The effect of platelet activating factor antagonist (BN 52021) on cerulein-induced acute pancreatitis with reference to oxygen radicals

Acute edematous pancreatitis was induced in Wistar male rats by iv infusion of cerulein (CR) in the dose of 5.10^-6g.kg^-1.h^-1 during 3 or 6 h. The effect of BN 52021—platelet activating factor (PAF) receptor antagonist, against this model of disease was examined. BN 52021 was applied iv as a bolus injection in the dose of 5.10^-3g.kg^-1 at 0 time. Treatment with this agent significantly ameliorates cerulein-induced acute pancreatitis in rats. The effect of BN 52021 was expressed by significant reduction of pancreas edema, diminution of hyperamylasemia, lack of superoxide dismutase activity depletion, and inhibition of lipid peroxidation in pancreatic tissue. These changes were accompanied by significant reduction of acinar cells vacuolization and remarkable inhibition of infiltration with inflammatory cells in the interacinar space. We suppose that beneficial effect of BN 52021 against cerulein-induced acute pancreatitis in rats depends on the prevention of inflammatory cells activation and subsequent generation of oxygen radicals within pancreatic tissue.     

The effect of platelet activating factor antagonist (BN 52021) on acute experimental pancreatitis with reference to multiorgan oxidative stress

Summary Acute hemorrhagic pancreatitis was induced in Wistar rats using a retrograde intraductal injection of 5% Na-taurocholate. Rats were treated with platelet-activating factor receptor (PAF) antagonist—BN 52021 (5 mg/kg) and sacrificed at 1 and 3 h after induction of acute pancreatitis. Malondialdehyde and sulfhydryl groups concentration were measured in pancreatic, lung, and liver tissue as a parameters of oxidant-antioxidant balance. We have shown that BN 52021 exerts only partial protecting effect against Na-TC-induced AP in rats. The positive effects of BN 52021 were expressed by: (1) Significant reduction of hyperamylasemia accompanied by lower malondialdehyde accumulation in pancreatic tissue; (2) Prevention of sulfl hydryl groups depletion in lung tissue; (3) Diminution of necrotic and inflammatory changes in pancreatic tissue; and (4) Improvement of survival rate. We suggest that these effects may depend on the inhibition of PAF-mediated activation and oxidant generation by phagocytes.     

Oxygen-derived free radicals in cerulein-induced acute pancreatitis

Conscious rats were treated with a supramaximal dose of 5.10(-6)g.kg-1.h-1 of cerulein for periods of 3 and 12 h. In both groups of animals typical features of acute oedematous pancreatitis were proved by biochemical and histologic examinations. The most important finding of our study was the decrease of superoxide dismutase (SOD) activity in pancreatic tissue, accompanied by a slight increase of this scavenger enzyme in serum of rats stimulated with cerulein during 3 h. Parallelly, evident elevation of malondialdehyde (MDA) concentration in pancreatic tissue was noted. After the 12-h infusion of cerulein we were not able to detect any SOD activity in pancreatic tissue, whereas this activity appeared in ascitic fluid of tested animals. Further increase of MDA concentration in pancreatic tissue, in comparison with 3-h pancreatitis, was found. These data suggest that in 3-h and 12-h cerulein-induced pancreatitis the oxygen-derived free radicals mediate the increased lipid peroxidation in pancreatic tissue. We think that the depletion of the scavenger enzyme SOD may be responsible for such a disturbance of lipid metabolism.     

Protective effect of human pancreatic secretory trypsin inhibitor on cerulein-induced acute pancreatitis in rats.

We examined the protective effect of human pancreatic secretory trypsin inhibitor (PSTI), a specific trypsin inhibitor secreted from pancreatic acinar cells into the pancreatic duct, on cerulein-induced acute pancreatitis in conscious rats. The protective effect of human PSTI-RS, an analogue of PSTI with Arg-44 to Ser substitution which has a longer half-life in vitro, was also examined. Intraperitoneal administration of a pharmacological dose of cerulein to conscious rats induced acute pancreatitis, characterized by light microscopy as cellular disorganization of the acini and interstitial edema. Intravenous infusion of human PSTI (10, 50 or 250 micrograms/rat/h) into rats with cerulein-induced acute pancreatitis decreased their pancreatic wet weight and plasma amylase concentration. It also caused a dose-dependent decrease in vacuoles in acinar cells and interstitial edema. Human PSTI-RS, which has a longer half-life in vivo, was more effective than native PSTI at the same dose rate (10 micrograms/rat/h) in reducing pancreatitis. These results suggest that human PSTI may have a beneficial effect on acute pancreatitis.     

