Background:Sepsis triggers dysregulated systemic inflammation and multiple-organ dysfunction, with the lungs being particularly susceptible to injury. Sepsis-induced acute respiratory distress syndrome (ARDS) is mainly driven by TLR4/NF-κB-mediated hyperinflammation and alveolar macrophage activation. Matrine, a bioactive alkaloid derived from Sophora flavescens, has been reported to modulate redox homeostasis and ferroptosis-associated lipid peroxidation. However, the target-specific mechanisms underlying its effects on ferroptosis and inflammatory signaling in sepsis-induced acute lung injury (SALI) remain incompletely understood. Purpose:This study aimed to evaluate the therapeutic effects of matrine in a cecal ligation and puncture (CLP)-induced SALI model and to determine whether its protective effects involve reinforcement of NQO1/SLC7A11/GPX4-associated anti-ferroptotic defenses and suppression of NF-κB-driven inflammation. Methods:We analyzed the single-cell RNA-sequencing (scRNA-seq) dataset GSE273924 to characterize CD45-enriched pulmonary immune-cell subsets in sham mice and mice with intratracheal Escherichia coli-induced pneumonia. Network pharmacology and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were performed to predict Kushen (KS)-related targets and pathways associated with SALI. Differential expression analysis and weighted gene co-expression network analysis (WGCNA) of GSE245013 were used to identify candidate targets. Matrine-NQO1 binding and intracellular target engagement were evaluated using molecular docking, molecular dynamics simulations, surface plasmon resonance (SPR), and the cellular thermal shift assay (CETSA). The therapeutic effects of matrine were assessed in mice with CLP-induced SALI and in lipopolysaccharide (LPS)-stimulated MH-S cells. Lung histopathology, inflammatory cytokine production, target protein expression, ferroptosis-associated indicators, and NF-κB activation were evaluated using molecular, biochemical, and histological assays. The functional contribution of NQO1 was further examined using the NQO1 inhibitor ES936. Results:scRNA-seq analysis of GSE273924 revealed substantial remodeling of the CD45-enriched pulmonary immune-cell landscape in mice with intratracheal E. coli-induced pneumonia, including macrophage transcriptional programs associated with ferroptosis and inflammatory signaling. Integrated network pharmacology and bioinformatics analyses prioritized NQO1 as a candidate target of matrine and identified NF-κB signaling as a potentially relevant pathway. Molecular docking, molecular dynamics simulations, SPR, and CETSA supported matrine-NQO1 binding and intracellular target engagement. Functionally, matrine improved survival, attenuated lung injury, reinforced NQO1/SLC7A11/GPX4-associated anti-ferroptotic defenses, and suppressed NF-κB activation in CLP mice. Similar protective effects were observed in LPS-stimulated MH-S cells. ES936 partially attenuated the matrine-mediated improvements in cell viability, redox homeostasis, ferroptosis-associated indicators, and NF-κB p65 phosphorylation, supporting a functional contribution of NQO1 to the protective effects of matrine. Conclusion:Matrine alleviates SALI by reinforcing NQO1/SLC7A11/GPX4-associated anti-ferroptotic defenses and attenuating NF-κB-driven inflammation.

Sepsis-induced acute lung injury (SALI) is a fatal complication of sepsis with limited therapeutic options. Karacoline, a diterpenoid alkaloid from Aconitum, shows potential protective effects, but its mechanism in SALI remains unclear. Network pharmacology and WGCNA identified core targets and pathways of Karacoline, with molecular docking confirming its binding to PPARγ. A cecal ligation and puncture mouse model was used to assess lung pathology, edema, barrier dysfunction, inflammatory cytokines, and apoptosis. In vitro, LPS-stimulated MH-S cells were used to evaluate apoptosis, ROS generation, mitochondrial function, and PPARγ/MAPK signaling. The PPARγ antagonist GW9662 was used to verify the involvement of PPARγ in the protective effects of Karacoline. Karacoline significantly alleviated lung injury, reduced the W/D ratio, decreased total protein and albumin levels in BALF, and suppressed IL-1β, IL-6, and TNF-α. Mechanistically, Karacoline upregulated PPARγ expression and inhibited ERK1/2 and JNK phosphorylation, thereby contributing to the reduction of pulmonary cell apoptosis. In vitro, Karacoline limited ROS generation, maintained mitochondrial membrane potential, and reduced LPS-induced apoptosis in MH-S cells. Moreover, GW9662 weakened the anti-apoptotic effect of Karacoline and partially reversed its inhibition of JNK/ERK activation, further supporting the involvement of PPARγ in Karacoline-mediated inhibition of JNK/ERK MAPK signaling and apoptosis. Karacoline protects against SALI by suppressing mitochondrial apoptosis, an effect closely associated with PPARγ-related inhibition of JNK/ERK MAPK signaling. These findings suggest that Karacoline may be a candidate compound worthy of further validation for sepsis-induced lung injury intervention.

