Intranasal Administration Of Lipopolysaccharide Research Articles

Microglial stimulation triggered by intranasal lipopolysaccharide administration produces antidepressant-like effect through ERK1/2-mediated BDNF synthesis in the hippocampus

We recently reported that reversing the chronic stress-induced decline of microglia in the dentate gyrus (DG) of the hippocampus by intraperitoneal injection of a low dose of lipopolysaccharide (LPS) ameliorated depression-like behavior in chronically stressed mice. In this study, we found that a single intranasal administration of LPS dose-dependently improved depression-like behavior in mice treated with chronic unpredictable stress (CUS), as evidenced by the reduction of immobility time in the tail suspension test (TST) and forced swimming test (FST) and by the increase of sucrose uptake in the sucrose preference test (SPT). The antidepressant effects of intranasal administration of LPS could be abolished by inhibition of brain-derived neurotrophic factor (BDNF) signaling by infusion of an anti-BDNF antibody, by knock-in of the mutant BDNF Val68Met allele, or by the BDNF receptor antagonist K252a. In addition, intranasal administration of LPS was found to exert antidepressant effects in a BDNF-dependent manner via promotion of BDNF synthesis mediated by extracellular signal-regulated kinase 1/2 (ERK1/2) signaling but not protein kinase B (Akt)-mammalian target of rapamycin (mTOR) signaling in DG. Inhibition of microglia by minocycline or depletion of microglia by PLX3397 was able to abolish the reversal effect of intranasal LPS administration on CUS-induced depression-like behaviors as well as the CUS-induced decrease in phospho-ERK1/2 and BDNF protein levels in DG. These results demonstrate that stimulation of hippocampal microglia by intranasal LPS administration can induce antidepressant effects via ERK1/2-dependent synthesis of BDNF protein, providing hope for the development of new strategies for the treatment of depression.

Intranasal administration of lipopolysaccharide reverses chronic stress-induced depression-like behavior in mice by microglial stimulation

We recently reported that intraperitoneal injection of a low dose of lipopolysaccharide (LPS) reversed depression-like behavior in mice induced by chronic stress by stimulating microglia in the hippocampus. In this study, we found that a single intranasal administration of LPS at a dose of 5 or 10 μg/mouse, but not at a dose of 1 μg/mouse, rapidly reversed depression-like behavior in mice stimulated with chronic unpredictable stress (CUS). In the time-dependent experiment, a single intranasal administration of LPS (10 μg/mouse) reversed CUS-induced depression-like behavior in mice 5 and 8 h but not 3 h after drug administration. The antidepressant effect of a single intranasal LPS administration (10 μg/mouse) lasted at least 10 days and disappeared 14 days after administration. Fourteen days after the first intranasal LPS administration, a second intranasal LPS administration (10 μg/mouse) still reversed the increased immobility time in TST and FST and the decreased sucrose uptake in SPT in CUS mice, which again exhibited depression-like behaviors 5 h after LPS administration. The antidepressant effect of intranasal LPS administration was dependent on microglial activation, because inhibition of microglia by pretreatment with minocycline (40 mg/kg) or depletion of microglia by pretreatment with PLX3397 (290 mg/kg) prevented the antidepressant effect of intranasal LPS administration in CUS mice. These results suggest that stimulation of the microglia-mediated innate immune response by intranasal administration of LPS can produce rapid and sustained antidepressant effects in animals under chronic stress conditions.

Human umbilical cord-derived мesenchymal stromal cells mitigate lipopolysaccharide-induced liver injury in rats

