Trypsinogen Activation Peptide Research Articles

Evolution of trypsinogen activation peptides.

The activation peptide of mammalian trypsinogens contains a highly conserved tetra-aspartate sequence (D19-D20-D21-D22) preceding the K23-I24 scissile peptide bond, which is hydrolyzed as the first step in the activation process. Here, we examined the evolution and function of trypsinogen activation peptides through integrating functional characterization of disease-associated mutations with comparative genomic analysis. Activation properties of three chronic pancreatitis-associated activation peptide mutants (the novel D19A and the previously reported D22G and K23R) were simultaneously analyzed, for the first time, in the context of recombinant human cationic trypsinogen. A dramatic increase in autoactivation of cationic trypsinogen was observed in all three mutants, with D22G and K23R exhibiting the most marked increases. The physiological activator enteropeptidase activated the D19A mutant normally, activated the D22G mutant very poorly, and stimulated activation of the K23R mutant. The biochemical and structural data, taken together with a comprehensive sequence comparison, indicates that the tetra-aspartate sequence in mammalian trypsinogen activation peptides has evolved not only for optimal enteropeptidase recognition in the duodenum but also for efficient inhibition of trypsinogen autoactivation within the pancreas. Moreover, the use of lysine instead of arginine at the P1 position of activation peptides also has an advantageous effect against trypsinogen autoactivation. Finally, fixed substitutions in the key residues of the trypsinogen activation peptide may suggest the evolution of new functions unrelated to digestion, as found in the group III trypsinogens of cold-adapted fishes.

Expression, purification, and characterization of human enteropeptidase catalytic subunit in Escherichia coli

Enteropeptidase (synonym: enterokinase, EC 3.4.21.9) is a heterodimeric serine protease of the intestinal brush border that activates trypsinogen by highly specific cleavage of the trypsinogen activation peptide following the sequence (Asp) 4-Lys. The DNA sequence encoding the light chain (catalytic subunit) of human enteropeptidase (GenBank Accession No. U09860) was synthesized from 26 oligonucleotides by polymerase chain reaction and cloned into plasmid pET-32a downstream to the gene of fusion partner thioredoxin immediately after the DNA sequence encoding enteropeptidase recognition site. The fusion protein thioredoxin/human enteropeptidase light chain was expressed in Escherichia coli BL21(DE3) strain in both soluble and insoluble forms. The soluble recombinant fusion protein failed to undergo autocatalytic cleavage and activation; however, autocatalytic cleavage and activation of recombinant human enteropeptidase light chain (L-HEP) were achieved by solubilization and renaturation of the fusion protein from inclusion bodies and the active L-HEP was purified on agarose-linked soybean trypsin inhibitor. The purified L-HEP cleaved the synthetic peptide substrate Gly-Asp-Asp-Asp-Asp-Lys-β-naphthylamide with kinetic parameters K m=0.16 mM and k cat=115 s −1 and small ester Z-Lys-SBzl with K m =140 μ M, k cat=133 s −1. L-HEP associated with soybean trypsin inhibitor slowly and small ester Z-Lys-SBzl cleavage was inhibited with K i *=2.3 nM. L-HEP digested thioredoxin/human epidermal growth factor fusion protein five times faster than equal activity units of bovine recombinant light chain (EKMax, Invitrogen) at the same conditions.

Trypsinogen-2 and trypsinogen activation peptide (TAP) in urine of patients with acute pancreatitis

