A revised model of platelet aggregation.

In this study we have examined the mechanism of platelet aggregation under physiological flow conditions using an in vitro flow-based platelet aggregation assay and an in vivo rat thrombosis model. Our studies demonstrate an unexpected complexity to the platelet aggregation process in which platelets in flowing blood continuously tether, translocate, and/or detach from the luminal surface of a growing platelet thrombus at both arterial and venous shear rates. Studies of platelets congenitally deficient in von Willebrand factor (vWf) or integrin alpha(IIb)beta(3) demonstrated a key role for platelet vWf in mediating platelet tethering and translocation, whereas integrin alpha(IIb)beta(3) mediated cell arrest. Platelet aggregation under flow appears to be a multistep process involving: (a) exposure of vWf on the surface of immobilized platelets; (b) a reversible phase of platelet aggregation mediated by the binding of GPIbalpha on the surface of free-flowing platelets to vWf on the surface of immobilized platelets; and (c) an irreversible phase of aggregation dependent on integrin alpha(IIb)beta(3). Studies of platelet thrombus formation in vivo demonstrate that this multistep adhesion mechanism is indispensable for platelet aggregation in arterioles and also appears to promote platelet aggregate formation in venules. Together, our studies demonstrate an important role for platelet vWf in initiating the platelet aggregation process under flow and challenge the currently accepted view that the vWf-GPIbalpha interaction is exclusively involved in initiating platelet aggregation at elevated shear rates.

The effect of branched-chain amino acid-enriched parenteral nutrition on gut permeability.

In situations of catabolic stress, the gut becomes atrophic and has a diminished barrier function as evidenced by an increased permeability to a variety of molecules. It is known that the parenteral administration of branched-chain amino acids (BCAA) reduce gut atrophy. The aim of this study was to examine the effect of BCAA-enriched solutions of parenteral nutrients on gut permeability. A secondary aim was to observe the association between gut permeability and variables that have been used to assess jejunal atrophy. Central venous lines were inserted into 30 rats before randomization to receive nutritional support with: (1) a conventional parenteral solution (CPN), (2) A 2.0% BCAA-enriched solution (BCAA), or (3) rat food ad lib (Rat Food). The rats were assessed after 7 d for nutritional status, gut morphology, and gut permeability ratio (ratio of the permeability to 14C raffinose and 3H mannitol). We found that rats in the Rat Food Group lost the least amount of weight, had the least amount of jejunal atrophy, and had better preservation of barrier function as determined by gut permeability. When compared with the CPN Group, the BCAA Group had better preservation of jejunal morphology and protein content (p < 0.05), but a similar gut permeability. A cross-correlation matrix demonstrated a significant negative correlation between permeability to mannitol and mucosal weight, mucosal protein content and mucosal DNA content. Branched-chain amino acid-enriched parenteral nutrition reduced gut atrophy but not the gut permeability associated with parenteral nutrition. In the parenterally nourished rat model, atrophy of the jejunum is associated with increased permeability to small molecules.

Peritoneal defences and peritoneum-associated lymphoid tissue.

The peritoneum is mainly protected by the innate immune system. This consists of mechanical clearance of the peritoneal cavity, activation of complement, and the actions of polymorphonuclear neutrophils and macrophages. The specific immune system, which is mediated by the activity of lymphocytes, provides a secondary amplification system that may be of great importance for patients with intraperitoneal sepsis. This review provides an overview of the relevant innate immune mechanisms and explores the possible role of peritoneum-associated lymphoid tissue.

Enteral branched-chain amino acids increase the specific activity of jejunal glutaminase and reduce jejunal atrophy.

Branched-chain amino acid (BCAA)-enriched nutrient solutions reduce gut atrophy associated with parenteral nutrition. We hypothesized that this effect was mediated by phosphate-dependent glutaminase. Thirty male Wistar rats (300-350 g) underwent a standardized surgical procedure and were then randomized into three groups to receive 6 days of ad libitum enteral nutrition. The animals were fed a solution of conventional nutrients, a solution of conventional nutrients enriched with 2.0% BCAA or a solution of conventional parenteral nutrients enriched with 2.5% glutamine. When compared with rats fed conventional nutrients, rats fed BCAA and glutamine had less jejunal atrophy (P < 0.05) and a greater specific activity of phosphate-dependent glutaminase in the jejunum (131%; P < 0.05). It is concluded that enteral BCAA reduce atrophy of the jejunum via the generation of glutamine.

The effect of minimum luminal nutrition on bacterial translocation and atrophy of the jejunum during parenteral nutrition.