Acute Respiratory Distress Syndrome (ARDS) is a severe clinical condition that can cause multi-organ dysfunction, including liver injury. Human umbilical cord-derived mesenchymal stromal cells (hUC-MSCs) have been shown to possess therapeutic potential for a variety of diseases due to their ability to differentiate into various cell types and their anti-inflammatory and immunomodulatory properties. Purpose: To investigate the potential of hUC-MSCs for treating lipopolysaccharide (LPS)-induced liver injury in rats. Materials and methods: 72 mature male Wistar rats were randomly assigned to nine groups: control, 3 days, 7 days, and 28 days after intranasal administration of LPS, 24 hours of LPS and 2 days of hUC-MSCs, 4 days of LPS and 3 days of hUC-MSCs, 14 days of LPS and 14 days of hUC-MSCs, 21 days of LPS and 7 days of hUC-MSCs injection, and control 3 days after hUC-MSCs injection. The isolation of MSCs from human umbilical cord tissue was performed using an enzymatic digestion method with collagenase I. hUC-MSCs were injected intraperitoneally at a dose of 1∙106 cells/kg body weight. Serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP) were measured using the kinetic method. The levels of hepatocellular necrosis, liver structural damage, hepatocyte vacuolation, inflammation and disseminated intravascular coagulation (DIC) were analyzed by histological scoring of sections stained with hematoxylin and eosin. The expression of TGF-β1 in the liver tissue was evaluated by immunohistochemistry. Results: The preclinical study demonstrated that treatment with hUC-MSCs significantly improved liver function and attenuated LPS-induced liver injury in rats. This was evidenced by a reduction in hepatocellular necrosis, liver structural damage, hepatocyte vacuolation, inflammation, signs of DIC, fibrosis and lower levels of serum liver markers ALT, AST and ALP in the hUC-MSCs-treated groups compared to the untreated groups. The study also revealed that the use of hUC-MSCs was more effective at the earlier stage of liver injury. Conclusions: Our findings suggest that hUC-MSCs therapy may hold promise as a potential treatment for LPS-induced liver injury. Further research is needed to better understand the underlying mechanisms and to determine the potential for hUC-MSCs therapy in clinical practice.

Periplaneta Americana Extract Ameliorates LPS-induced Acute Lung Injury Via Reducing Inflammation and Oxidative Stress.

Acute lung injury (ALI) is an acute clinical syndrome characterized by uncontrolled inflammation response, which causes high mortality and poor prognosis. The present study determined the protective effect and underlying mechanism of Periplaneta americana extract (PAE) against lipopolysaccharide (LPS)-induced ALI. The viability of MH-S cells was measured by MTT. ALI was induced in BALB/c mice by intranasal administration of LPS (5 mg/kg), and the pathological changes, oxidative stress, myeloperoxidase activity, lactate dehydrogenase activity, inflammatory cytokine expression, edema formation, and signal pathway activation in lung tissues and bronchoalveolar lavage fluid (BALF) were examined by H&E staining, MDA, SOD and CAT assays, MPO assay, ELISA, wet/dry analysis, immunofluorescence staining and Western blotting, respectively. The results revealed that PAE obviously inhibited the release of proinflammatory TNF-α, IL-6 and IL-1β by suppressing the activation of MAPK/Akt/NF-κB signaling pathways in LPS-treated MH-S cells. Furthermore, PAE suppressed the neutrophil infiltration, permeability increase, pathological changes, cellular damage and death, pro-inflammatory cytokines expression, and oxidative stress upregulation, which was associated with its blockage of the MAPK/Akt/NF-κB pathway in lung tissues of ALI mice. PAE may serve as a potential agent for ALI treatment due to its anti-inflammatory and anti-oxidative properties, which correlate to the blockage of the MAPK/NF-κB and AKT signaling pathways.

Anti-inflammatory effect of Gyeji-tang in a chronic obstructive pulmonary disease mouse model induced by cigarette smoke and lipopolysaccharide

Context Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory lung disease associated with respiratory symptoms and narrowing of airways. Gyeji-tang (GJT) is a traditional Asian medicine that has been used to relieve early-stage cold symptoms, headache, and chills. Objective We examined the effect and potential molecular action mechanism of GJT in a mouse model of COPD induced by cigarette smoke (CS) plus lipopolysaccharide (LPS). Materials and methods COPD was induced in C57BL/6J mice via daily exposure to CS for 1 h for 8 weeks and intranasal administration of LPS on weeks 1, 3, 5, and 7. GJT (100 or 200 mg/kg) or roflumilast (5 mg/kg) was administrated daily for the final 4 weeks of COPD induction. Results Administration of GJT significantly suppressed the CS/LPS-induced increases in: the numbers of total cells and macrophages in bronchoalveolar lavage fluid; the expression levels of tumour necrosis factor-α, interleukin (IL)-6, IL-1β, and IL-8; the activities (phosphorylation) of nuclear factor kappa B and signal transducer and activator of transcription 3; and the expression levels of the structural remodelling markers, transforming growth factor beta, matrix metallopeptidase (MMP)-7, and MMP-9. Discussion and conclusions These results demonstrate that GJT prevents the lung inflammation and airway remodelling induced by CS plus LPS exposure in mice, suggesting that GJT may have therapeutic potential for the treatment of COPD.

