Mouse Model Of Sepsis Research Articles

Synergistic Enhancement of Isoforskolin and Dexamethasone Against Sepsis and Acute Lung Injury Mouse Models

Synergistic Enhancement of Isoforskolin and Dexamethasone Against Sepsis and Acute Lung Injury Mouse Models

Retracted] Resveratrol reduces acute lung injury in a LPS‑induced sepsis mouse model via activation of Sirt1.

Following the publication of this paper, it was drawn to the Editor's attention by a concerned reader that the β‑actin control western blotting data shown in Fig.3D on p.1893 were very similar to the contol data shown in Fig.4A on p.1894; furthermore, the data shown for the MMP‑9 and the INOS protein bands in Fig.4C were remarkably similar to the data shown for the IL‑1β and IL‑6 proteins, respectively, albeit the backgrounds surrounding the bands were different. Moreover, various of the western blotting data shown in these figures were strikingly similar to data that had already been published in different form in other articles written by (largely) different authors at different research institutes. Owing to the fact that the contentious data in the above article had already been published prior to its submission to Molecular Medicine Reports, and due to the number of apparent duplications of strikingly similar data between Figs.3 and 4, the Editor has decided that this paper should be retracted from the Journal. The authors were asked for an explanation to account for these concerns, but the Editorial Office did not receive a satisfactory reply. The Editor apologizes to the readership for any inconvenience caused. [Molecular Medicine Reports 7: 1889‑1895, 2013; DOI: 10.3892/mmr.2013.1444].

Platelet CD47 Maintains Circulating Monocyte Immune Homeostasis

In addition to their well-studied hemostatic functions, platelets are immune cells. Platelets circulate at the interface between the vascular wall and white blood cells, and transient platelet-leukocyte complexes are found in both healthy and disease states, positioning platelets to provide physiologic cues of both vascular health and injury. Roles for activated platelets in inducing and amplifying immune responses have received an increasing amount of research attention, but our past studies also showed that normal platelet counts are needed in healthy conditions to maintain immune homeostasis. Circulating monocytes are a part of the first line of host defenses that recognize and clear pathogens in part by releasing inflammatory cytokines, including IL-6, IL-8 and TNFα, that trigger a robust inflammatory reaction. However, this needs to be tightly regulated and in disease conditions like sepsis, dysregulated monocyte responses contribute to a “cytokine storm” that leads to exaggerated inflammation, tissue damage and potentially mortality. The cellular and molecular mechanisms that regulate monocyte immune phenotypes are complex and include roles for platelets. Considering the immune regulatory role of platelets and that thrombocytopenia is associated with worse sepsis outcomes, we hypothesize that resting platelets maintain monocyte immune homeostasis in healthy conditions and thrombocytopenia independently leads to monocyte dysregulation that exacerbates sepsis responses in a manner that is dependent on direct platelet-monocyte CD47 homotypic interactions that regulate monocyte immunometabolism and gene expression. Compared to monocytes from mice with normal platelet counts, circulating monocytes from thrombocytopenic mice had increased toll-like receptor (TLR) induced cytokine responses. Furthermore, ex vivo co-incubation of resting platelets with platelet naïve bone marrow monocytes, induced monocyte metabolic programming skewed toward glycolysis, consistent with our in vitro and in vivo RNA-Seq results. Enhanced glycolysis was mediated by AKT/mTOR pathway activation and resulted in durable changes in TLR agonist responses. Assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-Seq) on monocytes from thrombocytopenic mice showed open chromatin at LPS response genes, and resting platelet interactions with monocytes induced histone H3K9 and H3K27 trimethylation in a CD47 dependent manner. Using mouse models of thrombocytopenia and sepsis, we found that normal platelet numbers limited monocyte immune dysregulation and systemic inflammation. This was dependent on platelet CD47 as only WT platelets, not CD47 -/- platelet transfusion rescued the exaggerated inflammatory responses in thrombocytopenic mice (Fig 1). Both plasma IL-6 and IL-8 and circulating monocyte IL6 and IL8 gene expression from human patients with sepsis and non-septic conditions inversely correlated with patient platelet counts, further suggesting a resting platelet role in maintaining monocyte immune quiescence. Overall, our studies demonstrate that resting platelets maintain monocyte immune homeostasis by limiting TLR responses through regulating monocyte immunometabolic processes that lead to epigenetic changes in monocyte immune response genes (Fig 2). Thrombocytopenia therefore independently leads to immune dysregulation in sepsis, exacerbating systemic inflammatory processes. This is the first demonstration of sterile endogenous cell interactions that regulate innate immune-metabolism and monocyte pathogen responses.

