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Protease activity in mesenteric lymph following splanchnic arterial occlusion

  • Author(s): Richter, Michael D.
  • et al.
Abstract

The lymphatics play a key role in the pathogenesis of multiple organ failure following circulatory shock. Understanding the cause of this organ dysfunction is a fundamentally unsolved problem in medicine. Current results show that biologically active molecules produced in the gut enter the circulation via the mesenteric lymph duct, and induce neutrophil activation, increased vascular permeability, and acute lung injury. Despite several attempts to determine the protein composition of mesenteric lymph, the factors responsible for this biological activity and specifically the organ damage are unknown. The aim of this study was to investigate the level and activity of pancreatic digestive proteases in post-shock lymph, which could be derived from the intestine and which are a potentially damaging mediator. Male Wistar rats were subjected to laparotomy, followed by mesenteric lymph duct cannulation and one hour of lymph fluid collection before shock. Five animals then underwent one hour of splanchnic arterial occlusion followed by reperfusion, while sham shock animals remained perfused. Post-shock lymph was collected for three hours, with the samples aliquoted every hour. Protease activity in the lymph was analyzed using a fluorescently quenched casein substrate as an indicator for general protease activity and separately charge-changing fluorescently quenched peptide substrates specific to pancreatic digestive proteases (trypsin, chymotrypsin). These measurements showed trypsin and chymotrypsin activity in the lymph, and the presence of trypsin was confirmed using Western analysis. Over the time course we see a base level of protease activity in the mesenteric lymph that increases following gut ischemia and reperfusion. These results indicate that during shock digestive protease escape from the splanchnic bed, raising the possibility that they are involved in the systemic injury following shock. Supported by NIH grant GM-8507

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