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Nature Protocols 2009Sepsis remains a prevalent clinical challenge and the underlying pathophysiology is still poorly understood. To investigate the complex molecular mechanisms of sepsis,...
Sepsis remains a prevalent clinical challenge and the underlying pathophysiology is still poorly understood. To investigate the complex molecular mechanisms of sepsis, various animal models have been developed, the most frequently used being the cecal ligation and puncture (CLP) model in rodents. In this model, sepsis originates from a polymicrobial infectious focus within the abdominal cavity, followed by bacterial translocation into the blood compartment, which then triggers a systemic inflammatory response. A requirement of this model is that it is performed with high consistency to obtain reproducible results. Evidence is now emerging that the accompanying inflammatory response varies with the severity grade of sepsis, which is highly dependent on the extent of cecal ligation. In this protocol, we define standardized procedures for inducing sepsis in mice and rats by applying defined severity grades of sepsis through modulation of the position of cecal ligation. The CLP procedure can be performed in as little as 10 min for each animal by an experienced user, with additional time required for subsequent postoperative care and data collection.
Topics: Animals; Cecum; Disease Models, Animal; Ligation; Mice; Rats; Sepsis
PubMed: 19131954
DOI: 10.1038/nprot.2008.214 -
Frontiers in Immunology 2021Sepsis is a complex syndrome promoted by pathogenic and host factors; it is characterized by dysregulated host responses and multiple organ dysfunction, which can lead... (Review)
Review
Sepsis is a complex syndrome promoted by pathogenic and host factors; it is characterized by dysregulated host responses and multiple organ dysfunction, which can lead to death. However, its underlying molecular mechanisms remain unknown. Proteomics, as a biotechnology research area in the post-genomic era, paves the way for large-scale protein characterization. With the rapid development of proteomics technology, various approaches can be used to monitor proteome changes and identify differentially expressed proteins in sepsis, which may help to understand the pathophysiological process of sepsis. Although previous reports have summarized proteomics-related data on the diagnosis of sepsis and sepsis-related biomarkers, the present review aims to comprehensively summarize the available literature concerning "sepsis", "proteomics", "cecal ligation and puncture", "lipopolysaccharide", and "post-translational modifications" in relation to proteomics research to provide novel insights into the molecular mechanisms of sepsis.
Topics: Animals; Biomarkers; Cecum; Disease Models, Animal; Humans; Lipopolysaccharides; Protein Processing, Post-Translational; Proteomics; Punctures; Sepsis
PubMed: 34745104
DOI: 10.3389/fimmu.2021.733537 -
Scientific Reports Oct 2021Influenza viruses cause severe respiratory infections in humans and birds, triggering global health concerns and economic burden. Influenza infection is a dynamic...
Influenza viruses cause severe respiratory infections in humans and birds, triggering global health concerns and economic burden. Influenza infection is a dynamic process involving complex biological host responses. The objective of this study was to illustrate global biological processes in ileum and cecal tonsils at early time points after chickens were infected with low pathogenic avian influenza virus (LPAIV) H9N2 through transcriptome analysis. Total RNA isolated from ileum and cecal tonsils of non-infected and infected layers at 12-, 24- and 72-h post-infection (hpi) was used for mRNA sequencing analyses to characterize differentially expressed genes and overrepresented pathways. Statistical analysis highlighted transcriptomic signatures significantly occurring 24 and 72 hpi, but not earlier at 12 hpi. Interferon (IFN)-inducible and IFN-stimulated gene (ISG) expression was increased, followed by continued expression of various heat-shock proteins (HSP), including HSP60, HSP70, HSP90 and HSP110. Some upregulated genes involved in innate antiviral responses included DDX60, MX1, RSAD2 and CMPK2. The ISG15 antiviral mechanism pathway was highly enriched in ileum and cecal tonsils at 24 hpi. Overall, most affected pathways were related to interferon production and the heat-shock response. Research on these candidate genes and pathways is warranted to decipher underlying mechanisms of immunity against LPAIV in chickens.
