Infliximab Does Not Promote the Presence of Collagenolytic Bacteria in a Mouse Model of Colorectal Anastomosis

BACKGROUND: Previous work from our group has suggested a pivotal role for collagenolytic bacteria in the development of anastomotic complications. Tumor necrosis factor antagonists are a mainstay of treatment for patients with inflammatory bowel disease. The reported impact of these agents on key surgical outcomes such as anastomotic leak has been inconsistent. The objective of this study is to assess the impact of infliximab on the anastomotic microbiome in a mouse model of colon resection. DESIGN: BALB/c mice underwent colon resection with primary anastomosis. Mice were randomly assigned to receive either an intraperitoneal dose of saline (control) or 10 mg/kg of infliximab for 8 weeks prior to surgery. On postoperative day 7, the animals were sacrificed. Anastomotic tissues were analyzed by histology with TUNNEL staining as a marker of epithelial apoptosis. In order to assess compositional and functional changes of the local microbiome, anastomotic tissues were further analyzed by 16S rRNA V4 region sequencing and for the presence of collagenolytic strains that may impair anastomotic healing. The main outcome measures were microbiome community structure and the presence of collagenolytic bacteria. RESULTS: Infliximab-treated mice demonstrated an increase in epithelial apoptosis, consistent with the expected drug effect. Although infliximab modified the perianastomotic microbiome, no increase in the presence of collagenolytic bacteria was observed. CONCLUSIONS: Infliximab did not promote the emergence of collagenolytic bacteria or demonstrably impair anastomotic healing in a mouse model of colon resection and anastomosis

Fecal microbiota transplant rescues mice from human pathogen mediated sepsis by restoring systemic immunity.

Death due to sepsis remains a persistent threat to critically ill patients confined to the intensive care unit and is characterized by colonization with multi-drug-resistant healthcare-associated pathogens. Here we report that sepsis in mice caused by a defined four-member pathogen community isolated from a patient with lethal sepsis is associated with the systemic suppression of key elements of the host transcriptome required for pathogen clearance and decreased butyrate expression. More specifically, these pathogens directly suppress interferon regulatory factor 3. Fecal microbiota transplant (FMT) reverses the course of otherwise lethal sepsis by enhancing pathogen clearance via the restoration of host immunity in an interferon regulatory factor 3-dependent manner. This protective effect is linked to the expansion of butyrate-producing Bacteroidetes. Taken together these results suggest that fecal microbiota transplantation may be a treatment option in sepsis associated with immunosuppression

Mice Fed an Obesogenic Western Diet, Administered Antibiotics, and Subjected to a Sterile Surgical Procedure Develop Lethal Septicemia with Multidrug-Resistant Pathobionts

Despite antibiotics and sterile technique, postoperative infections remain a real and present danger to patients. Recent estimates suggest that 50% of the pathogens associated with postoperative infections have become resistant to the standard antibiotics used for prophylaxis. Risk factors identified in such cases include obesity and antibiotic exposure. To study the combined effect of obesity and antibiotic exposure on postoperative infection, mice were allowed to gain weight on an obesogenic Western-type diet (WD), administered antibiotics and then subjected to an otherwise recoverable sterile surgical injury (30% hepatectomy). The feeding of a WD alone resulted in a major imbalance of the cecal microbiota characterized by a decrease in diversity, loss of Bacteroidetes, a bloom in Proteobacteria, and the emergence of antibiotic-resistant organisms among the cecal microbiota. When WD-fed mice were administered antibiotics and subjected to 30% liver resection, lethal sepsis, characterized by multiple-organ damage, developed. Notable was the emergence and systemic dissemination of multidrug-resistant (MDR) pathobionts, including carbapenem-resistant, extended-spectrum β-lactamase-producing Serratia marcescens, which expressed a virulent and immunosuppressive phenotype. Analysis of the distribution of exact sequence variants belonging to the genus Serratia suggested that these strains originated from the cecal mucosa. No mortality or MDR pathogens were observed in identically treated mice fed a standard chow diet. Taken together, these results suggest that consumption of a Western diet and exposure to certain antibiotics may predispose to life-threating postoperative infection associated with MDR organisms present among the gut microbiota. IMPORTANCE Obesity remains a prevalent and independent risk factor for life-threatening infection following major surgery. Here, we demonstrate that when mice are fed an obesogenic Western diet (WD), they become susceptible to lethal sepsis with multiple organ damage after exposure to antibiotics and an otherwise-recoverable surgical injury. Analysis of the gut microbiota in this model demonstrates that WD alone leads to loss of Bacteroidetes, a bloom of Proteobacteria, and evidence of antibiotic resistance development even before antibiotics are administered. After antibiotics and surgery, lethal sepsis with organ damage developed in in mice fed a WD with the appearance of multidrug-resistant pathogens in the liver, spleen, and blood. The importance of these findings lies in exposing how the selective pressures of diet, antibiotic exposure, and surgical injury can converge on the microbiome, resulting in lethal sepsis and organ damage without the introduction of an exogenous pathogen

