Protein Concentration In Lavage Fluid Research Articles

Notoginsenoside R1 protects against hypobaric hypoxia-induced high-altitude pulmonary edema by inhibiting apoptosis via ERK1/2-P90rsk-BAD ignaling pathway

High-altitude pulmonary edema (HAPE) is a potentially fatal disease. Notoginsenoside R1 is a novel phytoestrogen with anti-inflammatory, antioxidant and anti-apoptosis properties. However, its effects and underlying mechanisms in the protection of hypobaric hypoxia-induced HAPE rats remains unclear. This study aimed to explore the protective effects and underlying mechanisms of Notoginsenoside R1 in hypobaric hypoxia-induced HAPE. We found that Notoginsenoside R1 alleviated the lung tissue injury, decreased lung wet/dry ratio, and reduced inflammation and oxidative stress. Additionally, Notoginsenoside R1 ameliorated the changes in arterial blood gas, decreased the total protein concentration in bronchoalveolar lavage fluid, and inhibited the occurrence of apoptosis caused by HAPE. In the process of further exploration of the mechanism, it was found that Notoginsenoside R1 could promote the activation of ERK1/2-P90rsk-BAD signaling pathway, and the effect of Notoginsenoside R1 was attenuated after the use of ERK1/2 inhibitor U0126. Our study indicated that the protective effects of Notoginsenoside R1 against HAPE were mainly related to the inhibition of inflammation, oxidative stress, and apoptosis. Notoginsenoside R1 may be a potential candidate for preventing HAPE.

Tamibarotene targets heparin-binding protein for attenuating lung injury in sepsis.

Excessively active pulmonary inflammation is a hallmark of sepsis-induced lung damage. A synthetic retinoid drug called tamibarotene reduces inflammation in a variety of conditions, including acute promyelocytic leukemia (APL), renal fibrosis, and neuroinflammation. Its effect on sepsis-related lung injury, however, has not been explained. The purpose of the study was to investigate how tamibarotene affected lung damage induced by cecal ligation and puncture (CLP) procedure. A CLP sepsis mouse model was developed, and tamibarotene was pretreated to determine whether it improved lung injury and survival. The degree of lung injury was evaluated using the Hematoxylin and eosin staining and lung injury score. In order to determine pulmonary vascular permeability, measurements were taken for total protein and cell content of bronchoalveolar lavage fluid (BALF), wet/dry ratio of the lung, and Evans blue stain. The BALF inflammatory mediators, including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-1β, and IL-17A were discovered by enzyme-linked immunosorbent serologic assay (ELISA). Then, the levels of heparin-binding protein (HBP), and phospho-nuclear factor kappa-B (p-NF-κB) P65, and NF-κB P65 were determined using ELISA and Western blot analysis, respectively. Tamibarotene considerably increases survival and lessens lung damage stimulated by sepsis. Specifically, tamibarotene significantly relieves pulmonary vascular permeability and inhibits inflammation response in sepsis. Moreover, we further confirmed that these ameliorating effects of tamibarotene on sepsis may be exerted by targeting HBP and regulating the activation of NF-κB signaling pathway. These findings demonstrated that tamibarotene lessens sepsis-induced lung injury, and the effect could be exerted by targeting HBP and thereby deregulating the NF-κB signaling pathway.

A study of the role of myeloid-derived growth factor on acute lung injury in vitro and In vivo

Objectives: Bone marrow-derived mesenchymal stem cells (BMSCs) are considered to have potential clinical application value in the treatment of acute lung injury (ALI). Myeloid-derived growth factor (MYDGF) can promote the proliferation of stem cell. We hypothesized that MYDGF may play a role in reducing lung injury in vitro and in vivo through bone marrow mesenchymal stem cells. Methods: An in vitro model of lipopolysaccharide (LPS)(MLE-12) was established, which was divided into five groups: A: MLE-12; B: MLE-12+LPS; C: MLE-12+LPS + BMSCs; D: MLE-12+LPS + MYDGF; and E: MLE-12+LPS + BMSCs + MYDGF. A Cell Counting Kit-8 was used to detect the OD value. And an ALI model was constructed by inducing mice with a lipopolysaccharide. Forty male Balb/c mice were randomly divided into five groups: A control group; B: model group; C: LPS + BMSCs; D: LPS + MYDGF; E: LPS +BMSCs +MYDGF. Specimens were collected after 24 h. Hematoxylin-eosin (HE)-staining was performed on the tissue sections. The protein concentration in the alveolar lavage fluid was measure by bicinchoninic acid (BCA). The NF-κB, p-Akt, Bax, and Bcl-2 protein expression was detected through Western blotting, and Enzyme linked immunosorbent assay (ELISA) was used to measure the expression of serum interleukin-6, interleukin-10, and TNF-α. Results: Compared with the model group, BMSCs and MYDGF can alleviate the ALI induced by lipopolysaccharide in vitro and vivo ( p < .05). Conclusion: We found that the combined treatment effect of MYDGF and BMSCs was better than using MYDGF or BMSCs alone. We speculate that a pretreatment with MYDGF after ALI in mice may improve the survival and growth of transplanted MSCs, thereby improving the curative effect of cell transplantation.

