Targeting complement hyperactivation:a novel therapeutic approach for severe pneumonia induced by influenza virus/staphylococcus aureus coinfection

Dear Editor,Infections by the influenza virus are a significant and widespread global health threat,as these infections have an annual death toll ranging from 290,000 to 650,000.1 A significant proportion of these fatalities are attributed to secondary bacterial pneumonia,a severe complication commonly caused by ubiquitous respiratory pathogens such as Staphylococcus aureus(S.aureus).2 Of particular concern is the increased morbidity and mortality rates in individuals infected simultaneously with influenza virus and methicillin-resistant S.aureus(MRSA).3 The concurrent presence of bacteria and influenza virus usually causes acute respiratory distress syndrome(ARDS),which is associated with acute lung injury(ALI),severe lung tissue edema,and widespread inflammation.Nevertheless,determining the complex mechanisms underlying the synergistic interplay will require further investigation with a suitable coinfection mouse model.In our previous study,we utilized different sequential coinfections at various time points to model influenza A virus and MRSA coinfection.

Sustained induction of IP-10 by MRP8/14 via the IFNβ-IRF7 axis in macrophages exaggerates lung injury in endotoxemic mice

Background:As a damage-associated molecular pattern,the myeloid-related protein 8/14(MRP8/14)heterodimer mediates various inflammatory diseases,such as sepsis.However,how MRP8/14 promotes lung injury by regulating the inflammatory response during endotoxemia remains largely unknown.This study aims at illuminating the pathological functions of MRP8/14 in endotoxemia.Methods:An endotoxemic model was prepared with wild-type and myeloid cell-specific Mrp8 deletion(Mrp8MC)mice for evaluating plasma cytokine levels.Lung injury was evaluated by hematoxylin and eosin(H&E)staining,injury scoring and wet-to-dry weight(W/D)ratio.The dynamic profile of interferonγ(IFNγ)-inducible protein 10(IP-10)mRNA expression induced by macrophage MRP8/14 was determined by quantitative real-time polymerase chain reaction(qPCR).Immunoblotting was used to evaluate the increase in IP-10 level induced by activation of the JAK-STAT signaling pathway.Luciferase reporter assay was performed to detect the involvement of IRF7 in Ip-10 gene transcription.In vivo air pouch experiments were performed to determine the biological function of IP-10 induced by MRP8/14.Results:Experiments with Mrp8MC mice showed that MRP8/14 promoted the production of cytokines,including IP-10,in the bronchoalveolar lavage fluid(BALF)and lung injury in endotoxic mice.The result of qPCR showed sustained expression of Ip-10 mRNA in macrophages after treatment with MRP8/14 for 12 h.Neutralization experiments showed that the MRP8/14-induced Ip-10 expression in RAW264.7 cells was mediated by extracellular IFNβ.Western blotting with phosphorylation-specific antibodies showed that the JAK1/TYK2-STAT1 signaling pathway was activated in MRP8/14-treated RAW264.7 cells,leading to the upregulation of Ip-10 gene expression.IRF7 was further identified as a downstream regulator of the JAK-STAT pathway that mediated Ip-10 gene expression in macrophages treated with MRP8/14.In vivo air pouch experiments confirmed that the IFNβ-JAK1/TYK2-STAT1-IRF7 pathway was required for chemokine(C-X-C motif)receptor 3(CXCR3)+T lymphocyte migration,which promoted lung injury in the context of endotoxemia.Conclusions:In summary,our study demonstrates that MRP8/14 induces sustained production of IP-10 via the IFNβ-JAK1/TYK2-STAT1-IRF7 pathway to attract CXCR3+T lymphocytes into lung tissues and ultimately results in lung injury by an excessive inflammatory response in the context of endotoxemia.

Screening cytokine/chemokine profiles in serum and organs from an endotoxic shock mouse model by LiquiChip

Studying the cytokine profiles in animal models or patients with sepsis provides an experimental basis for the identification of diagnostic biomarkers and therapeutic targets. In this study, we used a liquid protein chip （LiquiChip）, also known as flexible multi-analyte profiling technology, to perform quantitative analyses of several cytokines and chemokines （e.g., IL-II3, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12, TNF-ct, IFN-7, granulocyte-macrophage colony-stimulating factor, keratinocyte chemoattractant, monocyte chemoattractant protein, monokine induced by gamma interferon, IFN-7-inducible protein 10, and macrophage inflammatory protein 1 alpha）. The levels of these cytokines and chemokines were determined both in the blood and in tissues, including the lung, liver, heart, kidney, spleen, brain, stomach, intestine and muscle, of mice challenged with LPS, Our data showed variable production levels of LPS-induced cytokines in different mouse organs, and the cytokine in the blood did not correlate with those in the organs. We also showed that the levels of most cytokines peaked within 1 to 6 h and decreased rapidly afterward. A variety of inflammatory cytokines might be related to the damage in different organs during septic shock. Our data also suggest that the spleen might be an important target organ in the development of systemic inflammatory response syndrome and sepsis.

Identification of heptapeptides targeting a lethal bacterial strain in septic mice through an integrative approach

Effectively killing pathogenic bacteria is key for the treatment of sepsis.Although various anti-infective drugs have been used for the treatment of sepsis,the therapeutic effect is largely limited by the lack of a specific bacterium-targeting delivery system.This study aimed to develop antibacterial peptides that specifically target pathogenic bacteria for the treatment of sepsis.The lethal bacterial strain Escherichia coli MSI001 was isolated from mice of a cecal ligation and puncture(CLP)model and was used as a target to screen bacterial binding heptapeptides through an integrative bioinformatics approach based on phage display technology and high-throughput sequencing(HTS).Heptapeptides binding to E.coli MSI001 with high affinity were acquired after normalization by the heptapeptide frequency of the library.A representative heptapeptide VTKLGSL(VTK)was selected for fusion with the antibacterial peptide LL-37 to construct the specific-targeting antibacterial peptide VTK-LL37.We found that,in comparison with LL37,VTK-LL37 showed prominent bacteriostatic activity and an inhibitive effect on biofilm formation in vitro.In vivo experiments demonstrated that VTK-LL37 significantly inhibited bacterial growth,reduced HMGB1 expression,alleviated lesions of vital organs and improved the survival of mice subjected to CLP modeling.Furthermore,membrane DEGP and DEGQ were identified as VTKbinding proteins by proteomic methods.This study provides a novel strategy for targeted pathogen killing,which is helpful for the treatment of sepsis in the era of precise medicine.

Irreversible electroporation plus allogenic Vγ9Vδ2 T cells enhances antitumor effect for locally advanced pancreatic cancer patients

Immunotherapy has limited efficacy against locally advanced pancreatic cancer(LAPC)due to the presence of an immunosuppressive microenvironment(ISM).Irreversible electroporation(IRE)can not only induce immunogenic cell death,but also alleviate immunosuppression.This study aimed to investigate the antitumor efficacy of IRE plus allogeneicγδT cells in LAPC patients.A total of 62 patients who met the eligibility criteria were enrolled in this trial,then randomized into two groups(A:n=30 and B:n=32).All patients received IRE therapy and after receiving IRE,the group A patients received at least two cycles ofγδT-cell infusion as one course continuously.Group A patients had better survival than group B patients(median OS:14.5 months vs.11 months;median PFS:11 months vs.8.5 months).Moreover,the group A patients treated with multiple courses ofγδT-cell infusion had longer OS(17 months)than those who received a single course(13.5 months).IRE combined with allogeneicγδT-cell infusion is a promising strategy to enhance the antitumor efficacy in LAPC patients,yielding extended survival benefits.