Infiltration of peripheral immune cells into the olfactory bulb in a mouse model of acute nasal inflammation

Chronic nasal inflammation induces robust olfactory bulb (OB) atrophy in mice. Here we examined initial events that occur in the OB after bilateral intranasal administration of lipopolysaccharide, focusing on the olfactory nerve fibers and meninges. We analyzed the time course of OB and meninges inflammation using histological and biochemical approaches. Within 12 h, we observed increased chemokine expression and transient infiltration of peripheral immune cells into the OB, resulting in the development of pro-inflammatory status in the OB. Meningeal immunity was activated. Resident microglia produced anti-inflammatory cytokines within 24 h. These could be the initial events that lead to OB atrophy.

PEP-sNASP Peptide Alleviates LPS-Induced Acute Lung Injury Through the TLR4/TRAF6 Axis.

Acute lung injury (ALI) is a severe inflammatory lung disease associated with macrophages. Somatic nuclear autoantigenic sperm protein (sNASP) is a negative regulator of Toll-like receptor (TLR) signaling that targets tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) in macrophages, which is required to maintain homeostasis of the innate immune response. In the present study, we generated a cell permeable PEP-sNASP peptide using the sNASP protein N-terminal domain, and examined its potential therapeutic effect in a mouse model of ALI induced by the intranasal administration of lipopolysaccharide (LPS) and elucidated the underlying molecular mechanisms in RAW 264.7 cells. In vivo, PEP-sNASP peptide treatment markedly ameliorated pathological injury, reduced the wet/dry (W/D) weight ratio of the lungs and the production of proinflammatory cytokines (interleukin (IL)-1β, IL-6, and TNF-α). In vitro, we demonstrated that when the PEP-sNASP peptide was transduced into RAW 264.7 cells, it bound to TRAF6, which markedly decreased LPS-induced proinflammatory cytokines by inhibiting TRAF6 autoubiquitination, nuclear factor (NF)-κB activation, reactive oxygen species (ROS) and cellular nitric oxide (NO) levels. Furthermore, the PEP-sNASP peptide also inhibited NLR family pyrin domain containing 3 (NLRP3) inflammasome activation. Our results therefore suggest that the PEP-sNASP may provide a potential protein therapy against oxidative stress and pulmonary inflammation via selective TRAF6 signaling.

Pathological consequences of chronic olfactory inflammation on neurite morphology of olfactory bulb projection neurons

Chronic olfactory inflammation (COI) in conditions such as chronic rhinosinusitis significantly impairs the functional and anatomical components of the olfactory system. COI induced by intranasal administration of lipopolysaccharide (LPS) results in atrophy, gliosis, and pro-inflammatory cytokine production in the olfactory bulb (OB). Although chronic rhinosinusitis patients have smaller OBs, the consequences of olfactory inflammation on OB neurons are largely unknown. In this study, we investigated the neurological consequences of COI on OB projection neurons, mitral cells (MCs) and tufted cells (TCs). To induce COI, we performed unilateral intranasal administration of LPS to mice for 4 and 10 weeks. Effects of COI on the OB were examined using RNA-sequencing approaches and immunohistochemical analyses. We found that repeated LPS administration upregulated immune-related biological pathways in the OB after 4 weeks. We also determined that the length of TC lateral dendrites in the OB significantly decreased after 10 weeks of COI. The axon initial segment of TCs decreased in number and in length after 10 weeks of COI. The lateral dendrites and axon initial segments of MCs, however, were largely unaffected. In addition, dendritic arborization and AIS reconstruction both took place following a 10-week recovery period. Our findings suggest that olfactory inflammation specifically affects TCs and their integrated circuitry, whereas MCs are potentially protected from this condition. This data demonstrates unique characteristics of the OBs ability to undergo neuroplastic changes in response to stress.