Fecal microbiota transplant rescues mice from sepsis due to multi-drug resistant healthcare pathogens by restoring systemic immunity

Death due to sepsis remains a persistent threat to critically ill patients confined to the intensive care unit and is characterized by colonization with multi-drug-resistant healthcare-associated pathogens. Here we report that sepsis in mice caused by a defined four-member pathogen community isolated from a patient with lethal sepsis is associated with the systemic suppression of key elements of the host transcriptome required for pathogen clearance and decreased butyrate expression. More specifically, these pathogens directly suppress interferon regulatory factor 3. Fecal microbiota transplant (FMT) reverses the course of otherwise lethal sepsis by enhancing pathogen clearance via the restoration of host immunity in an interferon regulatory factor 3-dependent manner. This protective effect is linked to the expansion of butyrate-producing Bacteroidetes. Taken together these results suggest that fecal microbiota transplantation may be a treatment option in sepsis associated with immunosuppression

SARS-CoV-2 and the sympathetic immune response: Dampening inflammation with antihypertensive drugs (Clonidine and Propranolol)

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Oral Polyphosphate Suppresses Bacterial Collagenase Production and Prevents Anastomotic Leak Due to Serratia marcescens and Pseudomonas aeruginosa

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Spatial Compartmentalization of the Microbiome between the Lumen and Crypts Is Lost in the Murine Cecum following the Process of Surgery, Including Overnight Fasting and Exposure to Antibiotics.

The cecum is a unique region in the mammalian intestinal tract in which the microbiome is localized to two compartments, the lumen and the crypts. The microbiome within crypts is particularly important as it is in direct contact with lining epithelial cells including stem cells. Here, we analyzed the microbiome in cecum of mice using multiple techniques including metagenomics. The lumen microbiome comprised Firmicutes and Bacteroidetes whereas the crypts were dominated by Proteobacteria and Deferribacteres, and the mucus comprised a mixture of these 4 phyla. The lumen microbial functional potential comprised mainly carbon metabolism, while the crypt microbiome was enriched for genes encoding stress resistance. In order to determine how this structure, assembly, and function are altered under provocative conditions, we exposed mice to overnight starvation (S), antibiotics (A), and a major surgical injury (partial hepatectomy [H]), as occurs with major surgery in humans. We have previously demonstrated that the combined effect of this "SAH" treatment leads to a major disturbance of the cecal microbiota at the bottom of crypts in a manner that disrupts crypt cell homeostasis. Here, we applied the SAH conditions and observed a loss of compartmentalization in both composition and function of the cecal microbiome associated with major shifts in local physicochemical cues including decrease of hypoxia, increase of pH, and loss of butyrate production. Taken together, these studies demonstrated a defined order, structure, and function of the cecal microbiome that can be disrupted under provocative conditions such as major surgery and its attendant exposures.IMPORTANCE The proximal colon and cecum are two intestinal regions in which the microbiome localizes to two spatially distinct compartments, the lumen and crypts. The differences in composition and function of luminal and crypt microbiome in the cecum and the effect of physiological stress on their compartmentalization remain poorly characterized. Here, we characterized the composition and function of the lumen-, mucus-, and crypt-associated microbiome in the cecum of mice. We observed a highly ordered microbial architecture within the cecum whose assembly and function become markedly disrupted when provoked by physiological stress such as surgery and its attendant preoperative treatments (i.e., overnight fasting and antibiotics). Major shifts in local physicochemical cues including a decrease in hypoxia levels, an increase in pH, and a loss of butyrate production were associated with the loss of compositional and functional compartmentalization of the cecal microbiome