Background and aims. There is an obvious clinical need for a simple test that can identify patients at risk of developing severe acute pancreatitis. In this work we compared urinary trypsinogen-2 with urinary trypsinogen activation peptide (TAP) and serum C-reactive protein (CRP) for early differentiation between mild and severe acute pancreatitis. Patients and methods. The study population consisted of 127 consecutive patients with acute pancreatitis of whom 29 had severe disease. Urinary trypsinogen-2 was measured by a quantitative immunofluorometric assay and TAP by a competitive immunoassay. Serum CRP was determined by immunoturbidimetry. Results. The sensitivity and specificity to identify severe acute pancreatitis on admission was 72% and 81% for urinary trypsinogen-2, 64% and 82% for urinary TAP, and 29% and 93% for serum CRP, respectively. At 24 h after admission, the values were 82% and 78% for urinary trypsinogen-2, 52% and 92% for urinary TAP, and 84% and 72% for serum CRP, respectively. Receiver-operating characteristics curve analysis showed that the area under the curve was larger for urinary trypsinogen-2 than for urinary TAP and serum CRP on admission and 24 h after admission. On admission the positive likelihood ration for urinary trypsiongen-2 was 3.7, for urinary TAP 3.6, and 4.3 for serum CRP, respectively. The corresponding negative likelihood ratios were 0.34, 0.43, and 0.76, respectively. Conclusion. Urinary trypsinogen-2 was superior to serum CRP and as god as or even better than urinary TAP and in the early prediction of disease severity in acute pancreatitis. These results suggest that it could be a valuable adjunct in the early assessment of the severity of acute pancreatitis.

Mechanism of kinin release during experimental acute pancreatitis in rats: evidence for pro- as well as anti-inflammatory roles of oedema formation.

1 Kinin B(2) receptor antagonists or tissue kallikrein (t-KK) inhibitors prevent oedema formation and associated sequelae in caerulein-induced pancreatitis in the rat. We have now further investigated the mechanism of kinin generation in the pancreas. 2 Kinins were elevated in the pancreatic tissue already before oedema formation became manifest. Peak values (421+/-59 pmol g(-1) dry wt) were reached at 45 min and remained elevated for at least 2 h; a second increase was observed at 24 h. Pretreatment with the B(2) receptor antagonist icatibant abolished kinin formation, while post-treatment was ineffective. 3 Total kininogen levels were very low in the pancreas of controls, but increased 75-fold during acute pancreatitis. This increase was absent in rats that were pretreated with icatibant. 4 During pancreatitis, t-KK-like and plasma kallikrein (p-KK)-like activity in the pancreas, as well as trypsinogen activation peptide (TAP) increased significantly. Icatibant pretreatment further augmented t-KK about 100-fold, while p-KK was significantly attenuated; TAP levels remained unaffected. 5 Endogenous protease inhibitors (alpha(1)-antitrypsin, alpha(2)-macroglobulin) were low in normal tissues, but increased 45- and four-fold, respectively, during pancreatitis. This increase was abolished when oedema formation was prevented by icatibant. 6 In summary, oedema formation is initiated by t-KK; the ensuing plasma protein extravasation supplies further kininogen and active p-KK to the tissue. Concomitantly, endogenous protease inhibitors in the oedema fluid inhibit up to 99% of active t-KK. Our data thus suggest a complex interaction between kinin action and kinin generation involving positive and negative feedback actions of the inflammatory oedema.

Influence of pancreatic enzyme content on experimental acute pancreatitis.

Some studies demonstrate the crucial role of proteases in the pathogenesis of acute pancreatitis (AP). Systemic release of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) has been demonstrated in AP, yet the mechanism of activation remains unclear. Furthermore, it is not known if the amount of pancreatic enzyme in the pancreas determines the production of proinflammatory cytokines. To determine whether there is a link between the pancreatic enzyme content and the production of cytokines and consequently the systemic lesions observed in AP. Forty-seven animals were divided into three groups: group I had a high pancreatic enzyme level (with and without AP), group II had a low pancreatic enzyme level (cerulein infusion: 0.133 microg x kg(-1) x h(-1)) (with and without AP), and group III were the controls. AP was induced by injection of 5% sodium taurocholate into the pancreatic duct. To evaluate the pancreatic enzyme contents before AP, trypsinogen and amylase analysis was carried out on pancreatic tissue collected after the animals were killed. Two hours after induction of AP, concentrations of pancreatic enzymes and trypsinogen activation peptide (TAP) in serum, ascitic fluid, and pancreatic tissue were determined. The ascitic fluid was assayed for TNF-alpha and the serum was assayed for IL-6 with ELISA kits. Systemic lesions were sought on the basis of hepatic mitochondrial respiratory function measured polarographically. The administration of physiological doses of cerulein diminishes the pancreatic enzyme and TAP levels, the production of proinflammatory cytokines, and the liver mitochondrial dysfunction observed in AP, suggesting that the pancreatic enzyme content is an important factor in the severity of AP.

