Neutrophil Extracellular Traps Provide a Grip on the Enigmatic Pathogenesis of Acute Pancreatitis
2015; Elsevier BV; Volume: 149; Issue: 7 Linguagem: Inglês
10.1053/j.gastro.2015.10.027
ISSN1528-0012
AutoresJuha T. Korhonen, Vikas Dudeja, Rajinder Dawra, Paul Kubes, Ashok K. Saluja,
Tópico(s)Abdominal vascular conditions and treatments
ResumoSee “Neutrophil extracellular traps induce trypsin activation, inflammation, and tissue damage in mice with severe acute pancreatitis,” by Merza M, Hartman H, Rahman M, et al, on page 1920. See “Neutrophil extracellular traps induce trypsin activation, inflammation, and tissue damage in mice with severe acute pancreatitis,” by Merza M, Hartman H, Rahman M, et al, on page 1920. James Hirsch reported in 1958 that histones can kill bacteria.1Hirsch J.G. Bactericidal action of histone.J Exp Med. 1958; 108: 925-944Crossref PubMed Scopus (253) Google Scholar He speculated that histones might be present in blood in the course of certain pathologic conditions, such as necrosis. The histones, which Hirsch used in his experiments, were extracted from neutrophils. Almost a half century later, Zychlinsky et al2Brinkmann V. Reichard U. Goosmann C. et al.Neutrophil extracellular traps kill bacteria.Science. 2004; 303: 1532-1535Crossref PubMed Scopus (6167) Google Scholar observed that neutrophils externalize granular proteins and nuclear components, including DNA and histones, in response to bacterial stimulation, which contribute to bacterial killing. These web-like structures became known as neutrophil extracellular traps (NETs). After these pioneering studies, a considerable amount of literature has been published on NETs, and the concept of NET-mediated bacterial killing is increasingly accepted.3Yipp B.G. Kubes P. NETosis: how vital is it?.Blood. 2013; 122: 2784-2794Crossref PubMed Scopus (583) Google Scholar Extracellular histones are not only detrimental to bacteria, but also toxic to eukaryotic hosts. Esmon et al4Xu J. Zhang X. Pelayo R. et al.Extracellular histones are major mediators of death in sepsis.Nat Med. 2009; 15: 1318-1321Crossref PubMed Scopus (1059) Google Scholar were first to demonstrate that histones discharged in sepsis can be lethal. It has been shown that neutrophils release NETs during sepsis and entrap bacteria in blood,4Xu J. Zhang X. Pelayo R. et al.Extracellular histones are major mediators of death in sepsis.Nat Med. 2009; 15: 1318-1321Crossref PubMed Scopus (1059) Google Scholar but it is still not known whether NETs or injured parenchymal tissue is the primary source of extracellular histones and culprit of organ failure in sepsis. Acute pancreatitis is an inflammatory disorder of pancreas for which no specific treatment is available. The most severe cases of the disease are associated with mortality, with acute respiratory distress syndrome being the most frequent cause of death in the early phase of the disease.5Pandol S.J. Saluja A.K. Imrie C.W. et al.Acute pancreatitis: bench to the bedside.Gastroenterology. 2007; 132: 1127-1151Abstract Full Text Full Text PDF PubMed Scopus (470) Google Scholar Decades of research have provided some insight into the pathogenesis of the disease, but our understanding of the early intra-acinar events as well as the events which lead to overflow of inflammation from local to systemic levels is evolving. Experimental animal models of acute pancreatitis have suggested that the pancreatic acinar cell is the site of injury initiation.6Saluja A.K. Lerch M.M. Phillips P.A. et al.Why does pancreatic overstimulation cause pancreatitis?.Annu Rev Physiol. 2007; 69: 249-269Crossref PubMed Scopus (148) Google Scholar For example, in response to the injurious stimulus, typically cerulein (a cholecystokinin analog) supramaximal stimulation, a plethora of intra-acinar events are initiated.6Saluja A.K. Lerch M.M. Phillips P.A. et al.Why does pancreatic overstimulation cause pancreatitis?.Annu Rev Physiol. 2007; 69: 249-269Crossref PubMed Scopus (148) Google Scholar There is also altered trafficking of zymogens with co-localization of the zymogens and lysosomes leading to formation of co-localization organelles. In these organelles the digestive enzymes come into contact of lysosomal enzymes.7Saluja A. Saluja M. Villa A. et al.Pancreatic duct obstruction in rabbits causes digestive zymogen and lysosomal enzyme colocalization.J Clin Invest. 1989; 84: 1260-1266Crossref PubMed Scopus (139) Google Scholar, 8Saluja A.K. Donovan E.A. Yamanaka K. et al.Cerulein-induced in vitro activation of trypsinogen in rat pancreatic acini is mediated by cathepsin B.Gastroenterology. 