Endogenous glucocorticoids decrease the acinar cell sensitivity to apoptosis during cerulein pancreatitis in rats

BACKGROUND & AIMS: We recently showed that activation of the hypothalamus-pituitary-adrenal axis may mitigate the progress of acute pancreatitis. To clarify the mechanism, the role of endogenous glucocorticoids in pancreatic acinar cell death was examined. METHODS: The occurrence of apoptosis was studied in adrenalectomized or sham-operated rats with or without cerulein-induced pancreatitis. The effects of RU38486, a glucocorticoid-receptor antagonist, on the survival of cultured acinar cells (AR42J) were also examined. RESULTS: Adrenalectomy caused increases in terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick-end labeling (TUNEL) of acinar nuclei depending on the time after adrenalectomy but not of other cell types in the pancreas and in other digestive organs. Electron microscopy showed the characteristic features of apoptosis in the TUNEL-labeled acinar cells. In cerulein pancreatitis of adrenalectomized rats, the TUNEL-labeled acinar nuclei increased remarkably depending on the time after cerulein infusion. Replacement of glucocorticoids blocked the occurrence of apoptosis in these experiments. RU38486 induced dose dependently the apoptosis of AR42J cells. CONCLUSIONS: These results provide evidence that endogenous glucocorticoids are an important factor for acinar cell survival. Endogenous glucocorticoids may protect acinar cells by decreasing their sensitivity to the induction of cell death during acute pancreatitis.     

Triflavin potentiates the antiplatelet activity of platelet activating factor receptor antagonist on activated neutrophil-induced platelet aggregation

In this study, specific platelet activating factor (PAF) receptor antagonist ginkgolide B (BN52021) was tested for its antiplatelet activity in zymosan activated polymorphonuclear neutrophil-induced platelet aggregation. Triflavin was also tested for its antiplatelet activity compared with PAF receptor antagonist. Triflavin, an Arg-Gly-Asp-containing disintegrin purified from venom peptide inhibited platelet aggregation by interfering with the interaction of fibrinogen with the glycoprotein IIb/IIIa complex. Furthermore, we also report an efficient high resolution method for quantitative analysis of PAF using high-performance capillary electrophoresis (HPCE). The supernatant of polymorphonuclear neutrophils after their activation by opsonized zymosan induces the aggregation of washed rabbit platelets. In rabbit platelets, BN52021 (100-1000 microM) only partially inhibited activated polymorphonuclear neutrophil-induced platelet aggregation, and its maximal inhibition was estimated to be about 79%. Triflavin also partially inhibited platelet aggregation about 82% induced by activated polymorphonuclear neutrophils. Furthermore, after treatment with a combination of triflavin (0.26 microM) with various concentrations of BN52021 (4-1000 microM), the inhibitory effect of platelet aggregation was almost completely. This inhibition was greater than that produced by the individual drugs alone. These results indicate that a combination of glycoprotein IIb/IIIa complex and PAF receptor antagonist could completely inhibit activated polymorphonuclear neutrophil-induced platelet aggregation. In addition, the amount of PAF released from zymosan (6 mg/ml)-activated polymorphonuclear neutrophils was accurately calculated about 11.8+/-1.5 ng/10(6) cells, and did not further increase even at a high concentration of zymosan (10 mg/ml). These results suggest that PAF play a major role in the interaction between platelets and polymorphonuclear neutrophils. This interaction may be important in the pathogenesis of thrombosis and inflammatory diseases. Our present findings support the hypothesis that combination therapy with glycoprotein IIb/IIIa complex antagonists and PAF receptor antagonists may represent a new approach to the treatment of ischemic disorders.     