In situations of catabolic stress, the gut becomes atrophic and may have diminished barrier function as evidenced by an increase in bacterial translocation. The aim of this study was to examine the effect of minimum luminal nutrition during parenteral nutrition on the extent of jejunal atrophy and rate of bacterial translocation. Central venous lines were inserted into 30 rats before they underwent randomization to receive nutritional support with: (a) conventional parenteral nutrition; (b) conventional parenteral nutrition with 3 g/day of rat food (i.e., minimum luminal nutrition); or (c) rat food ad libitum. The rats were assessed after 10 days for nutritional status, extent of jejunal atrophy, caecal flora, as well as the extent of bacterial translocation to the mesenteric lymph nodes, liver and spleen. Rats in the rat food ad libitum group lost the smallest amount of weight and had the least amount of jejunal atrophy, yet had a similar rate of bacterial translocation as the parenterally nourished groups. When compared with the conventional parenteral nutrition group, the minimum luminal nutrition group had better preservation of the weight of the small bowel and its isolated mucosa (P < 0.01), but had a similar rate of bacterial translocation. Minimum luminal nutrition reduced the extent of atrophy of the gut but did not affect the incidence of bacterial translocation. It is inferred that there is no direct relationship between the extent of mucosal atrophy and incidence of bacterial translocation.

Quantitation of glutaminase mRNA in enterocytes using competitive RT-PCR.

The concentration of mRNA within the intestinal mucosa is usually measured by either Northern blot analysis or semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). However, these methods are limited by a lack of valid internal controls, low sensitivity, and large differences in the concentration of the internal control and target gene. The authors present an alternative method using competitive RT-PCR to measure glutaminase mRNA in isolated enterocytes.

Review: Peyer's patches.

The function of Peyer's patches as antigenic sampling sites involves the complex interplay of a variety of mechanisms that aim to recognize luminal antigens, induce an immunological response and decrease the incidence of antigen translocation across the mucosal epithelium. This is achieved by M cells, which facilitate the uptake of luminal antigens, a vascular architecture that promotes the retention of absorbed antigens within the patch interstitium (allowing for maximal antigenic activation of lymphocytes) and the presence of lymphoid follicles that contain antigen-presenting cells and lymphocytes. Lymphocytes encountering antigen in the Peyer's patches proliferate, differentiate into fully mature antigen-specific effector cells and migrate to the mesenteric lymph nodes where they undergo final maturation. The mature lymphocytes then enter the systemic circulation and migrate throughout the other mucosa-associated lymphoid tissues of the body and "home' into the gut via high endothelial venules and gut-associated lymphoid tissue-specific adhesion molecules, providing antigen-specific lymphocytes at sites likely to re-encounter the antigen.

Warm ischaemia time in a model for small bowel transplantation.

Intestinal transplantation is associated with high rates of mortality and morbidity. This paper details our initial experience with 82 heterotopic small bowel transplants based upon the original rat model described by Monchik and Russell (Surgery 70:693-702, 1971). A key issue associated with mortality was a warm ischaemia time of more than 40 min (P < 0.01). Sixty-eight percent of the recipients (44/65) survived for more than 24 hr when the warm ischaemia time of the donor bowel was reduced to less than 40 min. Investigators establishing an animal model of heterotopic small bowel transplantation should pay particular attention to the warm ischaemia time of the donor bowel.

The pathobiology of peritonitis.

The peritoneum is more than a mechanical covering that allows for the easy gliding of opposed peritoneal surfaces. The peritoneal mesothelial cells facilitate the action of powerful innate immune mechanisms. In addition, the peritoneal-associated lymphoid tissues contain unique cells that may play a crucial role in the localization of intraperitoneal infection. A clearer understanding of the molecular and cellular events underlying peritoneal functions in both the unstimulated and stimulated state will aid future treatment of peritonitis.

The effect of minimum luminal nutrition on mucosal cellularity and immunity of the gut.

Many catabolic patients can only consume small volumes of enteral nutrients. The aim of this study was to evaluate markers of cellularity and immunity in the small intestine of rats randomized to receive 6 days of parenteral nutrition, 25% enteral and 75% parenteral nutrition (i.e. minimum luminal nutrition) or enteral nutrition. The same glutamine-enriched solution was used for both parenteral and enteral nutrition. Enteral nutrition was associated with the least amount of jejunal atrophy (P<0.01), with the results from the minimum luminal nutrition group approximating those of the parenteral nutrition group. Parenteral nutrition was associated with the greatest number of CD2+ cells (P< 0.05) and the lowest CD4/CD8 cell ratio (P< 0.01) in the jejunal mucosa. In essence, we failed to demonstrate that there are any appreciable benefits associated with the enteral consumption of 25% of a nutrient load.

Ischaemia-reperfusion injury to the intestine.

Ischaemia-reperfusion injury (IRI) is of obvious relevance in situations where there is an interruption of blood supply to the gut, as in vascular surgery, or in the construction of free intestinal grafts. It is now appreciated that IRI also underlies the guy dysfunction that occurs in early shock, sepsis, and trauma. The events that occur during IRI are complex. However, recent advances in cellular biology have started to unravel these underlying processes. The aim of this review is to provide an outline of current knowledge on the mechanisms and consequences of IRI. Initially, IRI appears to be mediated by reactive oxygen metabolites and, at a later stage, by the priming and activation of polymorphonuclear neutrophils (PMN). Ischaemia-reperfusion injury can diminish the barrier function of the gut, and can promote an increase in the leakage of molecules (intestinal permeability) or the passage of microbes across the wall of the bowel (bacterial translocation). Ischaemia-reperfusion injury to the gut can result in the generation of molecules that may also harm distant tissues.