Survival Advantage Associated with Heterozygous Factor V Leiden Mutation in Sars-Cov-2-Infected K18-hACE2 Mice

The pathogenesis of coronavirus disease 2019 (COVID-19) complications in critically ill patients involves virus-mediated cellular damage in infected tissue and secondary organ damage associated with a hyperinflammatory response. The proinflammatory milieu can result in endothelial dysfunction, predisposing patients to thrombosis leading to a life-threatening condition known as COVID-19-associated coagulopathy (CAC) which is driven by cellular and molecular mechanisms that are still unclear. We developed a mouse model based on the K18-hACE2 transgenic mouse recapitulating many of the CAC-associated pathological findings observed in human patients offering a reliable animal model for the study of SARS-CoV-2 pathogenesis. Plasma analysis revealed virus-induced upregulation of endothelial activation markers and alterations of all the measured coagulation factors' activities, resulting first in increased vascular leakage in the lung on day 5 post-infection, followed by the appearance of focal hemorrhagic lesions potentially associated with local thrombotic events on day 7. The infection-associated procoagulant phenotype appeared as early as day 3 post-infection (p.i.). Prothrombin time showed a 1/3 reduction in the normal clotting time in males. The overall procoagulant state was also reflected in the alteration of Factor(F) VIII and FX activity increasing to ~150% compared to uninfected controls. FIX showed the most significant alteration and was ~200% of the normal activity on day 3 p.i. and gradually increased over time, reaching ~350%. We also observed an increase in the activity of FV starting 3 days p.i. at ~210% of the normal level and persisting unchanged up to day 7 p.i. As reported during human infections, FV activity increased without significant changes in protein concentration or FVa levels. Natural anticoagulant and fibrinolytic mechanisms were dysregulated during virus infection resulting in a profound imbalance in thrombus formation and resolution. Infected K18-hACE2 mice showed downregulation of plasma protein C concentration which was not associated with any alteration of liver function, while plasma protein S concentration remained unaltered. Given the procoagulant environment generated after SARS-CoV-2 infection, reducing activated protein C (APC) anticoagulant and cytoprotective activities by blocking APC's active site with the SPC-54 monoclonal antibody worsened disease outcome and brought the mortality rate from 50% to 90%. In this observed procoagulant pathological context, where we observe increased activity of FV and where a deficiency of APC functions resulted in increased mortality, it wouldn't be surprising to observe an even more severe phenotype with the addition of APC resistance. To test this idea, we generated K18-hACE2 mice harboring the homologous human FV Leiden (FV L) mutation (R504Q). The Leiden mutation in FV eliminates one of several sites in FVa that are substrates for proteolysis by APC, resulting in diminished inactivation of FVa by APC and preventing the formation of the anticoagulant form of FV that serves as a cofactor for protein S-dependent inactivation of FVIIIa by APC. Upon SARS-CoV-2 infection, the presence of the homozygous FV L mutation in K18-hACE2 mice didn't affect the severity of the disease as measured by body weight loss during the course of the infection, and homozygous mutated mice had the same mortality rate as FV WT mice. Surprisingly, K18-hACE2 male mice carrying a heterozygous FV L mutation were extremely protected from SARS-CoV-2 infection, as their mortality dropped from 50% to only 15%. Histological analysis reveals how heterozygous FV L mice displayed a great reduction in immune infiltrate-associated fibrin deposition, suggesting an unexpected protective role of this prothrombotic mutation in the development of CAC. The protective effect of FV L heterozygosity may be attributable to enhanced thrombin formation which boosts inflammation and enhanced APC generation controlling proinflammatory and prothrombotic signaling, thus showing a delicate balance achieved by FV L heterozygosity that was lost for FV L homozygosity. As seen also in mouse models of bacterial sepsis, such favorable effects associated with FV L support the somewhat counterintuitive speculation that a partial increase in thrombin production in the early phase of the infection may support the resolution of COVID-19 complications.

ADENOSINE INFLUENCES FOXP3 EXPRESSION OF T REGS VIA THE A2AR/CREB PATHWAY IN A MOUSE MODEL OF SEPSIS.

