Neutrophils disrupt B-1a cell homeostasis by targeting Siglec-G to exacerbate sepsis.

B-1a cells, an innate-like cell population, are crucial for pathogen defense and the regulation of inflammation through their release of natural IgM and IL-10. In sepsis, B-1a cell numbers are decreased in the peritoneal cavity as they robustly migrate to the spleen. Within the spleen, migrating B-1a cells differentiate into plasma cells, leading to alterations in their original phenotype and functionality. We discovered a key player, sialic acid-binding immunoglobulin-like lectin-G (Siglec-G), which is expressed predominantly on B-1a cells and negatively regulates B-1a cell migration to maintain homeostasis. Siglec-G interacts with CXCR4/CXCL12 to modulate B-1a cell migration. Neutrophils aid B-1a cell migration via neutrophil elastase (NE)-mediated Siglec-G cleavage. Human studies revealed increased NE expression in septic patients. We identified an NE cleavage sequence in silico, leading to the discovery of a decoy peptide that protects Siglec-G, preserves peritoneal B-1a cells, reduces inflammation, and enhances sepsis survival. The role of Siglec-G in inhibiting B-1a cell migration to maintain their inherent phenotype and function is compromised by NE in sepsis, offering valuable insights into B-1a cell homeostasis. Employing a small decoy peptide to prevent NE-mediated Siglec-G cleavage has emerged as a promising strategy to sustain peritoneal B-1a cell homeostasis, alleviate inflammation, and ultimately improve outcomes in sepsis patients.

Self-objectification in Chinese pregnant women: The mixed role of functionality appreciation.

Extensions of objectification theory to pregnant women are few and continued research is needed to better understand the psychological consequences of significant changes to physical appearance during pregnancy. Specific interests in this area include functionality appreciation which may be particularly relevant to pregnancy. Research in this area is also lacking representation of non-Western cultural contexts. To this end, we employed an online survey to assess objectification theory and functionality appreciation in Chinese pregnant women (N = 345). Correlations showed that higher body surveillance and body shame were associated with higher disordered eating and psychological distress, and higher functionality appreciation was associated with lower body surveillance, body shame, and disordered eating. Mediation analyses suggested that higher body surveillance was associated with higher body shame which, in turn, was associated with higher disordered eating and psychological distress. Main effects suggested a negative association between functionality appreciation and body shame, but moderation analyses suggested that higher functionality appreciation strengthened the positive association between body surveillance and body shame. Findings underscore objectification theory as a useful framework to understand eating and body image disturbances and psychological distress in Chinese pregnant women and outline future directions to clarify the temporal nature of these associations and the precise role of functionality appreciation.

B-1a Cells Scavenge NETs to Attenuate Sepsis.

B-1a cells, a regulatory subset of B lymphocytes, produce natural IgM and IL-10. Neutrophil extracellular traps (NETs) play a crucial role in pathogen defense, but their excessive formation during sepsis can cause further inflammation and tissue damage. In sepsis, extracellular cold-inducible RNA-binding protein (eCIRP), a damage-associated molecular pattern, is released to induce NET formation. We hypothesize that B-1a cells clear NETs to prevent sepsis-induced injury. Sepsis in mice was induced by injecting 1 × 107 and 5 × 107 CFU E. coli intraperitoneally (i.p.). After 4 and 20 hours, we assessed the number of B-1a cells in the peritoneal cavity using flow cytometry. Our results showed that the number of peritoneal B-1a cells was significantly decreased in E. coli-sepsis mice. Importantly, replenishing B-1a cells via i.p. injection in sepsis mice significantly decreased NETs in peritoneal neutrophils. We also observed a decrease in serum inflammation and injury markers and a significant increase in overall survival rate in B-1a cell-treated septic mice. To understand the mechanism, we co-cultured bone marrow-derived neutrophils (BMDNs) with peritoneal B-1a cells in a contact or non-contact condition using an insert and stimulated them with eCIRP. After 4 hours, we found that eCIRP significantly increased NET formation in BMDNs. Interestingly, we observed that B-1a cells inhibited NETs by 67% in a contact-dependent manner. Surprisingly, when B-1a cells were cultured in inserts, there was no significant decrease in NET formation, suggesting that direct cell-to-cell contact is crucial for this inhibitory effect. We further determined that B-1a cells promoted NET phagocytosis and this was mediated through natural IgM, as blocking IgM receptor attenuated the engulfment of NETs by B-1a cells. Finally, we identified that following their engulfment, NETs were localized into the lysosomal compartment for lysis. Thus, our study suggests that B-1a cells decrease NET content in eCIRP-treated neutrophils and E. coli-sepsis mice.