Sepsis is a life-threatening organ dysfunction caused by dysregulated inflammatory and immune responses to infection. Its global incidence and mortality remain high, posing a severe threat to public health. Acute lung injury (ALI) is a common and serious complication of sepsis. Current understanding of the pathogenesis and effective therapeutic strategies for sepsis-induced acute lung injury (SI-ALI) remains insufficient. This study aims to investigate the role and underlying mechanisms of the deubiquitinase OTUD1 in sepsis-induced pulmonary injury. Using a mouse model of sepsis-induced lung injury combined with genetic knockout techniques and ferroptosis inhibitors, we systematically analyzed the protective effects of OTUD1 in sepsis-related lung damage and explored the regulatory roles of AMPK and GSK3β/β-catenin signaling pathways. Results demonstrated that OTUD1 gene deletion exacerbated lung tissue damage and inflammatory responses in septic mice while increasing ferroptosis levels; pretreatment with the ferroptosis inhibitor Ferrostatin-1 significantly ameliorated these effects. Further mechanistic studies revealed that OTUD1 may regulate ferroptosis levels in lung tissue by modulating the activation status of AMPK and GSK3β/β-catenin pathways. Specifically, OTUD1 may remove K63-linked ubiquitin chains from AMPK, altering its protein conformation and subsequently promoting AMPK phosphorylation to regulate the GSK3β/β-catenin signaling cascade. Collectively, this study provides the first systematic elucidation of OTUD1's protective role in sepsis-induced lung injury and its relationship with ferroptosis, offering novel molecular targets and theoretical foundations for the treatment of sepsis-associated pulmonary damage.

Neutrophil extracellular traps (NETs) are reticular structures released by neutrophils, and the process of their formation is called NETosis. NETs play a key role in the pathological process of sepsis. However, the specific regulatory mechanism has not been fully clarified. This study finds that the levels of NETs in peripheral blood are significantly elevated in clinical sepsis patients and cecal ligation and puncture (CLP) mouse models, and the expression of Acod1 is closely related to the generation of NETs. Acod1 knockout led to a further increase in NETs levels in CLP mice, aggravated the inflammatory response, worsened organ damage, and reduced the survival rate. Further studies indicate that E3 ubiquitin ligase UBR5 interacts with PAD4 (one of the core proteins for NETs generation). Acod1/itaconate (ITA) enhanced the enzymatic activity of UBR5 through alkylation modification, promoting the K48-linked polyubiquitination and degradation of PAD4, thereby inhibiting NETosis. In conclusion, this study combines transcriptomics, metabolomics, genetic engineering, and co-immunoprecipitation techniques to reveal the molecular mechanism of Acod1/ITA in regulating NETs, providing new potential targets and theoretical basis for the treatment of sepsis.

Sepsis-associated encephalopathy (SAE), a life-threatening neurological complication of systemic infection, contributes substantially to sepsis-related mortality. Accumulating evidence demonstrates that microglia-driven neuroinflammation emerges as a central pathogenic mechanism underlying SAE. Here, we identify ovarian tumor deubiquitinase 1 (OTUD1) as a critical mediator of SAE pathogenesis. We demonstrate that OTUD1 promotes hexokinase 2 (HK2) dissociation from mitochondria via selective K63-linked deubiquitination, triggering microglia pyroptosis and neuroinflammation. Our findings address a key knowledge gap by elucidating the OTUD1-HK2 axis as a novel regulatory pathway in SAE, offering potential therapeutic targets to mitigate cognitive deficits in sepsis. Single-cell RNA sequencing was used to identify SAE-specific microglia subpopulations and analyze the expression of deubiquitinases within these subpopulations. OTUD1 knockout mice were generated to investigate the role of OTUD1 in SAE. Both wild-type and OTUD1 knockout mice were subjected to cecal ligation and puncture to induce SAE. In vitro, primary microglia and BV2 cells were treated with LPS and nigericin to simulate inflammatory conditions. Cognitive function of the mice was assessed through behavioral tests. Neuronal and synaptic damage were evaluated using HE and Nissl staining, as well as transmission electron microscopy. ELISA and qPCR were used to detect neuroinflammation. Western blot and immunofluorescence were employed to analyze protein expression. Molecular docking, 3D confocal microscopy, and co-immunoprecipitation were conducted to detect the interaction between OTUD1 and HK2. Finally, the correlation between OTUD1 and SAE was evaluated by analyzing clinical samples. Through single-cell RNA seq and subpopulation analysis, we identified an SAE-associated microglia (SAM) subpopulation with high expression of pyroptosis-related genes. Deubiquitinase expression analysis showed significantly elevated OTUD1 expression in SAM. OTUD1 deficiency attenuated neural damage and cognitive dysfunction in SAE mice in vivo. Further experiments revealed that OTUD1 regulates pyroptosis in microglia, affecting the progression of SAE. Mechanistically, OTUD1 directly binds to the C-terminal domain of HK2 through its Ala-rich domain and selectively cleaves K63-linked polyubiquitin chains on HK2 to promote the dissociation of HK2 from mitochondria, thereby activating the NLRP3 inflammasome and pyroptosis. In SAE, OTUD1 deubiquitinates HK2, promoting its dissociation from mitochondria, which triggers microglia pyroptosis, leading to neuronal damage and cognitive impairment.