The anti-inflammatory effect of methylprednisolone occurs down-stream of nuclear factor-κB DNA binding in acute pancreatitis

Glucocorticoids are potent anti-inflammatory drugs. The molecular mechanisms underlying these effects have not yet been fully revealed. The aim of the present study was to establish whether methylprednisolone pretreatment is beneficial and if it can block the pancreatic DNA binding of the transcription factor nuclear factor-κB (NF-κB) and proinflammatory cytokine synthesis during cholecystokinin-octapeptide (CCK)-induced acute pancreatitis in rats. Additionally, we set out to investigate the potential effects of methylprednisolone and CCK on pancreatic heat shock protein (HSP) synthesis. The dose–response (5–40 mg/kg) and time–course (6–72 h) curves of methylprednisolone on pancreatic HSP60 and HSP72 synthesis were evaluated following methylprednisolone treatment. We demonstrated that methylprednisolone specifically and dose-dependently induced HSP72 in the pancreas of rats, while it did not have a significant effect on HSP60 expression. The pancreatitis was induced near the peak level of HSP72 synthesis (2×30 mg/kg body weight [b.w.] methylprednisolone i.m. at an interval of 12 h, followed by a 12-h recovery period after the second injection of methylprednisolone) by administering 2×100 μg/kg CCK subcutaneously at an interval of 1 h. The injections of CCK in the vehicle-pretreated group significantly elevated the levels of pancreatic HSP60 and HSP72 2–4 h after the second CCK injection. Methylprednisolone pretreatment ameliorated many of the examined laboratory (the pancreatic weight/body weight [p.w./b.w.] ratio, the serum amylase activity, the plasma trypsinogen activation peptide concentration, the pancreatic levels of tumor necrosis factor-α and interleukin-6, the degree of lipid peroxidation, protein oxidation, nonprotein sulfhydryl group content and the pancreatic myeloperoxidase activity) and morphological parameters of the disease. Methylprednisolone pretreatment did not influence pancreatic NF-κB DNA binding, but decreased proinflammatory cytokine synthesis in this acute pancreatitis model. The findings suggest that the anti-inflammatory effect of large doses of methylprednisolone in secretagogue-induced pancreatitis occurs downstream of NF-κB DNA binding, and that increased pancreatic HSP72 synthesis may play a role in the protective effect of the drug.

Assessing the severity of canine pancreatitis

The objective of this study was to determine whether laboratory testing currently available is able to provide prognostic information in canine pancreatitis. A prospective study of dogs with naturally occurring pancreatitis was undertaken. Twenty-two cases with histologically confirmed pancreatic inflammation were included in the study. Each dog had routine haematology parameters, serum biochemistry (including lipase and amylase), serum trypsin-like immunoreactivity and trypsinogen activation peptides (TAP) in urine and plasma measured. Twelve of the dogs were classified as having severe disease. These dogs had statistically significant increases in urinary TAP–creatinine ratio (UTCR) measurement, serum lipase, serum phosphate and serum creatinine concentrations. Additionally dogs with severe pancreatitis had significantly decreased urine specific gravity levels. The most sensitive and specific test to assess the severity of pancreatitis was the measurement of UTCR.

