390 Gut 2000;46:390–394 EVect of and

blockade on capillary permeability in Gut: first published as 10.1136/gut.46.3.390 on 1 March 2000. Downloaded from experimental pancreatitis

G Eibl, H G Hotz, J Faulhaber, M Kirchengast, H J Buhr, Th Foitzik

Abstract involve premature intrapancreatic protease Background—Capillary leakage with fluid activation and acinar cell injury causing a local loss into the third space contributes to and systemic inflammatory response mediated many of the early systemic complications by (pro)inflammatory and secondary in severe acute pancreatitis. There has mediators. Recent research has suggested that been increasing interest in endothelin as the degree to which these mediators escape one of the factors aVecting capillary into the circulation correlates significantly with permeability. disease severity.1–4 Disease severity not only Aim—To elucidate further the role of comprises local tissue injury but also (maybe endothelin in the development of capillary even more importantly) the systemic sequelae leakage in acute pancreatitis by investigat- associated with systemic inflammatory re- ing the eVect of exogenous endothelin sponse syndrome. Increased capillary perme- administration and endothelin receptor ability with subsequent fluid loss from the blockade in sham operated animals and intravascular compartment into the third space two models of acute pancreatitis. is believed to contribute substantially to these 5 Methods—Determination of capillary systemic sequelae. Therefore it is important to permeability in the pancreas and colonic identify the dominant vasoactive mediators mucosa by quantifying extravasation of that increase capillary permeability. Vasoactive fluorescein labelled dextran using a novel substances assumed to be involved include platelet activating factor, nitric oxide, brady- computer assisted video image analysis 6–9 system. , prostaglandins, and endothelin. In the Results—Pancreatic and colonic capillary light of previous findings showing that en- permeability increased stepwise from dothelin receptor blockade ameliorates disease severity,5 this study aimed to evaluate further mild to severe acute pancreatitis. En- http://gut.bmj.com/ dothelin increased pancreatic and colonic how capillary permeability is aVected by capillary permeability in healthy animals endothelin-1, the most abundant and potent member of the endothelin family. Our under- and animals with mild acute pancreatitis lying questions were as follows. ( ) Does but had no additional adverse eVect in a exogenous endothelin-1 increase capillary per- severe acute pancreatitis. Endothelin re- meability in healthy rats? ( ) Is the pancreas ceptor blockade decreased pancreatic b especially susceptible to endothelin-1, or are capillary permeability in sham operated other organs similarly aVected? ( ) Does exog- on September 30, 2021 by guest. Protected copyright. rats but had no eVect on the colon. In mild c enous endothelin-1 likewise increase capillary and severe acute pancreatitis, endothelin permeability in mild (associated with a normal receptor blockade stabilised increased endogenous endothelin-1 level and normal or capillary permeability in both the pan- slightly increased capillary blood flow and per- Department of creas and colon. Surgery, Benjamin meability) and severe (increased endogenous Conclusions—Endothelin plays an impor- endothelin-1, decreased capillary blood flow, Franklin Medical tant role in mediating capillary perme- Centre, Freie and increased permeability) acute pancreatitis? ability in the pancreas. In severe Universität Berlin, (d) How does endothelin receptor blockade Germany pancreatitis, it increases capillary perme- (endothelin-1 ) influence G Eibl ability even outside the pancreas, thereby capillary permeability in and outside the H G Hotz contributing to capillary leakage. En- pancreas in healthy rats and in those with mild J Faulhaber dothelin receptor blockade significantly H J Buhr and severe acute pancreatitis? Th Foitzik reduces capillary permeability in acute pancreatitis both in and outside the Knoll AG, pancreas, suggesting a therapeutic ap- Methods Ludwigshafen, proach to counteract capillary leakage in ANIMAL PREPARATION AND INDUCTION OF Germany severe acute pancreatitis. M Kirchengast PANCREATITIS (Gut 2000;46:390–394) All experiments were conducted in accordance Correspondence to: Keywords: pancreatitis; endothelin; endothelin with the national guidelines for the use and Dr Th Foitzik, Department care of laboratory animals and were approved of Surgery, Benjamin receptor antagonist; microcirculation; capillary Franklin Medical Centre, permeability; fluid sequestration by the local ethics committee. After overnight Freie Universität Berlin, fasting, male Sprague-Dawley rats (mean (SD) Hindenburgdamm 30, weight 330 (20)g) were anaesthetised with D-12200 Berlin, Germany Acute pancreatitis, a disease with high morbid- intraperitoneal pentobarbital (20 mg/kg) and Accepted for publication ity and mortality, can only be managed by sup- ketamine (40 mg/kg). Polyethylene catheters 5 October 1999 portive treatment. Early pathogenic events (internal diameter 0.5 mm) were inserted into Endothelin and capillary permeability in acute pancreatitis 391