Effect of recombinant platelet-activating factor acetylhydrolase on two models of experimental acute pancreatitis

BACKGROUND & AIMS: Recent reports suggest that platelet-activating factor (PAF) plays a role in pancreatitis and pancreatitis-associated lung injury. In this study, the effects on these processes of termination of PAF action by recombinant PAF-acetylhydrolase (rPAF-AH) were investigated. METHODS: Rats were given rPAF-AH and then infused with a supramaximally stimulating dose of cerulein to induce mild pancreatitis. Opossums underwent biliopancreatic duct ligation to induce severe pancreatitis, and rPAF-AH administration was begun 2 days later. RESULTS: In mild, secretagogue-induced pancreatitis, rPAF-AH given before the cerulein reduced hyperamylasemia, acinar cell vacuolization, and pancreatic inflammation but did not alter pancreatic edema or pulmonary microvascular permeability. In severe, biliopancreatic duct ligation-induced pancreatitis, rPAF-AH delayed and reduced the extent of inflammation and acinar cell injury/necrosis and completely prevented lung injury even though the rPAF-AH administration was begun after the onset of pancreatitis. CONCLUSIONS: PAF plays an important role in the regulation of pancreatic injury but not pancreatic edema or increased pulmonary microvascular permeability in mild, secretagogue-induced pancreatitis. PAF plays a critical role in the regulation of progression of pancreatic injury and mediation of pancreatitis-associated lung injury in severe biliary pancreatitis. Amelioration of pancreatitis and prevention of pancreatitis-associated lung injury can be achieved with rPAF-AH even if treatment is begun after pancreatitis is established.     

Histological study of saphenous vein graft disease after coronary artery bypass grafting

This study examines whether platelet-activating factor (PAF) is involved in the occurrence of vasospasm after subarachnoid hemorrhage (SAH). A vasospasm model was produced in rabbits, with animals in six experimental groups receiving two subarachnoid injections of autologous arterial blood with the addition of one of the following; saline (Control Group 1), 25% dimethyl sulfoxide (Control Group 2), PAF (1, 2.5, 5, or 10 micrograms), CV6209 (10 or 100 micrograms), BN52021 (10 or 100 micrograms), or anti-PAF immunoglobulin G (IgG, 50 or 500 micrograms). No significant differences were detected between Control Groups 1 and 2 with regard to neurological deterioration and basilar artery constriction after SAH was induced. Administration of PAF together with autologous blood aggravated neurological deficits in a dose-dependent manner (r = 0.724, p < 0.001) and produced basilar artery constriction at two doses each of 2.5 micrograms (p < 0.05), 5 micrograms (p < 0.01), and 10 micrograms (p < 0.01). Neurological deterioration was prevented in rabbits receiving an intracisternal administration of either PAF antagonist CV6209 or BN52021 or anti-PAF IgG (p < 0.01 at a total dose of 20 micrograms and p < 0.05 at a total dose of 200 micrograms CV6209, p < 0.01 at total doses of 20 and 200 micrograms BN52021, and p < 0.01 at total doses of 100 and 1000 micrograms anti-PAF IgG). A reduction in basilar artery constriction was achieved by the injection of anti-PAF IgG (p < 0.05 at total doses of 100 and 1000 micrograms). Histological examination at autopsy on Days 14 to 21 showed mainly ischemic changes in the brain, including selective neuronal necrosis and cerebral infarction. The control and PAF groups showed marked ischemic changes. On the other hand, no ischemic changes were noted in the anti-PAF IgG group, and only 9% of animals in the CV6209 group and 25% in the BN52021 group demonstrated selective neuronal necrosis or infarction. This study thus provides evidence to support the role of PAF in the pathogenesis of vasospasm after SAH.     

Effects of methylglyoxal on platelet hydrogen peroxide accumulation, aggregation and release reaction

Alpha-naphthylisothiocyanate (ANIT) is a cholangiolitic hepatotoxicant that causes periportal edema, hepatic parenchymal and biliary epithelial cell necrosis, and cholestasis in the rat. A hallmark of ANIT hepatotoxicity is periportal inflammation that includes neutrophil infiltration. Neutrophils are requisite for the expression of ANIT-induced liver injury; however, the mechanism(s) of neutrophil accumulation in the liver and the role of these cells in ANIT hepatotoxicity is incompletely understood. Platelet activating factor (PAF) is a proinflammatory agent that is both a chemoattractant for and an activator of neutrophils. Therefore, we evaluated the role of PAF in ANIT-induced liver injury. Rats were treated with the PAF receptor antagonist, WEB 2086 (WEB), to determine if it afforded protection from ANIT hepatotoxicity. In a separate study, a combination of WEB and the leukotriene synthesis inhibitor, Zileuton (ZIL), was used to address the possible interaction of PAF and leukotrienes in ANIT-induced liver injury. Treatment of rats with WEB, alone or in combination with Zileuton, did not attenuate ANIT-induced liver injury as assessed by increases in plasma alanine aminotransferase or gamma-glutamyl transferase activities. In addition, neither treatment ameliorated ANIT-induced cholestasis assessed as increased plasma bilirubin concentration. These results suggest that PAF, alone or in combination with products of 5-lipoxygenase, does not contribute to ANIT-induced liver injury.     