A novel opsonic eCIRP inhibitor for lethal sepsis.

Sepsis is a life-threatening inflammatory condition partly orchestrated by the release of various damage-associated molecular patterns such as extracellular cold-inducible RNA-binding protein (eCIRP). Despite advances in understanding the pathogenic role of eCIRP in inflammatory diseases, novel therapeutic strategies to prevent its excessive inflammatory response are lacking. Milk fat globule-epidermal growth factor-VIII (MFG-E8) is critical for the opsonic clearance of apoptotic cells, but its potential involvement in the removal of eCIRP was previously unknown. Here, we report that MFG-E8 can strongly bind eCIRP to facilitate αvß3-integrin-dependent internalization and lysosome-dependent degradation of MFG-E8/eCIRP complexes, thereby attenuating excessive inflammation. Genetic disruption of MFG-E8 expression exaggerated sepsis-induced systemic accumulation of eCIRP and other cytokines, and consequently exacerbated sepsis-associated acute lung injury. In contrast, MFG-E8-derived oligopeptide recapitulated its eCIRP binding properties, and significantly attenuated eCIRP-induced inflammation to confer protection against sepsis. Our findings suggest a novel therapeutic approach to attenuate eCIRP-induced inflammation to improve outcomes of lethal sepsis.

Picky eating is associated with lower life satisfaction and elevated psychological distress and psychosocial impairment in Chinese pregnant women.

OBJECTIVE: This study assessed picky eating in pregnant women by exploring whether picky eating is associated with pregnant women's well-being, including life satisfaction, psychological distress, and psychosocial impairment. METHOD: Data collected were from 345 Chinese pregnant women ( M age = 29.95 years, SD = 5.58). Pearson correlation analyses were conducted to examine zero-order correlations between picky eating and well-being variables (i.e., life satisfaction, psychological distress, and psychosocial impairment). Hierarchical multiple regressions were conducted to examine the unique associations of picky eating with well-being variables, adjusting for demographic and pregnancy-related characteristics and thinness-oriented disordered eating. RESULTS: Picky eating was significantly and negatively correlated with life satisfaction (r = -.24, p < .001) and positively correlated with psychological distress (r = .37, p < .001) and psychosocial impairment (r = .50, p < .001). When adjusting for covariates and thinness-oriented disordered eating, picky eating was still significantly associated with lower life satisfaction, higher psychological distress, and higher psychosocial impairment. DISCUSSION: The findings suggest that picky eating may be a significant correlate of pregnant women's poorer well-being. Future research with longitudinal designs is warranted to further examine the temporal associations between picky eating and pregnant women's well-being. PUBLIC SIGNIFICANCE: Picky eating behaviors are poorly understood in pregnant women. Our results revealed that higher picky eating behaviors were associated with lower life satisfaction and higher psychological distress and psychosocial impairment in Chinese pregnant women. Researchers and clinicians may consider picky eating in the assessment and treatment of mental health and disordered eating in pregnant women.

An engineered miRNA PS-OMe miR130 inhibits acute lung injury by targeting eCIRP in sepsis.

