism was more prominent in cells pretreated with immunometabolic modulators such as LPS and IFN-γ. In a dose-dependent manner, naltrexone also modulated mTOR/S6K expression, which underlies the cell metabolic phenotype regulating microglia immune properties and adaptation. Conclusion: By modulating the phenotypic features by metabolic switching of activated microglia, naltrexone was found to be an effective and powerful tool for immunometabolic reprogramming and could be a promising novel treatment for various neuroinflammatory conditions.

Activation of Neuroprotective Microglia and Astrocytes at the Lesion Site and in the Adjacent Segments Is Crucial for Spontaneous Locomotor Recovery after Spinal Cord Injury.

Microglia and astrocytes play an important role in the regulation of immune responses under various pathological conditions. To detect environmental cues associated with the transformation of reactive microglia (M1) and astrocytes (A1) into their polarization states (anti-inflammatory M2 and A2 phenotypes), we studied time-dependent gene expression in naive and injured spinal cord. The relationship between astrocytes and microglia and their polarization states were studied in a rat model after Th9 compression (40 g/15 min) in acute and subacute stages at the lesion site, and both cranially and caudally. The gene expression of microglia/macrophages and M1 microglia was strongly up-regulated at the lesion site and caudally one week after SCI, and attenuated after two weeks post-SCI. GFAP and S100B, and A1 astrocytes were profoundly expressed predominantly two weeks post-SCI at lesion site and cranially. Gene expression of anti-inflammatory M2a microglia (CD206, CHICHI, IL1rn, Arg-1), M2c microglia (TGF-β, SOCS3, IL4R α) and A2 astrocytes (Tgm1, Ptx3, CD109) was greatly activated at the lesion site one week post-SCI. In addition, we observed positive correlation between neurological outcome and expression of M2a, M2c, and A2 markers. Our findings indicate that the first week post-injury is critical for modulation of reactive microglia/astrocytes into their neuroprotective phenotypes.

Interrod spacing dependent angiogenesis and osseointegration of Na2TiO3 nanorods-patterned arrays via immunoregulation

Nanorods-arrayed coatings show topographical parameters-dependent immunomodulation that plays a key role in angiogenesis and consequent osseoinegration, while immunomodulation by interrod spacing, as a solely altered topographic parameter of an array, has not been clarified yet. In this context, four kinds of nanorods-arrayed coatings, appearing similar surface roughness and hydrophilia, were hydrothermally grown on Ti. The coatings comprise a bilayer structure with compact nanogranular Na2TiO3 as an inner layer and Na2TiO3 nanorods-patterned array as an outer layer. The nanorods in the coatings exhibit identical rod diameter and length but different interrod spacing values of ~ 45, 75, 140, and 245 nm, respectively. Compared to Ti, the arrayed coatings promote adhesion and filopodia formation of macrophages to accelarate their transformation from pro-inflammatory M1 phenotype to anti-inflammatory M2 phenotype. Moreover, the accelaration is more pronounced by the coatings with nanoscale interrod spacing (especially ~ 75 nm) compared to those with submicro-scale spacing. The rapid phenotypic switch of macrophages on the arrayed coatings trigers the increase in their secretion of pro-angiogenic factors, including PDGF-BB, ANG1 and SDF-1, to enhance recruitment and angiogenesis of human umbilical vein endothelial cells (HUVECs) in co-culture condition, and the enhancment is the most pronounced for HUVECs in response to macrophages on ~ 75 nm spaced nanorods-arrayed coating. Owing to the favorable immune microenvironment induced by the coating and consequently enhanced angiogenesis, ~ 75 nm spaced nanorods-arrayed coating exhibits the best osseoinetegration in rat femoral condyles. Our work provides insights into the surface structural design of an orthopedic implant favoring osseointegration.

Biological fabrication and electrostatic attractions of new layered silver/talc nanocomposite using Lawsonia inermis L. and its chitosan-capped inorganic/organic hybrid: Investigation on acceleration of Staphylococcus aureus and Pseudomonas aeruginosa infected wound healing