1997; 113: 304-310Abstract Full Text PDF PubMed Scopus (172) Google Scholar Experimental data suggest that in these co-localized organelles cathepsin B activates trypsinogen to trypsin.9Halangk W. Lerch M.M. Brandt-Nedelev B. et al.Role of cathepsin B in intracellular trypsinogen activation and the onset of acute pancreatitis.J Clin Invest. 2000; 106: 773-781Crossref PubMed Scopus (460) Google Scholar, 10Van Acker G.J. Saluja A.K. Bhagat L. et al.Cathepsin B inhibition prevents trypsinogen activation and reduces pancreatitis severity.Am J Physiol Gastrointest Liver Physiol. 2002; 283: G794-G800Crossref PubMed Scopus (114) Google Scholar The popular belief was that this activated trypsin, through some unclear mechanism, leads to acinar cell death. However, recent data suggest that once cathepsin B activates trypsin in co-localized organelles, this activated trypsin then permeabilizes the co-localized organelles leading to release of the contents of the lysosomal enzymes and zymogens into the cytosol. Intriguingly, once in the cytosol, cathepsin B but not trypsin then activates the cell death pathway (Figure 1). In this issue of Gastroenterology, Merza et al11Merza M. Hartman H. Rahman M. et al.Neutrophil extracellular traps induce trypsin activation, inflammation, and tissue damage in mice with severe acute pancreatitis.Gastroenterology. 2015; 149: 1920-1931Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar have reported that neutrophils aggravate inflammation in acute pancreatitis by expelling NETs and thus have found a link between these 2 phenomena. Using the L-arginine and sodium taurocholate–induced experimental animal models of acute pancreatitis, the authors demonstrate that during acute pancreatitis, neutrophils release extracellular DNA traps in the pancreas and at the sites of systemic injury, namely, the lungs. The authors detected NETs in the inflamed pancreas by ultrastructural imaging. This was also corroborated by increased concentrations of cell-free DNA in the circulation, as well as increase in extracellular histone levels in the pancreas. The authors further demonstrate that degradation of extracellular DNA by pretreatment with deoxyribonuclease (DNAse) or depletion of neutrophils by antineutrophil antibody reduced local acinar damage as well as systemic inflammation. Furthermore, administration of DNase decreased the concentrations of many inflammatory mediators in blood, including CXCL2, interleukin-6, HMGB1, and matrix metalloproteinase-9. Intriguingly, DNAse treatment decreased the local and systemic recruitment of the neutrophils and intra-acinar trypsinogen activation. The authors also demonstrate that NETs could activate the intra-acinar trypsin in vitro. The significance of these new findings need to be considered in the context of our current knowledge of the pathogenesis of acute pancreatitis and our increasing understanding of the role of trypsin activation as the central event in acute pancreatitis. Although these data are consistent with previous studies demonstrating the important role for neutrophils in the pathogenesis of the disease,5Pandol S.J. Saluja A.K. Imrie C.W. et al.Acute pancreatitis: bench to the bedside.Gastroenterology. 2007; 132: 1127-1151Abstract Full Text Full Text PDF PubMed Scopus (470) Google Scholar the authors show that the mechanism may involve NET production. Current evidence suggests that the infiltrating neutrophils increase the severity of injury and their depletion by antineutrophil serum or inhibiting their recruitment by targeting adhesion molecules has been shown to reduce the severity of acute pancreatitis.12Gukovskaya A.S. Vaquero E. Zaninovic V. et al.Neutrophils and NADPH oxidase mediate intrapancreatic trypsin activation in murine experimental acute pancreatitis.Gastroenterology. 2002; 122: 974-984Abstract Full Text Full Text PDF PubMed Scopus (209) Google Scholar Although multiple mechanisms of neutrophil induced injury are possible, studies suggest that infiltrating neutrophils can augment intra-acinar trypsinogen activation. For example, infiltrating neutrophils induce intra-acinar trypsin activation through products of NADPH oxidase, and both trypsin activation and pancreatic injury were reduced in NADPH oxidase-deficient mice.12Gukovskaya A.S. Vaquero E. Zaninovic V. et al.Neutrophils and NADPH oxidase mediate intrapancreatic trypsin activation in murine experimental acute pancreatitis.Gastroenterology. 