The adenosine concentration and forkhead box protein (Foxp3) expression in T regulatory cells (T regs ) are increased during sepsis. However, the mechanism by which adenosine induces Foxp3 expression is incompletely understood. A cecal ligation and puncture (CLP) model was constructed using C57BL/J mice. The plasma adenosine concentration and Foxp3 expression in splenic T regs were increased consistently for 15 days after sepsis onset. Analysis of the mean fluorescence intensity of Foxp3 and adenosine concentration in the same mice revealed a linear correlation. In the CLP model, adenosine 2a receptor (A2aR) blockade inhibited Foxp3 expression in T regs . In vitro activation of A2aR promoted Foxp3 expression in T regs and facilitated secretion of extracellular vesicles. Transcriptome sequencing revealed that A2aR blockade led to changes in cyclic adenosine monophosphate response element-binding protein (CREB) transcription in T regs in our sepsis model. Use of adenosine or A2aR agonists promoted CREB expression, CREB phosphorylation at S133, T reg expression of Foxp3, and enhanced inhibition of proliferation of cluster of differentiation (CD)4+ lymphocytes. A2aR blockade or inhibition of CREB expression inhibited Foxp3 expression in T regs . In the CLP model, use of CREB inhibitors could inhibit Foxp3 expression and reduce the bacterial load. In summary, adenosine in sepsis promotes CREB phosphorylation via A2aR which, in turn, upregulates Foxp3 expression in T regs .

Apelin alleviates sepsis-induced acute lung injury in part by modulating the SIRT1/NLRP3 pathway to inhibit endothelial cell pyroptosis

BackgroundSepsis, an intricate systemic inflammatory syndrome, gives rise to various life-threatening complications, with acute lung injury (ALI) being prominently encountered. ALI, clinically characterized by pulmonary infiltration, hypoxemia, and edema, stands as a prevailing consequence of sepsis. This work sought to elucidate the mechanism of Apelin in mitigating sepsis-induced ALI (siALI). MethodsA mouse sepsis model was constructed by cecal ligation and puncture surgery, followed utilizing histopathological analysis using HE staining. mRNA levels of inflammatory cytokines (IL-1β, IL-6, and TNF-α) were assessed utilizing qRT-PCR, while ELISA was employed to measure the levels of vWF, VEGF, IL-1β, and IL-18. Western blot was conducted to examine protein levels of NLRP3, Caspase-1 p20, GSDMD-N, and SIRT1. To evaluate the extent of endothelial cell (EC) pyroptosis, immunofluorescence co-staining of CD31, NLRP3, and Caspase-1 p20 was fulfilled. Furthermore, TUNEL staining was utilized to ascertain the degree of plasma membrane damage and cell death. ResultsApelin demonstrated its potential in ameliorating siALI in mice by diminishing mRNA expression levels of pro-inflammatory cytokines (IL-1β, IL-6, TNF-α) as well as expression levels of vWF and VEGF. Apelin inhibited protein expression of NLRP3, Caspase-1 p20, and GSDMD-N, indicating that EC pyroptosis was suppressed. Finally, Apelin could upregulate the protein expression of SIRT1. This upregulation led to the inhibition of protein expression of NLRP3, Caspase-1 p20, and GSDMD-N, consequently suppressing EC pyroptosis. As a result, a reduction in the expression of inflammatory cytokines IL-1β and IL-18 ultimately alleviated siALI. ConclusionApelin was confirmed to alleviate siALI partially by modulating SIRT1/NLRP3 pathway to inhibit EC pyroptosis, which dawned on the molecular mechanism of siALI and had important clinical significance for treating ALI effectively.

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.

Multimodal nanoparticle-containing modified suberoylanilide hydroxamic acid polymer conjugates to mitigate immune dysfunction in severe inflammation.

