Abstract
SUMMARYThe gut barrier, composed of a single layer of intestinal epithelial cells (IECs) held together by tight junctions, prevents the entrance of harmful microorganisms, antigens and toxins from the gut lumen into the blood. Small intestinal homeostasis is normally maintained by the rate of shedding of senescent enterocytes from the villus tip exactly matching the rate of generation of new cells in the crypt. However, in various localized and systemic inflammatory conditions, intestinal homeostasis can be disturbed as a result of increased IEC shedding. Such pathological IEC shedding can cause transient gaps to develop in the epithelial barrier and result in increased intestinal permeability. Although pathological IEC shedding has been implicated in the pathogenesis of conditions such as inflammatory bowel disease, our understanding of the underlying mechanisms remains limited. We have therefore developed a murine model to study this phenomenon, because IEC shedding in this species is morphologically analogous to humans. IEC shedding was induced by systemic lipopolysaccharide (LPS) administration in wild-type C57BL/6 mice, and in mice deficient in TNF-receptor 1 (Tnfr1−/−), Tnfr2 (Tnfr2−/−), nuclear factor kappa B1 (Nfκb1−/−) or Nfĸb2 (Nfĸb2−/−). Apoptosis and cell shedding was quantified using immunohistochemistry for active caspase-3, and gut-to-circulation permeability was assessed by measuring plasma fluorescence following fluorescein-isothiocyanate–dextran gavage. LPS, at doses ≥0.125 mg/kg body weight, induced rapid villus IEC apoptosis, with peak cell shedding occurring at 1.5 hours after treatment. This coincided with significant villus shortening, fluid exudation into the gut lumen and diarrhea. A significant increase in gut-to-circulation permeability was observed at 5 hours. TNFR1 was essential for LPS-induced IEC apoptosis and shedding, and the fate of the IECs was also dependent on NFκB, with signaling via NFκB1 favoring cell survival and via NFκB2 favoring apoptosis. This model will enable investigation of the importance and regulation of pathological IEC apoptosis and cell shedding in various diseases.
Highlights
The gut barrier consists of a single layer of intestinal epithelial cells (IECs) and the tight junctions between them
Utilizing the inflammatory response induced by systemic administration of lipopolysaccharide (LPS) in mice, the authors developed a new model for the investigation of IEC shedding
A threshold dose of ≥0.125 mg LPS/kg body weight delivered by intraperitoneal injection induced rapid and dynamic villus IEC apoptosis and shedding, which peaked at 1.5 hours post-administration
Summary
The gut barrier consists of a single layer of intestinal epithelial cells (IECs) and the tight junctions between them. It allows absorption of nutrients from the intestinal lumen into the circulation, while preventing the entry of injurious microorganisms, toxins and antigens. In mice, which exhibit whole IEC shedding similar to that which occurs in humans (Bullen et al, 2006), ~1400 IECs are estimated to be shed in this way from a single villus tip per day (Potten and Loeffler, 1990). Of the highest cell turnover rates in the body, with an estimated 1011 and 2×108 cells being shed per day from the small intestine of humans and mice, respectively (Potten and Loeffler, 1990). During the process of physiological cell shedding, the highly coordinated process of tight junction rearrangement that is required to allow the detachment and release of IECs from the epithelium maintains the gut barrier (Madara, 1990)
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