Sepsis-induced acute lung injury (ALI) is a life-threatening condition with high mortality rates, and its underlying mechanisms remain poorly understood. This study investigates the role of TNF-α-induced protein 8-like 2 (TIPE2) in modulating PANoptosis, an integrated form of programmed cell death that includes apoptosis, necroptosis, and pyroptosis, in the context of sepsis-induced lung injury. We utilized a cecal ligation and puncture (CLP) mouse model to examine the effects of TIPE2 knockout and overexpression on lung injury, inflammation, and cell death pathways. Our findings demonstrate that TIPE2 knockout exacerbates lung injury by promoting the abnormal activation of PANoptosis-related proteins, leading to increased inflammation and tissue damage. In contrast, overexpression of TIPE2 in macrophages significantly reduces these effects by inhibiting the ZBP1-dependent PANoptosis pathway via TRIF signaling. These results highlight the crucial role of TIPE2 in maintaining the balance between cell survival and death during sepsis and suggest that targeting TIPE2 could be a novel therapeutic strategy for treating sepsis-related lung injury.

BACKGROUND:Sepsis-associated encephalopathy (SAE) refers to acute brain dysfunction caused by sepsis without direct central nervous system infection, in which microglia plays a pivotal role. Microglial ferroptosis is one of the key drivers of SAE. Dihydroartemisinin (DHA) is a natural product with anti-inflammatory effects and associated with ferroptosis. However, no current studies indicating that DHA plays a role in SAE or microglial ferroptosis, and further exploration is needed. OBJECTIVE:To investigate therapeutic effects of DHA on SAE induced by caecal ligation and puncture (CLP), and its regulation of microglial ferroptosis. METHODS:Network pharmacology, transcriptome sequencing, and bioinformatics were used to identify potential pathways and core DHA targets for SAE treatment. Molecular docking, molecular dynamics simulations (MDS) and Surface plasmon resonance (SPR) were performed to validate these targets. Models were created in vitro and in vivo utilizing lipopolysaccharide (LPS)-stimulated BV2 cells and CLP, respectively, to assess the therapeutic benefits of DHA. The in vivo therapeutic effectiveness of DHA was assessed using behavioral tests, survival analysis, and hematoxylin and eosin and Nissl staining. To validate important pathways and targets, RT-qPCR, immunofluorescence, Western blotting, ELISA, and flow cytometry were used. RESULTS:Network pharmacology identified 70 key therapeutic targets for AAL-SAE. After transcriptome sequencing analysis, 10 potential core targets of DHA for SAE treatment were identified. Molecular docking, MDS and SPR indicated DHA exhibited strong binding energy with HIF1A and formed a stable complex. Liquid chromatography-mass spectrometry (LC/MS) indicated DHA pass through the blood-brain barrier (BBB) to hippocampus and exert its effects. Animal experiments demonstrated that DHA improved the survival rate and alleviated sepsis scores, cognitive dysfunction, and neuroinflammation in SAE mice. DHA suppressed increased HIF1A and HMOX1 expression and reduced that of SLC7A11 and GPX4 in SAE mice hippocampus. Cell experiments revealed that DHA inhibited pro-inflammatory cytokine secretion and reduced migration in BV2 cells. Furthermore, DHA inhibited LPS-induced ferroptosis (evidenced by lipid peroxidation, Fe²⁺, and ROS levels) and mitochondrial dysfunction (evidenced by TMRE and mtDNA content). Additionally, DHA suppressed LPS-induced HIF1A and HMOX1 upregulation while promoting SLC7A11 and GPX4 downregulation in microglia. CONCLUSION:DHA alleviates cognitive dysfunction in SAE mice by reducing iron accumulation and mitochondrial dysfunction in hippocampal microglia through HIF1A/HMOX1 downregulation pathway and upregulation of SLC7A11/GPX4 pathway.