Sequential Changes in Pancreatic Markers in Acute Pancreatitis

Background: Trypsinogen activation within acinar cells plays a crucial role in the pathogenesis of acute pancreatitis (AP). Our aim was to characterize temporal changes of trypsinogen-1, trypsinogen-2, complexes of trypsin-1- ! 1 -antitrypsin (T1-AAT) and trypsin-2- ! 1 -antitrypsin (T2-AAT), trypsinogen activation peptide (TAP) and pancreatic secretory trypsin inhibitor (PSTI) in patients with AP. Methods: The study comprised 64 consecutive patients with AP (19 with severe disease) and 32 controls. The concentrations of trypsinogen-1 and -2, PSTI, T1-AAT and T2-AAT were determined by time-resolved immunofluorometric assays (IFMA), and TAP was measured using a competitive enzyme immunoassay from serum and urine. Results: The concentrations of trypsinogen-1 and -2 in serum reflected similar patterns, but excretion of trypsinogen-1 into urine was markedly lower than that of trypsinogen-2, the concentration of which correlated strongly with disease severity. The concentrations of T1-AAT were no higher in severe AP than in mild AP, while T2-AAT concentrations were significantly higher in severe than in mild disease. PSTI increased over the course of several days, showing strong correlation with disease severity. The concentrations of plasma and urinary TAP decreased rapidly to undetectable levels. During the early phase of AP, TAP correlated with the disease severity in plasma and urine but there was no difference between controls and patients with mild AP. Conclusion: More pronounced changes in trypsinogen-2 and its complex with AAT than in those of trypsinogen-1 were demonstrated, suggesting that trypsinogen-2 might play a more important role in the pathogenesis of AP than earlier believed. Urinary PSTI showed features warranting further investigations as a marker of disease severity.

The pH modulator chloroquine blocks trypsinogen activation peptide generation in cerulein-induced pancreatitis.

We examined the effects of a weak base, chloroquine, on the trypsinogen processing in cerulein-induced pancreatitis. Immunofluorescence studies were performed using newly generated affinity-purified antibodies to the trypsinogen activation peptide (TAP). The present study showed that chloroquine pretreatment blocked intracellular TAP generation in cerulein-induced pancreatitis. These results indicate that intracellular trypsinogen activation, which plays an important role in acute pancreatitis, requires a low-pH compartment, as well as serine protease activity.

Urinary trypsinogen activation peptide is more accurate than hematocrit in determining severity in patients with acute pancreatitis: a prospective study

OBJECTIVE: The management of patients with acute pancreatitis is complicated by the inability to distinguish mild from severe disease during the early stages. It has been previously shown that urinary trypsinogen activation peptide (TAP) and hematocrit (Hct) may serve as early predictors of severity in patients with acute pancreatitis. To establish which marker is more accurate in the determination of severity in patients with acute pancreatitis, a prospective study was performed. METHODS: A consecutive series of patients admitted with pain consistent with acute pancreatitis and an amylase of three times the upper limit of normal were included. The admission and 24-h Hct was obtained. A urine sample was obtained within 12 h of admission. Urinary TAP was determined using a modified solid phase ELISA. Severity was defined by the Atlanta Symposium, as the presence of organ failure and/or pancreatic necrosis. RESULTS: Fifty-eight consecutive patients with acute pancreatitis participated. There were 33 men and 25 women with a mean age of 60 ± 19. Thirty-nine patients had mild disease; 19 had severe disease. Urinary TAP was elevated in 26 patients. All patients with severe pancreatitis were correctly identified as having severe disease by an elevated urinary TAP (sensitivity 100%, specificity 77%). The admission Hct was higher than 47 in only three patients, all with mild disease. Of the patients with a rise in Hct, eight had mild disease, and only one had severe disease. Using a Hct of 44 as a cutoff did not affect the accuracy. There was no association between a rise in Hct and failure of Hct to decrease in the determination of severity. CONCLUSIONS: In comparison to admission Hct, urinary TAP was more accurate in determining severity in patients with acute pancreatitis by Atlanta, APACHE II, and Ranson criteria. We conclude that urinary TAP is a more accurate predictor of severity in patients with acute pancreatitis compared with Hct. Urinary TAP should be used to determine severity in patients early in the course of acute pancreatitis.