The platelet-activating factor antagonist BN 52021 attenuates hypoxic-ischemic brain injury in the immature rat

Platelet-activating factor (PAF) is overproduced in ischemic brain. Although postischemic PAF antagonist administration protects the mature brain in some models, little is known about the effects of PAF antagonists in the immature brain. We hypothesized that the PAF antagonist BN 52021 would attenuate perinatal cerebral hypoxic-ischemic injury. To elicit focal hypoxic-ischemic brain injury, 7-d-old (P7) rats (n = 111) underwent right carotid ligation, followed by 2.5-3.25 h of hypoxia (fractional concentration of inspired O2 = 0.08). BN 52021 neuroprotection was evaluated in three groups of experiments: 1) 25 mg/kg/dose, 0 and 2 h posthypoxia; 2), 25 mg/kg/dose immediately before and 1 h after hypoxia; and 3) posthypoxia-ischemia treatment with BN 52021 12.5, 25, or 50 mg/kg/dose in 2 doses 0 and 2 h after hypoxia. All experiments included concurrent vehicle-injected controls. To quantitate severity of injury, bilateral regional cross-sectional areas (groups 1 and 2) or hemisphere weights (group 3) were evaluated on P12. Both pre- and posthypoxic treatment with BN 52021 (25 mg/kg/dose, two serial doses) decreased the incidence of cerebral infarction from 90% to about 30% (p < 0.02, Fisher's exact test). Measurement of cross-sectional areas confirmed neuroprotection and indicated some benefit of pre- over posthypoxic-ischemic treatment in hippocampus and cortex. Over the dose range tested, the neuroprotective effect of BN 52021 administration was not dose-dependent. In contrast, BN 52021 did not attenuate N-methyl-D-aspartate-induced hippocampal excitotoxic injury in P7 rats. Either prophylactic or "rescue" administration of PAF antagonists decreases the incidence and severity of brain injury associated with an episode of perinatal cerebral hypoxia-ischemia.     

Experimental model of cardiovascular post-resuscitation syndrome--no effect of platelet activating factor antagonism

The cardiovascular instability seen in the reperfusion phase after resuscitation from cardiac arrest may contribute to secondary brain injury. The aim of the present study was to characterise post-resuscitation cardiovascular instability in an experimental model of cardiac arrest and to test if cardiovascular stability could be improved by pre-treatment with the platelet activating factor (PAF) antagonist BN52021. Ten anaesthetised pigs received pre-treatment with BN52021 before induction of ventricular fibrillation (arrest/BN52021 group), while ten animals received only the vehicle (arrest/vehicle group). After a non-intervention interval of 9 min, resuscitation was attempted. Resuscitated animals were observed for 5 h and compared to a sham arrest group of seven animals. The haemodynamic situation after resuscitation was characterised by a low cardiac output that was inadequate in relation to the oxygen demand, as reflected by a low mixed venous oxygen saturation. The arterial blood pressure was to some extent reduced and the filling pressures in both the right and left heart were increased, but urinary output was not reduced. The severe haemodynamic compromise was not adequately reflected by standard monitoring variables such as arterial blood pressure and urinary output. Pre-treatment with BN52021 was unable to improve any aspect of short-term survival or haemodynamic stability.     