BACKGROUND: Sepsis is caused by the dysregulated immune response due to an initial infection and results in significant morbidity and mortality in humans. Extracellular cold inducible RNA binding protein (eCIRP) is a novel mediator identified in sepsis. We have previously discovered that microRNA 130b-3p inhibits eCIRP mediated inflammation. As RNA mimics are very unstable in vivo, we hypothesize that an engineered miRNA 130b-3p mimic named PS-OMe miR130, improves stability of the miRNA by protection from nuclease activity. We further hypothesize that PS-OMe miR130 reduces not only eCIRP-mediated inflammation and but also acute lung injury in a murine model of polymicrobial sepsis. METHODS: Single stranded PS-OMe miR130 was synthesized and the binding affinity to eCIRP was evaluated using surface plasmon resonance (SPR) and computational modeling. Macrophages were treated with PS-OMe miR130 with and without eCIRP and cell supernatant analyzed for cytokines. In vitro stability and the in vivo half-life of PS-OMe miR130 were also assessed. The effect of PS-Ome miR130 on eCIRP's binding to TLR4 was evaluated by SPR analysis and modeling. Finally, the effect of PS-OMe miR130 on inflammation and injury was assessed in a murine model of sepsis. RESULTS: We demonstrate via SPR and computational modeling that PS-OMe miR130 has a strong binding affinity to eCIRP. This engineered miRNA decreases eCIRP induced TNF-α and IL-6 proteins, and it is highly stable in vitro and has a long in vivo half-life. We further demonstrate that PS-OMe miR130 blocks eCIRP binding to its receptor TLR4. Finally, we show that PS-OMe miR130 inhibits inflammation and lung injury, and improves survival in murine sepsis. CONCLUSION: PS-OMe miR130 can be developed as a novel therapeutic by inhibiting eCIRP-mediated inflammation and acute lung injury in sepsis.

The Prevalence and Factors That Impact Frailty in Inflammatory Bowel Disease Patients Hospitalized in a Tertiary Hospital in China.

BACKGROUND: The prevalence of inflammatory bowel disease (IBD) is rapidly increasing in China. Beyond disease management, frailty is an important predictor of adverse outcomes in IBD patients, which has not been well investigated. This study aimed to assess frailty status and explore the impact factors in IBD inpatients. METHODS: A total of 372 IBD inpatients were recruited from July 2021 to November 2021 at Peking Union Medical College Hospital. All the participants were surveyed by face-to-face questionnaires, including demographic characteristics, disease conditions, lifestyle, psychology, social support, and frailty. The impact factors of frailty were further assessed by multinomial logistic regression analysis. RESULTS: The participants' median age was 31.00 (Q1: 24.00, Q3: 40.00) years. The overall prevalence of prefrailty and frailty in IBD patients was 59.4% (n = 221) and 14% (n = 52), respectively, and was higher for frailty in females (17.2%) than in males (12.3%). Increased body mass index (odds ratio (OR) 0.917; 95% confidence interval (CI) 0.860-0.978), increased erythrocyte sedimentation rate (OR 1.039; 95% CI: 1.002-1.077), sleep impairment (OR 5.160; 95% CI: 2.394-11.119), and depression (OR 9.480; 95% CI: 3.602-24.949) were independently significantly correlated with prefrailty (p < 0.05). Increased body mass index (OR 0.744; 95% CI: 0.654-0.848), increased erythrocyte sedimentation rate (OR 1.052; 95% CI: 1.011-1.096), sleep impairment (OR 5.832; 95% CI: 2.092-16.260), and depression (OR 10.041; 95% CI: 2.740-36.793) were independently significantly correlated with frailty. Among the factors, whether for frailty or prefrailty, the strongest impact factor was depression. CONCLUSIONS: IBD inpatients are prone to frailty. Comprehensive management focused on the prevention of frailty is warranted to improve the overall prognosis.

Necroptosis-Mediated eCIRP Release in Sepsis.