both jugular veins and the left carotid artery, the pancreas or the colonic mucosa (after subcutaneously tunnelled to the neck, and antimesenteric enterotomy13) were recorded. advanced through a steel tether, which allowed Subsequently 0.2 ml 5% fluorescein isothio- blood sampling and intravenous access in the cyanate labelled dextran (molecular mass Gut: first published as 10.1136/gut.46.3.390 on 1 March 2000. Downloaded from unrestrained animals. 150 000 Da; Sigma, Deisenhofen, Germany) Mild oedematous acute pancreatitis was were injected, and, after 30 minutes, the same induced by a continuous intravenous infusion three regions were recorded for oV line of 5 µg/kg/h cerulein (Farmitalia, Freiburg, determination of capillary permeability using a Germany) over six hours, and severe necrotis- commercially available video image analysis ing acute pancreatitis by cerulein hyperstimu- system (Cap-Image, V.6.01; Zeintl, Heidel- lation superimposed on a standardised retro- berg, Germany). This computer assisted sys- grade infusion of 0.5 ml of 10 mmol/l tem quantifies the increase in perivascular glycodeoxycholic acid (Sigma, St Louis, Mis- fluorescein intensity resulting from the extrava- souri, USA) into the biliopancreatic duct for 10 sation of the fluorescence labelled dextran over minutes. As previously described, this tech- time (here 30 minutes). nique requires mini-laparotomy for canulation The total time needed for the exposure and of the biliopancreatic duct by puncturing the recording of the microcirculatory beds was duodenum.10 For comparison, laparotomy and 60–90 minutes per animal. Heart rate, systolic canulation of the biliopancreatic duct was like- and mean arterial pressure, and blood gases wise performed in animals with mild pancrea- were measured before and after intravital micro- titis (intravenous cerulein, intraductal saline) scopy. We only included data from animals in and healthy control animals (intravenous and stable cardiorespiratory condition for analysis of intraductal saline infusion). As an improve- the microcirculatory variables to avoid bias pos- ment on the original technique,10 a special sibly resulting from systemic cardiorespiratory infusion pump (IVAC 770; Lilly Medisintech- derangement. Exclusion criteria were: mean

nik, Giessen, Germany) was used for the pres- arterial pressure <80 mm Hg; PO2 <80 mm Hg;

sure controlled (30 mm Hg), volume control- PCO2 >50 mm Hg; pH <7.3 or >7.5. led (0.5 cm3), and time controlled (10 minutes) intraductal glycodeoxycholic acid infusion. STATISTICAL ANALYSIS All results are expressed as mean (SEM). STUDY PROTOCOL AND TREATMENT Variables were tested for group diVerences with Six hours after sham operation (n = 30 rats; Student’s t test and analysis of variance as group 1) or induction of mild (n = 30; group 2) appropriate. p<0.05 was considered to be or severe (n = 45; group 3) acute pancreatitis, significant. animals were randomised into three groups for intravenous injection of normal saline 0.9% (A), 1.25 µg/kg/h endothelin-1 (B; provided by Results http://gut.bmj.com/ Knoll AG, Ludwigshafen, Germany), or 50 SHAM OPERATED ANIMALS mg/kg of the endothelin-1 receptor antagonist Endothelin-1 administration (group 1B) had no LU-135252 (C; provided by Knoll AG). measurable eVect on the systemic cardiorespi- Systemic cardiorespiratory variables were ratory variables except to produce an immedi- monitored over the next six hours. The animals ate transient decrease in systolic arterial blood were subsequently relaparotomised for intra- pressure (10–30 mm Hg) which normalised

