Sepsis-related Acute Lung Injury Research Articles

Heparin-Binding Protein Promotes Acute Lung Injury in Sepsis Mice by Blocking the Aryl Hydrocarbon Receptor Signaling Pathway.

This study aimed to explore the therapeutic effect and potential mechanism of heparin-binding protein (HBP) reduction on sepsis-related acute lung injury. We utilized a murine model of sepsis-induced by intraperitoneal injection of lipopolysaccharides (LPS) in C57BL/6J mice divided into four groups: Control, LPS, Anti-HBP, and ceftriaxone (CEF). Following sepsis induction, Anti-HBP or CEF treatments were administered, and survival rates were monitored for 48 h. We then used reverse-transcription quantitative PCR to analyze the expression levels of HBP in lung tissues, immunohistochemistry for protein localization, and Western blotting for protein quantification. Pulmonary inflammation was assessed using enzyme-linked immunosorbent assays of proinflammatory cytokines (tumor necrosis factor-α, interleukin [IL]-1β, IL-6, and interferon-γ). The activation state of the aryl hydrocarbon receptor (AhR) signaling pathway was determined via Western blotting, evaluating both cytoplasmic and nuclear localization of AhR and the expression of cytochrome P450 1A1 protein by its target gene. Anti-HBP specifically reduced HBP levels. The survival rate of mice in the Anti-HBP and CEF groups was much higher than that in the LPS group. The severity of lung injury and pulmonary inflammatory response in the Anti-HBP and CEF groups was significantly lower than that in the LPS group. AhR signaling pathway activation was observed in the Anti-HBP and CEF groups. Additionally, there was no significant difference in the above indices between the Anti-HBP and CEF groups. HBP downregulation in lung tissues significantly improved LPS-induced lung injury and the pulmonary inflammatory response, thereby prolonging the survival of sepsis mice, suggesting activation of the AhR signaling pathway. Moreover, the effect of lowering the HBP level was equivalent to that of the classical antibiotic CEF. Not applicable.

A neutrophil elastase inhibitor, sivelestat, attenuates sepsis-induced acute kidney injury by inhibiting oxidative stress

BackgroundSivelestat, a selective inhibitor of neutrophil elastase (NE), can mitigate sepsis-related acute lung injury. However, the role of sivelestat in inhibiting oxidative stress and attenuating sepsis-related acute kidney injury (AKI) remains unclear. Here, we reported the effects of sivelestat against oxidative stress-induced AKI by suppressing the production of oxidative stress indicators. Materials and methodsA male Sprague-Dawley rat model of sepsis was established by cecal ligation and puncture (CLP). Sivelestat or normal saline was administered into jugular vein with a sustained-release drug delivery system. Indicators of inflammation and AKI, including white blood cells (WBC), neutrophils, lymphocytes, C-reactive proteins (CRP), procalcitonin (PCT), blood urea nitrogen (BUN), creatinine (Cr) and uric acid (UA), were assessed at 24 h post-sivelestat treatment. Indicators of liver injury, including direct bilirubin (DBIL), indirect bilirubin (IBIL), aspartate aminotransferase (AST) and alanine aminotransferase (ALT), were also assessed at 24 h post-sivelestat treatment. Indicators of oxidative stress, including superoxide dismutase (SOD), malondialdehyde (MDA) and glutathione peroxidase (GSH-Px), were assessed at 12 h and 24 h post-sivelestat treatment. At 24 h post-sivelestat treatment, H&E staining of kidney and liver tissue was performed to observe pathological alterations. ResultsAt 24 h post normal saline or sivelestat (0.2 g/kg body weight) treatment, WBC, neutrophil, CRP, PCT, MDA, BUN, Cr, UA, AST, ALT, DBIL and IBIL were increased, while SOD and GSH-Px were decreased, in septic rats treated with normal saline compared with that in non-septic rats treated with normal saline (all p < 0.05). The changes of these indicators were reversed in septic rats treated with sivelestat compared with that in septic rats treated with normal saline (all p < 0.05). Similar results were found regarding the levels of oxidative stress indicators at 12 h post-sivelestat treatment. The degenerative histopathological changes in both kidney and liver tissues were ameliorated upon sivelestat treatment. ConclusionsSivelestat plays a protective role in sepsis-related AKI by inhibiting oxidative stress. Our study reveals a possible therapeutic potential of sivelestat for oxidative stress-induced AKI.