Compound K ameliorates airway inflammation and mucus secretion through the regulation of PKC signaling in vitro and in vivo

BackgroundCigarette smoke (CS) is considered a principal cause of chronic obstructive pulmonary disease (COPD) and is associated with mucus hypersecretion and airway inflammation. Ginsenoside compound K (CK), a product of ginsenoside metabolism, has various biological activities. Studies on the effects of CK for the treatment of COPD and mucus hypersecretion, including the underlying signaling mechanism, have not yet been conducted. MethodsTo study the protective effects and molecular mechanism of CK, phorbol 12-myristate 13-acetate (PMA)-induced human airway epithelial (NCI–H292) cells were used as a cellular model of airway inflammation. An experimental mouse COPD model was also established via CS inhalation and intranasal administration of lipopolysaccharide. Mucin 5AC (MUC5AC), monocyte chemoattractant protein-1, tumor necrosis factor-α (TNF-α), and interleukin-6 secretion, as well as elastase activity and reactive oxygen species production, were determined through enzyme-linked immunosorbent assay. Inflammatory cell influx and mucus secretion in mouse lung tissues were estimated using hematoxylin and eosin and periodic acid–schiff staining, respectively. PKCδ and its downstream signaling molecules were analyzed via western blotting. ResultsCK prevented the secretion of MUC5AC and TNF-α in PMA-stimulated NCI–H292 cells and exhibited a protective effect in COPD mice via the suppression of inflammatory mediators and mucus secretion. These effects were accompanied by an inactivation of PKCδ and related signaling in vitro and in vivo. ConclusionCK suppressed pulmonary inflammation and mucus secretion in COPD mouse model through PKC regulation, highlighting the compound's potential as a useful adjuvant in the prevention and treatment of COPD.

Nasal Administration of Lipopolysaccharide Exacerbates Allergic Rhinitis through Th2 Cytokine Production from Mast Cells

Background: Microbial infection or exposure to endotoxin later in life exacerbates established asthma. Mast cells are involved in the exacerbation of asthma. This exacerbation involves a toll-like receptor (TLR)–mediated response of mast cells. In the clinical practice of otolaryngology, otolaryngologists experience an exacerbation of nasal congestion when infectious rhinitis develops in patients with allergic rhinitis, but the mechanisms are unknown. Therefore, this study investigated the effect of lipopolysaccharide (LPS) on allergic rhinitis using a mouse allergic rhinitis model. Methods: Female BALB/c mice, TLR4 gene mutant C3H/HeJ mice or mast cell–deficient WBB6F1-W/Wv mice were sensitized intraperitoneally with ovalbumin (OVA)/alum, and were intranasal challenged with OVA and/or LPS. Nasal symptoms and histologic changes were examined. Cytokines in nasal tissue were examined by Western blot. The effects of LPS on degranulation and cytokine production of bone marrow–derived mast cells (BMMCs) were investigated. Results: Nasal administration of LPS together with the antigen exacerbated nasal symptoms, eosinophil infiltration of the nasal mucosa, and increased IL-5 production in the nasal mucosa. It was not observed in C3H/HeJ mice and WBB6F1-W/Wv mice. The addition of LPS increased the production of IL-5 from BMMCs in a dose-dependent manner, but no effect on degranulation was observed. Conclusions: Intranasal administration of LPS exacerbates allergic rhinitis through Th2 cytokine production from mast cells. This observation provides clues to the mechanism of exacerbation of allergic rhinitis caused by an infection in daily clinical practice.