Urinary trypsinogen activation peptide is more accurate than hematocrit in determining severity in patients with acute pancreatitis: a prospective study.

The management of patients with acute pancreatitis is complicated by the inability to distinguish mild from severe disease during the early stages. It has been previously shown that urinary trypsinogen activation peptide (TAP) and hematocrit (Hct) may serve as early predictors of severity in patients with acute pancreatitis. To establish which marker is more accurate in the determination of severity in patients with acute pancreatitis, a prospective study was performed. A consecutive series of patients admitted with pain consistent with acute pancreatitis and an amylase of three times the upper limit of normal were included. The admission and 24-h Hct was obtained. A urine sample was obtained within 12 h of admission. Urinary TAP was determined using a modified solid phase ELISA. Severity was defined by the Atlanta Symposium, as the presence of organ failure and/or pancreatic necrosis. Fifty-eight consecutive patients with acute pancreatitis participated. There were 33 men and 25 women with a mean age of 60 +/- 19. Thirty-nine patients had mild disease; 19 had severe disease. Urinary TAP was elevated in 26 patients. All patients with severe pancreatitis were correctly identified as having severe disease by an elevated urinary TAP (sensitivity 100%, specificity 77%). The admission Hct was higher than 47 in only three patients, all with mild disease. Of the patients with a rise in Hct, eight had mild disease, and only one had severe disease. Using a Hct of 44 as a cutoff did not affect the accuracy. There was no association between a rise in Hct and failure of Hct to decrease in the determination of severity. In comparison to admission Hct, urinary TAP was more accurate in determining severity in patients with acute pancreatitis by Atlanta, APACHE II, and Ranson criteria. We conclude that urinary TAP is a more accurate predictor of severity in patients with acute pancreatitis compared with Hct. Urinary TAP should be used to determine severity in patients early in the course of acute pancreatitis.

Cathepsin B cleavage of the trypsinogen activation peptide.

BackgroundCathepsin B is thought to play a central role in intrapancreatic trypsinogen activation and the onset of pancreatitis. A recent investigation of the cathepsin B mediated activability of wildtype trypsinogen and their mutations N29I, N29T and R122H, which are associated to hereditary pancreatitis, revealed no differences. This action seems to be restricted to the K23-I24 peptide bond, which is the trypsinogen activation bond. Here we investigated the influence of the mutations D22G and K23R of the trypsinogen activation peptide on the cleavability by cathepsin B.MethodsTo investigate the functional impact of the TAP mutations on cathepsin B mediated cleavage of the trypsinogen activating K23-I24 bond, the corresponding peptides pWT, APFDDDDKIVGG; pD22G, APFDDDGKIVGG; and pK23R, APFDDDDRIVGG were digested with cathepsin B for 30 min at pH 3.8 and 5.0, and the fragments were analysed by high-performance liquid chromatography.ResultsWithout cathepsin B, less than 1 % of the peptides were hydrolysed. After a 30-minute digestion with cathepsin B at pH 5, 96% of pWT, 48% of pK23R, but only 2.4% of pD22G were hydrolysed. At pH 3.8, the cathepsin B cleavage of pWT and pK23R was less than at pH 5, whereas the cleavage of pD22G was completely inhibitedConclusionsCathepsin B mediated trypsinogen activation seems not to be a crucial pathogenic step in hereditary pancreatitis patients with the trypsinogen mutations D22G and K23R.