Topics: Animals; Cecum; Chickens; Female; Gene Expression Profiling; Heat-Shock Proteins; Ileum; Immunity, Innate; Influenza A Virus, H9N2 Subtype; Influenza in Birds; Interferons; RNA, Messenger
PubMed: 34650121
DOI: 10.1038/s41598-021-99182-3 -
Methods in Molecular Biology (Clifton,... 2021Studying the pathophysiology of sepsis still requires animal models, and the mouse remains the most commonly used species. Here we discuss the "cecal slurry" (CS) model...
Studying the pathophysiology of sepsis still requires animal models, and the mouse remains the most commonly used species. Here we discuss the "cecal slurry" (CS) model of polymicrobial, peritoneal sepsis and compare and contrast it to other commonly used methods. Among the different murine models of sepsis, cecal ligation and puncture (CLP), and not the CS, is often considered the "gold standard" to induce polymicrobial sepsis in laboratory animals. CLP is a well-described model involving a simple surgical procedure that closely mimics the clinical course of intra-abdominal sepsis. However, CLP may not be an option for experiments involving newborn pups, where the cecum is indistinguishable from small bowel, where differences in microbiome content may affect the experiment, or where surgical procedures/anesthesia exposure needs to be limited. An important alternative method is the CS model, involving the intraperitoneal injection of cecal contents from a donor animal into the peritoneal cavity of a recipient animal to induce polymicrobial sepsis. Furthermore, CS is an effective alternative model of intraperitoneal polymicrobial sepsis in adult mice and can now be considered the "gold standard" for experiments in neonatal mice.
Topics: Abdomen; Animals; Animals, Newborn; Cecum; Disease Models, Animal; Female; Ligation; Mice; Peritonitis; Punctures; Sepsis
PubMed: 34048005
DOI: 10.1007/978-1-0716-1488-4_4 -
PloS One 2022Percutaneous decompression of the cecum is a procedure that could be considered for horses with cecal gas distension. The aim of this study was to identify complications...
Percutaneous decompression of the cecum is a procedure that could be considered for horses with cecal gas distension. The aim of this study was to identify complications such as peritonitis and clinically relevant peritonitis (CRP) after transabdominal cecal trocarization in healthy horses using a cattle trocar and a cecal needle. Mixed breed horses were assigned to three groups (n = 6): horses that underwent trocarization with a cecal needle (G1) or a cattle trocar (G2), and a control group (CG) without cecal trocarization. The same horses were used in each group, respecting a three-month washout period between studies. A physical examination, serial blood, and peritoneal fluid sampling were performed, prior to cecal trocarization and 2, 6 and 12 hours after the first collection and 1, 2, 3, 7, and 14 days after the procedure. Acute-phase proteins in blood and peritoneal fluid were analyzed by polyacrylamide gel electrophoresis. Horses with a high cell count in the peritoneal fluid (i.e., 10,000 cells/μl) were considered to have peritonitis and CRP if they met at least two of the following clinical criteria: anorexia, lethargy, tachycardia, tachypnea, fever, ileus, abnormal oral mucous membrane color, abnormal white blood cells count, or high blood fibrinogen concentration (> 5 g/L). All horses recovered from cecal trocarization and abdominocentesis with no major complications. Cecal trocarization caused cytologic evidence of peritonitis in G1 and G2 during the 14 days of evaluation. CRP was not observed, although a decrease in cecal motility was observed in G1 and G2 during the experimental period and three horses, one from G1 and two from G2, showed a single moment of fever. None of the groups showed leukopenia or leukocytosis, although blood neutrophil count decreased at D7 and D14 in G1 and at D14 in G2 (p ≤ 0.05). After cecal trocarization, an increase in the total nucleated cells count, total proteins, globulins, alkaline phosphatase and acute phase proteins were observed in the peritoneal fluid of G1 and G2 during the 14 days of evaluation (p ≤ 0.05), without causing clinically relevant peritonitis. Transcutaneous cecal trocarization promotes peritonitis, which is more intense with a cattle trocar than with a cecal needle. The cecal needle should be considered for cecal trocarization of horses with cecal tympany.
