Gut-derived endotoxin translocation is the main aggravating mechanism of acute severe pancreatitis

Severe acute pancreatitis (SAP) is a serious systemic disease. It exacerbates when complicated with multiple organ dysfunction syndrome or failure (MODS or MOF). However, the aggravating mechanism of SAP is still unknown up to now. Study showed that maintaining integrity of intestinal mucosal barrier function by given effective antibiotics, selective digestive decontamination (SDD) and enteral nutrition therapy to the patients with SAP could significantly reduce infection of pancreatic necrotic tissue and improve the patient's outcome. Combining the findings of gut-derived bacteria in animals' pancreas, liver, spleen, mesenteric lymph nodes with increasing concentration of inflammatory cytokines and endotoxin in plasma with SAP, we hypothesize that gut-derived endotoxin translocation is the main aggravating mechanism of SAP. The hypothesis holds potential as a target for therapeutic intervention.     

Anti‐inflammatory effects of melatonin in a rat model of caerulein‐induced acute pancreatitis

The purpose of our study was to evaluate the protective effect of melatonin in a rat model of caerulein‐induced acute pancreatitis. For the induction of experimental acute pancreatitis, four subcutaneous injections of caerulein (20 µg kg^–1 body weight) were given to Wistar rats at 2‐h intervals. Melatonin was injected intraperitoneally (25 mg kg^–1 body weight) 30 min before each caerulein injection. After 12 h, rats were sacrificed by decapitation. Blood and pancreas samples were collected and processed for serological and histopathological studies, respectively. Lipase, α‐amylase, corticosterone, total antioxidant power and cytokines interleukin (IL)‐1β, IL‐4 and tumour necrosis factor (TNF)‐α were determined using commercial kits. ANOVA and Tukey tests (P < 0.05) were performed for the statistical analysis of the results. Results showed that the administration of melatonin reduced histological damage induced by caerulein treatment as well as the hyperamylasemia and hyperlipidemia. Corticosterone and antioxidant total power were also reverted to basal activities. Furthermore, melatonin pre‐treatment reduced pro‐inflammatory cytokines IL‐1β and TNF‐α and increased the serum levels of anti‐inflammatory cytokine IL‐4. In conclusion, the findings suggest that the protective effect of melatonin in caerulein‐induced acute pancreatitis is mediated by the anti‐inflammatory ability of this indolamine. Thus, melatonin may have a protective effect against acute pancreatitis. Copyright © 2013 John Wiley & Sons, Ltd.     

肠道菌群失衡与急性胰腺炎易位感染的关系

目的探讨急性胰腺炎患者肠道菌群与胰腺炎易位感染的关系。方法前瞻性地募集重症急性胰腺炎患者171名,根据是否发生易位感染,分为急性胰腺炎非感染组(131名)和急性胰腺炎易位感染组(40名),所有入选者入院时收集粪便标本和血清,分别采用实时荧光定量PCR和分光光度法测定肠道菌群组成及肠道屏障功能,探讨肠道菌群与胰腺炎肠道屏障以及易位感染的关系。结果急性胰腺炎易位感染组患者入院时肠道菌群中的肠杆菌以及肠球菌含量较非感染组显著增加,实时荧光定量PCR结果发现感染组患者潜在致病菌如肠球菌、肠杆菌显著高于非感染组,与肠道黏膜屏障功能的内毒素水平、二胺氧化酶活性以及D-乳酸含量显著正相关,而双歧杆菌显著低于非感染组,与内毒素水平、二胺氧化酶活性以及D-乳酸含量显著负相关,结果提示肠道菌群失衡可能是急性胰腺炎易位感染发生的主要原因之一。结论肠道菌群失衡是急性胰腺炎易位感染发生的高危因素,与急性胰腺炎易位感染发生相关。     

The recovery of acute pancreatitis depends on the enzyme amount stored in zymogen granules at early stages