2002; 122: 974-984Abstract Full Text Full Text PDF PubMed Scopus (209) Google Scholar However, the mechanism by which neutrophils activated intra-acinar trypsinogen is not clear. In their current paper, Merza et al11Merza M. Hartman H. Rahman M. et al.Neutrophil extracellular traps induce trypsin activation, inflammation, and tissue damage in mice with severe acute pancreatitis.Gastroenterology. 2015; 149: 1920-1931Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar have proposed a novel mechanism for this phenomenon. Interestingly, incubation of pancreatic acinar cells with neutrophil-derived NETs in vitro led to trypsin activation in the acinar cells similar to that observed with supramaximal dose of the secretagogoue cerulein. Furthermore, inhibition of the NETs in vitro by treatment with DNAse abolished the trypsin activation. The authors also demonstrated that co-incubation of acinar cells with NETs led to intra-acinar STAT3 activation. The mechanism by which NETs induce intra-acinar cell changes is not clear, but the data in their article suggest that histones could induce these changes. Thus, this study provides an additional link between the neutrophilic infiltration and trypsin activation in the acinar cells. What is the significance of premature trypsin activation in acinar cell injury has always been a point of contention. For over a century now, pancreatitis has been considered a disease of autodigestion and premature activation of trypsin has been central to this hypothesis. This central role of trypsin is based on the earlier studies where chemical inhibition of trypsin prevented or reduced many of the phenotypic changes of pancreatitis in animal models. Additional studies with adenoviral mediated intra-acinar trypsin activation techniques have supported this notion.13Ji B. Gaiser S. Chen X. et al.Intracellular trypsin induces pancreatic acinar cell death but not NF-kappaB activation.J Biol Chem. 2009; 284: 17488-17498Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar Furthermore, the association of mutations in trypsinogen or trypsin inhibitor genes with hereditary pancreatitis has further solidified the “trypsin central” hypothesis. Although these studies have been fundamental to our current understanding, there are some drawbacks to this approach, ranging from the specificity of the inhibitors to the artificial nature of adenoviral-mediated overexpression. However, in trypsinogen isoform 7 (T7) knockout mice no trypsin activation is observed either in vitro or in vivo thus confirming that the activation of T7 isoform is responsible for the trypsin activity observed during experimental acute pancreatitis.14Dawra R. Sah R.P. Dudeja V. et al.Intra-acinar trypsinogen activation mediates early stages of pancreatic injury but not inflammation in mice with acute pancreatitis.Gastroenterology. 2011; 141: 2210-2217.e2Abstract Full Text Full Text PDF PubMed Scopus (179) Google Scholar In this model, the absence of T7 completely abolishes supramaximal cerulein-induced acinar cell injury in vitro, but only partially reduces acinar cell necrosis in acute pancreatitis in vivo.14Dawra R. Sah R.P. Dudeja V. et al.Intra-acinar trypsinogen activation mediates early stages of pancreatic injury but not inflammation in mice with acute pancreatitis.Gastroenterology. 2011; 141: 2210-2217.e2Abstract Full Text Full Text PDF PubMed Scopus (179) Google Scholar Surprisingly, no reduction in local and systemic inflammation and intra-acinar nuclear factor-κB activation is observed in T7 knockout as compared with wild-type animals in vivo, which counters the paradigm where trypsin is central in the pathogenesis of pancreatitis.14Dawra R. Sah R.P. Dudeja V. et al.Intra-acinar trypsinogen activation mediates early stages of pancreatic injury but not inflammation in mice with acute pancreatitis.Gastroenterology. 2011; 141: 2210-2217.e2Abstract Full Text Full Text PDF PubMed Scopus (179) Google Scholar Similarly, and upon induction of chronic pancreatitis, atrophy, histomorphologic features, and chronic inflammation in the T7 knockout mice are virtually indistinguishable from that in the wild-type mice, thus challenging the trypsin-centric hypothesis.15Sah R.P. Dudeja V. Dawra R.K. et al.Cerulein-induced chronic pancreatitis does not require intra-acinar activation of trypsinogen in mice.Gastroenterology. 