vital microscopic determination of capillary within two to three minutes, followed by an on September 30, 2021 by guest. Protected copyright. permeability. increase in arterial blood pressure (30–40 mm Hg), which returned to baseline within 10–15 DETERMINATION OF CAPILLARY PERMEABILITY minutes. Blood pressure at later time points was Capillary permeability was determined by not diVerent from sham operated animals given intravital microscopy using the same technique saline (group 1A). An increase in packed cell and equipment as in previous studies.11 12 volume developed in all endothelin-1 treated Briefly, the abdomen was opened by a small animals (baseline 48 (1) v 54 (1) six hours after midline incision. Either the duodenum to- the start of endothelin-1 administration). Cap- gether with the head of the pancreas or the illary permeability increased by 104% in the ascending colon were mobilised and placed on pancreas (fig 1A) and 82% in the colon (fig 1B). a special stage for intravital microscopy. After a Endothelin-1 receptor antagonist (group 1C) 10 minute stabilisation period, three randomly had no significant eVect on packed cell volume chosen regions (400 × 325 µm) in the head of and cardiorespiratory variables; capillary

300 300 A * Sal B 250 ET-1 250 * * * ET-RA 200 200 * * 150 * 150 *

100 100 * 50 50 Sham EP NP Sham EP NP Figure 1 EVect of endothelin-1 (ET-1) and endothelin receptor antagonist (ET-RA) on capillary permeability (measured as the increase in perivascular density (%) 30 minutes after injection of fluorescein isothiocyanate labelled dextran 150) in the pancreas (A) and colon (B) of sham operated animals and animals with mild (EP) or severe (NP) acute pancreatitis. *p<0.05 compared with saline injected animals (Sal). 392 Eibl, Hotz, Faulhaber, et al

permeability decreased in the pancreas (fig 1A) untreated sham operated animals and untreated but was not significantly aVected in the colon animals with mild pancreatitis, pancreatic and (fig 1B). colonic capillary permeability in rats with severe pancreatitis given saline were significantly in- Gut: first published as 10.1136/gut.46.3.390 on 1 March 2000. Downloaded from MILD PANCREATITIS creased (248 (17)% in the colon and 284 (16)% Compared with saline treated sham operated in the pancreas) (fig 2). Endothelin administra- controls (group 1A), animals with oedematous tion did not further increase capillary perme- acute pancreatitis given saline (group 2A) ability in the pancreas and colon (fig 1A,B). showed an increase in packed cell colume (48 However, endothelin receptor antagonist de- (1) v 51 (1)) and capillary permeability in both creased capillary permeability in the pancreas by the pancreas and colon (fig 2). This increase 63% (fig 1A) and in the colon by 90% (fig 1B). was more pronounced in the pancreas than in the colon (114% increase in the pancreas com- Discussion pared with 82% increase in the colon). Increased capillary permeability leading to fluid Cardiorespiratory variables remained un- loss from the intravascular space and fluid changed. Endothelin-1 (group 2B) further sequestration into the third space is a hallmark increased packed cell volume (51 (1) v 55 (1)) of severe acute pancreatitis. Clinically, capillary and significantly increased capillary permeabil- leakage is reflected by intravascular fluid loss ity by 40% in the pancreas and 69% in the leading to hypovolaemia (low central venous colon (fig 1). Endothelin-1 receptor antagonist pressure), haemoconcentration (high packed (group 2C) significantly reduced packed cell cell volume), and extravascular fluid volume (51 (1) v 47 (1)) and capillary perme- sequestration—for example, in the retroperito- ability by 69% in the pancreas (fig 1A) and neum, lungs, and pleural and abdominal 33% in the colon (fig 1B). Cardiorespiratory cavities. Factors believed to increase capillary variables did not diVer significantly between permeability in severe pancreatitis include the treatment groups. platelet activating factor, nitric oxide, brady- kinin, prostanoids, and endothelin.6–9 14 In this SEVERE PANCREATITIS study, we report our findings with endothelin-1 All animals in which severe acute pancreatitis and blockade was induced developed significant haemocon- (endothelin-1 receptor antagonist), which we centration (packed cell volume 56 (1)) and chose on the basis of previous investigations showed an increase in heart rate and decrease in showing that endothelin-1 aggravates acute mean arterial pressure (compared with sham pancreatitis whereas endothelin-1 receptor an- operated animals and animals with mild acute tagonist improves survival considerably.5 Meas- pancreatitis at six hours). However, no animal urements of packed cell volume and extravascu- had to be excluded at this point according to the lar fluid sequestration implied that these eVects http://gut.bmj.com/ defined criteria. After treatment, four animals were associated with the stablisation of in- treated with saline developed cardiorespiratory creased capillary permeability. Therefore this insuYciency and died before intravital micros- study aimed to evaluate further the eVect of copy. Five animals given endothelin-1 and two endothelin-1 and a specific endothelin-1 an- animals treated with endothelin-1 receptor tagonist (LU-135252) on capillary permeability antagonist died before intravital microscopic in two diVerent organ beds (pancreas and determination of capillary permeability. All colon) of two acute pancreatitis models using