Molecular mechanisms of transforming growth factor-beta signaling

The present experiments examined the effects of posttraining intrahippocampal injections of the degradative enzyme-resistant methylcarbamyl analog of the bioactive phospholipid platelet-activating factor (mc-PAF) and the platelet-activating factor (PAF) receptor antagonists BN52021 and BN 50730 on memory in male Long-Evans rats trained in a hidden platform version of the Morris water maze. Following an eight-trial training session, rats received a unilateral intrahippocampal injection of mc-PAF (0.5, 1.0, or 2.0 microgram/0.5 microliter), lyso-PAF (1.0 microgram/0.5 microliter), the cell surface PAF receptor antagonist BN 52021 (0.25, 0.5, or 1.0 micrigram/0.5 microliter/, the intracellular PAF receptor antagonist BN 50730 (2.0, 5.0, or 10.0 microgram/0.5 microliter), or vehicle (50% DMSO in 0.9% saline; 0.5 microliter). On a retention test conducted 24 h after training, the escape latencies of rats administered mc-PAF (1.0 or 2.0 microgram) were significantly lower than those of the vehicle-injected controls, demonstrating a memory-enhancing effect of mc-PAF. Injections of lyso-PAF, a structurally similar metabolite of PAF, had no influence on memory, indicating that the memory-enhancing effect of mc-PAF is not caused by membrane perturbation by the phospholipid. The retention test escape latencies of rats administered BN 52021 (0.5 microgram) and BN 50730 (5.0 or 10 microgram) were significantly higher than those of the controls, indicating a memory impairing effect of both PAF antagonists. When mc-PAF, BN 52021, or BN 50730 was administered 2 h posttraining, no effect on retention was observed, indicating a time-dependent effect of the neuroactive substances on memory storage. The findings suggest a role for endogenous PAF in hippocampal-dependent memory processes.     

Effects of ginkgolide B on isobaric hypoxic pulmonary hypertension in rats

We examined the effects of ginkgolide B (BN52021), a specific platelet activating factor (PAF) antagonist, on hypoxia-induced pulmonary hypertension and pulmonary vascular remodeling. Chronic hypoxia can cause pulmonary hypertension and pulmonary vascular remodeling in rats. Treated with BN52021, the rat's pulmonary hypertension, pulmonary vascular remodeling and right ventricular hypertrophy at 3 weeks of hypoxic exposure (FiO2 = 0,10) could be reduced. The results suggest that PAF contributes to the development of chronic hypoxic pulmonary hypertension and pulmonary vascular remodeling, and PAF antagonist may be an useful agent for preventing hypoxic pulmonary hypertension.     

Cell-density-dependent expression of Cyp1a2 gene in monolayer-cultured adult mouse hepatocytes

The effects of PAF antagonists, of substances which influence the arachidonic acid metabolism, and of dexamethasone and ketotifen were evaluated in an acute PAF-induced mortality model in female NMRI mice. We established a dependence of sensitivity to PAF on strain (AB mice showed no dose dependence) and on sex of the animals as well as on the PAF charges used in our experiments. PAF produced resistance in surviving animals against the PAF-induced death on repeated application. The PAF antagonists, WEB 2170 and WEB 2086, provided the best dose-dependent protection against PAF toxicity, followed by dexamethasone, by the COX/LOX synthetase inhibitor X 86 (a BW 755 C-analogue) and by the PAF receptor antagonist BN 52021. Particularly remarkable was the excellent prevention by aspirin. Aspirin may not only inhibit the cyclooxygenase pathway but also endogenous PAF synthesis. Other drugs, i.e. indomethacin, the thromboxane receptor antagonist, BM 13177, the thromboxane synthetase inhibitor, HOE 944, as well as the lipoxygenase inhibitors (NDGA, esculetin, SHAM and phenidone) exerted a dose-dependent protection only at high doses.     