Mechanism and dose-effect of Ginkgolide B on severe acute pancreatitis of rats

AIM:To determine the optimal dosage and mechanism of Ginkgolide B(BN52021) on severe acute pancreatitis(SAP) of rats.METHODS:Seventy male Wistar rats were randomly divided into seven groups(10 for each group).Shamoperation group(SO),SAP model group(SAP),dimethyl sulfoxide(DMSO) contrast group(DMSO),and groups treated with 2.5 mg/kg BN52021(BN1),5 mg/kg BN52021(BN2),10 mg/kg BN52021(BN3),and 20 μg/kg Sandostatin(SS).The SAP model was established in Wistar rats by injecting 5% sodium taurocholate retrogradely...     

Effect of hyperthermia on experimental acute pancreatitis

BACKGROUND: [corrected] Recent studies indicate that hyperthermia can change inflammatory mechanisms and protect experimental animals from deleterious effects of secretagogue-induced acute pancreatitis AIM: To evaluate the effects of hyperthermia post-treatment on cerulein-induced acute pancreatitis in rats METHODS: Twenty animals were divided in two groups: group I (n = 10), rats with cerulein-induced acute pancreatitis undergone hyperthermia, and group II (n = 10), animals with cerulein-induced acute pancreatitis that were kept normothermic. In all groups, amylase serum levels, histologic damage, vascular permeability and pancreatic water content were assessed. Acute pancreatitis was induced by administration of two cerulein injections (20 mcg/kg). A single dose of Evans' blue dye was administered along with the second dose of cerulein. All animals also received a subcutaneous injection of saline solution. After this process, animals undergone hyperthermia were heated in a cage with two 100 W lamps. Body temperature was increased to 39.5 degrees C and maintained at that level for 45 minutes. Normothermia rats were kept at room temperature in a second cage RESULTS: Control animals had typical edema, serum amylase activity and morphologic changes of this acute pancreatitis model. Hyperthermia post-treatment ameliorated the pancreatic edema, whereas the histologic damage and the serum amylase level remained unchanged CONCLUSIONS: The findings suggest a beneficial effect of the thermal stress on inflammatory edema in experimental acute pancreatitis.     

血小板活化因子在急性胰腺炎大鼠并发肾损伤中病理生理作用

目的:本文旨在观察血小板活化因子在急性出血坏死性胰腺炎(AHNP)大鼠肾损伤发生中作用。方法:159只SD大鼠随机分三组:假手术组、AHNP非治疗组和AHNP BN(52021)治疗祖。采用胰管内注入5％牛磺胆酸钠溶液诱导大鼠AHNP,应用~(86)Rb组织摄取法测定肾脏相对血流量及组织灌注量,并测定血小板聚集率(PAgR)。结果:与非治疗组相比较,BN(52021)治疗组肾脏相对血流量、组织灌注量显著提高;PAgR下降;肾脏病理损害减轻。结论:血小板活化因子参与了急性出血坏死性胰腺炎大鼠肾损害的发生。     

Involvement of endogenous cholecystokinin in pancreatic regeneration after cerulein-induced acute pancreatitis.