Introduction: Extracellular cold-inducible RNA-binding protein (eCIRP) is an endogenous pro-inflammatory mediator that exacerbates injury in inflammation and sepsis. The mechanisms in which eCIRP is released have yet to be fully explored. Necroptosis is a programmed cell death that is dependent on the activation of mixed lineage kinase domain-like pseudo kinase (MLKL) which causes the release of damage-associated molecular patterns. We hypothesize that eCIRP is released through necroptosis and intensifies inflammation in sepsis. Methods: RAW264.7 cells were treated with pan-caspase inhibitor z-VAD (15 µM) 1 h before stimulation with LPS (1 µg/mL). Necroptosis inhibitor, Necrostatin-1 (Nec-1) (10 µM) was added to the cells with LPS simultaneously. After 24 h of LPS stimulation, cytotoxicity was determined by LDH assay. eCIRP levels in the culture supernatants and phospho-MLKL (p-MLKL) from cell lysates were assessed by Western blot. p-MLKL interaction with the cell membrane was visualized by immunofluorescence. Sepsis was induced in C57BL/6 mice by cecal ligation and puncture (CLP). Mice were treated with Nec-1 (1 mg/kg) or DMSO. 20 h post-surgery, serum and peritoneal fluid levels of eCIRP, TNF-α and IL-6 were determined by ELISA. H&E staining of lung tissue sections was performed. Results: We found that in RAW264.7 cells, LPS+z-VAD induces necroptosis as evidenced by an increase in p-MLKL levels and causes eCIRP release. Nec-1 reduces both p-MLKL activation and eCIRP release in LPS+z-VAD-treated RAW264.7 cells. Nec-1 also inhibits the release of eCIRP, TNF-α and IL-6 in the serum and peritoneal fluid in CLP-induced septic mice. We predicted a transient interaction between eCIRP and MLKL using a computational model, suggesting that eCIRP may exit the cell via the pores formed by p-MLKL. Conclusion: Necroptosis is a novel mechanism of eCIRP release in sepsis. Targeting necroptosis may ameliorate inflammation and injury in sepsis by inhibiting eCIRP release.

Active Release of eCIRP via Gasdermin D Channels to Induce Inflammation in Sepsis.

Extracellular cold-inducible RNA binding protein (eCIRP) is an inflammatory mediator that causes inflammation and tissue injury in sepsis. Gasdermin D (GSDMD) is a protein that, when cleaved, forms pores in the cell membrane, releasing intracellular contents into the extracellular milieu to exacerbate inflammation. We hypothesize that eCIRP is released actively from viable macrophages via GSDMD pores. We found that LPS induced eCIRP secretion from macrophages into the extracellular space. LPS significantly increased the expression of caspase-11 and cleavage of the GSDMD, as evidenced by increased N-terminal GSDMD expression in RAW 264.7 cells and mouse primary peritoneal macrophages. GSDMD inhibitor disulfiram decreased eCIRP release in vitro. Treatment with glycine to prevent pyroptosis-induced cell lysis did not significantly decrease eCIRP release from LPS-treated macrophages, indicating that eCIRP was actively released and was independent of pyroptosis. Downregulation of GSDMD gene expression by siRNA transfection suppressed eCIRP release in vitro after LPS stimulation. Moreover, GSDMD-/- peritoneal macrophages and mice had decreased levels of eCIRP in the culture supernatants and in blood treated with LPS in vitro and in vivo, respectively. GSDMD inhibitor disulfiram inhibited serum levels of eCIRP in endotoxemia and cecal ligation and puncture-induced sepsis. We conclude that eCIRP release from living macrophages is mediated through GSDMD pores, suggesting that targeting GSDMD could be a novel and potential therapeutic approach to inhibit eCIRP-mediated inflammation in sepsis.

Exosome-Mediated eCIRP Release From Macrophages to Induce Inflammation in Sepsis.