Endothelial cell-derived extracellular vesicles expressing surface VCAM1 promote sepsis-related acute lung injury by targeting and reprogramming monocytes.

Acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) is a common life-threatening syndrome with no effective pharmacotherapy. Sepsis-related ARDS is the main type of ARDS and is more fatal than other types. Extracellular vesicles (EVs) are considered novel mediators in the development of inflammatory diseases. Our previous research suggested that endothelial cell-derived EVs (EC-EVs) play a crucial role in ALI/ARDS development, but the mechanism remains largely unknown. Here, we demonstrated that the number of circulating EC-EVs was increased in sepsis, exacerbating lung injury by targeting monocytes and reprogramming them towards proinflammatory macrophages. Bioinformatics analysis and further mechanistic studies revealed that vascular cell adhesion molecule 1 (VCAM1), overexpressed on EC-EVs during sepsis, activated the NF-κB pathway by interacting with integrin subunit alpha 4 (ITGA4) on the monocyte surface, rather than the tissue resident macrophage surface, thereby regulating monocyte differentiation. This effect could be attenuated by decreasing VCAM1 levels in EC-EVs or blocking ITGA4 on monocytes. Furthermore, the number of VCAM1+ EC-EVs was significantly increased in patients with sepsis-related ARDS. These findings not only shed light on a previously unidentified mechanism underling sepsis-related ALI/ARDS, but also provide potential novel targets and strategies for its precise treatment.

Cyasterone ameliorates sepsis-related acute lung injury via AKT (Ser473)/GSK3β (Ser9)/Nrf2 pathway

BackgroundAcute lung injury (ALI) is a severe disease that can lead to acute respiratory distress syndrome (ARDS), characterized by intractable hypoxemia, poor lung compliance, and respiratory failure, severely affecting patients' quality of life. The pathogenesis of ALI has not been fully elucidated yet, and sepsis is an important cause of ALI. Among the organ injuries caused by sepsis, the lungs are the earliest damaged ones. Radix cyathulae is reported to have analgesic, anti-inflammatory, and anti-aging effects. Cyasterone is extracted from Radix cyathulae. However, it is not known whether cyasterone has protective effects for ALI. This study aims to investigate the effect of cyasterone on sepsis-related ALI and its mechanism.MethodsWe used the cecal ligation peferation (CLP) method to establish a mouse sepsis model, and cyasterone was given intraperitoneally on days 1–3 to observe its preventive effect on sepsis-related acute lung injury. Primary murine peritoneal macrophages were used to investigate the molecular mechanism of cyasterone in vitro.ResultsCyasterone pretreatment inhibits pro-inflammatory cytokine production, NLRP3 inflammasome activation, and oxidative stress in vivo and in vitro. In addition, cyasterone attenuates sepsis-induced ALI by activating nuclear factor erythroid2-related factor (Nrf2), which may be associated with AKT(Ser473)/GSK3β(Ser9) pathway activation.ConclusionsCyasterone defends against sepsis-induced ALI by inhibiting inflammatory responses and oxidative stress, which depends heavily on the upregulation of the Nrf2 pathway through phosphorylation of AKT(Ser473)/GSK3β(Ser9). These results suggest cyasterone may be a valuable drug candidate for preventing sepsis-related ALI.

Exosomal miR-127-5p from BMSCs alleviated sepsis-related acute lung injury by inhibiting neutrophil extracellular trap formation

Neutrophil extracellular traps (NETs) play an important role in sepsis-related acute lung injury (ALI). Bone marrow mesenchymal stem cells (BMSCs)-derived exosomes and miRNA are becoming promising agents for the treatment of ALI. The current study aimed to elucidate the mechanism by BMSCs-derived exosomes carrying miR-127-5p inhibiting to the formation of NETs in sepsis-related ALI. We successfully isolated exosomes from BMSCs and confirmed that miR-127-5p was enriched in the exosomes. ALI mice treated with BMSCs-derived exosomes histologically improved, and the release of NETs and inflammatory factors in lung tissue and peripheral blood of mice also decreased compared with LPS group, while the protective effect of exosomes was attenuated after the knockdown of miR-127-5p. Using dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay, we identified CD64 as a direct target of miR-127-5p. Meanwhile, BMSCs-derived exosomes can synergize with anti-CD64 mab in ALI mice to reduce tissue damage, inhibit the release of inflammatory factors and NETs formation. The synergistic effect of exosomes was attenuated when miR-127-5p was down-regulated. These findings suggest that exosomal miR-127-5p derived from BMSCs is a potential therapeutic agent for treatment of sepsis-induced ALI through reducing NETs formation by targeting CD64.

