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Inflammation mediated by the peripheral infiltration of inflammatory cells plays an important role in intracerebral hemorrhage (ICH) induced secondary injury. Previous studies have indicated that regulatory T lymphocytes (Tregs) might reduce ICH-induced inflammation, but the precise mechanisms that contribute to ICH-induced inflammatory injury remain unclear. Our results show that the number of Tregs in the brain increases after ICH. Inducing Tregs deletion using a CD25 antibody or Foxp3 DTR -mice increased neurological deficient scores (NDS), the level of inflammatory factors, hematoma volumes, and neuronal degeneration. Meanwhile, boosting Tregs using a CD28 super-agonist antibody reduced the inflammatory injury. Furthermore, Tregs depletion shifted microglia/macrophage polarization toward the M1 phenotype while boosting Tregs shifted this transition toward the M2 phenotype. In vitro, a transwell co-culture model of microglia and Tregs indicated that Tregs changed the polarization of microglia, decreased the expression of MHC-II, IL-6, and TNF-α and increased CD206 expression. IL-10 originating from Tregs mediated the microglia polarization by increasing the expression of Glycogen Synthase Kinase 3 beta (GSK3β), which phosphorylates and inactivates Phosphatase and Tensin homologue (PTEN) in microglia, TGF-β did not participate in this conversion. Thus, Tregs ameliorated ICH-induced inflammatory injury by modulating microglia/macrophage polarization toward the M2 phenotype through the IL-10/GSK3β/PTEN axis.

Blood neutrophils perform an essential host-defense function by directly migrating to bacterial invasion sites to kill bacteria. The mechanisms mediating the transition from the migratory to bactericidal phenotype remain elusive. Here, we demonstrate that TRPM2, a trp superfamily member, senses neutrophil-generated reactive oxygen species and restrains neutrophil migration. The inhibitory function of oxidant sensing by TRPM2 requires the oxidation of Cys549, which then induces TRMP2 binding to formyl peptide receptor 1 (FPR1) and subsequent FPR1 internalization and signaling inhibition. The oxidant sensing-induced termination of neutrophil migration at the site of infection permits a smooth transition to the subsequent microbial killing phase., (Copyright © 2016 Elsevier Inc. All rights reserved.)

There are currently no effective treatments for sepsis and acute respiratory distress syndrome (ARDS). The repositioning of existing drugs is one possible effective strategy for the treatment of sepsis and ARDS. We previously showed that vascular repair and the resolution of sepsis-induced inflammatory lung injury is dependent upon endothelial HIF-1α/FoxM1 signaling. The aim of this study was to identify a candidate inducer of HIF-1α/FoxM1 signaling for the treatment of sepsis and ARDS. Employing high throughput screening of a library of 1200 FDA-approved drugs by using hypoxia response element (HRE)-driven luciferase reporter assays, we identified Rabeprazole (also known as Aciphex) as a top HIF-α activator. In cultured human lung microvascular endothelial cells, Rabeprazole induced HIF1A mRNA expression in a dose-dependent manner. A dose-response study of Rabeprazole in a mouse model of endotoxemia-induced inflammatory lung injury identified a dose that was well tolerated and enhanced vascular repair and the resolution of inflammatory lung injury. Rabeprazole treatment resulted in reductions in lung vascular leakage, edema, and neutrophil sequestration and proinflammatory cytokine expression during the repair phrase. We next used Hif1a/Tie2Cre knockout mice and Foxm1/Tie2Cre knockout mice to show that Rabeprazole promoted vascular repair through HIF-1α/FoxM1 signaling. In conclusion, Rabeprazole is a potent inducer of HIF-1α that promotes vascular repair and the resolution of sepsis-induced inflammatory lung injury via endothelial HIF-1α/FoxM1 signaling. This drug therefore represents a promising candidate for repurposing to effectively treat severe sepsis and ARDS. [ABSTRACT FROM AUTHOR]