Excessive immune activation and immunosuppression are opposing factors that contribute to the dysregulated innate and adaptive immune responses seen in severe inflammation and sepsis. Here, a novel analog of the histone deacetylase inhibitor (HDACi), suberoylanilide hydroxamic acid (SAHA-OH), was incorporated into immunomodulatory poly(lactic acid)-based nanoparticles (iNP-SAHA) by employing a prodrug approach through the covalent modification of poly(lactic-co-glycolic acid) (PLGA) with SAHA-OH. iNP-SAHA formulation allowed for controlled incorporation and delivery of SAHA-OH from iNP-SAHA and treatment led to multimodal biological responses including significant reductions in proinflammatory cytokine secretions and gene expression, while increasing the survival of primary macrophages under lipopolysaccharide (LPS) challenge. Using a lethal LPS-induced endotoxemia mouse model of sepsis, iNP-SAHA administration improved the survival of mice in a dose-dependent manner and tended to improve survival at the lowest doses compared to iNP control. Further, iNP-SAHA reduced the levels of plasma proinflammatory cytokines and chemokines associated with sepsis more significantly than iNP and similarly improved inflammation-induced spleen and liver toxicity as iNP, supporting its potential polypharmacological activity. Collectively, iNP-SAHA offers a potential drug delivery approach to modulate the multifaceted inflammatory responses observed in diseases such as sepsis.

Sepsis leads to lasting changes in phenotype and function of naïve CD8 T cells.

Sepsis, an amplified immune response to systemic infection, is characterized by a transient cytokine storm followed by chronic immune dysfunction. Consequently, sepsis survivors are highly susceptible to newly introduced infections, suggesting sepsis can influence the function and composition of the naïve CD8 T cell pool and resulting pathogen-induced primary CD8 T cell responses. Here, we explored the extent to which sepsis induces phenotypic and functional changes within the naïve CD8 T cell pool. To interrogate this, the cecal ligation and puncture (CLP) mouse model of polymicrobial sepsis was used. In normal, non-septic mice, we show type-I interferon (IFN I)-mediated signaling plays an important role in driving the phenotypic and functional heterogeneity in the naïve CD8 T cell compartment leading to increased representation of Ly6C+ naïve CD8 T cells. In response to viral infection after sepsis resolution, naïve Ly6C+ CD8 T cells generated more primary effector and memory CD8 T cells with slower conversion to a central memory CD8 T cell phenotype (Tcm) than Ly6C- naïve CD8 T cells. Importantly, as a potent inducer of cytokine storm and IFN I production, sepsis leads to increased representation of Ly6C+ naïve CD8 T cells that maintained their heightened ability to respond (i.e., effector and memory CD8 T cell accumulation and cytokine production) to primary LCMV infection. Lastly, longitudinal analyses of peripheral blood samples obtained from septic patients revealed profound changes in CD8 T cell subset composition and frequency compared to healthy controls. Thus, sepsis has the capacity to alter the composition of naïve CD8 T cells, directly influencing primary CD8 T cell responses to newly introduced infections.

Fangji Fuling Decoction Alleviates Sepsis by Blocking MAPK14/FOXO3A Signaling Pathway.

To examine the therapeutic effect of Fangji Fuling Decoction (FFD) on sepsis through network pharmacological analysis combined with in vitro and in vivo experiments. A sepsis mouse model was constructed through intraperitoneal injection of 20 mg/kg lipopolysaccharide (LPS). RAW264.7 cells were stimulated by 250 ng/mL LPS to establish an in vitro cell model. Network pharmacology analysis identified the key molecular pathway associated with FFD in sepsis. Through ectopic expression and depletion experiments, the effect of FFD on multiple organ damage in septic mice, as well as on cell proliferation and apoptosis in relation to the mitogen-activated protein kinase 14/Forkhead Box O 3A (MAPK14/FOXO3A) signaling pathway, was analyzed. FFD reduced organ damage and inflammation in LPS-induced septic mice and suppressed LPS-induced macrophage apoptosis and inflammation in vitro (P<0.05). Network pharmacology analysis showed that FFD could regulate the MAPK14/FOXO signaling pathway during sepsis. As confirmed by in vitro cell experiments, FFD inhibited the MAPK14 signaling pathway or FOXO3A expression to relieve LPS-induced macrophage apoptosis and inflammation (P<0.05). Furthermore, FFD inhibited the MAPK14/FOXO3A signaling pathway to inhibit LPS-induced macrophage apoptosis in the lung tissue of septic mice (P<0.05). FFD could ameliorate the LPS-induced inflammatory response in septic mice by inhibiting the MAPK14/FOXO3A signaling pathway.

GENIPOSIDE IMPROVES CLP-INDUCED SEPSIS MODEL PROGNOSIS BY UPREGULATING PPARγ TO MODULATE MONOCYTE PHENOTYPE AND CYTOKINE NETWORK.

