Questioning current concepts in acute pancreatitis: endotoxin contamination of porcine pancreatic elastase is responsible for experimental pancreatitis-associated distant organ failure.
The systemic inflammatory response syndrome 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-kappaB, and induces TNF-alpha secretion in myeloid cells via TLRs. In this study we demonstrate that a highly purified low endotoxin pancreatic elastase preparation (El-UP) failed both to activate NF-kappaB and to induce TNF-alpha release in RAW 264.7 cells and bone marrow-derived macrophages. In contrast, a less purified elastase preparation (El-IV) caused activation of NF-kappaB and was able to induce TNF-alpha release at very low concentrations. These effects were sensitive to pretreatment of the cells with polymyxin B and were resistant to heat inactivation. Endotoxin activity as determined by the Limulus amebocyte lysate assay was >3 orders of magnitude lower in the low endotoxin elastase preparation (El-UP) compared with less purified elastase preparations (El-IV). In contrast to contaminated elastase or LPS, elastase free of contamination (El-UP) failed to induce elevated serum TNF-alpha levels or pulmonary neutrophil infiltration after i.p. application in mice and did not induce lethality when coinjected with d-galactosamine. Failure of low endotoxin elastase (El-UP) 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. These results question current concepts of direct proinflammatory effects attributed to pancreatic elastase.
Geisler
Fabian
F
Department of Internal Medicine II, Klinikum Rechts der Isar, Technical University of Munich, Munich, Germany.
Algül
Hana
H
Riemann
Marc
M
Schmid
Roland M
RM
D000208:Acute Disease; D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D000943:Antigens, Differentiation / Q000235:genetics / Q000502:physiology; D001427:Bacterial Toxins / Q000008:administration & dosage / Q000009:adverse effects* / Q000302:isolation & purification; D002460:Cell Line; D002478:Cells, Cultured; D004340:Drug Contamination; D004731:Endotoxins / Q000008:administration & dosage / Q000009:adverse effects* / Q000302:isolation & purification; D005688:Galactosamine / Q000008:administration & dosage; D006358:Hot Temperature; D006868:Hydrolysis; D008264:Macrophages / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B / Q000378:metabolism; D010196:Pancreatic Elastase / Q000008:administration & dosage / Q000009:adverse effects* / Q000037:antagonists & inhibitors / Q000378:metabolism; D010195:Pancreatitis / Q000201:enzymology* / Q000276:immunology* / Q000473:pathology; D011112:Polymyxin B / Q000494:pharmacology; D011971:Receptors, Immunologic / Q000172:deficiency / Q000235:genetics / Q000502:physiology; D013552:Swine; D018746:Systemic Inflammatory Response Syndrome / Q000201:enzymology* / Q000276:immunology* / Q000401:mortality / Q000473:pathology; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D000943:Antigens, Differentiation; D001427:Bacterial Toxins; D004731:Endotoxins; C507780:Myd88 protein, mouse; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; D011971:Receptors, Immunologic; D014409:Tumor Necrosis Factor-alpha; C027952:salmonella toxin; D005688:Galactosamine; D010196:Pancreatic Elastase; D011112:Polymyxin B