2013; 144: 1076-1085 e2Abstract Full Text Full Text PDF PubMed Scopus (73) Google Scholar Although these data suggest that trypsin may have some role in acinar cell damage, inflammation, which is the primary hallmark of acute and chronic pancreatitis, is largely independent of trypsin. The pioneering study from Merza et al11Merza M. Hartman H. Rahman M. et al.Neutrophil extracellular traps induce trypsin activation, inflammation, and tissue damage in mice with severe acute pancreatitis.Gastroenterology. 2015; 149: 1920-1931Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar suggests that neutrophils could contribute to local and remote organ damage in acute severe pancreatitis by releasing NETs. One caveat when looking for NETs is the possibility of released chromatin from dying cells of pancreatic parenchyma. Notably, it has been shown recently that histones released from inflamed pancreas can aggravate the disease.16Kang R. Zhang Q. Hou W. et al.Intracellular Hmgb1 inhibits inflammatory nucleosome release and limits acute pancreatitis in mice.Gastroenterology. 2014; 146: 1097-1107Abstract Full Text Full Text PDF PubMed Scopus (153) Google Scholar A commonly used tool to identify a role for NETs is the use of DNAse, which could also degrade DNA released from dying pancreatic cells and thereby reduce neutrophil recruitment. Use of PAD4-deficient mice might need to be carried out to prove the specific role of NETs in acute pancreatitis. No doubt the new findings made by Merza et al11Merza M. Hartman H. Rahman M. et al.Neutrophil extracellular traps induce trypsin activation, inflammation, and tissue damage in mice with severe acute pancreatitis.Gastroenterology. 2015; 149: 1920-1931Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar will stimulate further studies to address these important questions. Taken together, the new evidence described in this issue of Gastroenterology11Merza M. Hartman H. Rahman M. et al.Neutrophil extracellular traps induce trypsin activation, inflammation, and tissue damage in mice with severe acute pancreatitis.Gastroenterology. 2015; 149: 1920-1931Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar suggests that DNA and histones mediate inflammation in severe acute pancreatitis and, as the authors propose, this could be owing to neutrophil activation and release of NETs in the inflamed pancreas. These new data suggest also a compelling mechanism by which neutrophils activate intra-acinar trypsin. However, although trypsin may contribute, rather indirectly, to acinar cell damage, it is not the master initiator or regulator of inflammation during pancreatitis, as it was once considered. It is possible that the presence of NETs ensures the availability of local high concentrations of secondary triggers for acinar cell damage. Future studies should lead to identification of these secondary triggers and the mechanisms by which these modify pancreatic injury. We congratulate Merza et al11Merza M. Hartman H. Rahman M. et al.Neutrophil extracellular traps induce trypsin activation, inflammation, and tissue damage in mice with severe acute pancreatitis.Gastroenterology. 2015; 149: 1920-1931Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar for providing this important piece of information that gets us a bit closer to solving the convoluted puzzle of the pancreatitis. Neutrophil Extracellular Traps Induce Trypsin Activation, Inflammation, and Tissue Damage in Mice With Severe Acute PancreatitisGastroenterologyVol. 149Issue 7PreviewNeutrophils are involved in the development of acute pancreatitis (AP), but it is not clear how neutrophil-induced tissue damage is regulated. In addition to secreting antimicrobial compounds, activated neutrophils eliminate invading microorganisms by expelling nuclear DNA and histones to form extracellular web-like structures called neutrophil extracellular traps (NETs). However, NETs have been reported to contribute to organ dysfunction in patients with infectious diseases. We investigated whether NETs contribute to the development of AP in mice. Full-Text PDF Covering the CoverGastroenterologyVol. 149Issue 7PreviewNonalcoholic fatty liver disease (NAFLD) is the most common form of chronic liver disease in the United States and progression to its more advanced form, nonalcoholic steatohepatitis, leads to progressive fibrosis, cirrhosis, and its complications, including hepatocellular carcinoma and the need for liver transplantation. Previous studies demonstrating a link between PNPLA-3 genotype and hepatic steatosis and features of nonalcoholic steatohepatitis have suggested a genetic predisposition to NAFLD. Full-Text PDF
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