Background : We explored the efficacy and main biological mechanism of geniposide intervention in sepsis. Methods : A sepsis model was established in male BALB/c mice through cecal ligation and puncture (CLP). Different doses of geniposide (20 or 40 mg/kg) were administered intravenously at 0 and/or 24 h after CLP surgery. The survival rate of different groups was observed. In addition, the expression levels of CD16 and major histocompatibility complex class II in monocytes were assessed using flow cytometry. The concentrations of TNF-α, IL-1β, IL-6, and IL-10 in the serum were measured by ELISA. We also observed the biological effects of geniposide on CD16 and MHC-II expression levels in RAW264.7 cells, as well as the secretion of TNF-α, IL-1β, IL-6, and IL-10 in the LPS-induced RAW264.7 cell model. The PPARγ levels were determined using western blot analysis. Results : Intravenous administration of 40 mg/kg of geniposide at 0 h after CLP significantly improved the survival outcomes in the septic mouse model, with no significant benefits from low dosing (20 mg/kg) or delayed administration (24 h). The effective dose of geniposide significantly decreased the serum cytokine TNF-α, IL-1β, IL-6, and IL-10 concentrations in septic mice ( P < 0.05). Notably, in vitro assays showed that geniposide specifically increased the IL-10 level. Geniposide significantly reduced the CD16 expression ( P < 0.05) and increased MHC-II expression in monocytes ( P < 0.05). In addition, geniposide elevated the PPARγ level in monocytes ( P < 0.05). Conclusions : High-dose early-stage geniposide administration significantly improved the survival rate in a CLP mouse sepsis model by modulating the monocyte phenotype and regulating the cytokine network (IL-6/IL-10 levels). The pharmacological mechanism of geniposide action might be exerted primarily through PPARγ upregulation.

CD137L Inhibition Ameliorates Hippocampal Neuroinflammation and Behavioral Deficits in a Mouse Model of Sepsis-Associated Encephalopathy

Anxiety manifestations and cognitive dysfunction are common sequelae in patients with sepsis-associated encephalopathy (SAE). Microglia-mediated inflammatory signaling is involved in anxiety, depression, and cognitive dysfunction during acute infection with bacterial lipopolysaccharide (LPS). However, the molecular mechanisms underlying microglia activation and behavioral and cognitive deficits in sepsis have not been in fully elucidated. Based on previous research, we speculated that the CD137 receptor/ligand system modulates microglia function during sepsis to mediate classical neurological SAE symptoms. A murine model of SAE was established by injecting male C57BL/6 mice with LPS, and cultured mouse BV2 microglia were used for in vitro assays. RT-qPCR, immunofluorescence staining, flow cytometry, and ELISA were used to assess microglial activation and the expression of CD137L and inflammation-related cytokines in the mouse hippocampus and in cultured BV2 cells. In addition, behavioral tests were conducted in assess cognitive performance and behavioral distress. Immunofluorescence and RT-qPCR analyses showed that hippocampal expression of CD137L was upregulated in activated microglia following LPS treatment. Pre-treatment with the CD137L neutralizing antibody TKS-1 significantly reduced CD137L levels, attenuated the expression of M1 polarization markers in microglia, and inhibited the production of TNF-α, IL-1β, and IL-6 in both LPS-treated mice and BV2 cells. Conversely, stimulation of CD137L signaling by recombinant CD137-Fc fusion protein activated the synthesis and release of pro-inflammatory cytokines in cultures BV2 microglia. Importantly, open field, elevated plus maze, and Y-maze spontaneous alternation test results indicated that TKS-1 administration alleviated anxiety-like behavior and spatial memory decline in mice with LPS-induced SAE. These findings suggest that CD137L upregulation in activated microglia critically contributes to neuroinflammation, anxiety-like behavior, and cognitive dysfunction in the mouse model of LPS-induced sepsis. Therefore, therapeutic modulation of the CD137L/CD137 signaling pathway may represent an effective way to minimize brain damage and prevent cognitive and emotional deficits associated with SAE.

Effect of cogon grass root ethanol extract on fatty acid binding protein 4 and oxidative stress markers in a sepsis mouse model.

