-
[show abstract]
[hide abstract]
ABSTRACT: Acute pancreatitis is an inflammatory disease involving acinar cell injury, and the rapid production and release of inflammatory cytokines, which play a dominant role in local pancreatic inflammation and systemic complications. Toll-like receptor 4 (TLR4) initiates a complex signalling pathway when it interacts with lipopolysaccharide (LPS), which ultimately results in a proinflammatory response. We hypothesised that TLR4 is important in the pathophysiology of acute pancreatitis, independently of LPS. Using two different models of acute pancreatitis, we investigated how genetic deletion of TLR4 or its co-receptor CD14 effects its progression and severity.
We induced acute pancreatitis by administering either caerulein or L-arginine to wild-type, TLR4(-/-), and CD14(-/-) mice. Control mice received normal saline injections. The severity of acute pancreatitis was determined by measuring serum amylase activity, quantifying myeloperoxidase (MPO) activity in the pancreatic tissue, and histologically assessing acinar cell injury.
It was found that administering caerulein and L-arginine to wild-type mice resulted in acute pancreatitis (as assessed by hyperamylasaemia, oedema, increased pancreatic MPO activity, and pancreatic necrosis) and associated lung injury. The same treatment to TLR4(-/-) or CD14(-/-) mice resulted in significantly less severe acute pancreatitis, and reduced lung injury. We found no evidence of either bacteria or LPS in the blood or in pancreatic tissue.
The severity of acute pancreatitis is ameliorated in mice that lack either TLR4 or CD14 receptors. Furthermore, these results indicate that TLR4 plays a significant pro-inflammatory role independently of LPS in the progression of acute pancreatitis.
Gut 03/2009; 58(6):813-9. · 10.11 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Intercellular adhesion molecule 1 (ICAM-1) and neutrophils play important roles in many inflammatory processes, but their importance in both acute pancreatitis and pancreatitis-associated lung injury has not been defined.
To address this issue, mice that do not express ICAM-1 were used and depleted of neutrophils by administration of antineutrophil serum. Pancreatitis was induced by administering either supramaximal doses of the secretagogue cerulein or feeding a choline-deficient, ethionine-supplemented diet. The severity of pancreatitis was evaluated by quantitating serum amylase, pancreatic edema, acinar cell necrosis, and pancreas myeloperoxidase activity (i.e., neutrophil content). Lung injury was evaluated by quantitating lung myeloperoxidase activity and pulmonary microvascular permeability. ICAM-1 was quantitated by enzyme-linked immunosorbent assay and was localized by light-microscopic immunohistochemistry.
It was found that serum, pancreas, and lung ICAM-1 levels increase during pancreatitis. Both pancreatitis and the associated lung injury are blunted, but not completely prevented, in mice deficient in ICAM-1. Neutrophil depletion also reduces the severity of both pancreatitis and lung injury. However, the combination of neutrophil depletion with ICAM-1 deficiency does not reduce the severity of pancreatitis or lung injury to a greater extent than either neutrophil depletion or ICAM-1 deficiency alone. Neither pancreatitis nor pancreatitis-associated lung injury are completely prevented by ICAM-1 deficiency, neutrophil depletion, or combined ICAM-1 deficiency plus neutrophil depletion.
The observations indicate that ICAM-1 plays an important, neutrophil-mediated, proinflammatory role in pancreatitis and pancreatitis-associated lung injury. The studies also indicate that ICAM-1 and neutrophil-independent events also contribute to the evolution of pancreatitis and lung injury in these models.
Gastroenterology 04/1999; 116(3):694-701. · 11.68 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: beta-Chemokines and their receptors mediate the trafficking and activation of a variety of leukocytes including the lymphocyte and macrophage. An array of no less than eight beta-chemokine receptors has been identified, four of which are capable of recognizing the chemokines MIP1alpha and RANTES. Genetic deletion of one of the MIP1alpha and RANTES receptors, CCR5, is associated with protection from infection with HIV-1 in humans, while deletion of the ligand MIP1alpha protects against Coxsackie virus-associated myocarditis. In this report we show that the deletion of another receptor for MIP1alpha and RANTES, the CCR1 receptor, is associated with protection from pulmonary inflammation secondary to acute pancreatitis in the mouse. The protection from lung injury is associated with decreased levels of TNF-alpha in a temporal sequence indicating that the activation of the CCR1 receptor is an early event in the systemic inflammatory response syndrome.