Exosomal PGE2 from M2 macrophages inhibits neutrophil recruitment and NET formation through lipid mediator class switching in sepsis

BackgroundExcess polymorphonuclear neutrophil (PMN) recruitment or excessive neutrophil extracellular trap (NET) formation can lead to the development of multiple organ dysfunction during sepsis. M2 macrophage-derived exosomes (M2-Exos) have exhibited anti-inflammatory activities in some inflammatory diseases to mediate organ functional protection, but their role in treating sepsis-related acute lung injury (ALI) remains unclear. In this study, we sought to investigate whether M2-Exos could prevent potentially deleterious inflammatory effects during sepsis-related ALI by modulating abnormal PMN behaviours.MethodsC57BL/6 wild-type mice were subjected to a caecal ligation and puncture (CLP) mouse model to mimic sepsis in vivo, and M2-Exos were administered intraperitoneally 1 h after CLP. H&E staining, immunofluorescence and immunohistochemistry were conducted to investigate lung tissue injury, PMN infiltration and NET formation in the lung. We further demonstrated the role of M2-Exos on PMN function and explored the potential mechanisms through an in vitro coculture experiment using PMNs isolated from both healthy volunteers and septic patients.ResultsHere, we report that M2-Exos inhibited PMN migration and NET formation, alleviated lung injury and reduced mortality in a sepsis mouse model. In vitro, M2-Exos significantly decreased PMN migration and NET formation capacity, leading to lipid mediator class switching from proinflammatory leukotriene B4 (LTB4) to anti-inflammatory lipoxin A4 (LXA4) by upregulating 15-lipoxygenase (15-LO) expression in PMNs. Treatment with LXA4 receptor antagonist attenuated the effect of M2-Exos on PMNs and lung injury. Mechanistically, prostaglandin E2 (PGE2) enriched in M2-Exos was necessary to increase 15-LO expression in PMNs by functioning on the EP4 receptor, upregulate LXA4 production to downregulate chemokine (C-X-C motif) receptor 2 (CXCR2) and reactive oxygen species (ROS) expressions, and finally inhibit PMN function.ConclusionsOur findings reveal a previously unknown role of M2-Exos in regulating PMN migration and NET formation through lipid mediator class switching, thus highlighting the potential application of M2-Exos in controlling PMN-mediated tissue injury in patients with sepsis.

Research Progress of the Association of the Neutrophil Elastase with Sepsis-related Acute Lung Injury

ABSTRACT Sepsis is considered a life-threatening organ dysfunction resulting from a dysregulation of the body's response to the infection, a syndrome of physiological, pathological, and biochemical abnormalities caused by infection. The prevention and treatment of sepsis-related acute lung injury has become a hot spot in sepsis research in recent years. Neutrophil elastase (NE) is a multifunctional serine protease released by neutrophils cells, which can participate in the regulation of the inflammatory response in the body and directly affect the occurrence and development of inflammation. This article reviews the pathogenesis of sepsis-related lung injury and the mechanism of action, clinical application and prospect of neutrophil elastase inhibitor (NEI).

The role of pyroptosis in endothelial dysfunction induced by diseases

Most organs in the body rely on blood flow, and vesicular damage is the leading cause of injury in multiple organs. The endothelium, as the barriers of vessels, play a critical role in ensuring vascular homeostasis and angiogenesis. The rapid development of risk factors in endothelial injuries has been seen in the past decade, such as smoking, infectious, and diabetes mellites. Pyroptotic endothelium is an inflammatory mode of governed endothelial cell death that depend on the metabolic disorder and severe infectious such as atherosclerosis, and sepsis-related acute lung injury, respectively. Pyroptotic endothelial cells need GSDMD cleaved into N- and C-terminal by caspase1, and the cytokines are released by a pore constructed by the N-terminal of GSDMD in the membrane of ECs, finally resulting in severe inflammation and pyroptotic cell death. This review will focus on the patho-physiological and pharmacological pathways of pyroptotic endothelial metabolism in diseases. Overall, this review indicates that pyroptosis is a significant risk factor in diseases and a potential drug target in related diseases.