Topics: Horses; Cattle; Animals; Horse Diseases; Cecum; Peritonitis; Leukocyte Count; Acute-Phase Proteins; Surgical Instruments
PubMed: 36417417
DOI: 10.1371/journal.pone.0277468 -
Cell Reports Dec 2021It is well established in the microbiome field that antibiotic (ATB) use and metabolic disease both impact the structure and function of the gut microbiome. But how host...
It is well established in the microbiome field that antibiotic (ATB) use and metabolic disease both impact the structure and function of the gut microbiome. But how host and microbial metabolism interacts with ATB susceptibility to affect the resulting dysbiosis remains poorly understood. In a streptozotocin-induced model of hyperglycemia (HG), we use a combined metagenomic, metatranscriptomic, and metabolomic approach to profile changes in microbiome taxonomic composition, transcriptional activity, and metabolite abundance both pre- and post-ATB challenge. We find that HG impacts both microbiome structure and metabolism, ultimately increasing susceptibility to amoxicillin. HG exacerbates drug-induced dysbiosis and increases both phosphotransferase system activity and energy catabolism compared to controls. Finally, HG and ATB co-treatment increases pathogen susceptibility and reduces survival in a Salmonella enterica infection model. Our data demonstrate that induced HG is sufficient to modify the cecal metabolite pool, worsen the severity of ATB dysbiosis, and decrease colonization resistance.
Topics: Animals; Anti-Bacterial Agents; Cecum; Diabetes Mellitus, Experimental; Drug Resistance, Bacterial; Dysbiosis; Female; Gastrointestinal Microbiome; Hyperglycemia; Male; Metabolome; Metagenome; Mice; Mice, Inbred C57BL; Microbiota; Salmonella Infections, Animal; Salmonella enterica; Transcriptome
PubMed: 34910917
DOI: 10.1016/j.celrep.2021.110113 -
Journal of Visualized Experiments : JoVE Jun 2019Sepsis, a severe and complicated life-threatening infection, is characterized by an imbalance between pro- and anti-inflammatory responses in multiple organs. With the...
Sepsis, a severe and complicated life-threatening infection, is characterized by an imbalance between pro- and anti-inflammatory responses in multiple organs. With the development of therapies, most patients survive the hyperinflammatory phase but progress to an immunosuppressive phase, which increases the emergence of secondary infections. Therefore, improved understanding of the pathogenesis underlying secondary hospital-acquired infections in the immunosuppressive phase during sepsis is of tremendous importance. Reported here is a model to test infectious outcomes by creating double-hit infections in mice. A standard surgical procedure is used to induce polymicrobial peritonitis by cecal ligation and puncture (CLP) and followed by intranasal infection of Staphylococcus aureus to simulate pneumonia occurring in immune suppression that is frequently seen in septic patients. This dual model can reflect the immunosuppressive state occurring in patients with protracted sepsis and susceptibility to secondary infection from nosocomial pneumonia. Hence, this model provides a simple experimental approach to investigate the pathophysiology of sepsis-induced secondary bacterial pneumonia, which may be used for discovering novel treatments for sepsis and its complications.
Topics: Animals; Cecum; Disease Models, Animal; Female; Immunosuppression Therapy; Ligation; Mice; Mice, Inbred C57BL; Nasal Surgical Procedures; Punctures; Sepsis
PubMed: 31259890
DOI: 10.3791/59386 -
Frontiers in Immunology 2020(.) is a common cause of pneumonia-derived sepsis in human and is associated with high morbidity and mortality. The microbiota promotes and maintains host immune...
(.) is a common cause of pneumonia-derived sepsis in human and is associated with high morbidity and mortality. The microbiota promotes and maintains host immune homeostasis during bacterial infections. However, the mechanisms by which the gut microbiota affects immune responses in the lung still remain poorly understood. Here, we performed cecal metabolomics sequencing and fecal 16s rRNA sequencing in -infected mice and uninfected controls and showed that infection led to profound alterations in the gut microbiome and thus the cecal metabolome. We observed that the levels of were significantly decreased in -infected mice. Spearman correlation analysis showed that alterations in the richness and composition of the gut microbiota were associated with profound changes in host metabolite concentrations. Further, short-chain fatty acids (SCFAs), including acetate, propionate, and butyrate, were detected in cecal contents and serum by gas chromatography-mass spectrometry (GC-MS). We observed that the concentrations of these three SCFAs were all lower in the infected groups than in the untreated controls. Lastly, oral supplementation with these three SCFAs reduced susceptibility to infections, as indicated by lower bacterial burdens in the lung and higher survival rates. Our data highlight the protective roles of gut microbiota and certain metabolites in -pneumonia and suggests that it is possible to intervene in this bacterial pneumonia by targeting the gut microbiota.