Journal of Clinical Investigation 11/1997; 100(8):2022-7. · 15.39 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Acute pancreatitis is characterized by hyperamylasemia, pancreatic edema, and the presence of activated digestive enzymes within the pancreas. The secretagogue-induced model of acute pancreatitis is also characterized by pancreatic acinar cell vacuolation, subcellular redistribution of lysosomal hydrolases, and a fall in pancreatic glutathione levels. We have performed time-dependence studies to determine the sequence with which these phenomena appear and to establish their cause-and-effect relationship. Evidence of lysosomal enzyme redistribution and trypsinogen activation within the pancreas could be detected within 10-15 min of the onset of supramaximal secretagogue stimulation, while hyperamylasemia (30 min), pancreatic edema (60 min), and acinar cell vacuolation (60 min) occurred at later times. Pancreatic glutathione levels were either unchanged (15 and 30 min) or elevated (60 min) during the early times of supramaximal stimulation and were only noted to be decreased at a later time. These results support the conclusion that intrapancreatic digestive enzyme activation, possibly occurring by a mechanism involving lysosomal hydrolase redistribution, is an early and likely a critical event in the evolution of secretagogue-induced pancreatitis but that glutathione depletion is neither early nor critical to the evolution of this model of pancreatitis.
The American journal of physiology 08/1996; 271(1 Pt 1):G20-6.
-
[show abstract]
[hide abstract]
ABSTRACT: The recent observation that the severity of pancreatitis is inversely related to the extent of apoptosis in five experimental models of the disease has suggested the possibility that apoptosis might protect against pancreatic injury in pancreatitis. This hypothesis was tested by inducing pancreatitis in mice during a phase of extensive apoptosis. Mice were fed a raw soya diet for five weeks to stimulate pancreatic growth and then switched to a regular chow diet for 27 hrs to permit involution of the hypertrophied gland. That involution is characterized by extensive apoptosis of acinar cells. Pancreatitis was induced, in either control mice or mice undergoing pancreatic involution, by repeated intraperitoneal administration of a supramaximally stimulating dose of caerulein (50 microg/kg given each hr for 12 hrs). The magnitude of hyperamylasemia, degree of inflammation, and extent of necrosis were reduced in the mice receiving caerulein during pancreatic involution. We conclude that induction of apoptosis may protect against acinar cell injury and reduce the severity of pancreatitis.
Biochemical and Biophysical Research Communications 04/1996; 220(3):875-8. · 2.48 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The events that characterise recovery from severe biliary pancreatitis have not been defined. This study used a reversible model of necrotising pancreatitis, induced by obstructing the opossum common bile pancreatic duct (CBPD), to evaluate this phenomenon. The CBPD of opossums was obstructed with a balloon tipped catheter for five days and then decompressed by removal of the catheter. Recovery was evaluated 0-90 days after relief of obstruction. Serum bilirubin and amylase values rapidly declined, reaching control values 7-14 days after removal of the obstructing catheter. Pancreatic protein and amylase values were transiently increased shortly after relief of obstruction but returned to control values 21 days after decompression. Pancreatic ornithine decarboxylase activity and incorporation of [3H]-thymidine into DNA were transiently increased 14 days after duct decompression suggesting that regeneration occurs at approximately that time. Foci of pancreatic necrosis involved roughly 40% of the gland at time of decompression but these foci gradually disappeared and the gland resembled that of control animals 60 days after decompression. Evidence of fibrosis or collagen deposition in the pancreas was not noted at any time. These studies show that recovery after necrotising biliary pancreatitis occurs comparatively rapidly and the restitution ad integrum occurs. Recovery from necrotising acute pancreatitis in this model is not associated with the development of chronic pancreatitis.
Gut 10/1995; 37(3):427-33. · 10.11 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Cerulein-induced pancreatitis causes a rapid increase in pancreatic enzyme levels in serum and decreases in pancreatic duct secretion and interstitial edema. One mechanism to explain these early events is disruption of the actin tight junction paracellular seal of acinar and intralobular pancreatic duct cells.
To examine the paracellular barrier of the proximal exocrine pancreas, rats were hyperstimulated with 5.0 micrograms.kg-1.h-1 of cerulein. Actin was visualized with rhodamine phalloidin and by electron microscopy and tight junctions were visualized with antibodies to the tight-junction protein ZO-1. Paracellular permeability was measured by movement of horseradish peroxidase from interstitium into duct or acinar lumens.