A patient’s recovery from lung inflammatory injury or development of multi-system organ failure is determined by the host’s ability to resolve inflammation and repair tissue damage, both of which require the clearance of apoptotic neutrophils by macrophages (efferocytosis). Here, we investigated the effects of isoflurane on macrophage efferocytosis and resolution of lung inflammatory injury. Treatment of murine bone marrow-derived macrophages (BMDMs) or alveolar macrophages with isoflurane dramatically enhanced phagocytosis of apoptotic neutrophils. Isoflurane significantly increased the surface expression of the receptor tyrosine kinase Mer in macrophages, but markedly decreased the levels of a soluble form of Mer protein in the medium. Isoflurane treatment also caused a decrease in a disintegrin and metalloproteinase 17 (ADAM17) on the cell surface and a concomitant increase in its cytoplasmic fraction. These responses induced by isoflurane were completely reversed by a pharmacological inhibitor or genetic deletion of AMP-activated protein kinase (AMPK). In a mouse model of lipopolysaccharide-induced lung injury, isoflurane accelerated the recovery of lung inflammation and injury that was coupled with an increase in the number of alveolar macrophages containing apoptotic bodies. In alveolar macrophage-depleted mice, administration of isoflurane-pretreated BMDMs facilitated resolution of lung inflammation following lipopolysaccharide challenge. Thus, isoflurane promoted resolution of lipopolysaccharide-induced lung inflammatory injury via enhancement of macrophage efferocytosis. Increased macrophage efferocytosis following isoflurane treatment correlates with upregulation of Mer surface expression through AMPK-mediated blockade of ADAM17 trafficking to the cell membrane. [ABSTRACT FROM AUTHOR]

Acute respiratory distress syndrome (ARDS) is characterized by acute hypoxemia respiratory failure, bilateral pulmonary infiltrates, and pulmonary edema of non-cardiac origin. Effective treatments for ARDS patients may arise from experimental studies with translational mouse models of this disease that aim to delineate the mechanisms underlying the disease pathogenesis. Mouse models of ARDS, however, can be limited by their rapid progression from injured to recovery state, which is in contrast to the course of ARDS in humans. Furthermore, current mouse models of ARDS do not recapitulate certain prominent aspects of the pathogenesis of ARDS in humans. In this study, we developed an improved endotoxemic mouse model of ARDS resembling many features of clinical ARDS including extended courses of injury and recovery as well as development of fibrosis following i.p. injection of lipopolysaccharide (LPS) to corn oil-preloaded mice. Compared with mice receiving LPS alone, those receiving corn oil and LPS exhibited extended course of lung injury and repair that occurred over a period of >2 weeks instead of 3–5days. Importantly, LPS challenge of corn oil-preloaded mice resulted in pulmonary fibrosis during the repair phase as often seen in ARDS patients. In summary, this simple novel mouse model of ARDS could represent a valuable experimental tool to elucidate mechanisms that regulate lung injury and repair in ARDS patients. [ABSTRACT FROM AUTHOR]

Inflammation mediated by the peripheral infiltration of inflammatory cells plays an important role in intracerebral hemorrhage (ICH) induced secondary injury. Previous studies have indicated that regulatory T lymphocytes (Tregs) might reduce ICH-induced inflammation, but the precise mechanisms that contribute to ICH-induced inflammatory injury remain unclear. Our results show that the number of Tregs in the brain increases after ICH. Inducing Tregs deletion using a CD25 antibody or Foxp3DTR-mice increased neurological deficient scores (NDS), the level of inflammatory factors, hematoma volumes, and neuronal degeneration. Meanwhile, boosting Tregs using a CD28 super-agonist antibody reduced the inflammatory injury. Furthermore, Tregs depletion shifted microglia/macrophage polarization toward the M1 phenotype while boosting Tregs shifted this transition toward the M2 phenotype. In vitro, a transwell co-culture model of microglia and Tregs indicated that Tregs changed the polarization of microglia, decreased the expression of MHC-II, IL-6, and TNF-α and increased CD206 expression. IL-10 originating from Tregs mediated the microglia polarization by increasing the expression of Glycogen Synthase Kinase 3 beta (GSK3β), which phosphorylates and inactivates Phosphatase and Tensin homologue (PTEN) in microglia, TGF-β did not participate in this conversion. Thus, Tregs ameliorated ICH-induced inflammatory injury by modulating microglia/macrophage polarization toward the M2 phenotype through the IL-10/GSK3β/PTEN axis. [ABSTRACT FROM AUTHOR]