Aortoiliac stent placement in patients treated for intermittent claudication

Inhaled nitric oxide (NO) is an important new therapeutic agent used to treat pulmonary arterial hypertension in a variety of disease states. However, the effects of NO on cells in the lung are uncertain. Previously, we have shown that NO gas depresses neutrophil oxidative cell function and increases neutrophil cell death. The purpose of this in vitro study was to determine the mechanism of neutrophil death. We hypothesized that NO hastened cell death by inducing apoptosis. To mimic the clinical environment of patients with respiratory failure, we also studied the effects of hyperoxia on neutrophil cell viability and apoptosis. Isolated human neutrophils were exposed to 80% O2 (O2), NO at 20 ppm in room air (NO/RA), 20 ppm NO blended with 80% O2 (NO/O2), or RA alone (control) for 2 to 24 h. Experiments were repeated with NO concentrations of 5 and 50 ppm and with 20 ppm in the presence of superoxide dismutase (SOD). Neutrophils were also incubated in the absence or presence of neutrophil stimulant fMLP (10 nM). Neutrophil cell viability was measured by fluorescence viability/cytotoxicity assay. Neutrophil apoptosis was assessed by cell death detection ELISA for histone-associated DNA fragments, TdT transferase-mediated fluorescence-labeled dUTP nick end labeling (TUNEL) assay, and DNA fragmentation gel electrophoresis. NO/O2-exposed neutrophils showed decreased viability at 2 h (31.7 +/- 3.7%, mean % viability +/- SD) compared with control (94.7 +/- 4.7%), O2 (75.6 +/- 9.3%), and NO/RA (62.8 +/- 14.9%; P < 0.05 by ANOVA; n = 9). Although control neutrophils demonstrated marked apoptosis at 24 h, there was no significant apoptosis at 2, 4, or 6 h (P < 0.001 by Kruskal-Wallis, n = 20) as assessed by ELISA and TUNEL assays. When compared with RA controls at 2 h, neutrophils exposed to NO/O2 showed significantly more apoptosis (292% of control, range: 106 to 2,488%, P < 0.001 by ANOVA and Kruskal-Wallis) but not with exposure to NO/RA or O2 alone. These findings were confirmed by TUNEL assay (n = 4, P < 0.05). NO/ RA and NO/O2-exposed neutrophils demonstrated both evidence of necrosis and enhanced DNA fragmentation at 2 h by gel electrophoresis (n = 2). Fifty parts per million NO produced similar findings, but exposure to 5 ppm NO did not induce significant DNA fragmentation. Coincubation with SOD inhibited NO/ O2-associated apoptosis, suggesting peroxynitrite contributed to cell death. Stimulation with fMLP did not alter apoptosis induced in neutrophils exposed to NO/RA or NO/O2. We conclude that exogenous NO gas, at clinically relevant concentrations under hyperoxic conditions, induces cell death in neutrophils in part by enhancing DNA fragmentation.     

The role of xanthine oxidase in platelet activating factor induced intestinal injury in the rat

BACKGROUND: Xanthine oxidase (XO) is an important source of reactive oxygen species in the small intestine. AIMS: To examine the interaction of platelet activating factor (PAF), XO, and neutrophils in mediating intestinal injury in rats. METHODS: Two doses of PAF were used to induce either reversible hypotension, or irreversible shock with intestinal necrosis. The activities of XO, and its precursor xanthine dehydrogenase (XD), in both the whole intestinal tissue and epithelial cells, were measured. XO was localised by histochemical staining. RESULTS: PAF dose dependently induced an increase in XO activity, predominantly in the ileal epithelium, without altering the total activity of XD+XO. Most of the XD to XO conversion was via proteolysis. PAF induced XO activation and intestinal injury were prevented by prior neutrophil depletion. PAF induced XO activation is probably not due to reperfusion, as XO activation preceded the recovery of mesenteric flow. Allopurinol pretreatment substantially inhibited intestinal neutrophil sequestration induced by high dose (but not low dose) PAF. CONCLUSIONS: PAF rapidly activates intestinal XO through proteolytic XD-XO conversion, predominantly in the ileal epithelium. This effect is mediated by neutrophils. XO activation promotes PAF induced polymorphonuclear leucocyte sequestration in the intestine.     

The learning ability paradox in adult metamemory research: where are the metamemory differences between good and poor learners?