Topics: Animals; Cecum; Fatty Acids, Volatile; Feces; Gastrointestinal Microbiome; Klebsiella Infections; Klebsiella pneumoniae; Metabolome; Mice; Mice, Inbred C57BL; Pneumonia, Bacterial; Sepsis
PubMed: 32849494
DOI: 10.3389/fimmu.2020.01331 -
Infection and Immunity Aug 2020A recent report by the National Institutes of Health on sepsis research has implied there is a trend to move away from mouse models of sepsis. The most commonly used... (Review)
Review
A recent report by the National Institutes of Health on sepsis research has implied there is a trend to move away from mouse models of sepsis. The most commonly used animal model to study the pathogenesis of human sepsis is cecal ligation and puncture (CLP) in mice. The model has been the mainstay of sepsis research for decades and continues to be considered the gold standard to inform novel pathways of sepsis physiology and its therapeutic direction. As there have been many criticisms of the model, particularly regarding its relevance to human disease, how this model might be repurposed to be more reflective of the human condition begs discussion. In this piece, we compare and contrast the mouse microbiome of the CLP model to the emerging science of the microbiome of human sepsis and discuss the relevance for mice to harbor the specific pathogens present in the human microbiome during sepsis, as well as an underlying disease process to mimic the characteristics of those patients with undesirable outcomes. How to repurpose this model to incorporate these "human factors" is discussed in detail and suggestions offered.
Topics: Animals; Anti-Bacterial Agents; Bacterial Typing Techniques; Cecum; Cytokines; Disease Models, Animal; Food, Formulated; Gastrointestinal Microbiome; Humans; Intraabdominal Infections; Ligation; Mice; Punctures; Sepsis; Survival Analysis
PubMed: 32571986
DOI: 10.1128/IAI.00942-19 -
ImmunoHorizons Jul 2022Sepsis, an amplified immune response to systemic infection that leads to life-threatening organ dysfunction, affects >125,000 people/day worldwide with 20% mortality....
Sepsis, an amplified immune response to systemic infection that leads to life-threatening organ dysfunction, affects >125,000 people/day worldwide with 20% mortality. Modest therapeutic progress for sepsis has been made, in part because of the lack of therapeutic translatability between mouse-based experimental models and humans. One potential reason for this difference stems from the extensive use of immunologically naive specific pathogen-free mice in preclinical research. To address this issue, we used sequential infections with well-defined BSL-2 pathogens to establish a novel immune-experienced mouse model (specific pathogen experienced [SPexp]) to determine the extent to which immunological experience and/or inflammation influences the host capacity to respond to subsequent infections, including sepsis. Consistent with their immunological experience, SPexp inbred or outbred mice had significant changes in the composition and activation status of multiple leukocyte populations known to influence the severity of cecal ligation and puncture-induced sepsis. Importantly, by varying the timing of sepsis induction, we found the level of basal inflammation controls sepsis-induced morbidity and mortality in SPexp mice. In addition, although a beneficial role of NK cells in sepsis was recently demonstrated in specific pathogen-free mice, NK cell depletion before cecal ligation and puncture induction in SPexp mice lead to diminished mortality, suggesting NK cells may have beneficial or detrimental roles in the response to septic insult dependent on host immune status. Thus, data highlight the importance of utilizing immune-experienced models for preclinical studies to interrogate the cellular/molecular mechanism(s) that could be therapeutically exploited during severe and dysregulated infection-induced inflammatory responses, such as sepsis.
Topics: Animals; Cecum; Disease Models, Animal; Humans; Inflammation; Ligation; Mice; Sepsis
PubMed: 35878936
DOI: 10.4049/immunohorizons.2200050