Sepsis-associated encephalopathy (SAE) is acute diffuse brain dysfunctional clinically caused by systemic infections originating outside the central nervous system (CNS), characterized by acute delirium, coma, and persistent cognitive dysfunction. Multiple studies have demonstrated a crucial role of microglia in the development of SAE. Bioinformatics analysis of the Human Protein Atlas and single-cell RNA sequencing datasets revealed that valosin-containing protein interaction protein 1 (VCPIP1) is expressed at the highest levels in hippocampal microglia, suggesting its potential involvement in neuroinflammatory regulation. In a murine cecal ligation and perforation (CLP) model of SAE, VCPIP1 was upregulated in hippocampal tissue. VCPIP1 deletion could exacerbate learning and memory impairment in CLP-induced conditions, along with suppressed microglial autophagy and increased proinflammatory cytokine production. Mechanistically, VCPIP1 deficiency activated the PI3K/AKT/mTOR signaling axis in microglia, thereby inhibiting autophagic flux. Inhibiting activation of PI3K by LY294002 reversed these effects, restoring autophagy, attenuating neuroinflammation, and mitigating neuronal injury. These results suggest that VCPIP1 improves autophagy by inhibiting the PI3K/AKT/mTOR pathway, highlighting its potential as a therapeutic target for SAE.

BackgroundSepsis refers to a systemic inflammatory response caused by infection, involving multiple organs. Sepsis-associated encephalopathy (SAE), as one of the most common complications in patients with severe sepsis, refers to the diffuse brain dysfunction caused by sepsis without central nervous system infection. However, there is no clear diagnostic criteria and lack of specific diagnostic markers.MethodsThe main active ingredients of coptidis rhizoma(CR) were identified from TCMSP and SwissADME databases. SwissTargetPrediction and PharmMapper databases were used to obtain targets of CR. OMIM, DisGeNET and Genecards databases were used to explore targets of SAE. Limma differential analysis was used to identify the differential expressed genes(DEGs) in GSE167610 and GSE198861 datasets. WGCNA was used to identify feature module. GO and KEGG enrichment analysis were performed using Metascape, DAVID and STRING databases. The PPI network was constructed by STRING database and analyzed by Cytoscape software. AutoDock and PyMOL software were used for molecular docking and visualization. Cecal ligation and puncture(CLP) was used to construct a mouse model of SAE, and the core targets were verified in vivo experiments.Results277 common targets were identified by taking the intersection of 4730 targets related to SAE and 509 targets of 9 main active ingredients of CR. 52 common DEGs were mined from GSE167610 and GSE198861 datasets. Among the 25,864 DEGs in GSE198861, LCN2 showed the most significant difference (logFC = 6.9). GO and KEGG enrichment analysis showed that these 52 DEGs were closely related to “inflammatory response” and “innate immunity”. A network containing 38 genes was obtained by PPI analysis, among which LCN2 ranked the first in Degree value. Molecular docking results showed that berberine had a well binding affinity with LCN2. Animal experiments results showed that berberine could inhibit the high expression of LCN2,S100A9 and TGM2 induced by CLP in the hippocampus of mice, as well as the high expression of inflammatory factors (TNFα, IL-6 and IL-1β). In addition, berberine might reduce inflammation and neuronal cell death by partially inhibiting NFκB/LCN2 pathway in the hippocampus of CLP models, thereby alleviating SAE.ConclusionOverall, Berberine may exert anti-inflammatory effects through multi-ingredients, multi-targets and multi-pathways to partially rescue neuronal death and alleviate SAE.

Macrophages are present in all tissues and body compartments under homeostatic physiological conditions. Importantly, they play a key role in pathological inflammatory processes when disturbed. They can quickly produce large amounts of inflammatory cytokines in response to danger signals. Macrophages can recognize muramyl dipeptide (MDP) through nucleotide-binding oligomerization domain (NOD)-like receptors, subsequently activating the NF-κB signaling pathway and producing proinflammatory cytokines. Erbin can bind to NOD2 and inhibit MDP-induced NF-κB activation, thus participating in the regulation of inflammatory response. Stabilizing or enhancing Erbin expression is essential for suppressing inflammatory responses. In this study, we used a deubiquitination enzyme plasmid library to screen for a key deubiquitinase, VCPIP1, which interacts with Erbin and influences its stability through deubiquitination modification. We investigated whether VCPIP1 affects inflammation using MDP-stimulated RAW 264.7 and BMDMs cells. The results showed that VCPIP1 deficiency reduced Erbin expression and increased NF-κB phosphorylation. Additionally, VCPIP1 deficiency promoted the release of inflammatory factors (IL-1β, IL-6, and TNF-α) in RAW 264.7 cells and BMDMs. This study further expands the role of deubiquitinases (DUBs) in inflammation, providing new insights for the prevention and treatment of sepsis, tumors, immune diseases, and other inflammatory reactions.

