Breakdown of Mucin as Barrier to Digestive Enzymes in the Ischemic Rat Small Intestine

Loss of integrity of the epithelial/mucosal barrier in the small intestine has been associated with different pathologies that originate and/or develop in the gastrointestinal tract. We showed recently that mucin, the main protein in the mucus layer, is disrupted during early periods of intestinal ischemia. This event is accompanied by entry of pancreatic digestive enzymes into the intestinal wall. We hypothesize that the mucin-containing mucus layer is the main barrier preventing digestive enzymes from contacting the epithelium. Mucin breakdown may render the epithelium accessible to pancreatic enzymes, causing its disruption and increased permeability. The objective of this study was to investigate the role of mucin as a protection for epithelial integrity and function. A rat model of 30 min splanchnic arterial occlusion (SAO) was used to study the degradation of two mucin isoforms (mucin 2 and 13) and two epithelial membrane proteins (E-cadherin and toll-like receptor 4, TLR4). In addition, the role of digestive enzymes in mucin breakdown was assessed in this model by luminal inhibition with acarbose, tranexamic acid, or nafamostat mesilate. Furthermore, the protective effect of the mucin layer against trypsin-mediated disruption of the intestinal epithelium was studied in vitro. Rats after SAO showed degradation of mucin 2 and fragmentation of mucin 13, which was not prevented by protease inhibition. Mucin breakdown was accompanied by increased intestinal permeability to FITC-dextran as well as degradation of E-cadherin and TLR4. Addition of mucin to intestinal epithelial cells in vitro protected against trypsin-mediated degradation of E-cadherin and TLR4 and reduced permeability of FITC-dextran across the monolayer. These results indicate that mucin plays an important role in the preservation of the mucosal barrier and that ischemia but not digestive enzymes disturbs mucin integrity, while digestive enzymes actively mediate epithelial cell disruption

An Alternate Explanation.

A 48-year-old man with long-standing type 2 diabetes mellitus (recent glycated hemoglobin level, 6.5%) and chronic kidney disease (baseline creatinine level, 3.3 mg per deciliter [292 μmol per liter]; glomerular filtration rate, 24 ml per minute per 1.73 m2 of body-surface area) presented to his primary care physician with a 3-month history of numbness, tingling, and faint violaceous discoloration of the tips of multiple fingers and toes. His physical examination showed reduced light-touch sensation in a glove-and-stocking distribution; the radial and pedal pulses were palpable. The vitamin B12 level was 260 pg per milliliter (192 pmol per liter; normal range, 190 to 950 pg per milliliter [140 to 701 pmol per liter]). He did not smoke tobacco, drink alcohol, or use illicit drugs. One month later, a nontraumatic wound developed on the left foot. The ankle–brachial index (ABI) was 1.2 on both sides (normal range, 0.91 to 1.3). Wound care was initiated for a presumed neuropathic ulcer

Cellular and molecular basis of Venous insufficiency

Chronic venous disease (CVD) has a range of clinical presentations, including tortuous, distended veins in lower extremities, increasing skin pigmentation, and in severe cases ulceration of the affected skin. Venous insufficiency, a precursor to CVD characterized by improper return of blood from the lower extremities to the heart, must be studied in its earliest stages at a time when preventative measures could be applied in man. This underscores the need for basic research into biomarkers and genetic predisposing factors affecting the progression of venous disease. Investigation over the past decade has yielded insight into these specific genetic, cellular and molecular mechanisms underlying the development of venous disease. Among the many advances include the elucidation of an increasing role for matrix metalloproteinases as important mediators of the degenerative process involved with venous insufficiency. This may be preceded by an inflammatory process which further contributes to venular degeneration and endothelial dysfunction seen in advanced presentation of disease. Furthermore, genomic analyses have shed light upon temporal expression patterns of matrix remodeling proteins in diseased tissue samples. In this review we examine some of the current findings surrounding cellular, molecular and genetic advances in delineating the etiology of chronic venous disease

In vivo analysis of intestinal permeability following hemorrhagic shock.

AimTo determine the time course of intestinal permeability changes to proteolytically-derived bowel peptides in experimental hemorrhagic shock.MethodsWe injected fluorescently-conjugated casein protein into the small bowel of anesthetized Wistar rats prior to induction of experimental hemorrhagic shock. These molecules, which fluoresce when proteolytically cleaved, were used as markers for the ability of proteolytically cleaved intestinal products to access the central circulation. Blood was serially sampled to quantify the relative change in concentration of proteolytically-cleaved particles in the systemic circulation. To provide spatial resolution of their location, particles in the mesenteric microvasculature were imaged using in vivo intravital fluorescent microscopy. The experiments were then repeated using an alternate measurement technique, fluorescein isothiocyanate (FITC)-labeled dextrans 20, to semi-quantitatively verify the ability of bowel-derived low-molecular weight molecules (< 20 kD) to access the central circulation.ResultsResults demonstrate a significant increase in systemic permeability to gut-derived peptides within 20 min after induction of hemorrhage (1.11 ± 0.19 vs 0.86 ± 0.07, P < 0.05) compared to control animals. Reperfusion resulted in a second, sustained increase in systemic permeability to gut-derived peptides in hemorrhaged animals compared to controls (1.2 ± 0.18 vs 0.97 ± 0.1, P < 0.05). Intravital microscopy of the mesentery also showed marked accumulation of fluorescent particles in the microcirculation of hemorrhaged animals compared to controls. These results were replicated using FITC dextrans 20 [10.85 ± 6.52 vs 3.38 ± 1.11 fluorescent intensity units (× 10(5), P < 0.05, hemorrhagic shock vs controls)], confirming that small bowel ischemia in response to experimental hemorrhagic shock results in marked and early increases in gut membrane permeability.ConclusionIncreased small bowel permeability in hemorrhagic shock may allow for systemic absorption of otherwise retained proteolytically-generated peptides, with consequent hemodynamic instability and remote organ failure

Amylase and trypsin activity in intestine homogenates is increased during SAO.

<p>Enzyme activity of amylase (<b>A</b>) and trypsin (<b>B</b>) in intestine homogenates of SHAM animals or animals subjected to SAO protocol with luminal inhibition using acarbose (ACA), tranexamic acid (TA) or nafamostat mesilate or without (NI). Activity of luminal contents of SHAM intestines for each enzyme is shown at the end of the graphs. Western blot for amylase, trypsin and β-actin in intestine homogenates of groups described above (<b>C</b>) with corresponding density levels measurements (<b>D, E</b>). Values are mean±SEM (n = 4)/group **P<0.001 compared to SHAM, †††P<0.0001 compared to SAO30, ‡P<0.05 and ‡‡P<0.001 compared to ACA, §§§ P<0.0001 compared to all the other groups.</p