Neuroinflammatory penumbra in Parkinson’s disease

To date, the etiology and pathogenesis of Parkinson’s disease (PD) are not fully understood. In this publication, we present a critical review of the available international data on the involvement of inflammation processes in the etiopathogenesis of the disease in the context of our research results. The data obtained by us during modeling of parkinsonian syndrome of neuroinflammatory origin (lipopolysaccharide endotoxin was used) in laboratory rats (basic group, n = 21; control group, n = 7) indicate that intranasal administration of lipopolysaccharide in all three studied doses (1, 10 and 100 μg/kg/ml) seven days after the end of the experiment (21 injections) causes the same moderate morphological degenerative and inflammatory changes in the histostructure of the substantia nigra, extending to the striatum area (p > 0.05). This fact can be best explained by the all-or-none law. The staged, sequential transition of primary reversible (neuroinflammatory) pathomorphological changes to secondary irreversible (neurodegenerative) ones in PD, as well as secondary activation of microglia during neuronal degeneration, allows proposing the term “neuroinflammatory penumbra”, which will best help understand the pathogenesis of the disease and the development of therapies that change the course of Parkinson’s disease. Our data on the use of mesenchymal multipotent stromal cells in rats with neurotoxic (rotenone) parkinsonian syndrome (n = 10) and in PD individuals (n = 10) demonstrate a decrease in motor impairment, overall improvement of patients and an effect on laboratory parameters compared with control groups. We associate the early positive effect, observed already in the first week after the administration of mesenchymal multipotent stromal cells, with their paracrine action, including on the neuroinflammatory penumbra.

Capsaicin Protects Against Lipopolysaccharide-Induced Acute Lung Injury Through the HMGB1/NF-κB and PI3K/AKT/mTOR Pathways.

PurposeCapsaicin (8-methyl-N-geranyl-6-nonamide; CAP) is an alkaloid isolated from chili peppers, which has complex pharmacological properties, including beneficial effects against various diseases. The aim of this study was to investigate the role of CAP in lipopolysaccharide (LPS)-induced acute lung injury (ALI), and the possible underlying mechanisms.Materials and MethodsALI was induced by intranasal administration of LPS (0.5 mg/kg), and CAP (1 mg/kg) injected intraperitoneally 3 days before exposure to LPS. Then, the histopathological changes were evaluated by hematoxylin and eosin staining. Enzyme-linked immunosorbent assay and qPCR were used to detect pro-inflammatory cytokines in serum and lung tissue. The expressions of HMGB1/NF-κB, PI3K/AKT/mTOR signaling pathways and apoptosis-associated molecules were determined by Western blot and/or qPCR. In addition, the lung cell apoptosis was analyzed by TUNEL staining, and the expression and location of cleaved caspase-3 were detected by immunofluorescence analysis.ResultsCAP pretreatment significantly protected mice from LPS-induced ALI, with reduced lung wet/dry weight ratio, lung histological damage, myeloperoxidase (MPO) activity, malondialdehyde (MDA) content and pro-inflammatory cytokine levels, and significant increased superoxide dismutase (SOD) activity. In addition, CAP pretreatment significantly inhibited the high-mobility group protein B1 (HMGB1) expression, nuclear factor-kappa B (NF-κB) activation, and the PI3K/AKT/mTOR signaling pathway. Furthermore, mice pre-treated with CAP exhibited reduced apoptosis of lung tissues, with associated down-regulation of caspase-3, cleaved caspase-3, and BAX expression, and up-regulation of BCL-2.ConclusionOur data demonstrate that CAP can protect against LPS-induced ALI by inhibiting oxidative stress, inflammatory responses and apoptosis through down-regulation of the HMGB1/NF-κB and PI3K/AKT/mTOR pathways.

Intranasal lipopolysaccharide administration prevents chronic stress-induced depression- and anxiety-like behaviors in mice