In controls, linear actin and ZO-1 staining occurred along the apical membrane of intralobular duct cells and extended to the apical pole of acinar cells. Hyperstimulation caused progressive disruption of the linear staining of f-actin and ZO-1. Actin disruption in duct cells was confirmed by electron microscopy. Horseradish peroxidase entered intralobular ducts and acinar lumens of hyperstimulated animals more frequently than those of controls.
The structure and function of the paracellular barrier of acinar and intralobular pancreatic duct cells are disrupted early during cerulein pancreatitis and may contribute to early clinical features.
Gastroenterology 07/1995; 108(6):1863-72. · 11.68 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The value of early endoscopic or surgical interventions to remove bile duct stones and decompress the biliopancreatic ductal system in gallstone pancreatitis is controversial.
To evaluate this issue, acute hemorrhagic necrotizing pancreatitis was induced in opossums by obstructing the biliopancreatic ductal system with a balloon catheter for 1, 3, or 5 days.
A progressive increase in the severity of pancreatitis, as manifested by inflammation, fat necrosis, hemorrhage, acinar cell vacuolization, in vitro lactate dehydrogenase release, and acinar cell necrosis, was noted in these obstructed animals. In contrast, decompression of the obstructed ductal system by removal of the balloon catheter after 1 or 3 days prevented the increase in severity of these parameters of pancreatic injury.
We concluded that the severity of biliary pancreatitis in this model is dependent upon the duration of ductal obstruction and that decompression of the ductal system can prevent progression of the disease. These observations support the practice of early attempts to remove obstructing stones in clinical gallstone pancreatitis.
Gastroenterology 08/1993; 105(1):157-64. · 11.68 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Pancreatic acinar cells possess both high low affinity receptors for cholecystokinin. The cholecystokinin analog caerulein, which exerts a trophic effect on the rat pancreas, acts as an agonist at both types of receptors. In contrast, the synthetic analog CCK-JMV-180, which also acts as an agonist at high affinity receptors, opposes the action of caerulein on the low affinity receptors. We report that infusion of either caerulein or CCK-JMV-180 into rats increases [3H]-thymidine incorporation into pancreatic DNA and causes the pancreatic weight as well as content of DNA, RNA, and protein to increase. CCK-JMV-180 also stimulates in-vitro incorporation of [3H]-thymidine into DNA of cultured rat acini. The finding that both caerulein and CCK-JMV-180 exert the same trophic effect on pancreatic acinar cells indicates that this effect is mediated via high affinity acinar cell cholecystokinin receptors.
Biochemical and Biophysical Research Communications 07/1993; 193(3):814-20. · 2.48 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: In vivo pancreatic secretion of the lysosomal hydrolase cathepsin B was found to be increased by infusion of the secretagogue caerulein. The basal as well as caerulein-stimulated in vivo rate of cathepsin B was further increased by infusion of either chloroquine or methylamine while neither the basal nor the secretagogue-stimulated rates of amylase secretion were altered by the lysosomotropic agents. These observations indicate that neutralization of the acidic prelysosomal compartment by administration of lysosomotropic agents results in lysosomal enzyme entry, by default, into the regulated secretory pathway. In vitro stimulation of pancreatic acini with caerulein was also found to stimulate cathepsin B secretion. That in vitro rate of cathepsin B secretion stimulated by caerulein was not increased in acini prepared from animals infused with caerulein, chloroquine, or methylamine, but the in vitro rate of cathepsin B secretion stimulated by caerulein was increased in acini prepared from animals infused with caerulein plus either chloroquine or methylamine. Under these conditions, redistribution of cathepsin B from the lysosome-enriched to the zymogen granule-enriched subcellular fraction was noted, and lysosomal enzyme-containing organelles became increasingly fragile. These observations indicate that in vivo secretagogue stimulation increases the degree of diversion of lysosomal hydrolases into the regulated secretory compartment when the prelysosomal compartment has been neutralized with lysosomotropic agents.
The American journal of physiology 04/1992; 262(3 Pt 1):G439-44.