Prevention of Anastomotic Leak Via Local Application of Tranexamic Acid to Target Bacterial-mediated Plasminogen Activation: A Practical Solution to a Complex Problem

OBJECTIVE: To investigate the role of bacterial- mediated plasminogen (PLG) activation in the pathogenesis of anastomotic leak (AL) and its mitigation by tranexamic acid (TXA). BACKGROUND: AL is the most feared complication of colorectal resections. The pathobiology of AL in the setting of a technically optimal procedure involves excessive submucosal collagen degradation by resident microbes. We hypothesized that activation of the host PLG system by pathogens is a central and targetable pathway in AL. METHODS: We employed kinetic analysis of binding and activation of human PLG by microbes known to cause AL, and collagen degradation assays to test the impact of PLG on bacterial collagenolysis. Further, we measured the ability of the antifibrinolytic drug TXA to inhibit this process. Finally, using mouse models of pathogen-induced AL, we locally applied TXA via enema and measured its ability to prevent a clinically relevant AL. RESULTS: PLG is deposited rapidly and specifically at the site of colorectal anastomoses. TXA inhibited PLG activation and downstream collagenolysis by pathogens known to have a causal role in AL. TXA enema reduced collagenolytic bacteria counts and PLG deposition at anastomotic sites. Postoperative PLG inhibition with TXA enema prevented clinically and pathologically apparent pathogen-mediated AL in mice. CONCLUSIONS: Bacterial activation of host PLG is central to collagenolysis and pathogen-mediated AL. TXA inhibits this process both in vitro and in vivo. TXA enema represents a promising method to prevent AL in high-risk sites such as the colorectal anastomoses

Prevention of Anastomotic Leak Via Local Application of Tranexamic Acid to Target Bacterial-mediated Plasminogen Activation A Practical Solution to a Complex Problem

Objective: To investigate the role of bacterial- mediated plasminogen (PLG) activation in the pathogenesis of anastomotic leak (AL) and its mitigation by tranexamic acid (TXA). Background: AL is the most feared complication of colorectal resections. The pathobiology of AL in the setting of a technically optimal procedure involves excessive submucosal collagen degradation by resident microbes. We hypothesized that activation of the host PLG system by pathogens is a central and targetable pathway in AL. Methods: We employed kinetic analysis of binding and activation of human PLG by microbes known to cause AL, and collagen degradation assays to test the impact of PLG on bacterial collagenolysis. Further, we measured the ability of the antifibrinolytic drug TXA to inhibit this process. Finally, using mouse models of pathogen-induced AL, we locally applied TXA via enema and measured its ability to prevent a clinically relevant AL. Results: PLG is deposited rapidly and specifically at the site of colorectal anastomoses. TXA inhibited PLG activation and downstream collagenolysis by pathogens known to have a causal role in AL. TXA enema reduced collagenolytic bacteria counts and PLG deposition at anastomotic sites. Postoperative PLG inhibition with TXA enema prevented clinically and pathologically apparent pathogen-mediated AL in mice. Conclusions: Bacterial activation of host PLG is central to collagenolysis and pathogen-mediated AL. TXA inhibits this process both in vitro and in vivo. TXA enema represents a promising method to prevent AL in high-risk sites such as the colorectal anastomoses