LncRNA CDKN2B-AS1 interacts with LIN28B to exacerbate sepsis-induced acute lung injury by inducing HIF-1α/NLRP3-mediated pyroptosis.

Sepsis-associated acute lung injury (ALI) is a life-threatening condition in intensive care units with high mortality. LncRNAs have been confirmed to participate in the underlying pathogenesis of septic ALI. This study investigated the biological functions of lncRNA CDKN2B-AS1 in septic ALI and its potential mechanism.BEAS-2B cells were challenged with lipopolysaccharide (LPS) and mice were subjected to caecal ligation and puncture (CLP) to induce septic ALI in vitro and in vivo. The expression levels of CDKN2B-AS1, LIN28B, HIF-1α, and pyroptosis-related molecules were assessed by qRT-PCR or Western blotting. The production of IL-1β and IL-18 was detected by ELISA. BEAS-2B cell pyroptosis was examined by flow cytometry. The interaction between LIN28B and CDKN2B-AS1/HIF-1α was validated by RIP and RNA pull-down assays. Colocalization of CDKN2B-AS1 and LIN28B was observed by FISH. ALI was determined by HE staining, the lung wet-to-dry (W/D) weight ratio, inflammatory cell numbers, and total protein concentration in bronchoalveolar lavage fluid (BALF). Caspase-1 expression in the lung tissues was examined by immunohistochemical staining.CDKN2B-AS1 was upregulated in BEAS-2B cells after LPS stimulation. CDKN2B-AS1 knockdown inhibited pyroptosis in LPS-exposed BEAS-2B cells in vitro and the lung tissues of septic mice in vivo. Mechanistically, CDKN2B-AS1 interacted with LIN28B to enhance HIF-1α stability. Rescue experiments showed that HIF-1α overexpression counteracted the inhibitory effect of sh-CDKN2B-AS1 on LPS-induced pyroptosis. CDKN2B-AS1 bound to LIN28B to trigger NLRP3-mediated pyroptosis by stabilizing HIF-1α, which promoted sepsis-induced ALI. CDKN2B-AS1 might be a novel therapeutic target for this disease.

Inhibition of the lectin pathway of complement activation reduces LPS-induced acute respiratory distress syndrome in mice.

Acute respiratory distress syndrome (ARDS) is a life-threatening disorder with a high rate of mortality. Complement activation in ARDS initiates a robust inflammatory reaction that can cause progressive endothelial injury in the lung. Here, we tested whether inhibition of the lectin pathway of complement could reduce the pathology and improve the outcomes in a murine model of LPS-induced lung injury that closely mimics ARDS in human. In vitro, LPS binds to murine and human collectin 11, human MBL and murine MBL-A, but not to C1q, the recognition subcomponent of the classical pathway. This binding initiates deposition of the complement activation products C3b, C4b and C5b-9 on LPS via the lectin pathway. HG-4, a monoclonal antibody that targets MASP-2, a key enzyme in the lectin pathway, inhibited lectin pathway functional activity in vitro, with an IC50 of circa 10nM. Administration of HG4 (5mg/kg) in mice led to almost complete inhibition of the lectin pathway activation for 48hrs, and 50% inhibition at 60hrs post administration. Inhibition of the lectin pathway in mice prior to LPS-induced lung injury improved all pathological markers tested. HG4 reduces the protein concentration in bronchoalveolar lavage fluid (p<0.0001) and levels of myeloid peroxide (p<0.0001), LDH (p<0.0001), TNFα and IL6 (both p<0.0001). Lung injury was significantly reduced (p<0.001) and the survival time of the mice increased (p<0.01). From the previous findings we concluded that inhibition of the lectin pathway has the potential to prevent ARDS pathology.

HYDROMORPHONE MITIGATES CARDIOPULMONARY BYPASS-INDUCED ACUTE LUNG INJURY BY REPRESSING PYROPTOSIS OF ALVEOLAR MACROPHAGES.