other animals were in stable cardiorespiratory direct intravital fluorescence microscopy and a on September 30, 2021 by guest. Protected copyright. condition at the time of intravital microscopy. novel computer assisted video image analysis There were no significant diVerences in cardio- system for quantitative assessment. respiratory variables between treatment groups. Packed cell volume, however, was significantly METHODOLOGICAL CONSIDERATIONS reduced in animals treated with the endothelin The two models of acute pancreatitis (mild/ receptor antagonist (51 (1) v 56 (1)). Adminis- oedematous and severe/necrotising) have been tration of endothelin did not further increase the extensively characterised histologically as well as packed cell volume. Two animals (one treated by biochemical variables, systemic disease se- with saline, one treated with endothelin) that quelae (including changes in fluid homoeosta- survived intravital microscopy could not be sis), and microcirculatory variables.10 15 Intravital included in the final analysis of capillary perme- microscopy and quantification of capillary per- ability, because the final assessment of arterial meability by Cap-Image are also well established blood gases indicated severe respiratory insuY- in our laboratory. As continuous extravasation of ciency (see exclusion criteria). Compared with fluorescein isothiocyanate labelled dextran Figure 2 Capillary causes brightness of the microscopic picture, permeability (measured as which compromises further measurements, cap- the increase in perivascular illary permeability could only be determined in density (%) 30 minutes one organ at a time. We therefore used five ani- after injection of fluorescein 300 † isothiocyanate labelled † mals per group to determine capillary perme- dextran 150) in the 250 ability in the pancreas and also in the colonic pancreas and colon of * Sham 200 mucosa. The latter was chosen as a major site of untreated sham operated * EP animals (no acute 150 NP fluid extravasation in capillary leakage syndrome pancreatitis) and animals because of its extensive surface and capillary with mild (EP) or severe 100 density. (NP) acute pancreatitis. 50 The endothelin-1 dose was chosen accord- *p<0.05 v sham operated; Pancreas Colon †p<0.05 v EP. ing to the manufacturer’s recommendations, Endothelin and capillary permeability in acute pancreatitis 393