In the present study, new-layered inorganic/organic hybrid of silver/talc nanocomposites (Ag/Tlc-NPs) and its chitosan-capped derivative (Ag/Tlc/Csn NCs) were biochemically synthesized utilizing Lawsonia inermis L. extract. The silver nanoparticles (Ag NPs) were synthesized employing green method on the exterior surface layer of talc mineral as a solid substrate. The negatively charged surface layer of talc might function as templates and can attract the chitosan cations from a solution to yield a layered hybrid structure, whose inorganic phase is formed by Si-O-Ag bonds. Our results revealed that Ag NPs were formed on the exterior surface of talc with a diameter with size of 124–215 nm. In addition, cytotoxicity, in vitro antibacterial activity, and clinical effects of wound-healing ointments containing talc were investigated. The results implied the successful synthesis of Ag/Tlc/Csn NCs using the extract. The structures were safe up to 0.50 mg/mL. In vitro studies confirmed antioxidant and antibacterial properties of Ag/Tlc/Csn NCs. In sum, our findings showed that the ointments improve wound healing process by inducing an anti-inflammatory M2 phenotype and bFGF, CD206, collagen1A, and IL-10 production that causes fibroblast migration and wound closure through influencing M2 macrophage. Ag/Tlc/Csn is suggested to be taken into consideration as a medical combination for improving infected wound healing and as a promising agent for clinical administration.

HAI-1 is an independent predictor of lung cancer mortality and is required for M1 macrophage polarization.

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death worldwide. Though immune checkpoint inhibitors (ICIs) have revolutionized lung cancer therapy in recent years, there are several factors limiting the therapeutic efficacy of ICI-based immunotherapy in lung cancer. Recent evidence suggests that one such mechanism is the phenotypic shift of tumor-infiltrating macrophages away from an anti-tumor M1 phenotype and towards an anti-inflammatory and tumor-permissive M2 phenotype. Though this phenomenon is well documented, the means through which the lung tumor microenvironment (TME) usurps macrophage function are poorly described. Hepatocyte growth factor (HGF) is a known driver of both lung cancer pathobiology as well as M2 polarization, and its signaling is antagonized by the tumor suppressor gene HAI-1 (SPINT1). Using a combination of genomic databases, primary NSCLC specimens, and in vitro models, we determined that patients with loss of HAI-1 have a particularly poor prognosis, hallmarked by increased HGF expression and an M2-dominant immune infiltrate. Similarly, conditioned media from HAI-1-deficient tumor cells led to a loss of M1 and increased M2 polarization in vitro, and patient NSCLC tissues with loss of HAI-1 showed a similar loss of M1 macrophages. Combined, these results suggest that loss of HAI-1 is a potential means through which tumors acquire an immunosuppressive, M2-dominated TME, potentially through impaired M1 macrophage polarization. Hence, HAI-1 status may be informative when stratifying patients that may benefit from therapies targeting the HGF pathway, particularly as an adjuvant to ICI-based immunotherapy.

Transplantation of Human Gingiva-Derived Mesenchymal Stem Cells Ameliorates Neurotic Erectile Dysfunction in a Rat Model.

Cavernous nerve injury (CNI) is the main cause of erectile dysfunction (ED) following pelvic surgery. Our previous studies have demonstrated that transplantation of different sources of mesenchymal stem cells (MSCs) was able to alleviate ED induced by CNI in rat models. However, little is known about the therapeutic effects of human gingiva-derived MSCs (hGMSCs) in CNI ED rats. Herein, we injected the hGMSCs around the bilateral major pelvic ganglia (MPG) in a rat model of CNI and evaluated their efficacy. The results showed that treatment of hGMSCs could significantly promote the recovery of erectile function, enhance smooth muscle and endothelial content, restore neuronal nitric oxide synthase (nNOS) expression, and attenuate cell apoptosis in penile tissue. Moreover, penile fibrosis was significantly alleviated after hGMSC administration. In addition, potential mechanism exploration indicated that hGMSCs might exert its functions via skewed macrophage polarity from M1 toward M2 anti-inflammatory phenotype. In conclusion, this study found that transplantation of hGMSCs significantly improved CNI-related ED, which might provide new clues to evaluate their pre-clinical application.

The osteoimmunomodulatory effect of nanostructured TiF x /TiO x coating on osteogenesis induction

Macrophages play a central role in the host response and the integration of implant materials. The nanostructured TiF x /TiO x coating (FOTi) on titanium surfaces has shown multiple properties, including antibacterial properties and bioactivity. However, little is known about the effects of these coatings on the regulation of macrophage activity and the subsequent immunomodulatory effects on osteogenesis. In this study, the behavior of macrophages on the FOTi samples was evaluated, and conditioned medium was collected and used to stimulate MC3T3-E1 cells in vitro. The results showed that the FOTi samples stimulated macrophage elongation and promoted the production of proinflammatory cytokines at 24 h, while induced macrophage polarization to the anti-inflammatory M2 phenotype at 72 h. Furthermore, the immune microenvironment generated by macrophage/ FOTi samples interactions effectively promoted the osteogenic differentiation of MC3T3-E1 cells, as evidenced by improved cell adhesion, enhanced alkaline phosphatase activity and extracellular matrix mineralization, and upregulated osteogenesis-related gene expression. In summary, the FOTi samples mediated macrophage phenotype behaviors and induced beneficial immunomodulatory effects on osteogenesis, which could be a potential strategy for the surface modification of bone biomaterials.