Little is known about the changes in pancreatic enzyme storage in acute pancreatitis. We have performed flow cytometric studies of zymogen granules from rats with acute pancreatitis induced by hyperstimulation with caerulein. A comparison was made with rats treated with hydrocortisone (10 mg/kg/day) over 7 days before inducing pancreatitis in order to find out whether the amount of enzymes stored in the pancreas plays a key role in the development of pancreatitis. The potentially therapeutic effect of L-364,718 (0.1 mg/kg/day, for 7 days), a CCK receptor antagonist, was assayed in the rats with caerulein-induced pancreatitis which had previously received the hydrocortisone treatment. A significant increase in the intragranular enzyme content was observed 5 h after hyperstimulation with caerulein. The highest values were reached in the rats previously treated with hydrocortisone. The greatest pancreatic enzyme load was parallel to the highest values in plasma amylase, edema and haematocrit observed. Acute pancreatitis was reversed seven days later. At this stage smaller granules appeared in the pancreas whose enzyme content was similar to that of controls when no treatment was applied after pancreatitis. In contrast, L-364,718 administration prevented the favourable evolution of pancreatitis since the antagonism exerted on CCK receptors induced a blockade of secretion of the large amounts of enzymes stored in the pancreas. Moreover, the enzyme content in zymogen granules was below normal values since the stimulatory CCK action on enzyme synthesis can be inhibited by L-364,718. Our results suggest that the efficiency of CCK antagonists, as potential therapy, would also depend on the load of enzymes in the pancreas when acute pancreatitis is produced.     

The role of oxygen radicals in experimental acute pancreatitis

Oxygen-derived free radicals mediate an important step in the initiation of experimental acute pancreatitis. Thereby, it seems that these reactive oxygen metabolites are generated at an early stage of disease. The source of the enhanced production of oxygen radicals still remains unclear. Experimentally, the efficiency of scavenger treatment varied between different models, whereby these differences depended on the experimental model and not on the form of pancreatitis which was induced. Most studies pretreated the experimental animals before inducing acute pancreatitis. This does not mirror the clinical reality, since patients are admitted to the hospital after onset of the disease. It was shown in Cerulein pancreatitis, however, that scavenger treatment also mitigated the pancreatic tissue damages after induction of acute pancreatitis. Moreover, antioxidant treatment also attenuated the extrapancreatic complications, thus improving the final outcome of the disease. The first indirect observations also suggest that in human acute recurrent and chronic pancreatitis, oxygen free radicals are generated and add to the damages seen. Therefore, well-defined controlled clinical studies with patients suffering from acute pancreatitis are needed to validate the role of oxygen radicals in this disease.     

Pro-inflammatory effects of hydrogen sulphide on substance P in caerulein-induced acute pancreatitis

Hydrogen sulphide (H(2)S), a novel gasotransmitter, has been recognized to play an important role in inflammation. Cystathionine-gamma-lyase (CSE) is a major H(2)S synthesizing enzyme in the cardiovascular system and DL-propargylglycine (PAG) is an irreversible inhibitor of CSE. Substance P (SP), a product of preprotachykinin-A (PPT-A) gene, is a well-known pro-inflammatory mediator which acts principally through the neurokinin-1 receptor (NK-1R). We have shown an association between H(2)S and SP in pulmonary inflammation as well as a pro-inflammatory role of H(2)S and SP in acute pancreatitis. The present study was aimed to investigate the interplay between pro-inflammatory effects of H(2)S and SP in a murine model of caerulein-induced acute pancreatitis. Acute pancreatitis was induced in mice by 10 hourly intraperitoneal injections of caerulein (50 (g/kg). PAG (100 mg/kg, i.p.) was administered either 1 hr before (prophylactic) or 1 hr after (therapeutic) the first caerulein injection. PAG, given prophylactically as well as therapeutically, significantly reduced plasma H(2)S levels and pancreatic H(2)S synthesizing activities as well as SP concentrations in plasma, pancreas and lung compared with caerulein-induced acute pancreatitis. Furthermore, prophylactic as well as therapeutic administration of PAG significantly reduced PPT-A mRNA expression and NK-1R mRNA expression in both pancreas and lung when compared with caerulein-induced acute pancreatitis. These results suggest that the pro-inflammatory effects of H(2)S may be mediated by SP-NK-1R pathway in acute pancreatitis.     

