Questioning Current Concepts in Acute Pancreatitis: Endotoxin Contamination of Porcine Pancreatic Elastase is Responsible for Experimental Pancreatitis-Associated Distant Organ Failure

Aims: The systemic inflammatory response syndrome (SIRS) is responsible for pancreatitis-associated mortality. Recent in vitro and in vivo studies have suggested that pancreatic elastase is one missing link between the localized inflammatory process in the pancreas and distant organ dysfunction and failure. It has been shown that pancreatic elastase activates transcription factors including NF-κB, induces TNF secretion in myeloid cells via toll-like receptors and causes a SIRS-like response in animal models. Methods: RAW264.7- and bone marrow derived macrophages (BMDM) were incubated with different pancreatic elastase preparations (El-IV–1, El-IV–2, El-I, all Sigma, and low endotoxin elastase, El-UP, Elastin Products). TNF content in supernatants, degradation of IκB proteins and activation of NF-κB were determined under various conditions. Systemic proinflammatory effects of pancreatic elastase after i.p. application in mice were assessed by TNF serum levels, lung MPO, histology and lethality after co-injection with D-galactosamine. Results: The highly purified low endotoxin pancreatic elastase preparation (El-UP) failed both to activate NF-κB and to induce TNF release in RAW264.7 cells and BMDM. In contrast, less purified elastase preparations (El-IV–1, El-IV–2, El-I) all caused activation of NF-κB and were able to induce TNF release at very low concentrations. These effects were sensitive to pretreatment with polymyxin B and resistant to heat inactivation. Endotoxin activity as determined by limulus amebocyte lysate (LAL)-assay was more than three orders of magnitude lower in the low endotoxin elastase preparation (El-UP) compared to less purified elastase preparations. In contrast to contaminated elastase or LPS, elastase free of contamination (El-UP) failed to induce elevated TNF serum levels or pulmonary neutrophil infiltration after intraperitoneal application in mice and did not induce lethality when co-injected with D-galactosamine. Failure of low endotoxin elastase to induce proinflammatory effects in vivo and in vitro was not due to functional inactivity of the elastase preparation as determined by elastase activity assay. Conclusion: These results question current concepts of direct proinflammatory effects attributed to pancreatic elastase.