Metabolic disorder has been found to be an important factor in the pathogenesis and progression of sepsis. However, the causation of such an association between serum metabolites and sepsis has not been established. We conducted a two-sample Mendelian randomization (MR) study. A genome-wide association study of 486 human serum metabolites was used as the exposure, whereas sepsis and sepsis mortality within 28 days were set as the outcomes. In MR analysis, 6 serum metabolites were identified to be associated with an increased risk of sepsis, and 6 serum metabolites were found to be related to a reduced risk of sepsis. Furthermore, there were 9 metabolites positively associated with sepsis-related mortality, and 8 metabolites were negatively correlated with sepsis mortality. In addition, “glycolysis/gluconeogenesis” (p = 0.001), and “pyruvate metabolism” (p = 0.042) two metabolic pathways were associated with the incidence of sepsis. This MR study suggested that serum metabolites played significant roles in the pathogenesis of sepsis, which may provide helpful biomarkers for early disease diagnosis, therapeutic interventions, and prognostic assessments for sepsis.

Ovarian tumor domain-containing protease 1 (OTUD1) is a critical negative regulator that promotes innate immune homeostasis and is extensively involved in the pathogenesis of sepsis. In this study, we performed a powerful integration of multiomics analysis and an experimental mechanistic investigation to elucidate the immunoregulatory role of OTUD1 in sepsis at the clinical, animal and cellular levels. Our study revealed the upregulation of OTUD1 expression and the related distinctive alterations observed via multiomics profiling in clinical and experimental sepsis. Importantly, in vivo and in vitro, OTUD1 was shown to negatively regulate inflammatory responses and play a protective role in sepsis-induced pathological lung injury by mechanistically inhibiting the activation of the transforming growth factor-beta-activated kinase 1 (TAK1)-mediated mitogenactivated protein kinase (MAPK) and nuclear factor kappa-B (NF-0B) signaling pathways in the present study. Subsequently, we probed the molecular mechanisms underlying OTUD1 ' s regulation of NF-0B and MAPK pathways by pinpointing the target proteins that OTUD1 can deubiquitinate. Drawing upon prior research conducted in our laboratory, it has been demonstrated that tumor necrosis factor-a-induced protein 8-like 2 (TIPE2) performs a protective function in septic lung injury and septic encephalopathy by suppressing the NF-0B and MAPK pathways. Hence, we hypothesized that TIPE2 might be a target protein of OTUD1. Additional experiments, including Co-IP, immunofluorescence co-localization, and Western blotting, revealed that OTUD1 indeed has the ability to deubiquitinate TIPE2. In summary, OTUD1 holds potential as an immunoregulatory and inflammatory checkpoint agent, and could serve as a promising therapeutic target for sepsis-induced lung injury.

Ovarian tumor domain-containing protease 1 (OTUD1) is a critical negative regulator that promotes innate immune homeostasis and is extensively involved in the pathogenesis of sepsis. In this study, we performed a powerful integration of multiomics analysis and an experimental mechanistic investigation to elucidate the immunoregulatory role of OTUD1 in sepsis at the clinical, animal and cellular levels. Our study revealed the upregulation of OTUD1 expression and the related distinctive alterations observed via multiomics profiling in clinical and experimental sepsis. Importantly, in vivo and in vitro, OTUD1 was shown to negatively regulate inflammatory responses and play a protective role in sepsis-induced pathological lung injury by mechanistically inhibiting the activation of the transforming growth factor-beta-activated kinase 1 (TAK1)-mediated mitogen-activated protein kinase (MAPK) and nuclear factor kappa-B (NF-κB) signaling pathways in the present study. Subsequently, we probed the molecular mechanisms underlying OTUD1's regulation of NF-κB and MAPK pathways by pinpointing the target proteins that OTUD1 can deubiquitinate. Drawing upon prior research conducted in our laboratory, it has been demonstrated that tumor necrosis factor-α-induced protein 8-like 2 (TIPE2) performs a protective function in septic lung injury and septic encephalopathy by suppressing the NF-κB and MAPK pathways. Hence, we hypothesized that TIPE2 might be a target protein of OTUD1. Additional experiments, including Co-IP, immunofluorescence co-localization, and Western blotting, revealed that OTUD1 indeed has the ability to deubiquitinate TIPE2. In summary, OTUD1 holds potential as an immunoregulatory and inflammatory checkpoint agent, and could serve as a promising therapeutic target for sepsis-induced lung injury.