Background: Sepsis causes several immunological and metabolic alterations that induce oxidative stress. The modulation of fatty acid-binding protein 4 (FABP4) has been shown to worsen this condition. Extract of cogon grass root (ECGR) contains flavonoids and isoeugenol compounds that exhibit anti-inflammatory and antioxidant properties. This study aimed to assess the effects of ECGR on FABP4 and oxidative stress-related factors in a sepsis mouse model. Methods: Twenty-nine male mice ( Mus musculus) of the Deutsche Denken Yoken strain were divided into four groups: group 1, control; group 2, mice treated with 10 μL/kg body weight (BW) lipopolysaccharide (LPS); and groups 3 and 4, mice pre-treated with 90 and 115 mg/kg BW, respectively, and then treated with 10 μL/kg BW LPS for 14 d. Blood, liver, lymph, and cardiac tissue samples were collected and subjected to histological and complete blood examinations. Antioxidant (Glutathione peroxidase 3 (GPx3) and superoxide dismutase), FABP4 levels, and immune system-associated biomarker levels (TNF-α, IL-6 and IL-1β) were measured. Results: Significant increases in platelet levels (p = 0.03), cardiomyocyte counts (p =0.004), and hepatocyte counts (p = 0.0004) were observed in group 4 compared with those in group 2. Conversely, compared with those in group 2, there were significant decreases in TNF-α expression in group 3 (p = 0.004), white pulp length and width in group 4 (p = 0.001), FABP4 levels in groups 3 and 4 (p = 0.015 and p = 0.012, respectively), lymphocyte counts in group 4 (p = 0.009), and monocyte counts (p = 0.000) and polymorphonuclear cell counts in the livers (p = 0.000) and hearts (p = 0.000) of groups 3 and 4. Gpx3 activity was significantly higher in group 3 than in group 1 (p = 0.04). Conclusions: ECGR reduces FABP4 level and modulating oxidative stress markers in sepsis mouse model.

Potentials of organic tellurium-containing compound AS101 to overcome carbapenemase-producing Escherichia coli.

The issue of carbapenem-resistant Escherichia coli was aggravated yearly. The previous studies reported the varied but critical epidemiology of carbapenem-resistant E. coli among which the carbapenemase-producing strains were regarded as one of the most notorious issues. AS101, an organic tellurium-containing compound undergoing clinical trials, was revealed with antibacterial activities. However, little is known about the antibacterial effect of AS101 against carbapenemase-producing E. coli (CPEC). The minimum inhibitory concentration (MIC) of AS101 against the 15 isolates was examined using a broth microdilution method. The scanning electron microscopy, pharmaceutical manipulations, reactive oxygen species level, and DNA fragmentation assay were carried out to investigate the antibacterial mechanism. The sepsis mouse model was employed to assess the invivo treatment effect. The blaNDM (33.3%) was revealed as the dominant carbapenemase gene among the 15 CPEC isolates, followed by the blaKPC gene (26.7%). The MICs of AS101 against the 15 isolates ranged from 0.5 to 32μg/ml, and 99.9% of bacterial eradication was observed at 8h, 4h, and 2h for 1×, 2×, and 4×MIC, respectively. The mechanistic investigations suggest that AS101 would enter the bacterial cell, and induce ROS generation, leading to DNA fragmentation. The invivo study exhibited that AS101 possessed a steady treatment effect in a sepsis mouse model, with an up to 83.3% of survival rate. The invitro activities, mechanisms, and invivo study of AS101 against CPEC were unveiled. Our finding provided further evidence for the antibiotic development of AS101.

Single-cell RNA sequencing reveals the effects of capsaicin in the treatment of sepsis-induced liver injury.

Sepsis is a difficult-to-treat systemic condition in which liver dysfunction acts as both regulator and target. However, the dynamic response of diverse intrahepatic cells to sepsis remains poorly characterized. Capsaicin (CAP), a multifunctional chemical derived from chilli peppers, has recently been shown to potentially possess anti-inflammatory effects, which is also one of the main approaches for drug discovery against sepsis. We performed single-cell RNA transcriptome sequencing on 86,830 intrahepatic cells isolated from normal mice, cecal ligation and puncture-induced sepsis model mice and CAP-treated mice. The transcriptional atlas of these cells revealed dynamic changes in hepatocytes, macrophages, neutrophils, and endothelial cells in response to sepsis. Among the extensive crosstalk across these major subtypes, KC_Cxcl10 shared strong potential interaction with other cells when responding to sepsis. CAP mitigated the severity of inflammation by partly reversing these pathophysiologic processes. Specific cell subpopulations in the liver act collectively to escalate inflammation, ultimately causing liver dysfunction. CAP displays its health-promoting function by ameliorating liver dysfunction induced by sepsis. Our study provides valuable insights into the pathophysiology of sepsis and suggestions for future therapeutic gain.