Effect of Hyperbaric Oxygen Therapy on Polarization Phenotype of Rat Microglia After Traumatic Brain Injury.

Background: The neurological defect caused by secondary damage following traumatic brain injury (TBI) is considered critical for the management of TBI. Microglia (MG) are a resident brain macrophage that could differentiate into M1 type or M2 type in response to injury and repair. It is known that the MG transition from M1 phenotype to anti-inflammatory M2 phenotype might reduce secondary injury of TBI. So, a TBI animal model was established and we compared biomarkers of M1 and M2MG between the controls and experimental animals receiving hyperbaric oxygen therapy (HBOT). This study aimed to explore whether HBOT was an effective method to improve neural functional recovery via promoting the polarization of MG into M2 after TBI.Methods: The rats were randomly divided into four groups: SH (Sham-operated), SH + HBO (hyperbaric oxygen), TBI, and TBI + HBO. Each group included 42 rats, and each of these were divided into the following groups: 1, 6, 12, 24, 72 h, 7, and 14 days. The expression of M1 biomarker inducible nitric oxide synthase (iNOS), M2 biomarker arginase 1 (Arg1), associated cytokine tumor necrosis factor-α (TNF-α), and transforming growth factor-β1 (TGF-β1) was evaluated after the observation time.Results: TBI significantly increased the expression levels of M1 marker iNOS and M2 markers Arg1 at different time points. The increased expression of iNOS was suppressed, while the expression level of Arg1 was enhanced by HBOT. Moreover, HBOT suppressed the pro-inflammatory TNF-α secreted by M1, and promoting the anti-inflammatory TGF-1β.Conclusions: In the present study, HBOT showed the effects on shift of M1 toward M2 phenotype with increased expression of M2 biomarkers and decreased expression of M1 biomarkers in the early stage after TBI.

Evaluation of the immunomodulatory effects of cobalt, copper and magnesium ions in a pro inflammatory environment

Biomaterials and scaffolds for Tissue Engineering are widely used for an effective healing and regeneration. However, the implantation of these scaffolds causes an innate immune response in which the macrophage polarization from M1 (pro-inflammatory) to M2 (anti-inflammatory) phenotype is crucial to avoid chronic inflammation. Recent studies have showed that the use of bioactive ions such as cobalt (Co2+), copper (Cu2+) and magnesium (Mg2+) could improve tissue regeneration, although there is limited evidence on their effect on the macrophage response. Therefore, we investigated the immunomodulatory potential of Co2+, Cu2+ and Mg2+ in macrophage polarization. Our results indicate that Mg2+ and concentrations of Cu2+ lower than 10 μM promoted the expression of M2 related genes. However, higher concentrations of Cu2+ and Co2+ (100 μM) stimulated pro-inflammatory marker expression, indicating a concentration dependent effect of these ions. Furthermore, Mg2+ were able to decrease M1 marker expression in presence of a mild pro-inflammatory stimulus, showing that Mg2+ can be used to modulate the inflammatory response, even though their application can be limited in a strong pro-inflammatory environment.

Anti-Inflammatory and Immunomodulatory Effects of the Grifola frondosa Natural Compound o-Orsellinaldehyde on LPS-Challenged Murine Primary Glial Cells. Roles of NF-κβ and MAPK.