Adult stem cell treatment is a potential novel therapeutic approach for acute respiratory distress syndrome. Given the extremely low rate of cell engraftment, it is believed that these cells exert their beneficial effects via paracrine mechanisms. However, the endogenous mediator(s) in the pulmonary vasculature remains unclear. Using the mouse model with endothelial cell (EC)-restricted disruption of FoxM1 ( FoxM1 CKO), here we show that endothelial expression of the reparative transcriptional factor FoxM1 is required for the protective effects of bone marrow progenitor cells (BMPC) against LPS-induced inflammatory lung injury and mortality. BMPC treatment resulted in rapid induction of FoxM1 expression in wild type (WT) but not FoxM1 CKO lungs. BMPC-induced inhibition of lung vascular injury, resolution of lung inflammation, and survival, as seen in WT mice, were abrogated in FoxM1 CKO mice following LPS challenge. Mechanistically, BMPC treatment failed to induce lung EC proliferation in FoxM1 CKO mice, which was associated with impaired expression of FoxM1 target genes essential for cell cycle progression. We also observed that BMPC treatment enhanced endothelial barrier function in WT but not in FoxM1-deficient EC monolayers. Restoration of β-catenin expression in FoxM1-deficient ECs normalized endothelial barrier enhancement in response to BMPC treatment. These data demonstrate the requisite role of endothelial FoxM1 in the mechanism of BMPC-induced vascular repair to restore vascular integrity and accelerate resolution of inflammation, thereby promoting survival following inflammatory lung injury. S tem C ells 2014;32:1855-1864 [ABSTRACT FROM AUTHOR]

Adult stem cell-based therapy is a promising novel approach for treatment of acute lung injury. Here we investigated the therapeutic potential of freshly isolated human umbilical cord blood CD34+ progenitor cells (fCB-CD34+ cells) in a mouse model of acute lung injury. At 3 h post-lipopolysaccharide (LPS) challenge, fCB-CD34+ cells were transplanted i.v. to mice while CD34− cells or PBS were administered as controls in separate cohorts of mice. We observed that fCB-CD34+ cell treatment inhibited lung vascular injury evident by decreased lung vascular permeability. In contrast, CD34− cells had no effects on lung vascular injury. Lung inflammation determined by myeloperoxidase activity, neutrophil sequestration and expression of pro-inflammatory mediators was attenuated in fCB-CD34+ cell-treated mice at 26 h post-LPS challenge compared to PBS or CD34− cell-treated controls. Importantly, lung inflammation in fCB-CD34+ cell-treated mice was returned to normal levels as seen in basal mice at 52 h post-LPS challenge whereas PBS or CD34− cell-treated control mice exhibited persistent lung inflammation. Accordingly, fCB-CD34+ cell-treated mice exhibited a marked increase of survival rate. Employing in vivo 5-bromo-2′-deoxyuridine incorporation assay, we found a drastic induction of lung endothelial proliferation in fCB-CD34+ cell-treated mice at 52 h post-LPS compared to PBS or CD34− cell-treated controls, which contributed to restoration of vascular integrity and thereby inhibition of lung inflammation. Taken together, these data have demonstrated the protective effects of fCB-CD34+ cell on acute lung injury induced by LPS challenge, suggesting fCB-CD34+ cells are an important source of stem cells for the treatment of acute lung injury. [ABSTRACT FROM AUTHOR]

Background: Disturbance of brain iron metabolism after intracerebral hemorrhage (ICH) results in oxidative brain injury and cognition impairment. Hepcidin plays an important role in regulating iron metabolism, and we have reported that serum hepcidin is positively correlated with poor outcomes in patients with ICH. However, the roles of hepcidin in brain iron metabolism after ICH remain largely unknown.Methods: Parabiosis and ICH models combined with in vivo and in vitro experiments were used to investigate the roles of hepcidin in brain iron metabolism after ICH.Results: Increased hepcidin-25 was found in serum and primarily in astrocytes after ICH. The brain iron efflux, oxidative brain injury, and cognition impairment were improved in Hepc-/- ICH mice but aggravated by the human hepcidin-25 peptide in C57BL/6 ICH mice. Data obtained in in vitro studies showed that increased hepcidin inhibited the intracellular iron efflux of brain microvascular endothelial cells but was rescued by a hepcidin antagonist, fursultiamine. Using parabiosis ICH models also shows that increased serum hepcidin prevents brain iron efflux. In addition, Toll-like receptor 4 (TLR4)/MyD88 signaling pathway increased hepcidin expression by promoting interleukin-6 expression and signal transducer and activator of transcription 3 phosphorylation. TLR4-/- and MyD88-/- mice exhibited improvement in brain iron efflux at 7, 14, and 28 days after ICH, and the TLR4 antagonist (6R)-6-[N-(2-chloro-4-fluorophenyl) sulfamoyl] cyclohex-1-ene-1-carboxylate significantly decreased brain iron levels at days 14 and 28 after ICH and improved cognition impairment at day 28.Conclusions: The results presented here show that increased hepcidin expression caused by inflammation prevents brain iron efflux via inhibition of the intracellular iron efflux of brain microvascular endothelial cells entering into circulation and aggravating oxidative brain injury and cognition impairment, which identifies a mechanistic target for muting inflammation to promote brain iron efflux and to attenuate oxidative brain injury after ICH. [ABSTRACT FROM AUTHOR]