based on large scale experiments with this there is no evidence from the literature or our agent in small rodents16 as well as on previous own histological studies that the capillaries are tests in our laboratory showing that doses lower injured, functional rather than morphological than 0.5 µg/kg have no eVect on blood pressure changes in the endothelium probably cause Gut: first published as 10.1136/gut.46.3.390 on 1 March 2000. Downloaded from or microcirculatory variables, whereas doses plasma extravasation. Although capillary perme- higher than 1.5 µg/kg cause pronounced ability was significantly higher in severe acute changes in both macrohaemodynamic vari- pancreatitis, pancreatic oedema was similar or ables and microcirculation. The dose of 1.25 even more pronounced in mild acute pancreati- µg/kg was finally chosen because it was the tis (as scored histologically5). This can be highest dose that had no appreciable eVects on explained by the observation that the increase in systemic cardiorespiratory variables except to capillary permeability is mainly confined to the cause an immediate transient fall in systolic pancreas in mild acute pancreatitis, whereas arterial blood pressure followed by a transient fluid also extravasates into the colon (and prob- increase in arterial blood pressure (see above). ably other capillary beds) in severe acute This intermittent depressor eVect, also ob- pancreatitis. Another reason for increased fluid served by other investigators,17–20 is thought to loss from the pancreas in oedematous acute result from the initial activation of non- pancreatitis may be capillary blood flow, which is selective endothelin B2 receptors located on increased in mild acute pancreatitis but de- the intraluminal side of endothelial cells medi- creased in severe acute pancreatitis.15 The fact ating vasodilatation. This eVect, however, that the increase in capillary permeability was seems to be immediately counteracted by more pronounced in the pancreas than in the either rapid degradation of intraluminal colon in mild acute pancreatitis agrees with the endothelin-1 or a compensatory pressor re- observation that the disease process is largely sponse when endothelin-1 reaches the high confined to the pancreas: neither animals nor aYnity endothelin A receptors on the vascular patients with mild acute pancreatitis develop smooth muscle cells. significant systemic disease sequelae or compli- Plasma endothelin-1 levels were not cations. In contrast, extremely increased colonic measured in this experiment because it is gen- capillary permeability in severe acute pancreati- erally accepted that it elicits its major eVects in tis seems to reflect the systemic reactions a paracrine manner, while endoluminal discussed above. endothelin-1 is rapidly degraded.17 21 In a The endothelin-1 induced increase in capil- previous study, we found significantly elevated lary permeability observed in otherwise healthy plasma endothelin-1 levels in the severe model sham operated rats suggests that endothelin-1 but not in mild oedematous acute pancreatitis.5 also plays a role in mediating capillary perme- Depending on the dose, LU-135252 first ability in other conditions such as trauma. This blocks endothelin A receptors mainly found on also agrees with other studies emphasising the http://gut.bmj.com/ vascular smooth muscle cells, which selectively eVect of endothelin on vascular permeability in bind endothelin-1 over endothelin-2 and various organs and tissues in conjunction with endothelin-3 to mediate contraction. In higher diseases and possibly under physiological con- doses, it also blocks the endothelin B receptors ditions as well.24 26 27 Failure of endothelin to mediating dilatation. Details on the properties increase capillary permeability further in severe of LU-135252 have been reported acute pancreatitis indicates that all endothelin elsewhere.16 22 23 We chose a dose of 50 mg/kg receptors were already saturated by endog-

for the present experiments, because doses as enous endothelin-1 released in the course of on September 30, 2021 by guest. Protected copyright. high as 100 mg/kg are well tolerated in rats and severe acute pancreatitis. This is in accordance do not cause any appreciable changes in with our previous findings that endothelin-1 in systemic haemodynamic variables (as shown in plasma is significantly increased in severe acute previous experiments5). This dose has been pancreatitis (but normal in mild acute shown to block endothelin-A receptors selec- pancreatitis).5 The fact that endothelin-1 has a tively (suggesting that the eVect of dose dependent eVect and is only harmful at endothelin-1 on capillary permeability is pri- higher concentrations28 would also explain the marily mediated by these receptors). This also contradictory results observed with various agrees with the findings of other groups show- doses of (exogenous) endothelin-1 in diVerent ing the inhibition of endothelin-1 induced models of acute pancreatitis (with or without plasma volume loss and reduced extravasation increased endogenous endothelin-1 levels or of Evans blue dye after treatment with other pre-existing microcirculatory disorders).29 endothelin A receptor blockers24 and with It is still unclear how endothelin-1 aVects recent studies of our group showing that vascular permeability and whether this eVect is endothelin A but not associated with its well known action as a vaso- blockade mediates capillary leakage in severe constrictor of precapillary and postcapillary acute pancreatitis.25 This also explains the vessels and venules.30 An endothelin-1 associ- diVerent results obtained with (non-selective) ated increase in postcapillary resistance may endothelin antagonists. elevate the hydrostatic pressure in the capillary bed, thereby enhancing fluid transfer from the INTERPRETATION OF RESULTS intra- to the extra-vascular space. Alternatively, Incremental extravasation of fluorescein isothio- it is conceivable that endothelin-1 aVects capil- cyanate labelled dextran in the pancreas from lary permeability by regulating interendothelial mild to severe acute pancreatitis indicates that gaps through the contraction of vascular the increase in capillary permeability correlates elements such as myosin and actin contained in with the magnitude of local tissue injury. As endothelial cells and surrounding pericytes.31 32 394 Eibl, Hotz, Faulhaber, et al