In response to foreign or endogenous stimuli, both microglia and astrocytes adopt an activated phenotype that promotes the release of pro-inflammatory mediators. This inflammatory mechanism, known as neuroinflammation, is essential in the defense against foreign invasion and in normal tissue repair; nevertheless, when constantly activated, this process can become detrimental through the release of neurotoxic factors that amplify underlying disease. In consequence, this study presents the anti-inflammatory and immunomodulatory properties of o-orsellinaldehyde, a natural compound found by an in silico approach in the Grifola frondosa mushroom, in astrocytes and microglia cells. For this purpose, primary microglia and astrocytes were isolated from mice brain and cultured in vitro. Subsequently, cells were exposed to LPS in the absence or presence of increasing concentrations of this natural compound. Specifically, the results shown that o-orsellinaldehyde strongly inhibits the LPS-induced inflammatory response in astrocytes and microglia by decreasing nitrite formation and downregulating iNOS and HO-1 expression. Furthermore, in microglia cells o-orsellinaldehyde inhibits NF-κB activation; and potently counteracts LPS-mediated p38 kinase and JNK phosphorylation (MAPK). In this regard, o-orsellinaldehyde treatment also induces a significant cell immunomodulation by repolarizing microglia toward the M2 anti-inflammatory phenotype. Altogether, these results could partially explain the reported beneficial effects of G. frondosa extracts on inflammatory conditions.

Annexin A1 protects against cerebral ischemia\u2013reperfusion injury by modulating microglia/macrophage polarization via FPR2/ALX-dependent AMPK-mTOR pathway

BackgroundCerebral ischemia–reperfusion (I/R) injury is a major cause of early complications and unfavorable outcomes after endovascular thrombectomy (EVT) therapy in patients with acute ischemic stroke (AIS). Recent studies indicate that modulating microglia/macrophage polarization and subsequent inflammatory response may be a potential adjunct therapy to recanalization. Annexin A1 (ANXA1) exerts potent anti-inflammatory and pro-resolving properties in models of cerebral I/R injury. However, whether ANXA1 modulates post-I/R-induced microglia/macrophage polarization has not yet been fully elucidated.MethodsWe retrospectively collected blood samples from AIS patients who underwent successful recanalization by EVT and analyzed ANXA1 levels longitudinally before and after EVT and correlation between ANXA1 levels and 3-month clinical outcomes. We also established a C57BL/6J mouse model of transient middle cerebral artery occlusion/reperfusion (tMCAO/R) and an in vitro model of oxygen–glucose deprivation and reoxygenation (OGD/R) in BV2 microglia and HT22 neurons to explore the role of Ac2-26, a pharmacophore N-terminal peptide of ANXA1, in regulating the I/R-induced microglia/macrophage activation and polarization.ResultsThe baseline levels of ANXA1 pre-EVT were significantly lower in 23 AIS patients, as compared with those of healthy controls. They were significantly increased to the levels found in controls 2–3 days post-EVT. The increased post-EVT levels of ANXA1 were positively correlated with 3-month clinical outcomes. In the mouse model, we then found that Ac2-26 administered at the start of reperfusion shifted microglia/macrophage polarization toward anti-inflammatory M2-phenotype in ischemic penumbra, thus alleviating blood–brain barrier leakage and neuronal apoptosis and improving outcomes at 3 days post-tMCAO/R. The protection was abrogated when mice received Ac2-26 together with WRW4, which is a specific antagonist of formyl peptide receptor type 2/lipoxin A4 receptor (FPR2/ALX). Furthermore, the interaction between Ac2-26 and FPR2/ALX receptor activated the 5’ adenosine monophosphate-activated protein kinase (AMPK) and inhibited the downstream mammalian target of rapamycin (mTOR). These in vivo findings were validated through in vitro experiments.ConclusionsAc2-26 modulates microglial/macrophage polarization and alleviates subsequent cerebral inflammation by regulating the FPR2/ALX-dependent AMPK-mTOR pathway. It may be investigated as an adjunct strategy for clinical prevention and treatment of cerebral I/R injury after recanalization. Plasma ANXA1 may be a potential biomarker for outcomes of AIS patients receiving EVT.

Pinosylvin Shifts Macrophage Polarization to Support Resolution of Inflammation.

Pinosylvin is a natural stilbenoid found particularly in Scots pine. Stilbenoids are a group of phenolic compounds identified as protective agents against pathogens for many plants. Stilbenoids also possess health-promoting properties in humans; for instance, they are anti-inflammatory through their suppressing action on proinflammatory M1-type macrophage activation. Macrophages respond to environmental changes by polarizing towards proinflammatory M1 phenotype in infection and inflammatory diseases, or towards anti-inflammatory M2 phenotype, mediating resolution of inflammation and repair. In the present study, we investigated the effects of pinosylvin on M2-type macrophage activation, aiming to test the hypothesis that pinosylvin could polarize macrophages from M1 to M2 phenotype to support resolution of inflammation. We used lipopolysaccharide (LPS) to induce M1 phenotype and interleukin-4 (IL-4) to induce M2 phenotype in J774 murine and U937 human macrophages, and we measured expression of M1 and M2-markers. Interestingly, along with inhibiting the expression of M1-type markers, pinosylvin had an enhancing effect on the M2-type activation, shown as an increased expression of arginase-1 (Arg-1) and mannose receptor C type 1 (MRC1) in murine macrophages, and C-C motif chemokine ligands 17 and 26 (CCL17 and CCL26) in human macrophages. In IL-4-treated macrophages, pinosylvin enhanced PPAR-γ expression but had no effect on STAT6 phosphorylation. The results show, for the first time, that pinosylvin shifts macrophage polarization from the pro-inflammatory M1 phenotype towards M2 phenotype, supporting resolution of inflammation and repair.