Introduction: Acute lung injury (ALI) is a devastating pulmonary illness with diffuse inflammatory responses. Hydromorphone (Hyd) is an opioid agonist used for relieving moderate-to-severe pain. The present work investigated the effect of Hyd on cardiopulmonary bypass (CPB)-induced ALI by regulating pyroptosis of alveolar macrophages (AMs). Methods: Rats were subjected to CPB, followed by Hyd treatment. The lung injury in rat lung tissues was appraised by the ratio of lung wet/dry weight (weight), histological staining, and the total protein concentrations in bronchoalveolar lavage fluid, and lung function was assessed by oxygenation index and respiratory index, and lung macrophage pyroptosis was observed by fluorescence staining. Alveolar macrophages were separated and pyroptosis was determined by western blot assay and enzyme-linked immunosorbent assay. The expression patterns of nuclear factor erythroid 2-related factor 2/heme oxygenase 1 (Nrf2/HO-1), nod-like receptor protein 3 (NLRP3), N-terminal gasdermin-D, and cleaved caspase-1 were examined by real-time quantitative polymerase chain reaction, western blot, and immunohistochemistry assays. The impact of NLRP3 or Nrf2 on pyroptosis of AMs and CPB-induced ALI was observed after treatment of nigericin (NLRP3 agonist) or ML385 (Nrf2 inhibitor). Results: Hyd attenuated CPB-induced lung injury as manifested by reductions in lung inflammation and edema, the scores of lung injury, the ratio of lung wet/dry weight, and the total protein concentrations in bronchoalveolar lavage fluid. Besides, Hyd repressed NLRP3 inflammasome-mediated pyroptosis of AMs after CPB treatment. Hyd upregulated Nrf2/HO-1 expression levels to repress NLRP3 inflammasome-mediated pyroptosis. Treatment of nigericin or ML385 counteracted the role of Hyd in ameliorating pyroptosis of AMs and CPB-induced ALI. Conclusions: Hyd alleviated NLRP3 inflammasome-mediated pyroptosis and CPB-induced ALI via upregulating the Nrf2/HO-1 pathway, which may be achieved by AMs.

Repurposing of carvedilol to alleviate bleomycin-induced lung fibrosis in rats: Repressing of TGF-β1/α-SMA/Smad2/3 and STAT3 gene expressions

Idiopathic pulmonary fibrosis (IPF) is the most widely studied interstitial lung disease. IPF eventually leads to respiratory insufficiency, lung cancer, and death. Carvedilol (CAR) is a third-generation β-adrenergic receptor antagonist with an α1-blocking effect. CAR demonstrates antifibrotic activities in various experimental models of organ fibrosis. Aims: This work is designed to explore the possible alleviating effects of CAR on bleomycin (BLM)-induced lung fibrosis in rats. Main methods: The BLM rat model of lung fibrosis was achieved by intratracheal delivery of a single dose of 5 mg/kg of BLM. Seven days following BLM injection, either prednisolone or CAR was orally administered at doses of 10 mg/kg once daily for 21 days to the rats. The actions of CAR were evaluated by lung oxidant/antioxidant parameters, protein concentration and total leucocyte count (TLC) in bronchoalveolar lavage fluid (BALF), fibrosis regulator-related genes along with the coexistent lung histological changes. Key findings: CAR effectively decreased lung malondialdehyde level, increased superoxide dismutase activity, declined both protein concentration and TLC in BALF, downregulated TGF-β1/α-SMA/Smad2/3 and STAT3 gene expressions, and repaired the damaged lung tissues. Significance: CAR conferred therapeutic potential against BLM-induced lung fibrosis in rats, at least in part, to its antioxidant, anti-inflammatory, and antifibrotic activities. CAR could be utilized as a prospective therapeutic option in patients with lung fibrosis in clinical practice.

Deletion of LCMR1 in alveolar type II cells induces lethal impairment of lung structure and function in adult mice.

Previous studies of lung cancer metastasis-related protein 1 (LCMR1) mainly focused on its relationship with cancer. However, the function of LCMR1 in normal tissues or cells is poorly understood. We aimed to investigate the effects of alveolar type II cell (AT2 cell)-specific LCMR1 deletion on lung structure and function in adult mice. Mice carrying the floxed LCMR1 allele with exons 2-4 flanked by loxP sites were constructed and then crossed with Sftpc-CreERT2 mice to obtain Sftpc-CreERT2 ; LCMR1 flox/flox for AT2 cell-specific LCMR1 deletion and LCMR1 flox/flox mice as littermate control. We observed the body weight change, histopathology, lung wet/dry weight ratio, pulmonary function, and survival of the mice, together with the protein concentration, inflammatory cells, and cytokine levels in bronchoalveolar lavage fluid. We also detected AT2 cell numbers and expression of pulmonary surfactant protein in the lung tissues. The apoptosis of AT2 cells was also assessed. We found that AT2 cell-specific LCMR1 deletion caused rapid weight loss and increased mortality in mice. Histopathological analysis revealed damaged lung structure, including inflammatory cell infiltration, alveolar hemorrhage, and edema. The lung wet/dry weight ratio was higher and bronchoalveolar lavage fluid (BALF) analysis revealed elevated protein concentration, inflammatory cell counts, and cytokine levels. Pulmonary function measurement showed increased airway resistance, decreased lung compacity, and compliance. We also found massive AT2 cell loss and altered expression of pulmonary surfactant protein. Deletion of LCMR1 promoted apoptosis in AT2 cells. We successfully generated an AT2 cell-specific LCMR1 conditional knockout mouse model and further revealed the crucial role of LCMR1 in maintaining AT2 cell homeostasis.