Sepsis-associated encephalopathy (SAE) is an acute brain dysfunction induced by systemic inflammation caused by sepsis and is one of the most common types of encephalopathy in intensive care units. Deteriorative neuroinflammation is closely related to the development of brain injury, which often transforms into common pathological manifestations in patients with severe sepsis. Therefore, taking necessary preventive and protective measures for potential brain injury and promptly reducing neuroinflammatory injury is necessary to improve the long-term prognoses of patients. Tumor necrosis factor-α-induced protein 8-like 2 (TIPE2) can play a significant protective role in septic lung injury, but studies on its expression and role in neurological diseases are rare. In the present study, we found that TIPE2 can expressed in microglia and ameliorate brain injury caused by SAE by suppressing neuroinflammation. The RhoA/ROCK2 pathway is the central coordinator of tissue injury response, and the activation of RhoA participates in the lipopolysaccharide-induced activation of the nuclear factor kappa B (NF-κB) signaling pathway. The activation of RhoA and phosphorylation of NF-κB was enhanced after TIPE2 deficiency. Importantly, TIPE2 negatively regulates inflammatory responses in vivo and in vitro and plays a protective role in SAE by inhibiting the activation of RhoA/ROCK2-NF-κB signaling pathways. The ultimate aim of our proposed project is to provide a theoretical basis for the development of a novel strategy for the early prevention and therapy of SAE.

Objective:To evaluate the role of tumor necrosis factor-alpha-induced protein-8 like-2(TIPE2) in sepsis-induced myocardial injury and the relationship with serine-threonine kinase(AKT)/glycogen synthase kinase-3β(GSK-3β)/β-catenin signaling pathway in mice.Methods:Sixteen male wild-type C57BL/6N mice and 16 TIPE2-gene knockout C57BL/6N mice, aged 6-8 weeks, with a body mass index of 20-25 g, were divided into 4 groups using a random number table method: wild-type+ sham operation group(group WT-sham), wild-type+ cecal ligation and perforation(CLP) group(group WT-CLP), TIPE2-gene knockout sham operation group(group KO-sham) and TIPE2-gene knockout CLP group(group KO-CLP), with 8 mice in each group. The model of myocardial injury induced by sepsis was developed by CLP in anesthetized animals. Blood samples from the inferior vena cava were collected at 24 h after surgery for determination of the concentrations of cardiac troponin I(cTnI) in serum by enzyme-linked immunosorbent assay. Then the mice were sacrificed and myocardial tissues were collected for determination of the pathological changes(by hematoxylin and eosin staining), expression of tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β) and IL-6 mRNA(by quantitative polymerase chain reaction), and expression of TIPE2, phosphorylated AKT(p-AKT), phosphorylated GSK-3β(p-GSK-3β) and β-catenin(by Western blot).Results:Compared with the corresponding Sham groups, the serum cTnI concentration was significantly increased, the expression of TNF-α, IL-1β and IL-6 mRNA and expression of p-AKT, p-GSK-3β and β-catenin in myocardial tissues were up-regulated, the expression of TIPE2 was down-regulated( P<0.05), and the pathological changes of myocardium were found in corresponding CLP groups. Compared with group WT-CLP, the serum cTnI concentration was significantly increased, the expression of TNF-α, IL-1β and IL-6 mRNA and expression of p-AKT, p-GSK-3β and β-catenin in myocardial tissues were up-regulated, the expression of TIPE2 was down-regulated( P<0.05), and the pathological changes of myocardium were aggravated in group KO-CLP( P<0.05). Conclusions:TIPE2 reduces the myocardial injury probably through inhibiting the AKT/GSK-3β/β-catenin signaling pathway in septic mice.