Macrophages and Stem Cells-Two to Tango for Tissue Repair?

Macrophages (MCs) are present in all tissues, not only supporting homeostasis, but also playing an important role in organogenesis, post-injury regeneration, and diseases. They are a heterogeneous cell population due to their origin, tissue specificity, and polarization in response to aggression factors, depending on environmental cues. Thus, as pro-inflammatory M1 phagocytic MCs, they contribute to tissue damage and even fibrosis, but the anti-inflammatory M2 phenotype participates in repairing processes and wound healing through a molecular interplay with most cells in adult stem cell niches. In this review, we emphasize MC phenotypic heterogeneity in health and disease, highlighting their systemic and systematic contribution to tissue homeostasis and repair. Unraveling the intervention of both resident and migrated MCs on the behavior of stem cells and the regulation of the stem cell niche is crucial for opening new perspectives for novel therapeutic strategies in different diseases.

α/β‐Hydrolase Domain‐6 Regulates Macrophage Polarization and its Inhibition Protects Against LPS‐Induced Inflammation

The activated M1 macrophages exhibit pro-inflammatory activity and contribute to diverse inflammatory disorders, such as microbial infection, sepsis, autoimmune diseases, and cardiometabolic disorders. Thus, targeting macrophage polarization towards an anti-inflammatory M2 phenotype may provide a strategy to treat inflammatory diseases. Deletion of the monoacylglycerol lipase α/β-hydrolase domain-6 (ABHD6) demonstrated the therapeutic potential of targeting ABHD6 against obesity, type-2-diabetes, and other inflammatory disorders. However, the role of ABHD6 in macrophage activation/polarization under inflammatory conditions remains to be ascertained. Using macrophage cell lines, a pharmacological approach, and whole-body ABHD6 KO mice, we investigated whether ABHD6 suppression is protective against lipopolysaccharide (LPS)-induced inflammation. Pharmacological inhibition of ABHD6 activity with a potent inhibitor (KT203) in RAW264.7 and J774A.1 macrophages attenuates LPS-induced expression of pro-inflammatory cytokines (Tnfa and Mcp1). In addition, ABHD6 inhibition results in decreased LPS-induced secretion of TNFα and MCP1 from RAW264.7 macrophages. Interestingly, absence of ABHD6 activity promotes LPS-primed macrophage polarization towards the M2 phenotype, as evidenced by the increased arginase-1 (Arg1) expression. Furthermore, whole-body ABHD6 KO mice, compared to the wild-type controls, show lower susceptibility to systemic LPS-induced inflammation as indicated by a decline in plasma TNFα and MCP-1 levels and reduced expression of pro-inflammatory markers (Cd68 and Il1b) in peripheral blood mononuclear cells and lung tissue. Finally, ABHD6 KO mediated protection against acute LPS-induced inflammation is more prominent in female versus male mice. Taken together, our data support a pro-inflammatory role for ABHD6 under LPS-induced condition. ABHD6 inhibition switches LPS-induced macrophage polarization towards an anti-inflammatory M2 state, and ABHD6 deletion in mice attenuates systemic response to LPS challenge. The results suggest the therapeutic potential of ABHD6 inhibition for the treatment of inflammation-associated disorders.