We studied morphologic changes in a rat model of acute hemorrhagic pancreatitis in order to investigate the mechanism by which water immersion stress injures the pancreas. Acute hemorrhagic pancreatitis was induced by two intraperitoneal injections of 40 micrograms/kg body weight of caerulein at 1-h intervals under water immersion stress for 5 h at 23 degrees C. Light microscopy showed interstitial edema with inflammatory cell infiltration, degeneration and necrosis of acinar cells, and bleeding. Electron microscopy showed large autophagic vacuoles, decreased zymogen granules, and dilated rough endoplasmic reticulum in acinar cells. Basolateral exocytosis of large vacuoles and phagocytosis of the degenerated acinar cells were observed. In addition, microvascular damage, including the destruction of the capillary endothelial cells, capillary thrombosis, and the extravasation of blood cells, was seen. In contrast, in a pancreatitis model induced by caerulein injection alone, there was no bleeding, no remarkable vascular change, and no thrombosis. Degeneration and necrosis of acinar cells were less severe. In the pancreas under stress alone, microvascular damage and degeneration of acinar cells were observed. These findings demonstrate that stress injures the pancreas and worsens the pancreatitis by causing microcirculatory disturbances, such as vascular damage, thrombosis, increased vascular permeability, and bleeding. These results suggest that chemical mediators, such as free radicals and platelet-activating factor (PAF), which are produced by vascular damage and thrombosis, may accelerate the activation of zymogen proteases in acinar cells in caerulein-induced pancreatitis, leading to hemorrhagic pancreatitis.     

Involvement of platelet-activating factor in cell death induced under ischemia/postischemia-like conditions in an immortalized hippocampal cell line

The involvement of platelet-activating factor (PAF) in cell damage induced by ischemia/postischemia-like conditions was studied in a hippocampus-derived cell line, HN33.11. Cells exposed to N2-saturated glucose-free HEPES-buffered saline (ischemia) for 5 h followed by 18 h of incubation in serum-free control medium (postischemia reincubation) remained 67.4 +/- 2.4% viable in comparison with sham-treated cells. Analysis of DNA fragmentation in combination with Hoechst 33258 staining indicates that apoptosis is the dominant mode of cell death in the present model. PAF level during 10 h of ischemia was unchanged. However, an increase in PAF accumulation was found early during the reincubation period that followed 5 h of ischemia. Peak PAF concentrations were noted at 2 h after initiation of reincubation and rapidly declined to control level after 7 h of reincubation. Consistent with a role of PAF in mediating cell death under ischemia/postischemia reincubation in this model, the PAF antagonist BN 50739 exerted a dose-dependent protective effect. Maximal protection (85.7 +/- 5.4%) of the cells from ischemia/reincubation-induced cell damage was achieved at 0.1 microM BN 50739. The PAF antagonist lacked any protective effect against ischemia-induced cell death. On the other hand, the addition of the stable PAF analogue 1-O-hexadecyl-2-N-methylcarbamyl-sn-glycero-3-phosphocholine (MC-PAF) at the onset of ischemia potentiated ischemia/reincubation-induced apoptosis--an effect that was blocked by BN 50739. Pretreatment of HN33.11 cells with the Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N,N-tetraacetic acid acetoxymethyl ester (BAPTA-AM) also provided a protective effect against ischemia/reincubation-induced cell damage. BAPTA-AM increased cell viability by 50%. Pretreatment with BAPTA-AM also decreased ischemia/reincubation-induced PAF accumulation in HN33.11 cells. The results suggest that PAF, acting via a PAF receptor, is at least in part mediating apoptosis under ischemia/postischemia-like conditions in HN33.11 cells.     

The routine analysis of breast milk for drugs of abuse in a clinical toxicology laboratory

BACKGROUND/AIMS: Contrast-enhanced computed tomography (CECT) is used to show areas of decreased pancreatic perfusion in severe acute pancreatitis (AP). To evaluate possible adverse effects of the contrast medium (CM) on the course of AP, the impact of intravenous CM in AP of graded severity in the rat was studied. METHODS: Pancreatitis of three levels of severity was induced in Sprague-Dawley rats with intravenous cerulein hyperstimulation plus time- and pressure-controlled intraductal infusion of saline or glycodeoxycholic acid. At 7 hours, control and pancreatitis animals received intravenous ionic CM, nonionic CM, or saline. The principal outcome measures were 24-hour survival, trypsinogen activation peptides (TAP) in ascites, and histological acinar necrosis score. RESULTS: There was no measurable effect of CM on the index features in control animals or animals with mild or moderate AP. In severe AP, CM caused a significant increase in mortality, ascites TAP, and necrosis score. CONCLUSIONS: Intravenous CM increases pancreatic injury when administered early in the course of severe experimental AP. Because CM may convert borderline ischemia to irreversible necrosis, CECT performed early in pancreatitis to show poor perfusion and predict areas of necrosis may depict a self-fulfilling prophecy. Early CECT should be reconsidered and perhaps avoided.     