Extracellular cold-inducible RNA-binding protein (eCIRP) is an important damage-associated molecular pattern (DAMP). Despite our understanding of the potentially harmful effects of eCIRP in sepsis, how eCIRP is released from cells remains elusive. Exosomes are endosome-derived extracellular vesicles, which carry proteins, lipids, and nucleic acids to facilitate intercellular communication and several extracellular functions. We hypothesized that eCIRP is released via exosomes to induce inflammation in sepsis. Exosomes isolated from the supernatants of LPS-treated macrophage culture and serum of endotoxemia and polymicrobial sepsis mice showed high purity, as revealed by their unique median sizes ranging between 70 and 126 nm in diameter. eCIRP levels of the exosomes were significantly increased after LPS treatment in the supernatants of macrophage culture, mouse serum, and cecal ligation and puncture (CLP)-induced sepsis mouse serum. Protease protection assay demonstrated the majority of eCIRP was present on the surface of exosomes. Treatment of WT macrophages and mice with exosomes isolated from LPS-treated WT mice serum increased TNFα and IL-6 production. However, treatment with CIRP-/- mice serum exosomes significantly decreased these levels compared with WT exosome-treated conditions. CIRP-/- mice serum exosomes significantly decreased neutrophil migration in vitro compared with WT exosomes. Treatment of mice with serum exosomes isolated from CIRP-/- mice significantly reduced neutrophil infiltration into the peritoneal cavity. Our data suggest that eCIRP can be released via exosomes to induce cytokine production and neutrophil migration. Thus, exosomal eCIRP could be a potential target to inhibit inflammation.

Identification of pathogen(s) in infectious diseases using shotgun metagenomic sequencing and conventional culture: a comparative study.

BACKGROUND: Early and accurate diagnosis of microorganism(s) is important to optimize antimicrobial therapy. Shotgun metagenomic sequencing technology, an unbiased and comprehensive method for pathogen identification, seems to potentially assist or even replace conventional microbiological methodology in the diagnosis of infectious diseases. However, evidence in clinical application of this platform is relatively limited. METHODS: To evaluate the capability of shotgun metagenomic sequencing technology in clinical practice, both shotgun metagenomic sequencing and conventional culture were performed in the PCR-positive body fluid specimens of 20 patients with suspected infection. The sequenced data were then analyzed for taxonomic identification of microbes and antibiotic resistance gene prediction using bioinformatics pipeline. RESULTS: Shotgun metagenomic sequencing results showed a concordance of 17/20 compared with culture results in bacterial detection, and a concordance of 20/20 compared with culture results in fungal detection. Besides, drug-resistant types annotated from antibiotic resistance genes showed much similarity with antibiotic classes identified by susceptibility tests, and more than half of the specimens had consistent drug types between shotgun metagenomic sequencing and culture results. CONCLUSIONS: Pathogen identification and antibiotic resistance gene prediction by shotgun metagenomic sequencing identification had the potential to diagnose microorganisms in infectious diseases, and it was especially helpful for multiple microbial co-infections and for the cases where standard culture approached failed to identify microorganisms.

Extracellular CIRP activates STING to exacerbate hemorrhagic shock.

Stimulator of IFN genes (STING) activates TANK-binding kinase 1 (TBK1) and IFN regulatory factor 3 (IRF3) to produce type I IFNs. Extracellular cold-inducible RNA-binding protein (eCIRP) is released from cells during hemorrhagic shock (HS). We hypothesized that eCIRP activates STING to induce inflammation and acute lung injury (ALI) after HS. WT and STING-/- mice underwent controlled hemorrhage by bleeding, followed by fluid resuscitation. Blood and lungs were collected at 4 hours after resuscitation. Serum ALT, AST, LDH, IL-6, and IFN-ß were significantly decreased in STING-/- mice compared with WT mice after HS. In STING-/- mice, the levels of pTBK1 and pIRF3, and expression of TNF-α, IL-6, and IL-1ß mRNAs and proteins in the lungs, were significantly decreased compared with WT HS mice. The 10-day mortality rate in STING-/- mice was significantly reduced. I.v. injection of recombinant mouse CIRP (rmCIRP) in STING-/- mice showed a significant decrease in pTBK1 and pIRF3 and in IFN-α and IFN-ß mRNAs and proteins in the lungs compared with rmCIRP-treated WT mice. Treatment of TLR4-/-, MyD88-/-, and TRIF-/- macrophages with rmCIRP significantly decreased pTBK1 and pIRF3 levels and IFN-α and IFN-ß mRNAs and proteins compared with WT macrophages. HS increases eCIRP levels, which activate STING through TLR4/MyD88/TRIF pathways to exacerbate inflammation.