Overexpression of FoxM1 optimizes the therapeutic effect of bone marrow mesenchymal stem cells on acute respiratory distress syndrome

BackgroundInjury of alveolar epithelial cells and capillary endothelial cells is crucial in the pathogenesis of acute lung injury/acute respiratory distress syndrome (ALI/ARDS). Mesenchymal stem cells (MSCs) are a promising cell source for ALI/ARDS treatment. Overexpression of Fork head box protein M1 (FoxM1) facilitates MSC differentiation into alveolar type II (AT II) cells in vitro. Moreover, FoxM1 has been shown to repair the endothelial barrier. Therefore, this study explored whether overexpression of FoxM1 promotes the therapeutic effect of bone marrow-derived MSCs (BMSCs) on ARDS by differentiation of BMSCs into AT II cells or a paracrine mechanism.MethodsA septic ALI model was established in mice by intraperitoneal administration of lipopolysaccharide. The protective effect of BMSCs-FoxM1 on ALI was explored by detecting pathological variations in the lung, total protein concentration in bronchoalveolar lavage fluid (BALF), wet/dry (W/D) lung weight ratio, oxidative stress levels, cytokine levels, and retention of BMSCs in the lung. In addition, we assessed whether FoxM1 overexpression promoted the therapeutic effect of BMSCs on ALI/ARDS by differentiating into AT II cells using SPC−/− mice. Furthermore, the protective effect of BMSCs-FoxM1 on lipopolysaccharide-induced endothelial cell (EC) injury was explored by detecting EC proliferation, apoptosis, scratch wounds, tube formation, permeability, and oxidative stress, and analyzing whether the Wnt/β-catenin pathway contributes to the regulatory mechanism in vitro using a pathway inhibitor.ResultsCompared with BMSCs-Vector, treatment with BMSCs-FoxM1 significantly decreased the W/D lung weight ratio, total BALF protein level, lung injury score, oxidative stress, and cytokine levels. With the detected track of BMSCs-FoxM1, we observed a low residency rate and short duration of residency in the lung. Notably, SPC was not expressed in SPC−/− mice injected with BMSCs-FoxM1. Furthermore, BMSCs-FoxM1 enhanced EC proliferation, migration, and tube formation; inhibited EC apoptosis and inflammation; and maintained vascular integrity through activation of the Wnt/β-catenin pathway, which was partially reversed by XAV-939.ConclusionOverexpression of FoxM1 enhanced the therapeutic effect of BMSCs on ARDS, possibly through a paracrine mechanism rather than by promoting BMSC differentiation into AT II cells in vivo, and prevented LPS-induced EC barrier disruption partially through activating the Wnt/β-catenin signaling pathway in vitro.

Irigenin attenuates lipopolysaccharide-induced acute lung injury by inactivating the mitogen-activated protein kinase (MAPK) signaling pathway.

Acute lung injury (ALI) is a serious pulmonary inflammation disease with high mortality. Irigenin, an isoflavone from rhizomes of the Belamcanda chinensis, has been reported to exert anti-inflammatory, anti-oxidative, and anti-apoptotic activities in several diseases. However, it is still unclear whether irigenin can exert a beneficial effect in ALI. A network pharmacology method was utilized to predict the hub targets and potential therapeutic mechanisms of irigenin against ALI. Lipopolysaccharide (LPS) was used to establish the mice model of ALI for evaluating the effects of irigenin. According to the protein-protein interaction (PPI) network, we identified EGFR, HRAS, AKT1, SRC, and HSP90AA1 as the top five significant genes. Gene Ontology (GO) and Kyoto Encyclopaedia of Genes and Genomes (KEGG) enrichment assays showed that irigenin might affect inflammatory response, cytokine production, and cell death by the mitogen-activated protein kinase (MAPK) signaling pathway. In vivo experiment results manifested that irigenin decreased pathological changes, lung Wet/Dry weight ratio, and total protein content in bronchoalveolar lavage fluid (BALF). Irigenin also reduced the production of inflammatory cytokines, including tumor necrosis factor-a (TNF-a), interleukin-1β (IL-1β), interleukin-6 (IL-6), and interleukin-18 (IL-18), and neutrophil infiltration. Additionally, irigenin inhibited pulmonary apoptosis in LPS-treated ALI mice. Moreover, LPS-induced phosphorylation of p38, JNK, and ERK was significantly abated due to the treatment of irigenin. In summary, irigenin ameliorates LPS-induced ALI by suppressing pulmonary inflammation and apoptosis via inactivation of the MAPK signaling pathway. These findings indicated the therapeutic potential of irigenin in ALI.