The role of pyroptosis in endothelial dysfunction induced by diseases.

Most organs in the body rely on blood flow, and vesicular damage is the leading cause of injury in multiple organs. The endothelium, as the barriers of vessels, play a critical role in ensuring vascular homeostasis and angiogenesis. The rapid development of risk factors in endothelial injuries has been seen in the past decade, such as smoking, infectious, and diabetes mellites. Pyroptotic endothelium is an inflammatory mode of governed endothelial cell death that depend on the metabolic disorder and severe infectious such as atherosclerosis, and sepsis-related acute lung injury, respectively. Pyroptotic endothelial cells need GSDMD cleaved into N- and C-terminal by caspase1, and the cytokines are released by a pore constructed by the N-terminal of GSDMD in the membrane of ECs, finally resulting in severe inflammation and pyroptotic cell death. This review will focus on the patho-physiological and pharmacological pathways of pyroptotic endothelial metabolism in diseases. Overall, this review indicates that pyroptosis is a significant risk factor in diseases and a potential drug target in related diseases.

Circ_0001498 contributes to lipopolysaccharide-induced lung cell apoptosis and inflammation in sepsis-related acute lung injury via upregulating SOX6 by interacting with miR-574-5p.

Circular RNAs (circRNAs) have important regulation in in sepsis-related acute lung injury (ALI). Circ_0001498 was significantly overexpressed in sepsis-induced acute respiratory distress syndrome. The aims of this study were to explore role and mechanism of circ_0001498 in lipopolysaccharide (LPS)-treated WI-38 cells. Human samples were collected from 56 sepsis patients and 46 healthy volunteers at Liyang People's Hospital. Circ_0001498, microRNA-574-5p (miR-574-5p) or sex-determining region Y-related high-mobility-group box 6 (SOX6) levels were detected via reverse transcription-quantitative polymerase chain reaction assay. Cell viability was determined through Cell Counting Kit-8 assay. Apoptosis rate was examined by flow cytometry. Western blot was used for measurement of proteins. Inflammatory cytokines were detected via enzyme-linked immunosorbent assay. Target relation was analyzed via dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. Circ_0001498 was overexpressed in sepsisrelated ALI patients and LPS-treated WI-38 cells. Silencing circ_0001498 reduced LPS-induced cell apoptosis and inflammation. Circ_0001498 interacted with miR-574-5p. The regulation of circ_0001498 knockdown was abolished by miR-574-5p inhibitor. Furthermore, miR-574-5p directly targeted SOX6 and circ_0001498 upregulated SOX6 via targeting miR-574-5p. Overexpression of miR-574-5p alleviated LPS-induced cell injury by downregulating SOX6. This research identified that circ_0001498 facilitated sepsis-related ALI progression by targeting miR-574-5p to upregulate SOX6.

Pyk2 regulates sepsis-induced lung injury via ferroptosis.

The onset of sepsis represents a hyper-inflammatory condition that can lead to organ failure and mortality. Recent findings suggest a potential beneficial effect of protein tyrosine kinase Pyk2 inhibitor on sepsis in a mouse model. In this study, we investigated the regulatory role of Pyk2 inhibitor in ferroptosis and sepsis-associated acute lung injury (ALI). A Pyk2 inhibitor or a ferroptosis regulator were injected into mice sustaining sepsis-induced ALI and the effects on lung injury and pro-inflammatory response were evaluated. Clinically, Pyk2 expression was determined in serum samples of patients with sepsis. Further, the association between serum Pyk2 levels and clinical features was determined. Experimental mouse models revealed that treatment with Pyk2 inhibitor TAE226 can significantly alleviate lung injury, downregulate pro-inflammatory responses and decrease markers of ferroptosis, which were induced by LPS. Both upregulation and downregulation of ferroptosis can lead to the loss of TAE226 function, indicating that Pyk2 promotes inflammation via ferroptosis induction. Analysis of clinical samples revealed that the serum Pyk2 levels were significantly increased in patients with sepsis. The serum Pyk2 levels were associated with APACHE2 scores and 30-day mortality. Further, we found a negative correlation between serum Pyk2 and Fe3+ levels, which was consistent with the mechanism identified in the mouse model. Pyk2 inhibitor of ferroptosis is a promising therapeutic candidate against sepsis-related ALI.