Serum free prostate-specific antigen and zinc levels in experimental acute pancreatitis

Serum free prostate-specific antigen (fPSA) is the most useful tumor marker for prostatic cancer screening. However, recently, fPSA has also been detected in sera from patients with pancreatic diseases. In addition, it has been shown that zinc (Zn) concentration might change in both serum and tissues in pancreatic disease. In the present study, we measured serum concentrations of fPSA and Zn as possible markers and prognostic factors in an experimental acute-pancreatitis model. Twenty-five female Wistar albino rats were divided into two groups: the control group (n=10) and the experimental group (n=15). Acute pancreatitis was induced by injection of ethyl alcohol into the common biliary duct. The animals were sacrificed 24 h later to detect the concentrations of serum fPSA and Zn. fPSA values were detected to be significantly higher in the experimental group p<0.001). There was also a significant decrease in the serum Zn level of the acute-pancreatitis group (p<0.001). In conclusion, these findings suggested that a combination of these parameters might represent a significant improvement on the diagnostic value of each of them separately and provide a powerful tool for differential diagnosis and prognosis in pancreatic diseases.     

Age-dependent vulnerability to experimental acute pancreatitis is associated with increased systemic inflammation and thrombosis

The severity and mortality rates of acute pancreatitis (AP) are significantly elevated in the elderly population. However, due to a lack of appropriate animal models, the underlying mechanisms for this age‐dependent vulnerability remain largely unknown. The purpose of this study was to characterize a murine model of AP, which displays age‐associated severity, and to use this model to identify pathophysiologies that are distinctive of the aged with AP. AP was induced in young (4–5 months), middle‐aged (12–13 months), and aged (23–25 months) C57BL/6 mice by repeated injection of caerulein, a homologue of the gastrointestinal hormone cholecystokinin. Approximately 10% of aged mice died during AP, while young and middle‐aged mice showed no mortality. Although both young and aged mice exhibited early signs of edema and inflammation in the pancreas, kidney, and lung, young mice showed signs of recovery within 24 h, while aged mice exhibited increasingly severe tissue damage and cell death. There was a significant age‐dependent increase in pancreatic neutrophil activation and systemic inflammation as assessed by pancreatic myeloperoxidase and plasma interleukin‐6 (IL‐6) concentration, respectively. Importantly, aged but not young mice with AP showed significantly elevated thrombosis in the lung and kidney as well as a marked increase in plasma concentration of plasminogen activator inhibitor‐1 (PAI‐1), a primary inhibitor of the fibrinolytic system. These results demonstrate that aging is associated with increased severity of AP characterized by augmented and prolonged pancreatic inflammation and the presence of multiple extra‐pancreatic sequelae including thrombosis.     

Kynurenine monooxygenase regulates inflammation during critical illness and recovery in experimental acute pancreatitis

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Proteomic profiling in an animal model of acute pancreatitis

Acute pancreatitis (AP) is an inflammatory disease of the pancreas, which evolves in approximately 20% of the patients to a severe illness associated with a high mortality rate. In this study, we performed a comparative proteomic analysis of pancreatic tissue extracts from rats with AP and healthy rodent controls in order to identify changes in protein expression related to the pathobiological processes of this disease. Pancreatic extracts from diseased and controls rats were analyzed by 2-DE and MS/MS. A total of 125 proteins were identified from both samples. Comparative analysis allowed the detection of 42 proteins or protein fragments differentially expressed between diseased and control pancreas, some of them being newly described in AP. Interestingly, these changes were representative of the main pathobiological pathways involved in this disease. We observed activation of digestive proteases and increased expression of various inflammatory markers, including several members of the alpha-macroglobulin family. We also detected changes related to oxidative and cell stress responses. Finally, we highlighted modifications of 14-3-3 proteins that could be related to apoptosis regulation. These results showed the interest of proteomic analysis to identify changes characterizing pancreatic tissue damage and, therefore, to highlight new potential biomarkers of AP.     

Pathobiology of experimental acute pancreatitis.