Exercise‐linked FNDC5/Irisin attenuates macrophage‐mediated renal inflammation in type 1 diabetes in aged mice

Diabetes is associated with chronic low-grade inflammation in the kidney. Both classically and alternately activated macrophages play key roles in diabetic nephropathy by mediating inflammatory processes and anti-inflammatory responses respectively. Physical activity is known to modulate innate immune function, indeed exercise training is reported to reduce low-grade inflammation and improve insulin sensitivity in obese subjects. FNDC5 is a myokine secreted by skeletal muscles during exercise and the cleavage of its extracellular domain yields Irisin. In vitro studies have demonstrated that Irisin exhibits anti-inflammatory properties in adipose tissues and RAW 264.7 cells. The purpose of our study was to investigate the effect of exercise induced FNDC5/Irisin on inflammation and damage of aged kidney in type I diabetes. C57BL/6-Ins2Akita/J (Type 1 diabetes, Akita) and C57BL/6J (Wild type, WT) mice aged 54 - 56 weeks underwent 16-week exercise training of 280 m run on treadmill at speed of 7m/min. for 5 days/week for 2 weeks and increased to 12 m/min. for the remaining 14 weeks. FNDC5/Irisin was upregulated in the kidneys of WT and Akita exercise groups but to a greater magnitude in the WT. Sedentary Akita mice kidneys showed increased macrophage recruitment and M1 macrophages (CCL2, CXCL2, and CD86+/CD40+ cells) and inflammatory cytokine production. In contrast, Akita mice undergoing exercise training increased anti-inflammatory profile and M2 phenotype polarization (CD206+ cells). Exercise induced changes were associated with robust activation of AMPK and MAPK signaling in Akita mice. In comparison to sedentary group, exercise training reduced renal resistive index, improved cortical blood flow and glomerular filtration rate in WT and Akita mice. Taken together, our results suggest that exercise training and Irisin secretion protects the aged kidney from type 1 diabetes induced damage by reducing macrophage-mediated inflammation.

Imaging of Inflammation in Spinal Cord Injury: Novel Insights on the Usage of PFC-Based Contrast Agents.

Labeling of macrophages with perfluorocarbon (PFC)-based compounds allows the visualization of inflammatory processes by 19F-magnetic resonance imaging (19F-MRI), due to the absence of endogenous background. Even if PFC-labeling of monocytes/macrophages has been largely investigated and used, information is lacking about the impact of these agents over the polarization towards one of their cell subsets and on the best way to image them. In the present work, a PFC-based nanoemulsion was developed to monitor the course of inflammation in a model of spinal cord injury (SCI), a pathology in which the understanding of immunological events is of utmost importance to select the optimal therapeutic strategies. The effects of PFC over macrophage polarization were studied in vitro, on cultured macrophages, and in vivo, in a mouse SCI model, by testing and comparing various cell tracking protocols, including single and multiple administrations, the use of MRI or Point Resolved Spectroscopy (PRESS), and application of pre-saturation of Kupffer cells. The blood half-life of nanoemulsion was also investigated by 19F Magnetic Resonance Spectroscopy (MRS). In vitro and in vivo results indicate the occurrence of a switch towards the M2 (anti-inflammatory) phenotype, suggesting a possible theranostic function of these nanoparticles. The comparative work presented here allows the reader to select the most appropriate protocol according to the research objectives (quantitative data acquisition, visual monitoring of macrophage recruitment, theranostic purpose, rapid MRI acquisition, etc.). Finally, the method developed here to determine the blood half-life of the PFC nanoemulsion can be extended to other fluorinated compounds.

Camptothecin Regulates Microglia Polarization and Exerts Neuroprotective Effects via Activating AKT/Nrf2/HO-1 and Inhibiting NF-κB Pathways In Vivo and In Vitro.

Microglia, the main immune cells in the brain, participate in the innate immune response in the central nervous system (CNS). Studies have shown that microglia can be polarized into pro-inflammatory M1 and anti-inflammatory M2 phenotypes. Accumulated evidence suggests that over-activated M1 microglia release pro-inflammatory mediators that damage neurons and lead to Parkinson’s disease (PD). In contrast, M2 microglia release neuroprotective factors and exert the effects of neuroprotection. Camptothecin (CPT), an extract of the plant Camptotheca acuminate, has been reported to have anti-inflammation and antitumor effects. However, the effect of CPT on microglia polarization and microglia-mediated inflammation responses has not been reported. In our study we found that CPT improved motor performance of mice and reduced the loss of neurons in the substantia nigra (SN) of the midbrain in LPS-injected mice. In the mechanism study, we found that CPT inhibited M1 polarization of microglia and promotes M2 polarization via the AKT/Nrf2/HO-1 and NF-κB signals. Furthermore, CPT protected the neuroblastoma cell line SH-SY5Y and dopaminergic neuron cell line MN9D from damage mediated by microglia activation. In conclusion, our results demonstrate that CPT regulates the microglia polarization phenotype via activating AKT/Nrf2/HO-1 and inhibiting NF-κB pathways, inhibits neuro-inflammatory responses, and exerts neuroprotective effects in vivo and in vitro.