Silencing long non-coding RNA SNHG3 repairs the dysfunction of pulmonary microvascular endothelial barrier by regulating miR-186-5p/Wnt axis

Barrier permeability changes of human pulmonary microvascular endothelial cells (HPMVECs) are important in sepsis-related acute lung injury (ALI) pathogenesis. Long non-coding small nucleolar RNA host gene 3 (SNHG3) mediates the cell-biological phenotype of lung cancer cells and affects the progression of lung cancer, but its role in regulating functions of lung non-malignant cells is still rarely reported. Therefore, we evaluated the regulatory effect of SNHG3 on the function of PMVECs in sepsis-related ALI. Small interference RNA (siRNA)-mediated deletion of SNHG3 promoted the proliferation of PMVECs, reduced apoptosis and barrier permeability, and increased the expression of tight junction proteins claudin-5 and ZO-1. Knockdown of SNHG3 increased the miR-186-5p expression, while overexpression of SNHG3 upregulated the level of wnt5a. Through a dual luciferase reporter assay, we confirmed the binding between SNHG3 and miR-186-5p, miR-186-5p and wnt5a. We further found that knockout of miR-186-5p could inhibit cell proliferation, increase apoptosis and barrier permeability, and down-regulate claudin-5 and ZO-1. Importantly, silencing miR-186-5p and activating Wnt signal pathway could eliminate the barrier repair effect caused by down-regulation of SNHG3. To sum up, our results suggested that knockdown of long non-coding RNA SNHG3 repaired the dysfunction of pulmonary microvascular endothelial barrier through the miR-186-5p/Wnt axis.

The dipeptidyl peptidase-4 inhibitor linagliptin ameliorates LPS-induced acute lung injury by maintenance of pulmonary microvascular barrier via activating the Epac1/AKT pathway

Pulmonary microvascular endothelial cells (PMVECs) barrier dysfunction is a main pathophysiological feature of sepsis-related acute lung injury (ALI). This study aimed to investigate whether the dipeptidyl peptidase (DPP)-4 inhibitor linagliptin could protect against LPS-induced PMVECs barrier disruption and its underlying molecular mechanisms. A classical ALI animal model and LPS-treated PMVECs were applied and all were treated with or without linagliptin. Cellular experiments demonstrated that linagliptin could mitigate LPS-induced PMVECs hyperpermeability and intercellular junction (VE-cadherin, β-catenin, and ZO-1) disruption in a dose-dependent manner. Correspondingly, it was observed that linagliptin pretreatment distinctly relieved LPS-induced lung injury, oxidative stress, and pulmonary edema in vivo. Furthermore, we found that the inhibition of oxidative stress by linagliptin may be achieved by reversing impaired mitochondrial function. Mechanistically, linagliptin administration promoted the activation of the Epac1 pathway and its downstream AKT pathway, while inhibition of the Epac1/Akt signaling pathway significantly alleviated the above-mentioned protective effect of linagliptin on the PMVECs barrier. Taken together, these data suggest that linagliptin can effectively reserve PMVECs barrier dysfunction and inhibit oxidative stress to protect against ALI via activating the Epac1/AKT signaling pathway, and thus may become a potential clinical therapeutic strategy for ALI.

Electroacupuncture preconditioning improves pulmonary function via inhibiting inflammatory response and up-regulating expression of ACE2 and Ang (1-7) in lipopolysaccharide-induced acute lung injury rats