PD-L1 Blockade Improves Survival in Sepsis by Reversing Monocyte Dysfunction and Immune Disorder

Monocyte dysfunction is critical to sepsis-induced immunosuppression. Programmed death ligand-1 (PD-L1) has shown a close relationship with inflammatory disorder among animal models and patients. We aimed to investigate the potential beneficial immunologic mechanisms of anti-PD-L1 on monocyte dysfunction of mice with sepsis. Firstly, we assessed the potential association between PD-L1 expression on monocyte subsets and sepsis severity as well as 28-day mortality. In this study, 52 septic patients, 28 septic shock patients, and 40 healthy controls were enrolled and their peripheral whole blood was examined by flow cytometry. Then, cecal ligation and puncture (CLP) were performed for establishing the mouse sepsis model. Subsequently, effects of anti-PD-L1 antibody on monocyte subset, major histocompatibility complex II (MHC II) expression, cytokine production, and survival were investigated. PD-L1 expression on the classical monocytes (CD14 + + CD16 −) was significantly upregulated among septic shock patients and the 28-day death group than non-septic shock group and 28-day survival group (P < 0.05). Compared to septic mice, anti-PD-L1-treated mice had significantly elevated percentages of major histocompatibility complex (MHC) II on peripheral Ly6chi monocyte at 24 h after CLP. Our results showed that the anti-PD-L1 antibody markedly decreased the level of serum inflammatory cytokines interleukin (IL)-6, tumor necrosis factor (TNF)-α, and IL-10 in sepsis mice at 24 h, 48 h, and 72 h, respectively (P < 0.05). The survival rate of CLP mice was significantly improved by anti-PD-L1 antibody treatment. Classical monocytes with high expression of PD-L1 were thought to be connected with sepsis progression. The PD-L1 blockade protects from sepsis, at least partially by inhibiting the reversal of monocyte dysfunction.

DLK1 overexpression improves sepsis-induced cardiac dysfunction and fibrosis in mice through the TGF-β1/Smad3 signaling pathway and MMPs.

Sepsis is a serious inflammatory disease caused by bacterial infection. Cardiovascular dysfunction and remodeling are serious complications of sepsis, which can significantly affect sepsis patients' mortality. Delta-like homologue 1 (DLK1) has been reported could inhibit cardiac myofibroblast differentiation. However, the function of DLK1 in sepsis is unknown. In the present study, the DLK1 expression was first identified based on the online dataset GSE79962 analysis and cecal ligation and puncture (CLP)-induced sepsis mouse model. DLK1 expression was significantly reduced in septic heart tissues. In septic mouse heart, CLP operation decreased the fractional shortening (EF) (%) and ejection fraction (FS) (%) and caused significant edema, disordered myofilament arrangement, and degradation and necrosis in myocardial cells; CLP operation also increased collagen deposition and elevated the protein levels of fibrotic markers (α-SMA and F-actin). DLK1 overexpression in septic mice could effectively increase EF (%) and FS (%), attenuate CLP-caused ECM degradation and deposition and partially inhibit the CLP-induced TGF-β1/Smad signaling activation. In conclusion, DLK1 expression was poorly expressed in the CLP-induced septic mouse heart. DLK1 overexpression partially alleviated sepsis-induced cardiac dysfunction and fibrosis, with the involvement of the TGF-β1/Smad3 signaling pathway and MMPs.