Comparison of Celsior and UW Solution in Experimental Pancreas Preservation

Background. The University of Wisconsin solution (UW) is the gold standard for pancreas preservation. Celsior (CEL) was formulated specifically for heart preservation. Recently, experimental and clinical experience has been reported on the application of CEL to abdominal organs. In this animal study, pancreas preservation with CEL was compared with that in UW solution.Patients and Materials. Heterotopic, allogeneic pancreaticoduodenal transplantation was performed in female Göttingen Minipigs (n = 12 donors, n = 12 recipients). The grafts were flushed and stored for 6 h at 4°C in UW or CEL. The recipients were randomized into two groups receiving either UW (n = 6)- or CEL (n = 6)-preserved grafts with a follow-up of 5 days. Blood flow (laser Doppler), partial oxygen tension, histological changes, endothelin-1 (plasma, immunohistochemistry), lipase, amylase, trypsinogen activation peptide, and C-reactive protein (CRP) were measured.Results. Partial oxygen tension was lower in the CEL group (P < 0.05). However, blood flow did not differ between UW- and CEL-preserved organs. The histomorphologic analysis of the pancreatic grafts revealed significantly less edema in the UW-preserved organs. Serum levels of amylase, lipase, CRP, and TAP taken from the central venous blood were comparable in the two groups, except for higher amylase values 36 h after reperfusion in the CEL group compared to the UW group (P < 0.05). Likewise, TAP taken from the portal venous effluent of the graft was found to be higher in the CEL group than in UW (P < 0.05). Endothelin-1 serum levels rose significantly during reperfusion without differences between the two groups. ET-1 immunohistochemistry revealed increased local ET-1 during reperfusion in all grafts. However, the ET-1 immunostaining in the CEL group was more pronounced than that in the UW group (P < 0.05).Conclusions. Our results suggest that CEL solution is not as effective in preventing pancreatic ischemia/reperfusion damage as the standard UW solution in experimental pancreas transplantation. Increased ET-1 immunostaining and reduced ptiO2 in the CEL group indicate increased microcirculatory damage in the CEL group.

Nontoxic heat shock protein coinducer BRX-220 protects against acute pancreatitis in rats

Background: Nontoxic heat shock protein (HSP) inducer compounds open up promising therapeutic possibilities by activating one of the natural and highly conserved defense mechanisms of the organism. Aims: In the present experiments, we examined the effects of a HSP coinducer drug-candidate, BRX-220, on the cholecystokinin-octapeptide (CCK)-induced acute pancreatitis in rats. Methods: Male Wistar rats weighing 240 to 270 g were divided into two groups. In group B, 20 mg/kg BRX-220 was administered orally, followed by 75 μg/kg CCK subcutaneously three times, after 1, 3, and 5 h. This whole procedure was repeated for 5 d. The aminals in group ∅B received physiological saline orally instead of BRX-220, but otherwise the protocol was the same as in group B. The rats were exsanguinated through the abdominal aorta 12 h after the last administration of CCK. We determined the serum amylase activity, the plasma trypsinogen activation peptide concentration, the pancreatic weight/body weight ratio, the DNA and total protein contents of the pancreas, the levels of pancreatic HSP60 and HSP72, the activities of pancreatic amylase, lipase, trypsinogen, and free radical scavenger enzymes (superoxide dismutase, catalase, and glutathione peroxidase), the degree of lipid peroxidation, protein oxidation, and the reduced glutathione level. Histopathological investigation of the pancreas was also performed in all cases. Results: Repeated CCK treatment resulted in the typical laboratory and morphological changes of experimentally induced pancreatitis. The pancreatic levels of HSP60 and HSP72 were significantly increased in the animals treated with BRX-220. In group B, the pancreatic total protein content and the amylase and trypsinogen activities were significantly higher vs. group ∅B. The plasma trypsinogen activation peptide concentration, and the pancreatic lipid peroxidation, protein oxidation, and the activity of Cu/Zn-superoxide dismutase were significantly decreased in group B vs. group ∅B, whereas the glutathione peroxidase activity was increased. The morphological damage in group B was significantly lower than that in group ∅B. Conclusion: The HSP coinducer BRX-220, administered for 5 d, has a protective effect against CCK-induced acute pancreatitis.

Trypsin activity is not involved in premature, intrapancreatic trypsinogen activation.