Microglia pyroptosis-mediated neuroinflammation is thought to be the crucial pathogenesis of sepsis-associated encephalopathy (SAE). Erbin has been reported to be associated with various inflammatory diseases. However, the role of Erbin in SAE and the relationship between Erbin and microglia pyroptosis are unknown. In this study, we investigated the promising role and underlying molecular mechanism of Erbin in the regulation of microglia pyroptosis. WT and Erbin knockout mice underwent cecum ligation perforation (CLP) to induce SAE. Primary mouse microglia and BV2 cells were treated with LPS/nigericin in vitro. Behavioral tests were performed to evaluate cognitive function. Nissl staining and transmission electron microscopy were used to assess histological and structural lesions. ELISA and qPCR were carried out to detect neuroinflammation. Western blot and immunofluorescence were used to analyze protein expression. Flow cytometry and confocal microscopy were utilized to observe the Ca2+ changes in the cytoplasm and endoplasmic reticulum (ER). To further explore the underlying mechanism, STF083010 was administered to block the IRE1α/Xbp1s pathway. Erbin deletion resulted in more pronounced neuronal damage and cognitive impairment in mice that underwent CLP. Erbin knockout promoted microglial pyroptosis and inflammatory cytokines secretion in vivo and in vitro, which was mediated by activation of the IRE1α/Xbp1s. Treatment with the selective inhibitor STF083010 significantly inhibited IRE1α/Xbp1s pathway activity, decreased intracytoplasmic Ca2+, attenuated microglial pyroptosis, reduced pro-inflammatory cytokine secretion, lessened neuronal damage, and improved cognitive function. In SAE, Erbin inhibits IRE1/Xbp1s pathway activity and reduces the ER Ca2+ influx to the cytoplasm, reducing microglial pyroptosis.

Objective:To evaluate the role of tumor necrosis factor-alpha-induced protein-8 like-2 (TIPE2) in endogenous protective mechanism of acute lung injury (ALI) in septic mice and the relationship with triggering receptor expressed on myeloid cells-1 (TREM-1)/NOD-like receptor protein 3 (NLRP3) inflammasome signaling pathway.Methods:Twenty male wild-type mice and 20 TIPE2 knockout mice were divided into 4 groups using a random number table method: wild-type+ sham operation group (group WT-sham), wild-type+ ALI group (group WT-ALI), TIPE2-knockout+ sham operation group (group KO-sham) and TIPE2-knockout+ CLP group (group KO-ALI), with 10 mice in each group.The ALI model was established by cecal ligation and perforation (CLP) in septic mice.Mice were sacrificed after blood samples were obtained from the abdominal aorta at 24 h after CLP, and lung tissue specimens were obtained for microscopic examination of pathological changes (with a light microscope) which were scored and for determination of wet/dry weight ratio (W/D ratio), myeloperoxidase (MPO) activity, expression of TIPE2, TREM-1, NLRP3, caspase-1 and GSDMD (by Western blot), and concentrations of interleukin-1beta (IL-1β) and IL-18 in serum (by enzyme-linked immunosorbent assay).Results:Compared with group WT-sham, the lung injury score, W/D ratio, MPO activity and concentrations of IL-1β and IL-18 in serum were significantly increased, the expression of TREM-1, NLRP3, caspase-1 and GSDMD was up-regulated, and the expression of TIPE2 was down-regulated in group WT-ALI and group KO-ALI ( P<0.05). Compared with group WT-ALI, the lung injury score, W/D ratio, MPO activity and concentrations of IL-1β and IL-18 in serum were significantly increased, the expression of TREM-1, NLRP3, caspase-1 and GSDMD was up-regulated, and the expression of TIPE2 was down-regulated in group KO-ALI ( P<0.05). Conclusion:TIPE2 is involved in endogenous protective mechanism of ALI in septic mice, which is related to inhibition of activation of TREM-1/NLRP3 inflammasome signaling pathway.

Objective:To evaluate the role of tumour necrosis factor-α-induced protein 8-like 2 (TIPE2) in the acute lung injury (ALI) induced by endotoxin in mice.Methods:Forty SPF healthy adult male BALB/c mice, aged 6-8 weeks, weighing 20-25 g, were divided into 4 groups ( n=10 each) using a random number table method: vehicle plasmid group (VP group), vehicle plasmid plus ALI group (VP+ ALI group), TIPE2 adeno-associated virus overexpression group (T group) and TIPE2 adeno-associated virus overexpression plus ALI group (T+ ALI group). The mice in VP and VP+ ALI groups were injected with empty adeno-associated virus, while the mice in T and T+ ALI groups were intratracheally given adeno-associated virus carrying TIPE interference sequence.Three weeks later, the model of endotoxin-induced ALI was established.Lipopolysaccharide (LPS) 5 mg/kg was intratracheally given in VP+ ALI and T+ ALI groups, and the equal volume of phosphate buffered saline (PBS) was given in VP and T groups.Blood samples were obtained from the abdominal aorta at 24 h after injection of LPS for blood gas analysis, oxygenation index (OI) was calculated, and tumor necrosis factor-alpha (TNF-α) in serum were detected by enzyme-linked immunosorbent assay.The animals were then sacrificed, and lung tissues were removed for examination of pathological changes which were scored after haematoxylin and eosin staining, for calculation of the wet/dry weight ratio (W/D ratio) and for determination of myeloperoxidase (MPO) activity and the expression of TIPE2, phosphorylated c-Jun N-terminal kinase (p-JNK) and nuclear factor kappa B(NF-κB) (by Western blot). Results:Compared with VP group, the lung injury score, W/D ratio, MPO activity and concentration of serum TNF-α were significantly increased, PaO 2 and OI were decreased, expression of TIPE2 was down-regulated and expression of p-JNK and NF-κB was up-regulated in VP+ ALI group ( P<0.05). Compared with VP+ ALI group, the lung injury score, W/D ratio, MPO activity and concentration of serum TNF-α were significantly decreased, PaO 2 and OI were increased, expression of TIPE2 was up-regulated and expression of p-JNK and NF-κB was down-regulated in T+ ALI group ( P<0.05). Conclusion:The down-regulation of TIPE2 expression is involved in the process of ALI induced by endotoxin in mice.