This study was undertaken to determine the involvement of endogenous cholecystokinin (CCK) in the regeneration of pancreatic tissue after cerulein-induced acute pancreatitis treated by the CCK receptor antagonist L364,718. Acute pancreatitis was induced in rats by s.c. injections of cerulein in gelatin (12 micrograms/kg) three times a day for 2 days with controls receiving saline in gelatin. Rats were then divided into four treatment groups: saline-dimethyl sulfoxide (DMSO) (SD), saline-L364,718 (SA), cerulein-pancreatitis-DMSO (CD), and cerulein-pancreatitis-L364,718 (CA). In the first experiment, rats were treated for 3 or 10 days with DMSO or L364,718 (0.1 mg/kg, twice a day). In the second experiment, rats were treated for 13 days with DMSO or L364,718 (1.0 mg/kg, twice a day). After the rats were killed, pancreata were weighed and evaluated for their total protein, amylase, chymotrypsin, RNA, and DNA. We found that destruction of the pancreatic tissue occurred after cerulein-induced pancreatitis and that regeneration of the tissue was in progress but incomplete after 10 days; the low dose of L364,718 did not prevent regeneration. After 13 days, regeneration was still incomplete but the 1-mg dose of L364,718 strongly inhibited spontaneous regeneration. These data suggest that endogenous CCK is an important and potent trophic factor in the regeneration process of pancreatic tissue following an episode of acute pancreatitis.     

Ischemic preconditioning inhibits development of edematous cerulein-induced pancreatitis： Involvement of cyclooxygenases and heat shock protein 70

AIM: To determine whether ischemic preconditioning (IP) affects the development of edematous cerulein-induced pancreatitis and to assess the role of cyclooxygenase-1 (COX-1), COX-2, and heat shock protein 70 (HSP 70) in this process, METHODS: In male Wistar rats, IP was performed by clamping of celiac artery (twice for 5 min at 5-min intervals). Thirty minutes after IP or sham operation, acute pancreatitis was induced by cerulein. Activity of COX-1 or COX-2 was inhibited by resveratrol or rofecoxib, respectively (10 mg/kg). RESULTS: IP significantly reduced pancreatic damage in cerulein-induced pancreatitis as demonstrated by the improvement of pancreas histology, reduction in serum lipase and poly-C ribonuclease activity, and serum concentration of pro-inflammatory interleukin (IL)-lp. Also, IP attenuated the pancreatitis-evoked fall in pancreatic blood flow and pancreatic DNA synthesis. Serum level of anti-inflammatory IL-10 was not affected by IP. Cerulein-induced pancreatitis and IP increased the content of HSP 70 in the pancreas. Maximal increase in HSP 70 was observed when IP was combined with cerulein-induced pancreatitis. Inhibition of COXs, especially COX-2, reduced the protective effect of IP in edematous pancreatitis. CONCLUSION: Our results indicate that IP reduces pancreatic damage in cerulein-induced pancreatitis and this effect, at least in part, depends on the activity of COXs and pancreatic production of HSP 70.     

Intracellular action of an exogenous low-molecular-weight synthetic protease inhibitor, E3123, in cerulein-induced acute pancreatitis in rats

The intracellular distribution and action of a new synthetic protease inhibitor, E3123, were studied in cerulein-induced acute pancreatitis in rats. Acute pancreatitis was induced by a 4-h iv infusion of a supramaximal dose of cerulein, and was treated by prophylactic (pretreatment) or therapeutic (posttreatment) continuous administration of E3123. Pancreatic edema and hyperamylasemia were ameriolated only by prophylactic treatment. A subcellular fractionation study showed that the activities of cathepsin-B and trypsin in the zymogen granule-enriched fraction of the cerulein-pancreatitis group were remarkably increased. Both prophylactic and therapeutic treatment significantly prevented the elevation of these enzyme activities. These effects were accompanied by amelioration of pancreatic histopathological features, including intracellular vacuolization and fat necrosis. A microscopic autoradiographic study using²H-labeled E3123 showed diffuse intracellular distribution of E3123, and the radioactivity of²H-E3123 in the posttreatment group was three times greater than that in the pretreatment group. This study provides the first experimental evidence that, even when administered therapeutically, exogenous protease inhibitors are transported into pancreatic acinar cells, thereby reducing the seventy of early intracellular alterations in cerulein-induced acute pancreatitis.     