Vitexin attenuates smoke inhalation induced acute lung injury in rats by inhibiting oxidative stress via PKC β/p66Shc signaling pathway

Purpose: To investigate the protective effect of vitexin on smoke inhalation-induced acute lung injury (SI-ALI), and the underlying mechanism of action.Methods: The ALI rat model was established by inhalation of smoke in a closed smoke chamber. Survival rate, arterial blood gas analysis, wet-to-dry weight ratio of lung tissues, bronchoalveolar lavage fluid protein concentration, lung tissue histology, and oxidative stress and inflammation level were evaluated. Expressions of protein kinase C β (PKC β), p66Shc, and phosphorylated p66Shc were determined by western blot or quantitative reverse transcription-polymerase chain reaction.Results: Compared with smoke inhalation group, vitexin alleviated the decline in arterial partial pressure of oxygen (p &lt; 0.05), reduced lung tissue exudation and pathological lung tissue damage, inhibited the expression of PKC β/p66Shc signaling pathway proteins, downregulated the level of oxidative stress and inflammation, and ultimately improved the survival rate in SI-ALI rats (p &lt; 0.05).Conclusion: Vitexin attenuates SI-ALI in rats by alleviating oxidative stress via inhibition of PKC β/p66Shc signaling pathway. Thus, this compound is a potential agent for the treatment of SI-ALI.

Effects of electroacupuncture pretreatment on NLRP3 inflammasome in mice with ventilator-induced lung injury

To observe the effect of electroacupuncture (EA) pretreatment on inflammatory response in ven-tilator-induced lung injury (VILI) mice, so as to explore the underlying mechanism of EA pretreatment on prevention of VILI. C57BL/6 mice were randomly divided into sham-operation group, model group, EA group and sham-acupoint group，with 8 mice in each group. The VILI model was established by ventilation with high tidal volume. Mice in the EA group and sham-acupoint group were given EA at "Zusanli" (ST36)and "Feishu"(BL13) or non-acupoints (located at 1-2 cm on both sides of the tail root of the proximal trunk) before mechanical ventilation, 30 min each time, once a day for 5 days. Arterial blood was collec-ted for blood gas analysis, the total protein content in bronchoalveolar lavage fluid (BALF) was detected by BCA method. The contents of interleukin-1β (IL-1β) and interleukin-18 (IL-18) in BALF were detected by ELISA. Lung injury score was determined after HE staining. The protein expression levels of nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3), apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC) and Caspase-1 in lung tissue was detected by Western blot. Compared with the sham-operation group, the arterial partial pressure of oxygen and oxygenation index were decreased(P<0.05), the levels of total protein, IL-1β and IL-18 in BALF, the W/D value and the pathological injury score of lung tissue and the protein expression levels of NLRP3, Caspase-1 and ASC were increased(P<0.05)in the model group. Following the interventions, the above mentioned increased or decreased indicators were reversed(P<0.05) in the EA group rather than in the sham-acupoint group. EA pretreatment of ST36 and BL13 can reduce the damage of lung tissue caused by mechanical ventilation, which may be related to its effect in reducing the expression of NLPR3 inflammasome related proteins, reducing the activation of inflammasome, and thereby reducing the inflammatory response.

Anti-Inflammatory Effect of Dendrobium Huoshanense Polysaccharides on Carrageenan-Induced Air Pouch Synovitis in Mice

Objective: The purpose was to observe the anti-inflammatory effects of Dendrobium huoshanense polysaccharides (DHPs) on carrageenan-induced sterile air pouch synovitis. Materials and Methods: A total of 30 Institute of Cancer Research (ICR) mice were randomly and equally assigned to the control, carrageenan-induced air pouch synovitis model (model), and carrageenan-induced air pouch synovitis model + DHP (200 mg/kg of body weight) (model + DHP) groups. Mice in the model + DHP group were intragastrically administered 200 mg/kg BW of DHP solution daily for 10 days. Mice in the control and model groups were intragastrically administered the same amount of distilled water. Two hours after intragastric administration on day 10, 1 mL of a 1% carrageenan solution in a sterile 0.9% saline solution was injected into the air pouch of mice in the model and model + DHP groups. Six hours later, the mice were sacrificed and 4 mL of ice-cold sterile 0.9% saline solution was injected into the air pouch to fully wash its inner wall. The lavage fluid was collected to observe the color and turbidity of the lavage fluid, as well as the appearance of the backside of the air pouch. The exudate volume, total number of leukocytes, protein content, levels of malondialdehyde (MDA), interleukin-1b (IL-1β), and tumor necrosis factor-alpha (TNF-α), and total superoxide dismutase (T-SOD) activity of the lavage fluid were analyzed. Results: The results showed that pretreatment with DHPs reduced the carrageenan-induced exudate volume (P &lt; 0.01), total leukocytes (P &lt; 0.05), and protein content (P &lt; 0.01) in the air pouch lavage fluid. Furthermore, mice in the model + DHP group had significantly higher (P &lt; 0.01) T-SOD activity and lower MDA content (P &lt; 0.05), IL-1β (P &lt; 0.05), and TNF-α (P &lt; 0.01) levels in the air pouch lavage fluid compared with the model group. Conclusion: It is concluded that DHPs partially alleviated carrageenan-induced sterile inflammation, and its mechanism may be related to reducing exudation and scavenging oxygen-free radicals, inhibiting lipid peroxidation, and reducing the level of the proinflammatory factors, such as IL-1β and TNF-α.