Free radical enhancement promotes leucocyte recruitment through a PAF and LTB4 dependent mechanism

In the present investigation we studied the concerted role of superoxide anion, platelet activating factor (PAF) and leukotriene B4 (LTB4) in the mechanism that results in polymorphonuclear leucocyte accumulation induced by oxygen free radicals in rat pancreas. This was done by comparing the effects of a PAF antagonist (BN-52021), a LTB4 inhibitor (MK-886) and superoxide dismutase (SOD) in a experimental rat model of inflammation elicited by the oxygen free radicals induced via infusion of xanthine/xanthine oxidase. Also, the effect of independent LTB4 infusion has been studied. The results show that increases in polymorphonuclear cell infiltration (evaluated by tissue histology), myeloperoxidase and LTB4 levels induced in pancreas by infusion of xanthine/xanthine oxidase were abolished by the administration of either the PAF antagonist, the LTB4 inhibitor, or SOD. The fact that BN-52021 could prevent neutrophil recruitment and LTB4 synthesis suggests that PAF is a necessary step for subsequent LTB4 synthesis and polymorphonuclear leucocyte accumulation.     

Chemokine gene expression in rat pancreatic acinar cells is an early event associated with acute pancreatitis

BACKGROUND & AIMS: The molecular mechanisms underlying pancreatitis are largely unknown. The goal of this study was to identify an early genetic event that correlated with pancreatitis. METHODS: Differential display of messenger RNAs (mRNAs) was conducted on normal pancreas vs. those of animals with secretagogue-induced pancreatitis. Northern blots from normal animals and animals with experimental acute pancreatitis were probed with cloned complementary DNAs for chemokines. Pancreatitis was induced with cerulein and by retrograde injection of bile salts. Immunocytochemistry was used to identify the source of chemokine expression. Pyrrolidine dithiocarbamate was tested for effects on chemokine expression and pancreatitis. RESULTS: A differentially amplified band was consistently observed early after cerulein hyperstimulation. This band was identified as a portion of the mob-1 gene, an alpha-chemokine. Northern analysis indicated that mRNAs for mob-1 and another chemokine, mcp-1, were induced after cerulein hyperstimulation in vivo. mob-1 mRNA was also induced by retrograde injection of bile salts and by cerulein in acinar cells in vitro. mob-1 protein was localized to exocrine cells in pancreata of diseased animals. Pyrrolidine dithiocarbamate inhibited both chemokine gene expression and early inflammatory characteristics of pancreatitis. CONCLUSIONS: Chemokines are induced in acinar cells by treatments that induce pancreatitis and may play an important role in the early stages of the disease.     

Role of substance P and the neurokinin 1 receptor in acute pancreatitis and pancreatitis-associated lung injury

Substance P, acting via the neurokinin 1 receptor (NK1R), plays an important role in mediating a variety of inflammatory processes. However, its role in acute pancreatitis has not been previously described. We have found that, in normal mice, substance P levels in the pancreas and pancreatic acinar cell expression of NK1R are both increased during secretagogue-induced experimental pancreatitis. To evaluate the role of substance P, pancreatitis was induced in mice that genetically lack NK1R by administration of 12 hourly injections of a supramaximally stimulating dose of the secretagogue caerulein. During pancreatitis, the magnitude of hyperamylasemia, hyperlipasemia, neutrophil sequestration in the pancreas, and pancreatic acinar cell necrosis were significantly reduced in NK1R-/- mice when compared with wild-type NK1R+/+ animals. Similarly, pancreatitis-associated lung injury, as characterized by intrapulmonary sequestration of neutrophils and increased pulmonary microvascular permeability, was reduced in NK1R-/- animals. These effects of NK1R deletion indicate that substance P, acting via NK1R, plays an important proinflammatory role in regulating the severity of acute pancreatitis and pancreatitis-associated lung injury.