-
[show abstract]
[hide abstract]
ABSTRACT: To investigate the possible secretion of lysosomal enzymes into pancreatic juice during stimulation with a pancreatic secretagogue under both physiological and pathological conditions, we measured the amount of cathepsin B, a lysosomal enzyme, in the pancreatic juice during the infusion of 6 different concentrations of caerulein (0.02, 0.05, 0.2, 0.5, 1.0, and 2.0 micrograms/kg. hr). In one group of rabbits the pancreatic duct was only cannulated (free-flow group); in others the pancreatic duct was obstructed for 7 hours and secretin was infused at 0.2 CU/kg. hr (obstructed group). In addition, we evaluated the effect of the intraduodenal instillation of a liquid meal (2 g/kg) on the secretion of lysosomal enzymes into pancreatic juice. Caerulein stimulated the secretion of cathepsin B into pancreatic juice in a dose-dependent manner, as it did that of amylase, and at higher concentrations of caerulein (1.0 and 2.0 micrograms/kg. hr), both cathepsin B output and amylase output were decreased. There was a significant positive correlation between cathepsin B output and amylase output into pancreatic juice during stimulation with caerulein. Blockage of the pancreatic duct for 7 hours caused a significant rise in serum amylase levels and a redistribution of cathepsin B activity in the pancreatic subcellular fractions, as a result of which an increased amount of cathepsin B was recovered in the pellet obtained by 1000 x g centrifugation for 15 min, which contained many zymogen granules. These changes noted after short-term pancreatic duct obstruction are very similar to those previously noted in the early stage of diet-and caerulein-induced experimental pancreatitis, suggesting the colocalization of lysosomal enzyme and digestive enzymes. In the duct-obstructed animals, the secretion of cathepsin B stimulated by caerulein was significantly greater than in the free-flow group. Furthermore, the intraduodenal instillation of a liquid meal caused the secretion of cathepsin B into the pancreatic juice along with amylase. These results indicate that under physiological conditions, such as food intake, lysosomal enzymes are secreted into the pancreatic juice in response to stimulation by gut hormones in the same manner as classical pancreatic digestive enzymes. Moreover, zymogen colocalized with lysosomal enzymes in duct-obstructed animals is secreted into pancreatic juice in increased amounts together with digestive enzymes; this finding suggests that lysosomal enzymes play important pathophysiological roles in pancreatic juice and that acinar cells are altered to maintain cellular organization by secreting the potentially dangerous lysosomal enzymes. This pancreatic duct-obstructed rabbit model should be useful in clarifying the early events of acute pancreatitis.
Nippon geka hokan. Archiv für japanische Chirurgie 04/1992; 61(2):103-24.
-
[show abstract]
[hide abstract]
ABSTRACT: The complex events by which digestive enzyme zymogens and lysosomal hydrolases are segregated from each other and differentially transported to their respective membrane-bound intracellular organelles in the pancreas have been noted to be disturbed during the early stages of several models of experimental pancreatitis. As a result, lysosomal hydrolases such as cathepsin B are redistributed to the subcellular zymogen granule-rich fraction and lysosomal hydrolases as well as digestive enzyme zymogens are colocalized within large cytoplasmic vacuoles. The current study was designed to create an in vitro system that would reproduce this redistribution phenomenon. Our results indicate that cathepsin B redistribution occurs when rat pancreatic fragments are incubated with a supramaximally stimulating concentration of the cholecystokinin analogue caerulein along with plasma from an animal subjected to in vivo supramaximal caerulein stimulation. Neither the plasma nor a supramaximally stimulating concentration of caerulein, alone, is sufficient to induce in vitro cathepsin B redistribution. The ability of the plasma to induce in vitro cathepsin redistribution is dependent upon its content of a 10,000-30,000-D protein and is lost by exposure to protease inhibitors. In vitro cathepsin B redistribution also occurs when rat pancreatic fragments are incubated with plasma obtained from opossums with hemorrhagic necrotizing pancreatitis caused by bile/pancreatic duct ligation.
Journal of Clinical Investigation 05/1991; 87(4):1280-5. · 15.39 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Lysosomal hydrolases such as cathepsin B are apically secreted from rabbit pancreatic acinar cells via a regulated as opposed to a constitutive pathway. Intravenous infusion of the cholecystokinin analogue caerulein results in highly correlated apical secretion of digestive and lysosomal enzymes, suggesting that they are discharged from the same presecretory compartment (zymogen granules). Lysosomal enzymes appear to enter that compartment as a result of missorting. After 7 h of duct obstruction is relieved, caerulein-stimulated apical secretion of cathepsin B and amylase is increased, but the ratio of cathepsin B to amylase secretion is not different than that following caerulein stimulation of animals never obstructed. These findings indicate that duct obstruction causes an increased amount of both lysosomal and digestive enzymes to accumulate within the secretagogue releasable compartment but that duct obstruction does not increase the degree of lysosomal enzyme missorting into that compartment. Pancreatic duct obstruction causes lysosomal hydrolases to become colocalized with digestive enzymes in organelles that, in size and distribution, resemble zymogen granules but that are not subject to secretion in response to secretagogue stimulation. These organelles may be of importance in the development of pancreatitis.