β-Sitosterol Enhances Lung Epithelial Cell Permeability by Suppressing the NF-κB Signaling Pathway.

The dysregulation between pro-inflammatory and anti-inflammatory responses during sepsis is a crucial factor in driving sepsis progression. Acute lung injury (ALI) resulting from excessive production and accumulation of inflammatory mediators in the lungs contributes to impaired lung barrier function. The activation of the NF-κB signaling pathway during inflammation leads to the transcriptional activation of multiple inflammatory genes. Given the plausible impact of NF-κB signaling suppression in mitigating lung injury, substantive evidence demonstrates beta-sitosterol (BS)'s proficient ability to block NF-κB activation. Therefore, the aim of the present investigation was to delve into the impacts of BS in the context of sepsis-induced acute lung injury, employing both a mouse model and a model involving lung epithelial cells. Sepsis-induced lung injury was simulated in mice through cecum ligation and puncture (CLP). To emulate injury in murine lung epithelial (MLE-12) cells, an experiment involving lipopolysaccharide (LPS) was administered. Evaluation of alterations in lung tissue permeability encompassed techniques such as lung wet/dry (W/D) mass ratio, Evans blue staining, and quantification of total protein concentration in bronchoalveolar lavage fluid (BALF). Lung tissue histopathological shifts were ascertained via hematoxylin and eosin (HE) staining. Additionally, the concentrations of inflammatory cytokines IL-6 and TNF-α were quantified in every lung tissue and cell group by implementing enzyme-linked immunosorbent assay (ELISA). Protein quantification for signal biomarkers was carried out using Western blotting and immunofluorescence methodologies. In tandem, the assessment of MLE-12 cell permeability was conducted by evaluating fluorescein isothiocyanate (FITC)-dextran extravasation. BS mitigated lung tissue pathologies, reduced inflammatory factors, and lowered tissue and cell permeability. BS inhibited NF-κB signaling and increased claudin-4 and claudin-5 expression, enhancing septic lung epithelial cell permeability. Through suppressing the NF-κB signaling cascade, BS effectively curtails the levels of inflammatory mediators. Simultaneously, it orchestrates the modulation of claudin-4 and claudin-5 expression, culminating in the augmentation of lung epithelial cell barrier competence, thus improving sepsis-induced lung injury.

Activation of cannabinoid-2 receptor protects against Pseudomonas aeruginosa induced acute lung injury and inflammation

BackgroundBacterial pneumonia is a major risk factor for acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). Pseudomonas aeruginosa (PA), an opportunistic pathogen with an increasing resistance acquired against multiple drugs, is one of the main causative agents of ALI and ARDS in diverse clinical settings. Given the anti-inflammatory role of the cannabinoid-2 receptor (CB2R), the effect of CB2R activation in the regulation of PA-induced ALI and inflammation was tested in a mouse model as an alternative to conventional antibiotic therapy.MethodsIn order to activate CB2R, a selective synthetic agonist, JWH133, was administered intraperitoneally (i.p.) to C57BL/6J mice. Furthermore, SR144528 (a selective CB2R antagonist) was administered in combination with JWH133 to test the specificity of the CB2R-mediated effect. PA was administered intratracheally (i.t.) for induction of pneumonia in mice. At 24 h after PA exposure, lung mechanics were measured using the FlexiVent system. The total cell number, protein content, and neutrophil population in the bronchoalveolar lavage fluid (BALF) were determined. The bacterial load in the whole lung was also measured. Lung injury was evaluated by histological examination and PA-induced inflammation was assessed by measuring the levels of BALF cytokines and chemokines. Neutrophil activation (examined by immunofluorescence and immunoblot) and PA-induced inflammatory signaling (analyzed by immunoblot) were also studied.ResultsCB2R activation by JWH133 was found to significantly reduce PA-induced ALI and the bacterial burden. CB2R activation also suppressed the PA-induced increase in immune cell infiltration, neutrophil population, and inflammatory cytokines. These effects were abrogated by a CB2R antagonist, SR144528, further confirming the specificity of the CB2R-mediated effects. CB2R-knock out (CB2RKO) mice had a significantly higher level of PA-induced inflammation as compared to that in WT mice. CB2R activation diminished the excess activation of neutrophils, whereas mice lacking CB2R had elevated neutrophil activation. Pharmacological activation of CB2R significantly reduced the PA-induced NF-κB and NLRP3 inflammasome activation, whereas CB2KO mice had elevated NLRP3 inflammasome.ConclusionOur findings indicate that CB2R activation ameliorates PA-induced lung injury and inflammation, thus paving the path for new therapeutic avenues against PA pneumonia.