We recently reported that intraperitoneal injection of a low dose of lipopolysaccharide (LPS) prevents chronic stress-induced depression-like behaviors in mice. In this study, we reported that a single intranasal LPS administration (10 μg/mouse) one day prior to stress exposure produced prophylactic effects on chronic social defeat stress (CSDS)-induced depression-like behaviors, which was indicated by the reduction in social interaction time in the social interaction test and the decrease in immobility time in the tail suspension test and forced swimming test. The single intranasal LPS administration prior to stress exposure was also found to prevent CSDS-induced anxiety-like behaviors, including prevention of CSDS-induced decrease in the time spent in open arms in the elevated plus maze test, decrease in the time spent in lit side in the light-dark test, and decrease in the time spent in central regions in the open field test, along with no changes in locomotor activity. Further analysis showed that the single intranasal LPS administration one day prior to stress exposure prevented CSDS-induced increase in levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and IL-1β mRNA in the hippocampus and prefrontal cortex. Inhibition of innate immune stimulation by minocycline pretreatment not only abrogated the preventive effect of intranasal LPS administration on CSDS-induced depression- and anxiety-like behaviors, but also abrogated the preventive effect of intranasal LPS administration on CSDS-induced neuroinflammatory responses in the hippocampus and prefrontal cortex. These results demonstrate that intranasal administration of innate immune stimulants could be a potential approach for the prevention of depression and anxiety.

Proteomic analysis of a chronic obstructive pulmonary disease mouse model to determine the efficacy of treatment using Guben Zhike decoction

Guben Zhike decoction (GBZKD) is derived from the experience of Professor Enxiang Chao, an esteemed master of Chinese medicine, while treating chronic obstructive pulmonary disease (COPD). GBZKD reinforces the healthy qi and consolidates defensive qi. This study explored the efficacy and potential mechanism of action of GBZKD in a COPD mouse model using proteomics. A COPD mouse model was established through cigarette smoke exposure and intranasal lipopolysaccharide administration. The model was verified through lung function test and lung histopathological observation. Label-free quantitative proteomics was used to detect the lung tissue proteins of mice from the GBZKD, COPD, and control groups. GBZKD markedly improved the lung function and associated pathological conditions in the COPD mouse model. Proteomic analysis identified 4316 proteins, of which 3696 were quantitative proteins. We highlighted 287 and 184 proteins with significant regulatory roles in the lung tissues of COPD mice and GBZKD-treated mice, respectively. These proteins participated in multiple functions, including complement/coagulation cascade, immune response, and metabolic pathways. GBZKD exhibits multitarget and multipathway therapeutic effects in a COPD mouse model.

Prospects for the use of oxidized dextran for the prevention of viral pneumonia and pulmonary fibrosis

Study background. The relevance of the study arises from the large number of fibrotic complications in viral interstitial pneumonia in COVID-19. Objective: to evalu-ate the therapeutic efficacy of inhaling 2% oxidised dex-tran solution in an in vivo model simulating interstitial pneumonia in COVID-19. Materials and methods. The studies were performed on ACR mice. Interstitial pneu-monia in mice was induced by intranasal administra-tion of Escherichia coli lipopolysaccharide (LPS). After intranasal administration of LPS to mice, 2% solution of oxidised dextran or control (saline) solution were inhaled via ultrasonic inhaler. One day later, the animals were killed, and standard histological sections of lung tissue were prepared with subsequent morphometric evalua-tion of the results. Results with data. inhalation of a 2% solution of oxidised dextran makes it possible to almost halve all morphological manifestations of LPS-induced pneumonia in terms of morphometric parameters: the numerical density of vessels with signs of hemocircula-tory disorders, the numerical density of thrombosed vessels, the percentage of thrombosed vessels, the volu-metric density of hemorrhages, the volumetric density atelectasis, volumetric density of cellular inflammatory infiltrate. Conclusion. Inhalation of oxidised dextran so-lutions may be a very promising method for the preven-tion of interstitial pneumonia and pulmonary fibrosis in COVID-19.

Vascular endothelial growth factor contributes to lung vascular hyperpermeability in sepsis-associated acute lung injury.