To observe the effect of electroacupuncture (EA) at "Zusanli"(ST36) pretreatment on lung functions, inflammatory response, and levels of angiotensin-converting enzyme 2 (ACE2) and angiotensin (1-7) ［Ang (1-7)］ in rats with sepsis-induced acute lung injury (ALI), so as to explore its mechanisms underlying improvement of ALI. Thirty male SD rats were randomly divided into normal, model and EA groups (n=10 in each group). The sepsis-related ALI model was established by intraperitoneal injection of lipopolysaccharide (LPS, 5 mg/kg). Rats of the EA group received EA (4 Hz/20 Hz, 1-3 mA) stimulation at bilateral ST36 for 30 min, once each day, for 7 days before modeling. The lung functions including forced vital capacity (FVC), forced expiratory volume at 0.1 second (FEV0.1) and FEV0.3 were detected using a respiratory function detector for small animals at 3 h after modeling. The bronchoalveolar lavage fluid (BALF) was collected for assaying the contents of Ang (1-7), tumor necrosis factor-α (TNF-α) and interleukin-1 β (IL-1β) using ELISA. The lung wet/dry weight (W/D) ratio, FEV0.1/FVC, and FEV0.3/FVC were calculated. The histopathological changes of lung tissues were displayed by hematoxylin-eosin (H.E.) staining. The expression of ACE2 and mitochondrial assembly receptor (MasR) mRNAs and proteins in the lung tissue was detected by fluorescence quantitative real-time PCR and Western blot, separately. Following modeling, the levels of FVC, FEV0.1, FEV0.3, ratio of FEV0.1/FVC and FEV0.3/FVC, content of Ang (1-7) in the BALF, and the expression levels of ACE2 and MasR mRNAs and proteins in the lung tissue were significantly decreased (P<0.01), while the level of W/D ratio and TNF-α and IL-1β contents in the BALF significantly increased (P<0.01) in the model group relevant to the normal group. In comparison with the model group, the levels of FVC, FEV0.1, FEV0.3, ratio of FEV0.1/FVC and FEV0.3/FVC, content of Ang (1-7) in the BALF, and expression levels of ACE2 and MasR mRNAs and proteins in the lung tissue were significantly increased (P<0.05, P<0.01), whereas the level of W/D ratio, and TNF-α and IL-1β contents in the BALF were significantly decreased (P<0.05, P<0.01) in the EA group. H.E. staining showed pulmonary interstitial edema and alveolar septum thickening with severe inflammatory cell infiltration in the model group, which was relatively milder in the EA group. EA preconditioning at ST36 can improve pulmonary function in sepsis-related ALI rats, which may be related to its effects in inhibiting inflammatory response and up-regulating ACE2 and MasR expression and Ang (1-7) content in the lung tissue.

Preterm sepsis is associated with acute lung injury as measured by pulmonary severity score.

Sepsis related acute lung injury (ALI) is established in adults but has not been investigated in premature infants. Herein, we used pulmonary severity score (PSS) trajectories and C-reactive protein (CRP) to examine the relation between sepsis and ALI in premature infants. This retrospective study identified 211 sepsis and 123 rule out (RO) events in 443 infants born <31 weeks and <1500 grams. The PSS was calculated prior to, at the time of, and up to 1 week after each event. Initial and peak CRP values were collected for each event. PSS significantly increased at 0 h from baseline (-72h) and remained increased at all subsequent time points (all p < 0.002) in sepsis events. Mean PSS in sepsis episodes were also higher compared to RO events at +24 h, +48 h, +72 h, and +168 h (all p < 0.004). A positive correlation was noted between peak CRP values in sepsis events and PSS at 0 h, +24 h, +48 h, and +72 h. The temporal PSS trends and correlation with CRP levels observed in sepsis but not in RO events supports the hypothesis that neonatal sepsis is associated with ALI and contributes to the accumulating evidence that neonatal ARDS occurs. To evaluate pulmonary severity scores and c-reactive protein values over time to establish an association between preterm neonatal sepsis and acute lung injury (ALI). Though sepsis is well established as the most common indirect cause of ALI leading to acute respiratory distress syndrome (ARDS) in adults and pediatrics, this phenomenon remains undefined in neonates. This study validates the proposal by the Neonatal ARDS Project that ARDS also occurs in neonates by demonstrating acute and sustained changes in markers of pulmonary injury temporally related to a diagnosis of neonatal sepsis in preterm infants.

Knockdown of PFKFB2 Alleviates Oxidative Stress and Inflammation in LPS-Induced Alveolar Epithelial Cells by Reducing Glycolysis