Amelioration of bleomycin induced pulmonary fibrosis by administration of Salvianolic acid B in mice.

Pulmonary fibrosis is the end‑stage manifestation of wide range of respiratory diseases and during pulmonary fibrosis, pulmonary inflammation and epithelial‑mesenchymal transition (EMT) play important roles. Salvianolic acid B (SAB) from the herb Salviae miltiorrhiza has been reported to possess an excellent anti‑inflammatory, antifibrotic and antioxidant activity. The present study aimed to investigate the ameliorative effect of SAB on bleomycin induced pulmonary fibrosis in mice. Adult albino mice were divided as SHAM/control group (saline alone), BLM group (bleomycin @ 1mg/kg intratracheally once) and SAB groups (BLM challenged once and SAB administration in three dosages @ 5, 10 and 15 mg/kg intraperitoneally daily for 30 days). Lungs wet/dry ratio and protein concentration in bronchoalveolar lavage fluid, MPO activity, oxidative stress markers, hydroxyproline assay, levels of inflammatory cytokines (TNF‑α, IL‑6 and TGF‑β1), NF‑κB activity, histopathology, immunostaining (E‑cadherin, vimentin and alpha ‑smooth muscle actin) and ultrastructural changes were studied. SAB showed anti‑inflammatory and anti‑fibrotic effects through inhibition of inflammatory cell infiltration, alveolar structure disruption, and collagen deposition and the expression of several fibrogenic cytokines. SAB also up‑regulate E‑cadherin and down‑regulated vimentin and alpha‑smooth muscle actin expression. In conclusion, Salvianolic acid B is effective in alleviating the BLM induced lung fibrosis through suppression of oxidative stress, inflammation, histological, ultrastructural changes and EMT.

Optimizing antidotal treatment with the oral HSP90 inhibitor TAS-116 against hydrochloric acid-induced pulmonary fibrosis in mice.

Exposure to high concentrations of hydrochloric acid (HCl) can lead to severe acute and chronic lung injury. In the aftermath of accidental spills, victims may be treated for the acute symptoms, but the chronic injury is often overlooked. We have developed a mouse model of acute and chronic lung injury, in which the peak of acute lung injury occurs on the day 4 after HCl exposure. We have also demonstrated that HSP90 inhibitors are effective antidotes when administered starting 24h after HCl. In this study we examined the hypothesis that the novel oral HSP90 inhibitor TAS-116 can effectively ameliorate HCl-induced lung injury even when treatment starts at the peak of the acute injury, as late as 96h after HCl. C57BI/6J mice were intratracheally instilled with 0.1N HCl. After 24 or 96h, TAS-116 treatment began (3.5, 7 or 14mg/kg, 5 times per week, p. o.) for either 2,3 or 4 orweeks. TAS-116 moderated the HCl-induced alveolar inflammation, as reflected in the reduction of white blood cells and total protein content in bronchoalveolar lavage fluid (BALF), overexpression of NLRP3 inflammasome, and inhibited the activation of pro-fibrotic pathways. Furthermore, TAS-116 normalized lung mechanics and decreased the deposition of extracellular matrix proteins in the lungs of mice exposed to HCl. Delayed and shortened treatment with TAS-116, successfully blocked the adverse chronic effects associated with acute exposure to HCl.

Isolation & identification of anti-inflammatory constituents of Randia dumetorum lamk. fruit: Potential beneficial effects against acute lung injury

Ethnopharmacological relevanceRandia dumetorum Lamk. is an Indian traditional medicinal plant that has been used for the treatment of various disorders including respiratory ailments. Aim of the studyIn continuation of our recent report that the Ethanol soluble fraction (ESF) of Randia dumetorum fruit had potent anti-inflammatory activity against acute lung injury (ALI) in mice, the present work was undertaken to unveil the key bioactive constituents possessing anti-inflammatory action against ALI by employing bioactivity-guided fractionation of ESF. Material and methodsDifferent fractions/sub-fractions obtained by column chromatography of ESF were subjected to bioactivity studies by analyzing total and differential count, and protein content in broncho-alveolar lavage fluid (BALF) procured from mice. The most bioactive sub-fraction F3.2 was analyzed for the assessment of various inflammatory mediators using molecular techniques like ELISA, PCR, and western blotting. Further, an attempt was made to separate the key compounds in F3.2 using solvents of differential polarities; and isolated compounds were validated for their anti-inflammatory activity followed by their characterization using spectral techniques like 1HNMR, 13CNMR, FT-IR, and ESIMS Mass Spectrometry. ResultsThe column chromatography of ESF yielded four fractions (F1, F2, F3, and F4) and data revealed that maximum activity resides in F3. Further fractionation of F3 yielded sub-fractions F3.1, F3.2, F3.3, and F3.4 which when tested for anti-inflammatory potential, showed F3.2 as the most active one. Moreover, the effect of F3.2 on oxidative stress parameters and inflammatory mediators analyzed via biochemical assays, PCR, and ELISA revealed the proficiency of this fraction in amelioration of ALI. F3.2 was then subjected to recrystallization using different solvents and two pure compounds were isolated which were characterized as D-Mannitol and Oleanolic acid (OA). D-Mannitol did not display any bioactivity, but OA showed potent anti-inflammatory activity. ConclusionConsidering the ethnopharmacological role of R. dumetorum in respiratory ailments, OA as an aglycone moiety seems to be the main active principle possessing anti-inflammatory potential against ALI.