The aim of the present study was to investigate the effect of penehyclidine hydrochloride (PHC) pretreatment on mice with lipopolysaccharide (LPS)‑induced acute lung injury (ALI) and its possible underlying mechanisms. Mice were randomly separated into six groups: i) Sham group; ii) LPS group; iii) LPS + PHC group; iv) tumor necrosis factor a‑induced protein 8‑like protein 2 (TIPE2) group; v) LPS + TIPE2 group; and vi) LPS + TIPE2 + PHC group. The ALI model was induced using LPS through intratracheal injection. The mice received adenovirus gene to induce the overexpression of TIPE2. After mice were sacrificed, lung injury indices were assessed, and arterial blood, bronchoalveolar lavage fluid and lung tissues were collected for subsequent assays. Expression levels of related proteins were detected by using western blotting. It was found that compared with the sham group, the mice treated with LPS showed increased lung injury and dysfunctions of gas exchange. However, these trends were significantly ameliorated in the LPS + PHC group. Evaluation of protein expression in lung tissues showed that the increased expression of nuclear NF‑κB p65 and p‑c‑Jun N‑terminal kinase (JNK) induced by LPS were suppressed in the LPS + PHC group and the expression of TIPE2 was increased. The mice that received adenovirus gene to induce TIPE2 overexpression could also showed protective effects compared with the mice in the LPS group. However, the expression of TIPE2 decreased rather than increased in LPS group. In the mice pretreated with PHC, the expression of TIPE2 increased in mice with LPS‑induced ALI. To conclude, PHC pretreatment could inhibit the occurrence of inflammation and apoptosis in LPS‑induced ALI. This process may be related to the activation of TIPE2 and the inhibition of NF‑κB and JNK signaling pathway in the lungs of mice.

Objective:To evaluate the effects of esketamine on pyrolysis in lung tissues of rats with endotoxin-induced acute lung injury (ALI).Methods:SPF healthy adult male Sprague-Dawley rats, weighing 200-220 g, aged 8 weeks, were divided into 3 groups ( n=10 each) using a random number table method: control group (group C), endotoxin-induced ALI group (group ALI) and esketamine group (group E). Lipopolysaccharide (LPS) 10 mg/kg was intraperitoneally injected to establish the model of endotoxin-induced ALI model.The equal volume of 0.9% sodium chloride injection was intraperitoneally injected in group C. Esketamine 10 mg/kg was intraperitoneally injected at 30 min of injection of LPS in group E. Lung tissues were removed after blood samples were collected from hearts at 24 h after injection of LPS for determination of concentrations of serum interleukin-1beta (IL-1β) and IL-8 (by enzyme-linked immunosorbent assay), the wet/dry weight ratio (W/D ratio), activities of myeloperoxidase (MPO) (by colorimetric assay) and the expression of nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3 (NLRP3), caspase-1 and gasdermin D (GSDMD) (by Western blot) and for examination of pathological changes which were scored after haematoxylin and eosin staining and ultrastructure (using an electron microscope). Results:Compared with group C, the lung injury score, W/D ratio, MPO activity, expression of NLRP3, caspase-1 and GSDMD and concentrations of IL-1β and IL-18 in serum were significantly increased in ALI and E groups ( P<0.05). Compared with group ALI, the lung injury score, W/D ratio, MPO activity, expression of NLRP3, caspase-1 and GSDMD and concentrations of IL-1β and IL-18 in serum were significantly decreased in group E ( P<0.05). Conclusion:The mechanism by which esketamine reduces endotoxin-induced ALI is related with inhibition of pyrolysis in lung tissues of rats.