Glycolysis is the most important mode of energy metabolism in endothelial cells and has been shown to be involved in the pathological processes of acute and chronic inflammatory diseases. Phosphofructokinase 2/fructose-2, 6-bisphosphatase 2 (PFKFB2) exerts an important regulatory factor in the process of glycolysis by catalyzing the synthesis and degradation of fructose 2,6-bisphosphate. There is still unclear however, whether PFKFB2 can play a role in sepsis-related acute lung injury by regulating glycolysis. This research examines the role and mechanism of PFKFB2 in LPS-induced alveolar epithelial cells. In this study, the detection of mRNA expressions of PFKFB2, glycolysis and inflammation-related proteins employed quantitative real-time PCR (RTqPCR). Western blot was applied to examine the expressions of all proteins. The viability of A549 cells was assessed with the use of cell counting kit (CCK)-8. The expressions of related factors were quantified by commercial assay kits, respectively. The experimental results showed that the expression of PFKFB2 was increased in sepsis. Knockdown of PFKFB2 alleviated glycolysis in LPS-induced A549 cells. Additionally, knockdown of PFKFB2 reduced LPS-induced oxidative stress and inflammation through glycolysis. Knockdown of PFKFB2 also mitigated LPS-induced oxidative stress and inflammation in alveolar epithelial cells by reducing glycolysis. Hence, PFKFB2 may be served as an effective target for the treatment of sepsis-related acute lung injury.

MicroRNA-1224-5p Aggravates Sepsis-Related Acute Lung Injury in Mice.

Oxidative stress and inflammation are implicated in the development of sepsis-related acute lung injury (ALI). MicroRNA-1224-5p (miR-1224-5p) plays critical roles in regulating inflammatory response and reactive oxygen species (ROS) production. The present study is aimed at investigating the role and underlying mechanisms of miR-1224-5p in sepsis-related ALI. Mice were intratracheally injected with lipopolysaccharide (LPS, 5 mg/kg) for 12 h to induce sepsis-related ALI. To manipulate miR-1224-5p level, mice were intravenously injected with the agomir, antagomir, or matched controls for 3 consecutive days. Murine peritoneal macrophages were stimulated with LPS (100 ng/mL) for 6 h to further validate the role of miR-1224-5p in vitro. To inhibit adenosine 5′-monophosphate-activated protein kinase alpha (AMPKα) or peroxisome proliferator activated receptor-gamma (PPAR-γ), compound C or GW9662 was used in vivo and in vitro. We found that miR-1224-5p levels in lungs were elevated by LPS injection, and that the miR-1224-5p antagomir significantly alleviated LPS-induced inflammation, oxidative stress, and ALI in mice. Conversely, the miR-1224-5p agomir aggravated inflammatory response, ROS generation, and pulmonary dysfunction in LPS-treated mice. In addition, the miR-1224-5p antagomir reduced, while the miR-1224-5p agomir aggravated LPS-induced inflammation and oxidative stress in murine peritoneal macrophages. Further findings revealed that miR-1224-5p is directly bound to the 3′-untranslated regions of PPAR-γ and subsequently suppressed PPAR-γ/AMPKα axis, thereby aggravating LPS-induced ALI in vivo and in vitro. We demonstrate for the first time that endogenous miR-1224-5p is a critical pathogenic factor for inflammation and oxidative damage during LPS-induced ALI through inactivating PPAR-γ/AMPKα axis. Targeting miR-1224-5p may help to develop novel approaches to treat sepsis-related ALI.

CircEXOC5 promotes ferroptosis by enhancing ACSL4 mRNA stability via binding to PTBP1 in sepsis-induced acute lung injury

BackgroundSepsis causes severe acute lung injury (ALI). Circular RNA is involved in the regulation of sepsis-related ALI progression. The regulation mechanism of circEXOC5 in sepsis-induced ALI is still unclear. Whether circEXOC5 is involved in the regulation of ferroptosis remains to be explored. MethodsWe constructed a mouse model of sepsis through cecal ligation and puncture (CLP). LPS induced mouse lung microvascular endothelial cells (MPVECs) to construct a sepsis cell model. The expression of circEXOC5 in the sepsis model was detected by qPCR. The extent of lung injury in mice was analyzed by HE staining. The contents of GSH/GSSG, iron, MDA and 4HNE in mice lung tissues and cells were detected by the kit. And further the ROS content was detected in the cells. Finally, the binding relationship between circEXOC5 and PTBP1 was detected by RIP and RNA pulldown. ResultsOur results showed that the circEXOC5 expression was significantly increased in the in vivo and in vitro models of sepsis. And after inhibiting circEXOC5, it improved the lung injury of septic mice. It was confirmed in cell models that ROS levels and ferroptosis in cells were reduced after knocking down circEXOC5. In addition, the expressions of ACSL4 and Gpx4 proteins were regulated by the level of circEXOC5. Finally, we also found that circEXOC5 had a direct binding relationship with PTBP1. ConclusionOur study found that the expression of cell ferroptosis and circEXOC5 increased in ALI induced by sepsis, and circEXOC5 aggravated ferroptosis in septic cells by regulating the PTBP1/ACSL4 axis.

