Mechanisms of Intestinal Barrier Dysfunction in Sepsis

Abstract

Intestinal barrier dysfunction is thought to contribute to the development of multiple organ dysfunction syndrome in sepsis. Although there are similarities in clinical course following sepsis, there are significant differences in the host response depending on the initiating organism and time course of the disease, and pathways of gut injury vary widely in different preclinical models of sepsis. The purpose of this study was to determine whether the timecourse and mechanisms of intestinal barrier dysfunction are similar in disparate mouse models of sepsis with similar mortalities. FVB/N mice were randomized to receive cecal ligation and puncture (CLP) or sham laparotomy, and permeability was measured to fluoresceinisothiocyanate conjugated-dextran (FD-4) six to 48 h later. Intestinal permeability was elevated following CLP at all timepoints measured, peaking at 6 to 12 h. Tight junction proteins claudin 1, 2, 3, 4, 5, 7, 8, 13, and 15, Junctional Adhesion Molecule-A (JAM-A), occludin, and ZO-1 were than assayed by Western blot, real-time polymerase chain reaction, and immunohistochemistry 12 h after CLP to determine potential mechanisms underlying increases in intestinal permeability. Claudin 2 and JAM-A were increased by sepsis, whereas claudin-5 and occludin were decreased by sepsis. All other tight junction proteins were unchanged. A further timecourse experiment demonstrated that alterations in claudin-2 and occludin were detectable as early as 1 h after the onset of sepsis. Similar experiments were then performed in a different group of mice subjected to Pseudomonas aeruginosa pneumonia. Mice with pneumonia had an increase in intestinal permeability similar in timecourse and magnitude to that seen in CLP. Similar changes in tight junction proteins were seen in both models of sepsis although mice subjected to pneumonia also had a marked decrease in ZO-1 not seen in CLP. These results indicate that two disparate, clinically relevant models of sepsis induce a significant increase in intestinal permeability mediated through a common pathway involving alterations in claudin 2, claudin 5, JAM-A, and occludin although model-specific differences in ZO-1 were also identified.

Conflict of interest statement

Conflicts of Interest: Craig Coopersmith was the President of the Society of Critical Care Medicine when this work was submitted.

Figures

FIG. 1. Intestinal permeability by FD-4 following CLP

FD-4 levels in the bloodstream were increased compared to baseline at 6 (p

FIG. 2. Intestinal permeability by everted gut sacs following CLP

Permeability was higher in the proximal small intestine (10 cm distal to the duodenum) in septic mice vs. sham mice (A, p

FIG. 3. Intestinal tight junctions 12 hours after CLP

Levels of claudin-2 (A, p

FIG. 4. Histomicrographs of jejunal tight junctions 12 hours following CLP

Levels of claudin-2 in the crypt (A) and JAM-A in the villus and crypt (B) were increased while claudin-5 in the upper crypt and lower villus (C) and occludin in the lower villus (D) were decreased in representative histomicrographs. Each tight junction mediator stains red and is highlighted with black arrows in the histomicrographs. Magnification 20×.

FIG. 5. Intestinal tight junctions 1 hour after CLP

Claudin-2 (A, p

FIG. 6. Intestinal permeability following pneumonia

FD-4 levels in the bloodstream were increased compared to baseline at 6 (p

FIG. 7. Intestinal tight junctions 12 hours after pneumonia

Levels of claudin-2 (A, p