A premature and intracellular activation of digestive zymogens is thought to be responsible for the onset of pancreatitis. Because trypsin has a critical role in initiating the activation cascade of digestive enzymes in the gut, it has been assumed that trypsin also initiates intracellular zymogen activation in the pancreas. We have tested this hypothesis in isolated acini and lobules from rat pancreas. Intracellular trypsinogen activation was induced by supramaximal secretagogue stimulation and measured using either specific trypsin substrates or immunoreactivity of the trypsinogen activation peptide (TAP). To prevent a trypsin-induced trypsinogen activation, we used the cell-permeant, highly specific, and reversible inhibitor Nalpha-(2-naphthylsulfonyl)-3-amidinophenylalanine-carboxymethylpiperazide (S124), and to prevent cathepsin-induced trypsinogen activation, we used the cysteine protease inhibitor E-64d. Incubation of acini or lobules in the presence of S124 completely prevented the generation of trypsin activity in response to supramaximal caerulein but had no effect whatsoever on the generation of TAP. Conversely, when trypsin activity was recovered at the end of the experiment by either washout of S124 from acini or extensive dilution of lobule homogenates, it was up to 400% higher than after caerulein alone and corresponded, in molar terms, to the generation of TAP. Both trypsin activity and TAP release were inhibited in parallel by E-64d. We conclude that caerulein-induced trypsinogen activation in the pancreas is caused by an E-64d-inhibitable mechanism such as cathepsin-induced trypsinogen activation, and neither involves nor requires intracellular trypsin activity. Specific trypsin inhibition, on the other hand, prevents 80% of trypsin inactivation or autodegradation in the pancreas.

Prognostic factors in acute pancreatitis.

The assessment of the severity of acute pancreatitis (AP) is a critical early step in its management, as severity of AP predicts prognosis. A range of options are available for assessment of severity in AP, including clinical evaluation, standardized prognostic criteria, computed tomography (CT), and biochemical markers. Clinical assessment has limited accuracy for predicting severity early in the course of AP. Therefore, additional assessment using biochemical and radiologic criteria in combination with standardized criteria is appropriate to determine severity and prognosis in AP; a strategy emphasizing daily assessment of severity should be used. The APACHE II is the scoring system of choice for evaluating severity in AP, although it remains an imperfect tool. Computed tomographic grading of AP and the development of the CT severity index allow for heightened accuracy in the prediction of severity. C-Reactive protein is the standard for serum marker assessment of severity and prognosis in AP; other markers, including interleukin-6, polymorphonuclear elastase, and trypsinogen activation peptide, hold promise. The focus of this review is to examine the role of diagnostic tests in evaluating severity and predicting prognosis among patients with AP and to provide a diagnostic algorithm for initial management.

Urinary trypsinogen activation peptide is more accurate than hematocrit in determining severity in patients with acute pancreatitis. A prospective study

Urinary trypsinogen activation peptide is more accurate than hematocrit in determining severity in patients with acute pancreatitis. A prospective study

Regional effects of nafamostat, a novel potent protease and complement inhibitor, on severe necrotizing pancreatitis

Background. We evaluated the effect of the novel protease inhibitor nafamostat on rat necrotizing pancreatitis through different routes of administration. Methods. Three hours after the induction of severe pancreatitis, the rats received intravenous gabexate or intravenous or local mesenteric intra-arterial nafamostat. At 9 hours, ascites and bronchoalveolar lavage fluid were collected for the evaluation of capillary leakage (Evans blue extravasation). Pancreas and lung were excised for histologic features, myeloperoxidase, and trypsinogen activation peptide. Twenty-four hour survival was evaluated. Results. Only the intravenous infusion of nafamostat significantly reduced myeloperoxidase (11.7 ± 2.3 vs 18.3 ± 1.8 mU/mg; P <.05) and capillary leakage in lungs (Evans blue dye, 1.6 ± 0.3 vs 2.6 ± 0.3; P <.05). Only intra-arterial infusion of nafamostat significantly diminished capillary peritoneal leakage (Evans blue dye, 3.6 ± 0.9 vs 9.4 ± 0.4; P <.01). Typsinogen activation peptide levels were significantly reduced in all groups, but only intra-arterial infusion did so to baseline. Histologic inflammation in the pancreas was most significantly reduced after intra-arterial infusion (0.92 ± 0.08 vs 2.91 ± 0.06; P <.05). No form of protease inhibition reduced mortality rates. Conclusions. The effects of protease inhibition depend on the route of administration. Nafamostat has maximal effects on the pancreas and peritoneal capillary leakage when delivered by way of local intra-arterial infusion, and shows a greater reduction of lung leukocyte infiltration and capillary leakage by the intravenous route. Nafamostat is more effective than gabexate. (Surgery 2001;130:175-81.)