The Relationship between the Family Functioning of Individuals with Drug Addiction and Relapse Tendency: A Moderated Mediation Model.

To explore the relationship between family functioning, psychological capital, life history strategy, and relapse tendency of individuals with drug addiction, 842 individuals with drug addiction completed a questionnaire. The results showed that (1) there was a significant negative correlation between the family functioning of individuals with drug addiction and their relapse tendency; (2) psychological capital played an intermediary role between family functioning and relapse tendency; and (3) life history strategy regulated the mediating effect of psychological capital. The results of this study suggest that family members should collaborate with drug addiction treatment centers and participate in the education and treatment process to help reduce drug relapse tendency. Increasing the psychological capital and self-efficacy of individuals with drug addiction through group psychological counseling and psychological education courses could also reduce drug relapse tendency.

Inhibition of aerobic glycolysis alleviates sepsis­induced acute kidney injury by promoting lactate/Sirtuin 3/AMPK­regulated autophagy.

Metabolism reprogramming influences the severity of organ dysfunction, progression to fibrosis, and development of disease in acute kidney injury (AKI). Previously we showed that inhibition of aerobic glycolysis improved survival rates and protected septic mice from kidney injury. However, the underlying mechanisms remain unclear. In the present study, it was revealed that sepsis or lipopolysaccharide (LPS) enhanced aerobic glycolysis as evidenced by increased lactate production and upregulated mRNA expression of glycolysis­related genes in kidney tissues and human renal tubular epithelial (HK­2) cells. The aerobic glycolysis inhibitor 2­deoxy­D­glucose (2­DG) downregulated glycolysis, and improved kidney injury induced by sepsis. 2­DG treatments increased the expression of sirtuin 3 (SIRT3) and phosphorylation­AMP­activated protein kinase (p­AMPK), following promoted autophagy and attenuated apoptosis of tubular epithelial cells in septic mice and in LPS­treated HK­2 cells. However, the glycolysis metabolite lactate downregulated SIRT3 and p­AMPK expression, inhibited autophagy and enhanced apoptosis in LPS­treated HK­2 cells. Furthermore, pharmacological blockade of autophagy with 3­methyladenine (3­MA) partially abolished the protective effect of 2­DG in sepsis­induced AKI. These findings indicated that inhibition of aerobic glycolysis protected against sepsis­induced AKI by promoting autophagy via the lactate/SIRT3/AMPK pathway.

The vitals of NETs.

Neutrophils produce neutrophil extracellular traps (NETs) by expelling their extracellular chromatin embedded with citrullinated histone H3, myeloperoxidase, and other intracellular molecules. Since their discovery in 2004, numerous articles have demonstrated the mechanism of NET formation and their function in innate immunity and inflammation. NET components often play an antimicrobial role, but excessive NETs are deleterious and can cause inflammation and tissue damage. This review highlights recent advancements in the identification of novel pathways and mechanisms of NET formation. We also focus on the specific damaging impact of NETs in individual organs. We then discuss the progress and limitations of various NET detection assays. Collectively, these vital aspects of NETs significantly improve our understanding of the pathobiology of NETs and future diagnostics and therapeutic tools for examining and modulating NETs in inflammatory diseases.

Extracellular CIRP Induces Macrophage Extracellular Trap Formation Via Gasdermin D Activation.