Icilin, a cool/cold-inducing agent, alleviates lipopolysaccharide-induced septic sickness responses in mice

Sepsis is a significant global public health challenge, resulting in millions of human deaths annually. Transient receptor potential melastatin 8 (TRPM8), a non-selective ion channel, is the primary cold sensor in humans; however, its effects on endotoxin-induced inflammation remain unclear. We previously reported that TRPM8 knockout mice exhibited more severe physiological and behavioral endotoxemia responses upon a high-dose injection with lipopolysaccharide (LPS). In the present study, we investigated whether icilin, a TRPM8 agonist, was a target for the suppression of sickness responses using a mouse model of LPS-induced sepsis. A peripheral high-dose injection of LPS at 5 mg/kg showed a maximal body temperature decrease of 5.1 °C in mice subcutaneously pretreated with vehicle and 1.5 °C in icilin-pretreated animals. The decline in locomotor activity was attenuated in icilin-pretreated mice and its recovery was faster; however, the high-dose LPS injection rapidly decreased locomotor activity regardless of the icilin pretreatment. Furthermore, the icilin pretreatment attenuated LPS-induced decreases in body weight and food and water intakes and accelerated recovery from these sickness responses. Therefore, the present results demonstrated that the icilin pretreatment alleviated LPS-induced sickness responses or decreases in body temperature, locomotor activity, body weight loss, and food and water intakes, suggesting its potential as a therapeutic target for sepsis.

TCF7 and LEF-1 downregulation in sepsis promotes immune suppression by inhibiting CD4+ T cell proliferation

BackgroundPrevious studies have shown that sepsis is implicated in a reduction in the number and function of CD4+ T cells. TCF7 and LEF-1 facilitate early T cell development and lineage selection of CD4+ T cells. However, the function and mechanism of TCF7 and LEF-1 in sepsis are uncharacterized. This study intended to delineate effect of TCF7 and LEF-1 on sepsis and the impact on proliferation of CD4+ T cells in sepsis. MethodsA mouse sepsis model was constructed by cecal ligation and puncture (CLP) method. Expression of TCF7 and LEF-1 in sepsis was investigated using bioinformatics analysis and molecular experiments. We then constructed TCF7 and LEF-1 overexpression cell lines to investigate their effects on proliferation, apoptosis, effector activation, and immunosuppressive molecules of CD4+ T cells in sepsis. ResultsTCF7 and LEF-1 were downregulated in sepsis. As the duration of sepsis induction increased, the levels of TCF7 and LEF-1 gradually decreased, as did the number of CD4+ T cells. Cell experiments showed that overexpression of TCF7 and LEF-1 enhanced proliferation and effector activation of CD4+ T cells, reduced apoptosis, decreased PD-1 and LAG3 expression, and promoted immune response in sepsis. ConclusionIn conclusion, this study confirmed that downregulation of TCF7 and LEF-1 expression in sepsis inhibited proliferation of CD4+ T cells, leading to immune suppression. This finding suggested that TCF7 and LEF-1 were potential biological targets for sepsis and indicated that immunotherapy aimed at improving CD4+ T cell proliferation may be a new strategy for immune therapy in sepsis patients.

The effect of artesunate to reverse CLP-induced sepsis immunosuppression mice with secondary infection is tightly related to reducing the apoptosis of T cells via decreasing the inhibiting receptors and activating MAPK/ERK pathway

T cells play an important role in regulating immune system balance. Sepsis-associated immunosuppression causes apoptosis of T cells and a decrease in their number. Previously, artesunate was found to have an immunomodulatory effect on immunosuppression in model mice with cecal ligation and puncture (CLP)-induced sepsis. In the present study, mouse sepsis models of CLP and CLP with secondary infection were established and treated with artesunate in order to examine the effect of artesunate on adaptive immune response in sepsis-related immunosuppression. The results showed that artesunate treatment could increase the survival rate of CLP mice with secondary Pseudomonas aeruginosa infection, increase the bacterial clearance rate, and also increase the level of the pro-inflammatory cytokine TNF-α. In addition, artesunate resulted in an increase in the number of T cells, CD4+ T cells and CD8+ T cells, and inhibited CD4+ and CD8+ T-cell apoptosis. Artesunate was also found to inhibit the expression of the inhibitory receptors of PD-1, CTLA-4, and BTLA, but it did not affect the expression of Tim-3. Additionally, artesunate significantly increased the phosphorylated ERK level of CD4+ T cells and CD8+ T cells and inhibited mitochondrial pathway-mediated apoptosis in CLP mice with Pseudomonas aeruginosa infection. These findings reveal that artesunate has an immunomodulatory effect on the adaptive immune response in sepsis. These effects include an increase in the numbers of T cells, CD4+ T cells, and CD8+ T cells through inhibition of the expression of inhibitory receptors and promotion of the MAPK/ERK pathway.