Protease-activated receptor 2 exerts local protection and mediates some systemic complications in acute pancreatitis

BACKGROUND & AIMS: Protease-activated receptor 2 can be stimulated by interstitially released trypsin during acute inflammation of the pancreas. In this study, we investigated the roles of pancreatic and circulatory protease-activated receptor 2 in the pathogenesis of acute pancreatitis by using in vitro and in vivo model systems. METHODS: Physiological and pathologic effects of protease-activated receptor 2 activation were measured in isolated pancreatic cells and in rats with experimental pancreatitis. Consequences of protease-activated receptor 2 activation on the systemic and inflammatory responses were measured after treatments with trypsin or protease-activated receptor 2-activating peptide. RESULTS: Stimulation of protease-activated receptor 2 in rat pancreatic acinar cells activated short-lasting (Ca(2+) signaling) and long-lasting (extracellular signal-related kinase) signaling pathways and protected the cells against bile-induced cell damage. More importantly, protease-activated receptor 2 activation ameliorated the pathologic effects observed in the in vivo model of cerulein-induced pancreatitis. Trypsin in the circulation of rats with taurocholate-induced severe acute pancreatitis reached levels sufficient to activate endothelial and immune cells to stimulate nitric oxide and interleukin-8 production, respectively. Most notably, activation of systemic protease-activated receptor 2 by circulating protease-activated receptor 2 agonists induced a hemodynamic response pattern similar to that observed in rats with severe acute pancreatitis. The effects of protease-activated receptor 2 agonists and acute pancreatitis were not additive. CONCLUSIONS: These findings suggest that protease-activated receptor 2 may have a dual role in acute pancreatitis: protecting acinar and duct cells against pancreatitis-induced cell damage while mediating or aggravating the systemic complications of acute pancreatitis, which are the major cause of mortality in the early phase of necrotizing pancreatitis.     

Effect of BN52021 on NFκ-Bp65 expression in pancreatic tissues of rats with severe acute pancreatitis

AIM: To investigate dynamic changes and significance of expression of NF-κBp65 in pancreatic tissues of rats with severe acute pancreatitis (SAP), as well as BN52021 effects.METHODS: Wistar male rats were randomly divided into negative control group (NC group, n = 60), SAP-model group (SAP group, n = 60), and BN52021-treated group (BN group, n = 60), and each of the above groups was respectively divided into 6 subgroups at different time points after operation (1 h, 2 h, 3 h, 6 h, 12 h, and 24 h) (n = 10). By RT-PCR and Western blot, NF-KBp65 mRNA and its protein expression in pancreatic tissues of rats were detected respectively.RESULTS: The expression of NF-KBp65 mRNA dynamically changed in both SAP groups and BN groups. The mRNA level was higher in SAP groups than NC groups at 2 h, 3 h, 12 h, and 24 h after operation (P < 0.05), higher in BN groups than NC groups at all time points (P < 0.05), and higher in BN groups than SAP group at 1 h (P < 0.05). The NF-jcBp65 protein level was higher in SAP groups than NC groups at 1 h, 3 h, and 6 h (P < 0.01), and 2 h, 12 h, and 24 h (P < 0.05), higher in BN groups than NC groups at all time points (P < 0.05), and lower in BN groups than SAP groups at 1 h, 3 h, and 6 h (P < 0.05).CONCLUSION: The expression of NF-icBp65 in pancreatic tissues is dynamically changed and the changes play an important role in pathogenesis of SAP. BN52021 exerts therapeutic effects through reducing the expression level of NF-κBp65 protein in the early stage of SAP.     

Effect of BN52021 on NFκ-Bp65 expression in pancreatic tissues of rats with severe acute pancreatitis