Extracellular cold-inducible RNA-binding protein (eCIRP) is a damage-associated molecular pattern promoting inflammation and tissue injury. During bacterial or viral infection, macrophages release DNA decorated with nuclear and cytoplasmic proteins known as macrophage extracellular traps (METs). Gasdermin D (GSDMD) is a pore-forming protein that has been involved in extracellular trap formation in neutrophils. We hypothesized that eCIRP induces MET formation by activating GSDMD. Human monocytic cell line THP-1 cells were differentiated with phorbol 12-myristate 13-acetate (PMA) and treated with recombinant murine (rm) CIRP. The MET formation was detected by three methods: time-lapse fluorescence microscopy (video imaging), colorimetry, and ELISA. Cleaved forms of GSDMD, and caspase-1 were detected by Western blotting. Treatment of THP-1 cells with rmCIRP increased MET formation as revealed by SYTOX Orange Staining assay in a time- and dose-dependent manner. METs formed by rmCIRP stimulation were further confirmed by extracellular DNA, citrullinated histone H3, and myeloperoxidase. Treatment of THP-1 cells with rmCIRP significantly increased the cleaved forms of caspase-1 and GSDMD compared to PBS-treated cells. Treatment of macrophages with caspase-1, and GSDMD inhibitors z-VAD-fmk, and disulfiram, separately, significantly decreased rmCIRP-induced MET formation. We also confirmed rmCIRP-induced MET formation using primary cells murine peritoneal macrophages. These data clearly show that eCIRP serves as a novel inducer of MET formation through the activation of GSDMD and caspase-1.

HSF1 Alleviates Microthrombosis and Multiple Organ Dysfunction in Mice with Sepsis by Upregulating the Transcription of Tissue-Type Plasminogen Activator.

Sepsis is a life-threatening complication of infection closely associated with coagulation abnormalities. Heat shock factor 1 (HSF1) is an important transcription factor involved in many biological processes, but its regulatory role in blood coagulation remained unclear. We generated a sepsis model in HSF1-knockout mice to evaluate the role of HSF1 in microthrombosis and multiple organ dysfunction. Compared with septic wild-type mice, septic HSF1-knockout mice exhibited a greater degree of lung, liver, and kidney tissue damage, increased fibrin/: fibrinogen deposition in the lungs and kidneys, and increased coagulation activity. RNA-seq analysis revealed that tissue-type plasminogen activator (t-PA) was upregulated in the lung tissues of septic mice, and the level of t-PA was significantly lower in HSF1-knockout mice than in wild-type mice in sepsis. The effects of HSF1 on t-PA expression were further validated in HSF1-knockout mice with sepsis and in vitro in mouse brain microvascular endothelial cells using HSF1 RNA interference or overexpression under lipopolysaccharide stimulation. Bioinformatics analysis, combined with electromobility shift and luciferase reporter assays, indicated that HSF1 directly upregulated t-PA at the transcriptional level. Our results reveal, for the first time, that HSF1 suppresses coagulation activity and microthrombosis by directly upregulating t-PA, thereby exerting protective effects against multiple organ dysfunction in sepsis.

HSF1 Attenuates LPS-Induced Acute Lung Injury in Mice by Suppressing Macrophage Infiltration.

Heat shock factor 1 (HSF1) is a transcription factor involved in the heat shock response and other biological processes. We have unveiled here an important role of HSF1 in acute lung injury (ALI). HSF1 knockout mice were used as a model of lipopolysaccharide- (LPS-) induced ALI. Lung damage was aggravated, and macrophage infiltration increased significantly in the bronchoalveolar lavage fluid (BALF) and lung tissue of HSF-/- mice compared with the damage observed in HSF1+/+ mice. Upon LPS stimulation, HSF-/- mice showed higher levels of monocyte chemoattractant protein-1 (MCP-1) in the serum, BALF, and lung tissue and increased the expression of MCP-1 and chemokine (C-C motif) receptor 2 (CCR2) on the surface of macrophages compared with those in HSF1+/+. Electrophoretic mobility shift assays (EMSA) and dual luciferase reporter assays revealed that HSF1 could directly bind to heat shock elements (HSE) in the promoter regions of MCP-1 and its receptor CCR2, thereby inhibiting the expression of both genes. We concluded that HSF1 attenuated LPS-induced ALI in mice by directly suppressing the transcription of MCP-1/CCR2, which in turn reduced macrophage infiltration.