Background: Disturbance of brain iron metabolism after intracerebral hemorrhage (ICH) results in oxidative brain injury and cognition impairment. Hepcidin plays an important role in regulating iron metabolism, and we have reported that serum hepcidin is positively correlated with poor outcomes in patients with ICH. However, the roles of hepcidin in brain iron metabolism after ICH remain largely unknown.Methods: Parabiosis and ICH models combined with in vivo and in vitro experiments were used to investigate the roles of hepcidin in brain iron metabolism after ICH.Results: Increased hepcidin-25 was found in serum and primarily in astrocytes after ICH. The brain iron efflux, oxidative brain injury, and cognition impairment were improved in Hepc−/− ICH mice but aggravated by the human hepcidin-25 peptide in C57BL/6 ICH mice. Data obtained in in vitro studies showed that increased hepcidin inhibited the intracellular iron efflux of brain microvascular endothelial cells but was rescued by a hepcidin antagonist, fursultiamine. Using parabiosis ICH models also shows that increased serum hepcidin prevents brain iron efflux. In addition, Toll-like receptor 4 (TLR4)/MyD88 signaling pathway increased hepcidin expression by promoting interleukin-6 expression and signal transducer and activator of transcription 3 phosphorylation. TLR4−/− and MyD88−/− mice exhibited improvement in brain iron efflux at 7, 14, and 28 days after ICH, and the TLR4 antagonist (6R)-6-[N-(2-chloro-4-fluorophenyl) sulfamoyl] cyclohex-1-ene-1–carboxylate significantly decreased brain iron levels at days 14 and 28 after ICH and improved cognition impairment at day 28.Conclusions: The results presented here show that increased hepcidin expression caused by inflammation prevents brain iron efflux via inhibition of the intracellular iron efflux of brain microvascular endothelial cells entering into circulation and aggravating oxidative brain injury and cognition impairment, which identifies a mechanistic target for muting inflammation to promote brain iron efflux and to attenuate oxidative brain injury after ICH. [ABSTRACT FROM AUTHOR]

Background: The integrity of endothelial monolayer is a sine qua non for vascular homeostasis and maintenance of tissue-fluid balance. However, little is known about the signaling pathways regulating regeneration of the endothelial barrier after inflammatory vascular injury.Methods and Results: Using genetic and pharmacological approaches, we demonstrated that endothelial regeneration selectively requires activation of p110γPI3K signaling, which thereby mediates the expression of the endothelial reparative transcription factor Forkhead box M1 (FoxM1). We observed that FoxM1 induction in the pulmonary vasculature was inhibited in mice treated with a p110γ-selective inhibitor and in Pik3cg(-/-) mice after lipopolysaccharide challenge. Pik3cg(-/-) mice exhibited persistent lung inflammation induced by sepsis and sustained increase in vascular permeability. Restoration of expression of either p110γ or FoxM1 in pulmonary endothelial cells of Pik3cg(-/-) mice restored endothelial regeneration and normalized the defective vascular repair program. We also observed diminished expression of p110γ in pulmonary vascular endothelial cells of patients with acute respiratory distress syndrome, suggesting that impaired p110γ-FoxM1 vascular repair signaling pathway is a critical factor in persistent leaky lung microvessels and edema formation in the disease.Conclusions: We identify p110γ as the critical mediator of endothelial regeneration and vascular repair after sepsis-induced inflammatory injury. Thus, activation of p110γ-FoxM1 endothelial regeneration may represent a novel strategy for the treatment of inflammatory vascular diseases. [ABSTRACT FROM AUTHOR]