The endothelin-1 receptor antagonist in- tion and the acute-phase response in patients with acute pancreatitis. Pancreas 1995;10:347–53. duced decrease in capillary permeability ob- 5 Foitzik T, Faulhaber J, Hotz HG, et al. Endothelin receptor served in the pancreas but not in the colon of blockade improves fluid sequestration, pancreatic capillary

blood flow and survival in severe experimental pancreatitis. Gut: first published as 10.1136/gut.46.3.390 on 1 March 2000. Downloaded from otherwise healthy sham operated rats suggests Ann Surg 1998;228:670−5. that endogenous endothelin-1 was released 6 Yamanaka K, Saluja AK, Brown GE, et al. Protective eVects during organ exposure for intravital micros- of prostaglandin E1 on acute lung injury of caerulein- induced acute pancreatitis in rats. Am J Physiol 1997;272: copy and that the pancreas was more suscepti- G23–30. ble to this manipulation, endothelin itself or 7 Weidenbach H, Lerch MM, Gress TM, et al. Vasoactive mediators and the progression from edematous to necrotiz- both. Several investigators have previously ing experimental acute pancreatitis. Gut 1995;37:434–40. shown that the pancreas is especially suscepti- 8 Dobosz M, Hac S, Wajda Z. Does nitric oxide protect from microcirculatory disturbances in experimental acute pan- ble to endothelin-1. For example, exogenous creatitis in rats? Int J Microcirc Clin Exp 1996;16:221–6. endothelin-1 in doses not aVecting systemic 9 Yotsumoto F, Manabe T, Ohshio G, et al. Role of pancreatic blood flow and vasoactive substances in the development of haemodynamic variables significantly reduced canine acute pancreatitis. J Surg Res 1993;55:531–6. pancreatic perfusion whereas other organs— 10 Schmidt J, Rattner DW, Lewandrowski K, et al. A better model of acute pancreatitis for evaluating therapy. Ann Surg for example, heart, lungs, liver—were not 1992;215:44–56. 33 aVected or even showed increased blood flow. 11 Mithöfer K, Schmidt J, Gebhard MM, et al. Measurement This was explained by di erent expression of of blood flow in pancreatic exchange capillaries with V FITC-labelled erythrocytes. Microvasc Res 1995;49:33–48. endothelin receptors. 12 Hotz HG, Schmidt J, Ryschich EW, et al. Isovolemic Capillary permeability in acute pancreatitis hemodilution with dextran prevents contrast medium induced impairment of pancreatic microcirculation in after endothelin-1 receptor antagonist treatment necrotizing pancreatitis in the rat. Am J Surg 1995;169: did not reach the values measured in healthy 161–6. 13 Hotz HG, Foitzik T, Rohweder J, et al. Intestinal animals, which indicates that endothelin-1 is not microcirculation and gut permeability in acute pancreatitis. the only mediator of capillary permeability in Early changes and therapeutic implications. J Gastrointest Surg 1998;2:518−25. acute pancreatitis. This underlines the need to 14 Bloechle C, Kusterer K, Kuehn RM, et al. Inhibition of study further the eVects of other vasoactive sub- B2 receptor preserves microcirculation in experimental pancreatitis in rats. Am J Physiol 1998;274: stances and mediators on capillary permeability G42–51. in future experiments. 15 Klar E, Schratt W, Foitzik T, et al. 