Extracellular CIRP Induces Inflammation in Alveolar Type II Cells via TREM-1.

Extracellular cold-inducible RNA-binding protein (eCIRP) induces acute lung injury (ALI) in sepsis. Triggering receptor expressed on myeloid cells-1 (TREM-1) serves as a receptor for eCIRP to induce inflammation in macrophages and neutrophils. The effect of eCIRP on alveolar epithelial cells (AECs) remains unknown. We hypothesize that eCIRP induces inflammation in AECs through TREM-1. AECs were isolated from C57BL/6 mice and freshly isolated AECs were characterized as alveolar type II (ATII) cells by staining AECs with EpCAM, surfactant protein-C (SP-C), and T1 alpha (T1α) antibodies. AECs were stimulated with recombinant murine (rm) CIRP and assessed for TREM-1 by flow cytometry. ATII cells from WT and TREM-1-/- mice were stimulated with rmCIRP and assessed for interleukin-6 (IL-6) and chemokine (C-X-C motif) ligand 2 (CXCL2) in the culture supernatants. ATII cells from WT mice were pretreated with vehicle (PBS), M3 (TREM-1 antagonist), and LP17 (TREM-1 antagonist) and then after stimulating the cells with rmCIRP, IL-6 and CXCL2 levels in the culture supernatants were assessed. All of the freshly isolated AECs were ATII cells as they expressed EpCAM and SP-C, but not T1α (ATI cells marker). Treatment of ATII cells with rmCIRP significantly increased TREM-1 expression by 56% compared to PBS-treated ATII cells. Stimulation of WT ATII cells with rmCIRP increased IL-6 and CXCL2 expression, while the expression of IL-6 and CXCL2 in TREM-1-/- ATII cells were reduced by 14 and 23%, respectively. Pretreatment of ATII cells with M3 and LP17 significantly decreased the expression of IL-6 by 30 and 47%, respectively, and CXCL2 by 27 and 34%, respectively, compared to vehicle treated ATII cells after stimulation with rmCIRP. Thus, eCIRP induces inflammation in ATII cells via TREM-1 which implicates a novel pathophysiology of eCIRP-induced ALI and directs a possible therapeutic approach targeting eCIRP-TREM-1 interaction to attenuate ALI.

Inhibition of Aerobic Glycolysis Promotes Neutrophil to Influx to the Infectious Site Via CXCR2 in Sepsis.

Recent evidences suggest that metabolic reprogramming plays an important role in the regulation of innate inflammatory response; however, the specific mechanism is unclear. In this study, we found that glycolytic inhibitor 2-deoxyglucose (2-DG) significantly improved the survival rate in cecal ligation and puncture (CLP)-induced septic mice. 2-DG-treated mice developed increased neutrophil migration to the infectious site and more efficient bacterial clearance than untreated mice. 2-DG reversed the down-regulation of chemokine receptor 2 (CXCR2) and the impaired chemotaxis induced by CLP in mice or lipopolysaccharides (LPS) in human neutrophils. Furthermore, 2-DG reversed the down-regulation of CXCR2 in neutrophils by decreasing the expression of G protein-coupled receptor kinase-2 (GRK2), a serin-threonine protein kinase that mediated the internalization of chemokine receptors, which was induced via the inhibition of extracellular regulated protein kinases (ERK) phosphorylation and the promotion of P38 phosphorylation. Finally, SB225002, a CXCR2 antagonist, partially blocked the protective effects of 2-DG in sepsis. Together, we found a novel mechanism for the migration of neutrophils regulated by metabolism and suggested that aerobic glycolysis might be a potential target of intervention in sepsis.