AIM： To investigate dynamic changes and significance of expression of NF-κBp65 in pancreatic tissues of rats with severe acute pancreatitis （SAP）, as well as BN52021 effects.
METHODS： Wistar male rats were randomly divided into negative control group （NC group, n = 60）, SAP-model group （SAP group, n = 60）, and BN52021-treated group （BN group, n = 60）, and each of the above groups was respectively divided into 6 subgroups at different time points after operation （1 h, 2 h, 3 h, 6 h, 12 h, and 24 h） （n = 10）. By RT-PCR and Western blot, NF-κBp65 mRNA and its protein expression in pancreatic tissues of rats were detected respectively.
RESULTS： The expression of NF-κBp65 mRNA dynamically changed in both SAP groups and BN groups. The mRNA level was higher in SAP groups than NC groups at 2 h, 3 h, 12 h, and 24 h after operation （P 〈 0.05）, higher in BN groups than NC groups at all time points （P 〈 0.05）, and higher in BN groups than SAP group at 1 h （P 〈 0.05）. The NF-κBp65 protein level was higher in SAP groups than NC groups at 1 h, 3 h, and 6 h （P 〈 0.01）, and 2 h, 12 h, and 24 h （P 〈 0.05）, higher in BN groups than NC groups at all time points （P 〈 0.05）, and lower in BN groups than SAP groups at 1 h, 3 h, and 6 h （P 〈 0.05）.
CONCLUSION： The expression of NF-κBp65 in pancreatic tissues is dynamically changed and the changes play an important role in pathogenesis of SAR BN52021 exerts therapeutic effects through reducing the expression level of NF-κBp65 protein in the early stage of SAR     

Endogenous hypertriglyceridemia intensifies the course of cerulein-induced pancreatitis in rat: relation with changes in the VLDL composition

AIMS: To study if the course of cerulein-induced pancreatitis in rats changes in a state of triglyceride-rich lipoprotein metabolism alteration. METHODS: Two groups of rats received control diet during a 90-day period (A) and sucrose-rich diet to induce endogenous hypertriglyceridemia (B). Subgroups A2 and B2 received i.p. 45 microg cerulein/kg body weight (to induce acute pancreatitis). Histological examination of pancreas tissue, serum pancreatic lipase, lipoprotein profile and VLDL chemical composition were assessed. Then, pancreatic lipase hydrolytic activity on VLDL-triglycerides was evaluated in vitro. RESULTS: Cellular vacuolization was observed in all of the cerulein-injected rats, but only in subgroup B2 fat necrosis was present. Serum triglycerides were higher in subgroup B1 than in subgroup A1 (mean +/- SEM, mg/dl 123,77 +/- 25.7 vs. 65.8 +/- 7, p < 0.01). Triglycerides from rats fed with sucrose-rich diet, decreased after cerulein-induced pancreatitis (80.38 +/- 11.3 vs. 123,77 +/- 25.7, p < 0.02). Moreover, the endogenous hypertriglyceridemic rats showed an increment of VLDL triglyceride content, which decreased when rats were injected with cerulein. A negative correlation was found between VLDL-triglyceride content and serum pancreatic lipase activity (r = 0.58, p < 0.02). The in vitro assay showed a decrease in VLDL-triglyceride content post incubation with pancreatic lipase enriched serum (mean +/- SD: 59.2 +/- 27.7%, p < 0.01). CONCLUSIONS: The endogenous hypertriglyceridemia intensifies the course of cerulein-induced pancreatitis and it could be related to the decrease in VLDL-triglycerides as a consequence of pancreatic lipase hydrolytic activity.     

Influence of PMN Leukocyte-Mediated Pancreatic Damage on the Systemic Immune Response in Severe Acute Pancreatitis in Rats

The outcome of severe acute pancreatitis is determined by the development of the systemic inflammatory response and subsequent multiorgan dysfunction. Using the taurocholate-induced model of acute pancreatitis in rats, we investigated the relationship between early polymorphonuclear (PMN)-mediated pancreatic tissue damage and the systemic inflammatory response. The respiratory burst of PMN leukocytes was increased in animals with acute pancreatitis and was reduced by anti-ICAM-1 antibody and oxygen radical scavenger treatment after 24 hr. In acute pancreatitis a reduced number of peripheral helper T cells was evident, most likely due to L-selectin-mediated increased lymphocyte homing. After 24 hr the CD45RC(high)/CD45RC(low) ratio of helper T cells, a critical factor in T cell-mediated disease was increased due to a reduction of regulatory CD45RC(low) cells. Only the treatment with anti-ICAM-1 mAb affected these changes, indicating that immunological changes in necrotizing pancreatitis are only in part affected by early PMN leukocyte-mediated pancreatic damage.     

Cholecystokinin antagonist prevents hyperamylasemia and improves pancreatic exocrine function in cerulein-induced acute pancreatitis.