Vascular endothelial growth factor (VEGF) is a prime regulator of vascular permeability. Acute lung injury (ALI) is characterized by high-permeability pulmonary edema in addition to refractory hypoxemia and diffuse pulmonary infiltrates. In this study, we examined whether VEGF can be implicated as a pulmonary vascular permeability factor in sepsis-associated ALI. We found that a great increase in lung vascular leak occurred in mice instilled intranasally with lipopolysaccharide (LPS), as assessed by IgM levels in bronchoalveolar lavage fluid. Treatment with the VEGF-neutralizing monoclonal antibody bevacizumab significantly reduced this hyperpermeability response, suggesting active participation of VEGF in non-cardiogenic lung edema associated with LPS-induced ALI. However, this was not solely attributable to excessive levels of intrapulmonary VEGF. Expression levels of VEGF were significantly reduced in lung tissues from mice with both intranasal LPS administration and cecal ligation and puncture (CLP)-induced sepsis, which may stem from decreases in non-endothelial cells-dependent VEGF production in the lungs. In support of this assumption, stimulation with LPS and interferon-γ (IFN-γ) significantly increased VEGF in human pulmonary microvascular endothelial cells (HPMECs) at mRNA and protein levels. Furthermore, a significant rise in plasma VEGF levels was observed in CLP-induced septic mice. The increase in VEGF released from HPMECs after LPS/IFN-γ challenge was completely blocked by either specific inhibitor of mitogen-activated protein kinase (MAPK) subgroups. Taken together, our results indicate that VEGF can contribute to the development of non-cardiogenic lung edema in sepsis-associated ALI due to increased VEGF secretion from pulmonary vascular endothelial cells through multiple MAPK-dependent pathways.

Ficolin A derived from local macrophages and neutrophils protects against lipopolysaccharide-induced acute lung injury by activating complement.

Ficolins are important and widely distributed pattern recognition molecules that can induce lectin complement pathway activation and initiate the innate immune response. Although ficolins can bind lipopolysaccharide (LPS) in vitro, the sources, dynamic changes and roles of local ficolins in LPS-induced pulmonary inflammation and injury remain poorly understood. In this study, we established a ficolin knockout mouse model by clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) technology, and used flow cytometry and hematoxylin and eosin staining to study the expressions and roles of local ficolins in LPS-induced pulmonary inflammation and injury. Our results show that besides ficolin B (FcnB), ficolin A (FcnA) is also expressed in leukocytes from the bone marrow, peripheral blood, lung and spleen. Further analyses showed that macrophages and neutrophils are the main sources of FcnA and FcnB, and T and B cells also express a small amount of FcnB. The intranasal administration of LPS induced local pulmonary inflammation with the increased recruitment of macrophages and neutrophils. LPS stimulation induced increased expression of FcnA and FcnB in neutrophils at the acute stage and in macrophages at the late stage. The severity of the lung injury and local inflammation of Fcna-/- mice was increased by the induction of extracellular complement activation. The recovery of LPS-induced local lung inflammation and injury was delayed in Fcnb-/- mice. Hence, these findings suggested that the local macrophage- and neutrophil-derived FcnA protects against LPS-induced acute lung injury by mediating extracellular complement activation.

Ficolin A Derived from Local Macrophages and Neutrophils Protects Against LPS-Induced Acute Lung Injury by Activating Complement

Background: Ficolins are important and widely distributed secreted pattern recognition molecules that can induce lectin complement pathway activation and initiate the innate immune response. Although ficolins can bind lipopolysaccharide (LPS) in vitro, the sources, dynamic changes and roles of ficolins in LPS-induced local pulmonary inflammation and injury remain poorly understood. Methods: C57BL/6 and Ficolins-KO mice were intranasally administrated 10 μg LPS to induce acute lung inflammation and injury. Flow cytometry and immunofluorescence were performed to reveal the expression of ficolins. Lung injury was analyzed by hematoxylin & eosin staining. Complement activation was confirmed by immunohistochemistry and sandwich ELISA. Findings: In this study, we found that ficolin A and ficolin B are expressed in leukocytes from the bone marrow, peripheral blood, lung and spleen. Further analyses showed that macrophages and neutrophils are the main sources of ficolin A and ficolin B, and T and B cells also express a small amount of ficolin B. The intranasal administration of LPS induced local pulmonary inflammation with the increased recruitment of macrophages and neutrophils. LPS stimulation induced increased expression of ficolin A and ficolin B in neutrophils at the acute stage, and in macrophages at the late stage. The severity of the lung injury and local inflammation of Fcna−/− mice was increased via the induction of extracellular complement activation. The recovery of LPS-induced local lung inflammation and injury was delayed in Fcnb−/− mice. Interpretation: The local macrophage- and neutrophil-derived ficolin A protects against LPS-induced acute lung injury by mediating extracellular complement activation. These findings suggested that ficolins might be novel therapeutic targets for gram-negative bacteria infection induced pneumonia. Funding Statement: This work was supported by National Science Grant for Distinguished Young Scholars (81425001/H0104), National Natural Science Foundation of China, China (81373114), Beijing Municipal Natural Science Foundation, China (7182013), CAMS Innovation Fund for Medical Sciences, China (CIFMS 2018-I2M-1-003, CIFMS 2016-2M-1-014, 2016-12M-006), the Chinese National Major S & T Project (2017ZX10304402-001). Declaration of Interests: All authors declare no conflicts of interest associated with this manuscript. Ethics Approval Statement: All experiments were conducted with the approval of the Institutional Animal Care and Use Committee (IACUC) of Capital Medical University.