A 4-Benzene-Indol Derivative Alleviates LPS-Induced Acute Lung Injury Through Inhibiting the NLRP3 Inflammasome.

Acute lung injury (ALI) is a common complication of critical illness that could frequently lead to acute respiratory distress syndrome and other serious clinical consequences. Sepsis is one of the major and most common inducements among all causes of ALI. Due to its high incidence and mortality rate and also the complexity in treatment, sepsis-related ALI has become an urgent clinical problem waiting to be solved effectively. At present, only the protective ventilation strategy, restrictive fluid management, and antibiotics application are measures that can improve the prognosis with evidence-based medical proof. No pharmacological treatment is currently available to protect or significantly reverse the prognosis. Seeking for effective interventions measures for sepsis-related ALI is one of the most necessitous research directions. In this research, a conspicuous discovery of treatment-related translational use for a 4-benzene-indol derivative was elaborated by screening a large number of chemical compounds. The results showed that 4-benzene-indol derivative could not only suppress the activation of NLRP3 inflammasome both in vitro and alleviate LPS-induced ALI in vivo but also suppress the NLRP3 inflammasome in human myeloid leukemia mononuclear cells (THP-1) cell lines. Mechanistically, 1,2-diol blocks the NLRP3 inflammasome activation by disrupting NLRP3–NEK7 interaction and the subsequent NLRP3 inflammasome assembly and activation. To summarize, this research indicated that the newly-discovered 4-benzene-indol derivative targets NLRP3 inflammasome signaling, which consequently alleviates sepsis-related ALI. Collectively, the 4-benzene-indol derivative may serve as a potential therapeutic drug and NLRP3 inflammasome signaling would be a novel pharmaceutical target for clinical treatment of sepsis-related ALI.

Exosomal miR-30d-5p of neutrophils induces M1 macrophage polarization and primes macrophage pyroptosis in sepsis-related acute lung injury

BackgroundPolymorphonuclear neutrophils (PMNs) play an important role in sepsis-related acute lung injury (ALI). Accumulating evidence suggests PMN-derived exosomes as a new subcellular entity acting as a fundamental link between PMN-driven inflammation and tissue damage. However, the role of PMN-derived exosomes in sepsis-related ALI and the underlying mechanisms remains unclear.MethodsTumor necrosis factor-α (TNF-α), a key regulator of innate immunity in sepsis-related ALI, was used to stimulate PMNs from healthy C57BL/6J mice in vitro. Exosomes isolated from the supernatant were injected to C57BL/6J wild-type mice intraperitoneally (i.p.) and then examined for lung inflammation, macrophage (Mϕ) polarization and pyroptosis. In vitro co-culture system was applied where the mouse Raw264.7 macrophages or bone marrow-derived macrophages (BMDMs) were co-cultured with PMN-derived exosomes to further confirm the results of in vivo animal study and explore the potential mechanisms involved.ResultsExosomes released by TNF-α-stimulated PMNs (TNF-Exo) promoted M1 macrophage activation after in vivo i.p. injection or in vitro co-culture. In addition, TNF-Exo primed macrophage for pyroptosis by upregulating NOD-like receptor 3 (NLRP3) inflammasome expression through nuclear factor κB (NF-κB) signaling pathway. Mechanistic studies demonstrated that miR-30d-5p mediated the function of TNF-Exo by targeting suppressor of cytokine signaling (SOCS-1) and sirtuin 1 (SIRT1) in macrophages. Furthermore, intravenous administration of miR-30d-5p inhibitors significantly decreased TNF-Exo or cecal ligation and puncture (CLP)-induced M1 macrophage activation and macrophage death in the lung, as well as the histological lesions.ConclusionsThe present study demonstrated that exosomal miR-30d-5p from PMNs contributed to sepsis-related ALI by inducing M1 macrophage polarization and priming macrophage pyroptosis through activating NF-κB signaling. These findings suggest a novel mechanism of PMN-Mϕ interaction in sepsis-related ALI, which may provide new therapeutic strategies in sepsis patients.