Supramaximal cerulein administration induces acute pancreatitis, which markedly impairs pancreatic secretion in conscious rats. We hypothesized that pretreatment with the potent cholecystokinin antagonist, L-364,718, improves the pancreatic secretory impairment associated with cerulein-induced acute pancreatitis. Rats were surgically prepared with gastric, duodenal, bile, and pancreatic fistulas and jugular vein catheters. On postoperative day 4, groups of rats were administered (a) L-364,718 1 mg/kg intraduodenally, (b) cerulein 5 micrograms/kg/h for 6 h intravenously, (c) L-364,718 1 mg/kg intraduodenally followed by cerulein 5 micrograms/kg/h for 6 h intravenously, and (d) safflower oil carrier intraduodenally. On postoperative day 5, we studied cholecystokinin (CCK)-stimulated pancreatic secretion. Plasma amylase was measured at the time of surgery and at the conclusion of experiments on postoperative days 4 and 5. The duodenally administered CCK antagonist had no effect, 24 h later, on CCK-evoked protein secretion and prevented the pancreatic exocrine impairment and hyperamylasemia caused by supramaximal cerulein administration. These observations suggest that cerulein-induced acute pancreatitis is mediated by a CCK-receptor mechanism.     

大鼠急性胰腺炎模型建立的探究

目的 建立一种简单易行和稳定可靠的L-精氨酸诱导的急性胰腺炎模型,以便对治疗急性胰腺炎的疗效进行观察和探讨.方法 选择36只健康SD大鼠,采用15％和20％不同浓度的L-精氨酸溶液（用0.9％的生理盐水配制而成）1.5 g/kg腹腔内注射入大鼠体内,分别于不同的时间点取材,分别检测大鼠的存活率、血生化指标中的血清淀粉酶和脂肪酶以及大鼠胰腺的病理变化.结果 15％L-精氨酸溶液腹腔注射大鼠的生存率明显高于20％组,实验组的动物12h后血清淀粉酶和血清脂肪酶水平明显升高,显著高于对照组（P ＜0.0001）,但两实验组之间差异无统计学意义（P＞0.05）,病理结果显示胰腺腺泡结构紊乱,大量腺泡细胞水肿、坏死,出现核固缩,胞内空泡形成,小叶间隙增宽并伴有炎性细胞浸润及出血.20％L-精氨酸组病理改变较严重,腺泡和导管消失并伴有皂化斑形成.结论 应用15％ ～ 20％的L-精氨酸溶液1.5 g/kg的剂量可致不同程度的比较稳定的急性胰腺炎模型.并且其浓度大小决定胰腺损伤的程度.15％L-精氨酸所致急性胰腺炎是比较理想的模型.     

Therapeutic effect of basic fibroblast growth factor on experimental pancreatitis in rat

Basic fibroblast growth factor (bFGF) is one of the mitogens that facilitate endothelial proliferation and angiogenesis. This study was designed to examine the therapeutic effect of bFGF on experimental pancreatitis in rat. Edematous pancreatitis was induced by intraperitoneal injections of cerulein (50 microg/kg) at hourly intervals. BFGF (70 nmol/kg) was administered intraperitoneally after induction of pancreatitis. DNA synthesis of isolated pancreatic acinar cells of normal rats was determined as the uptake of 5-bromo-2'-deoxyuridine (BrdU) into the cells. Immunohistochemical staining of DNA synthesis in acinar cells during cerulein-induced pancreatitis was also examined with BrdU labeling in vivo technique. Cerulein administration increased serum amylase, lipase level, and wet weight of pancreatic tissue. Treatment with bFGF markedly ameliorated all these parameters. In primary culture system of isolated pancreatic acinar cells of normal rats, bFGF caused a dose-dependent increase in BrdU incorporation into DNA, showing an EC50 value of 0.8 nmol/L and a maximum response of 2.5-fold increase at a concentration of 400 nmol/L. bFGF treatment (70 nmol/kg) markedly increased BrdU labeling in the nucleus of acinar cells of the pancreatitis rats group in immunohistochemical examination when compared with control without bFGF treatment. Treatment with bFGF may represent a promising therapeutic concept for patients with acute pancreatitis.