Pancreatic duct obstruction, even in the absence of biliary obstruction and/or bile reflux into the pancreatic duct, can trigger acute hemorrhagic necrotizing pancreatitis. The earliest changes are seen within acinar cells. Early derangements in acinar cell biology include inhibition of digestive enzyme secretion and the co-localization of lysosomal hydrolases with digestive enzyme zymogens. Under appropriate conditions, this co-localization could lead to digestive enzyme activation within acinar cells.     

Protective effect of CR 1409 (cholecystokinin antagonist) on experimental pancreatitis in rats and mice

CR 1409, a glutaramic acid derivative with competitive cholecystokinin-antagonistic activity, was administered IP and evaluated in comparison with proglumide (the model CCK-receptor antagonist), gabexate (protease inhibitor) and PGE₂ (cytoprotective) on two different models of experimental pancreatitis. Acute pancreatitis was induced in mice by six IP injections of 50 μg/kg caerulein at hourly intervals. The drugs were administered 30 minutes before each caerulein administration. Blood samples and pancreata were collected 3 hours after the last caerulein injection. In the second experiment, pancreatitis was induced in rats by injecting 0.3 ml 6% sodium taurocholate interstitially into the pancreas. The drugs were administered twice, 30 minutes before and 3 hours after taurocholate. The animals were killed 6 hours after laparotomy and blood samples and pancreata were collected. CR 1409 exhibited on both pancreatitis models a protective effect in a dose range of 0.3–10 mg/kg. Proglumide exhibited a protective activity at higher doses (200–400 mg/kg). Gabexate and PGE₂ were effective only in pancreatitis induced by taurocholate in a dose range of 30–60 mg/kg and 60–130 μg/kg respectively. These results, showing a high protective effect of CR 1409 on different models of acute pancreatitis, suggest an important role of CCK in the pathogenesis of pancreatitis.     

Central role of neutrophil in the pathogenesis of severe acute pancreatitis

Severe acute pancreatitis (SAP) is an acute abdominal disease with the strong systemic inflammatory response, and rapidly progresses from a local pancreatic damage into multiple organ dysfunction. For many decades, the contributions of neutrophils to the pathology of SAP were traditionally thought to be the chemokine and cytokine cascades that accompany inflammation. In this review, we focus mainly on those recently recognized aspects of neutrophils in SAP processes. First, emerging evidence suggests that therapeutic interventions targeting neutrophils significantly lower tissue damage and protect against the occurrence of pancreatitis. Second, trypsin activation promotes the initial neutrophils recruitment into local pancreas, and subsequently neutrophils infiltration in turn triggers trypsin production. Finally, neutrophils have the unique ability to release neutrophil extracellular traps even in the absence of pathogens.     

苦碟子注射液细菌内毒素检查法的探讨

目的：建立苦碟子注射液静脉点滴用药时细菌内毒素的检查方法。方法：采用2个厂家的鲎试剂对苦碟子注射液进行干扰试验。结果：鲎试剂浓度为0．25EU／mL时,苦碟子注射液稀释至1／30对细菌内毒素测定无干扰。结论：鲎试剂法检测苦碟子注射液静脉点滴用药时的细菌内毒素是可行的。     

Changes of serum glycans during sepsis and acute pancreatitis

Acute inflammatory response is a complex process associated with the production of both pro- and anti-inflammatory mediators. Although it is generally considered to be a single homeostatic mechanism, there are differences associated with the nature and the site of inflammation. We examined the changes of N-linked glycans released from the serum of a patient with sepsis and a patient with acute pancreatitis during the first eight days of the disease. Sera were taken from patients at the time of reporting to hospital and then three more times. The blood from a healthy individual was drawn on one occasion only. Glycans were released using N-glycosidase F and were subjected to normal phase and weak anion exchange high-performance liquid chromatography, exoglycosidase digestions, and mass spectrometry. The levels of identified structures have been followed through the course of disease and compared to the control levels. Changes in serum glycans were found to occur very early in acute inflammation. The most prominent differences include the increase in ratio of outer arm to core fucose, increase in the amount of tetrasialylated structures, changes in the levels of mannose structures, and in the degree of branching. The relative proportions of different glycans changed daily and some differences were also observed between sepsis and pancreatitis, probably reflecting that in these two conditions, the acute phase response is triggered by a different stimulus that is associated with different patterns of production of cytokines.