Linkage of oxidative and nonoxidative ethanol metabolismin the pancreas and toxicity of nonoxidative ethanol metabolites for pancreatic acinar cells

Background. Alcohol abuse is a major cause of pancreatic damage. Recent experimental evidence suggests that fatty acid ethyl esters (FAEE), nonoxidative ethanol metabolites, injure pancreatic acinar cells. Linkage between oxidative and nonoxidative metabolism of ethanol in the pancreas may contribute to increased FAEE levels. Methods. To study the association between oxidative and nonoxidative ethanol metabolism, FAEE concentration and FAEE synthase activity in rat pancreatic and liver homogenates incubated with ethanol were evaluated with and without inhibitors of oxidative ethanol metabolism. For toxicity studies, trypsinogen activation peptide synthesis as a measure of pancreatic cell injury was quantitated in unstimulated and cerulein-stimulated isolated pancreatic acinar cells incubated with ethanol or FAEE. Results. Inhibition of oxidative ethanol metabolism results in a 2- to 3-fold increase in nonoxidative ethanol metabolism to FAEE in pancreas and in liver. Both ethanol and FAEE induce increased intracellular trypsinogen activation by more than 50% in the presence of physiologic concentrations of cerulein in vitro. Conclusions. These findings demonstrate that the inhibition of oxidative ethanol metabolism results in an increase in flux through the nonoxidative pathway and support the proposition that alcohol-induced pancreatic injury is mediated at least in part by FAEE, which are important products of pancreatic ethanol metabolism.(Surgery 2001;129:736-44.)

Endothelin mediates local and systemic disease sequelae in severe experimental pancreatitis.

Endothelin-1 has been shown to reduce pancreatic blood flow and cause focal acinar cell necrosis similar to those seen in acute pancreatitis (AP), whereas therapy with endothelin receptor antagonists enhanced pancreatic capillary blood flow (PCBF) and decreased mortality rates. The current study evaluated the role of endothelin in the development of severe AP. Trypsinogen activation peptides, acinar cell necrosis, and PCBF were used as local indicators of disease severity, fluid sequestration, cardiorespiratory and renal parameters, and colonic capillary blood flow as systemic disease indicators. The following groups of animals were examined: 1) rats with mild edematous AP and 2) severe necrotizing AP treated with and without endothelin, 3) transgenic rats overexpressing endothelin with severe AP, and 4) rats with severe AP prophylactically treated with endothelin receptor antagonists. The following observations were made: endothelin superimposed on mild AP caused hemoconcentration, a decrease in PCBF, and necrosis and ascites not seen in this model without endothelin exposure. Endothelin superimposed on severe AP had no significant effects. After induction of severe AP, less PCBF and more acinar cell necrosis were observed in transgenic rats than in their normal littermates. Prophylactic endothelin receptor antagonists improved local (acinar necrosis, PCBF) and systemic parameters (ascites, urine production, colonic capillary blood flow) of disease severity in animals with severe AP. These observations underscore the role of endothelin as a mediator of disease severity in AP and suggest that endothelin receptor blockade may become a promising therapeutic tool in this disease.