Background: The blood-brain barrier (BBB) formed by brain endothelial cells interconnected by tight junctions is essential for the homeostasis of the central nervous system. Although studies have shown the importance of various signaling molecules in BBB formation during development, little is known about the molecular basis regulating the integrity of the adult BBB.Methods and Results: Using a mouse model with tamoxifen-inducible endothelial cell-restricted disruption of ctnnb1 (iCKO), we show here that endothelial β-catenin signaling is essential for maintaining BBB integrity and central nervous system homeostasis in adult mice. The iCKO mice developed severe seizures accompanied by neuronal injury, multiple brain petechial hemorrhages, and central nervous system inflammation, and all had postictal death. Disruption of endothelial β-catenin induced BBB breakdown and downregulation of the specific tight junction proteins claudin-1 and -3 in adult brain endothelial cells. The clinical relevance of the data is indicated by the observation of decreased expression of claudin-1 and nuclear β-catenin in brain endothelial cells of hemorrhagic lesions of hemorrhagic stroke patients.Conclusions: These results demonstrate the prerequisite role of endothelial β-catenin in maintaining the integrity of adult BBB. The results suggest that BBB dysfunction secondary to defective β-catenin transcription activity is a key pathogenic factor in hemorrhagic stroke, seizure activity, and central nervous system inflammation. [ABSTRACT FROM AUTHOR]

Summary: Hepatocellular carcinoma (HCC) is an aggressive malignancy mainly due to metastases or postsurgical recurrence. We postulate that metastases are influenced by the liver microenvironment. Here, we show that a unique inflammation/immune response-related signature is associated with noncancerous hepatic tissues from metastatic HCC patients. This signature is principally different from that of the tumor. A global Th1/Th2-like cytokine shift in the venous metastasis-associated liver microenvironment coincides with elevated expression of macrophage colony-stimulating factor (CSF1). Moreover, a refined 17 gene signature was validated as a superior predictor of HCC venous metastases in an independent cohort, when compared to other clinical prognostic parameters. We suggest that a predominant humoral cytokine profile occurs in the metastatic liver milieu and that a shift toward anti-inflammatory/immune-suppressive responses may promote HCC metastases. [Copyright &y& Elsevier]

Sepsis patients that exhibit lung vascular injury and high mortality often suffer from severe pulmonary thrombosis (e.g. disseminated intravascular coagulation), but the beneficial impact of anti-coagulants in clinical trials of these patients is modest or absent. Although the regulation of sepsis-induced thrombosis has been intensely investigated, the converse impact of thrombosis on post-sepsis vascular injury is incompletely understood. The main aims of this study were to reveal the impact of varying severities of pulmonary thrombosis on sepsis-induced lung injury, and to determine whether hypoxia-inducible factor (HIF) 1α in endothelial and myeloid cells moderates inflammatory lung injury following thrombotic insult. We employed a mouse model of lung thrombosis in which different numbers of polystyrene microbeads are intra-venously seeded into the lung microvasculature, before or after i.p. lipopolysaccharide challenge. In microbead-free mice challenged with lipopolysaccharide, platelet depletion resulted in reduced pulmonary thrombosis and enhanced lung injury and inflammation. Administration of i.v. polystyrene microbeads, however, restored the level of pulmonary thrombosis to that found in platelet-replete mice, and protected against depletion-induced lung injury and inflammation. We then showed in wild type mice, that while severe increases in the level of pulmonary micro-occlusion worsened post-sepsis lung injury and inflammation, induction of a mild level of pulmonary micro-occlusion instead improved post-sepsis lung injury and inflammation. We also showed that the beneficial impact of this mild level of pulmonary thrombosis on post-sepsis lung injury and inflammation is dependent upon HIF1α. Furthermore, the same mild level of pulmonary thrombosis could even be induced after sepsis challenge to accelerate the resolution of lung inflammation, while induction of severe levels of pulmonary thrombosis after sepsis challenge delayed lung recovery. The experimental models used in our study enable for the quantitative control of thrombosis in the lung, while the administration of i.v. microbeads represents a unique strategy for the induction of a beneficial level of micro-occlusion to stabilize HIF1α in the lung vasculature and reduce inflammatory lung injury. In summary, our study provides evidence that the induction of a mild level of thrombotic occlusion protects against inflammatory lung injury, a finding that could help to explain the unconvincing efficacy of anti-coagulation in sepsis patients. [ABSTRACT FROM AUTHOR]