Antithrombin plus alpha-1 protease inhibitor does not affect coagulation and inflammation in two murine models of acute lung injury

BackgroundIn acute respiratory distress syndrome (ARDS), uncontrolled production of activators of coagulation and proinflammatory mediators results in a shift from an adequate local innate immune response to hypercoagulability and inflammation. This study aimed to investigate whether the protease inhibitors antithrombin (AT) and alpha-1 protease inhibitor (A1PI) may attenuate an exaggerated pulmonary immune response.MethodsLung injury was induced either by single intranasal administration of lipopolysaccharide (LPS) (5 mg/kg) in BALB/c mice or by combination of an intravenous injection of LPS (10 mg/kg) with subsequent injurious ventilation using high tidal volumes (12–15 ml/kg) for 4 h in RccHan Wistar rats. Animals received either a single bolus of AT (250 IU/kg) or A1PI (60 mg/kg) alone or in combination, with or without intravenous low-dose heparin (100 U/kg). Control animals received saline. Additional controls received neither LPS, nor ventilation, nor treatment. Endpoints were local and systemic markers of coagulation, e.g., thrombin–antithrombin complexes (TATc), and inflammation, e.g., interleukin-6.ResultsBoth lung injury models resulted in a pronounced immune response within the pulmonary compartment shown by elevated levels of markers of coagulation and inflammation. The two-hit lung injury model also induced profound systemic coagulopathy and inflammation. Monotherapy with AT or A1PI did not reduce pulmonary coagulopathy or inflammation in any lung injury model. Nor did combination therapy with AT and A1PI result in a decrease of coagulation or inflammatory parameters. AT markedly reduced systemic levels of TATc in the two-hit lung injury model. Systemic inflammation was not affected by the different interventions. Additional administration of heparin did not lead to macroscopic bleeding incidences.ConclusionsIn two different murine models of acute lung injury, neither single therapy with AT or A1PI nor combination of both agents attenuates the pronounced pulmonary coagulation or inflammatory response.

Recombinant chemotaxis inhibitory protein of Staphylococcus aureus (CHIPS) protects against LPS-induced lung injury in mice

Acute lung injury (ALI) and/or acute respiratory distress syndrome (ARDS) are clinical conditions caused by trauma, lung infection or sepsis. ALI/ARDS is associated with massive recruitment of neutrophils into the lung with release of reactive oxygen species and excessive inflammatory response that damage alveolar tissue. Here we report the successful use of a potent recombinant chemotaxis inhibitory protein (rCHIPS) derived from Staphylococcus aureus in reducing the severity of ALI/ARDS. Treatment with rCHIPS reduces pulmonary inflammation and permeability in mice after intranasal administration of lipopolysaccharide (LPS). rCHIPS treatment significantly reduces lung myeloperoxidase (MPO) activity, pro-inflammatory cytokines, broncho-alveolar lavage (BAL) fluid protein content as well as histopathological changes. In addition, treatment with rCHIPS significantly diminishes neutrophils and leukocytes recruitment into lung tissue after LPS administration and hence protects mice from reactive oxygen species mediated lung injury. Our finding reveals potential therapeutic benefits of using rCHIPS for the treatment of ALI/ARDS.