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Regional vasodilatation is a in capillary permeability is more pronounced in prominent feature of the hemodynamic response to the pancreas than in the colon (suggesting that endothelin in anesthetized spontaneously hypertensive rats. Eur J Pharmacol 1988;155:201–3. http://gut.bmj.com/ the pancreas is especially susceptible to endothe- 20 Winquist RJ, Bunting PB, Garsky VM, et al. Prominent lin). (d) Exogenous endothelin-1 does not depressor response to endothelin in spontaneously hyper- tensive rats. Eur J Pharamcol 1989;163:199–203. further increase capillary permeability in severe 21 Wagner OF, Christ G, Wojta J, et al. Polar secretion of acute pancreatitis (suggesting that all endothelin-1 by cultured endothelial cells. J Biol Chem 1992;267:16066–8. endothelin-1 receptors are already occupied by 22 Raschak M, Unger L, Riechers H, et al. Receptor selectivity endogenous endothelin-1). 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tant role for endothelin-1 in regulating pancre- tion of ETA receptors. Br J Pharmacol 1993;109:880–6. atic microcirculation). 25 Foitzik T, Eibl G, Forgacs B, et al. Therapy of microcircula- tory disorders in severe acute pancreatitis. Endothelin-A In conclusion, these results underline the but not endothelin-B receptor blockade reduces capillary role of the endothelin system in mediating cap- leakage. Pancreas 1998;17:433. 26 Filep JG, Sirois MG, Rousseau A, et al.EVects of illary permeability in acute pancreatitis and endothelin-1 on vascular permeability in the conscious rat: suggest that endothelin-1 blockade by specific interactions with platelet-activating factor. Br J Pharmacol 1991;104:797–804. endothelin receptor antagonists may become a 27 Dahlöf B, Gustafsson D, Hedner T, et al. Regional hemody- powerful tool in the treatment of acute namic eVects of endothelin-1 in rat and man: unexpected adverse reactions. J Hypertens 1990;8:811–17. pancreatitis because they stabilise increased 28 Brain SD, Crossman DC, Buckley TL, et al. Endothelin-1: capillary permeability which contributes to demonstration of potent eVects of the microcirculation of humans and other species. J Cardiovasc Pharmacol many of the systemic sequelae in this disease. 1989;13(suppl 5):S147–9. 29 Kogire M, Inoue K, Higashide SI, et al. Protective eVects of endothelin-1 on acute pancreatitis in rats. Dig Dis Sci 1995; This work was presented in part at the European Pancreas Club, 40:1207–12. Thessaloniki, Greece, June 10–13, 1998. 30 Yang ZH, Bühler FR, Diederich D, et al.DiVerent eVects of endothelin-1 on cAMP- and cGMP-mediated vascular 1 Kingsnorth A. Role of cytokines and their inhibitors in acute relaxation in human arteries and veins: comparison with pancreatitis. Gut 1997;40:1–4. norepinephrine. J Cardiovasc Pharmacol 1989;13(suppl 2 Gross V, Leser HG, Heinisch A, et al. Inflammator 5):S129–31. mediators and cytokines. New aspects of the pathophysiol- 31 Becker CG, Nachman RL. Contractile of endothe- ogy and assessment of severity of acute pancreatitis? lial cells, platelets and smooth muscles. Am J Pathol 1973; Hepato-gastroenterol 1993;40:522–30. 71:1–22. 3 De Beaux AC, Goldie AS, Ross JA, et al. Serum concentra- 32 LeBeux YJ, Willemot J. Actin- and myosin-like filaments in tions of inflammatory mediators related to organ failure in rat brain pericytes. Anat Rec 1978;190:811–26. patients with acute pancreatitis. Br J Surg 1996;83:349–53. 33 Takaori K, Inoue K, Kogire M, et al.EVects of endothelin 4 Heath DI, Cruickshank A, Gudgeon AM, et al. The on microcirculation of the pancreas. Life Sci 1992;51:615– relationship between pancreatic enzyme release and activa- 22.