Sevoflurane alleviates lung injury and inflammatory response compared with propofol in a rat model of VV ECMO.

Although venovenous extracorporeal membrane oxygenation (VV ECMO) is a reasonable salvage treatment for acute respiratory distress syndrome (ARDS), it requires sedating the patient. Sevoflurane and propofol have pulmonary protective and immunomodulatory properties. This study aimed to compare the effectiveness of sevoflurane and propofol on rats with induced ARDS undergoing VV ECMO. Fifteen sprague-dawley (SD) rats were randomly divided into three groups: Con group, sevoflurane (Sevo) group and propofol (Pro) group. Arterial blood gas tests were performed at time pointsT0 (baseline), T1 (the time to ARDS), and T2 (weaning from ECMO). Oxygenation index (PaO2/FiO2) was calculated, and lung edema assessed by determining the lung wet:dry ratio. The protein concentration in bronchial alveolar lavage fluid (BALF) was determined by using bicinchoninic acid assay. Haematoxylin and eosin staining was used to evaluate the lung pathological scores in each group. IL-1β and TNF-α were also measured in the BALF, serum and lung. Oxygenation index showed improvement in the Sevo group versus Pro group. The wet:dry ratio was reduced in the Sevo group compared with propofol-treated rats. Lung pathological scores were substantially lower in the Sevo group versus the Pro group. Protein concentrations in the BALF and levels of IL-1β and TNF-α in the Sevo group were substantially lower versus Pro group. This study demonstrates that compared with propofol, sevoflurane was more efficacious in improving oxygenation and decreasing inflammatory response in rat models with ARDS subject to VV ECMO treatment.

Differential modulation of lung aquaporins among other pathophysiological markers in acute (Cl2 gas) and chronic (carbon nanoparticles, cigarette smoke) respiratory toxicity mouse models.

Inhaled toxic chemicals and particulates are known to disrupt lung homeostasis causing pulmonary toxicity and tissue injury. However, biomarkers of such exposures and their underlying mechanisms are poorly understood, especially for emerging toxicants such as engineered nanoparticles and chemical threat agents such as chlorine gas (Cl2). Aquaporins (AQPs), commonly referred to as water channels, are known to play roles in lung homeostasis and pathophysiology. However, little is known on their regulation in toxicant-induced lung injuries. Here, we compared four lung toxicity models namely, acute chemical exposure (Cl2)-, chronic particulate exposure (carbon nanotubes/CNT)-, chronic chemical exposure (cigarette smoke extract/CSE)-, and a chronic co-exposure (CNT + CSE)- model, for modulation of lung aquaporins (AQPs 1, 3, 4, and 5) in relation to other pathophysiological endpoints. These included markers of compromised state of lung mucosal lining [mucin 5b (MUC5B) and surfactant protein A (SP-A)] and lung-blood barrier [protein content in bronchoalveolar lavage (BAL) fluid and, cell tight junction proteins occludin and zona-occludens]. The results showed toxicity model-specific regulation of AQPs measured in terms of mRNA abundance. A differential upregulation was observed for AQP1 in acute Cl2 exposure model (14.71-fold; p = 0.002) and AQP3 in chronic CNT exposure model (3.83-fold; p = 0.044). In contrast, AQP4 was downregulated in chronic CSE model whereas AQP5 showed no significant change in any of the models. SP-A and MUC5B expression showed a decreasing pattern across all toxicity models except the acute Cl2 toxicity model, which showed a highly significant upregulation of MUC5B (25.95-fold; p = 0.003). This was consistent with other significant pathophysiological changes observed in this acute model, particularly a compromised lung epithelial-endothelial barrier indicated by significantly increased protein infiltration and expression of tight junction proteins, and more severe histopathological (structural and immunological) changes. To our knowledge, this is the first report on lung AQPs as molecular targets of the study toxicants. The differentially regulated AQPs, AQP1 in acute Cl2 exposure versus AQP3 in chronic CNT nanoparticle exposure, in conjunction with the corresponding differentially impacted pathophysiological endpoints (particularly MUC5B) could potentially serve as predictive markers of toxicant type-specific pulmonary injury and as candidates for future investigation for clinical intervention.