Journal of Clinical Investigation 04/1991; 87(3):865-9. · 15.39 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The short-term effects of rat pancreatic duct obstruction were evaluated and compared with those recently reported to follow obstruction of the rabbit pancreatic duct. In both species pancreatic edema and hyperamylasemia are noted, and the lysosomal hydrolase cathepsin B is redistributed from the lysosome-enriched to the zymogen granule-enriched subcellular fraction. Theoretically, this redistribution phenomenon might lead to digestive enzyme activation because cathepsin B is known to be capable of activating trypsinogen. The hyperamylasemia and pancreatic edema (but not the cathepsin B redistribution) that follow rat pancreatic duct obstruction were increased by infusion of a submaximally stimulating dose of the cholecystokinin analogue cerulein. Administration of the cholecystokinin-receptor antagonist L-364,718 reduced the hyperamylasemia but did not alter the pancreatic edema or cathepsin B redistribution. These observations indicate that cholecystokinin may modulate some but not all of the effects of duct obstruction. Secretin administration increased the degree of pancreatic edema and had no demonstrable protective effect. The rat duct-obstruction model described in this report may prove particularly useful in future studies designed to clarify the early events underlying the development of acute pancreatitis.
Gastroenterology 02/1991; 100(1):196-202. · 11.68 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The nonhydrolyzable guanyl nucleotide GTP gamma S stimulated phosphoinositidase C activity in two preparations obtained from mouse pancreatic acini labeled with myo[2-3H]inositol: a cell-free membrane fraction and intact electropermeabilized acini. This action was dose-dependent, was shared by other nonhydrolyzable guanyl nucleotides such as GMP-phencyclidine hydrochloride and GMP-PMP, as well as by fluoride, and was calcium-independent. Contrarily, no effect was observed even at doses of GTP gamma S as high as 10 microM when the same protocol was repeated on identical acinar preparations from mice fed a choline-deficient, ethionine-supplemented diet. This regimen is known to uncouple secretagogue-receptor occupancy from inositol 1,4,5-trisphosphate generation in pancreatic acinar cells and lead to necrotizing hemorrhagic pancreatitis. These data lead us to conclude that the ethionine-induced inactivation of guanyl nucleotide-dependent pancreatic phosphoinositidase C in pancreatic acinar cells is not the result of either a decrease in GTP level or a decrease in GTP availability. These findings further confirm previous work from this laboratory, which has shown that the biochemical lesion induced by this diet occurs after the agonist-receptor binding step. The diet-induced lesion could be either at the level of the G-protein that couples the enzyme with the receptor or at the level of the phospholipase itself.
Journal of Pharmacology and Experimental Therapeutics 06/1990; 253(2):847-50. · 3.83 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The effects of hemorrhagic shock, aspirin, and ethanol on the biochemical and morphologic changes of experimental pancreatitis were evaluated. Pancreatitis was induced by infusing rats with a supramaximally stimulating dose (5 micrograms/kg/h) of caerulein. Hemorrhagic shock was established by removing sufficient blood to reduce mean arterial pressure by 30%, where it was maintained for 30 min. Aspirin (25 mg/kg) and ethanol (2 g/kg) were administered through an orogastric tube at 8-h intervals for 48 h. Hemorrhagic shock did not alter the degree of hyperamylasemia, pancreatic edema, cathepsin B subcellular redistribution, or in vitro LDH leakage that characterize this model of pancreatitis. Hemorrhagic shock did, however, worsen the morphologic evidence of pancreatic injury. Administration of aspirin with ethanol did not alter the degree of hyperamylasemia, pancreatic edema, or subcellular cathepsin B redistribution. Aspirin-ethanol pretreatment also did not alter the morphologic severity of pancreatitis. These observations indicate that hemorrhagic shock worsens the microscopic evidence of pancreatitis induced by supramaximal secretagogue stimulation. In contrast, aspirin-ethanol pretreatment, which might have been expected to increase pancreatic ductal permeability, did not alter the severity of this model of experimental pancreatitis.
International journal of pancreatology: official journal of the International Association of Pancreatology 05/1990; 6(3):207-17.
-
[show abstract]
[hide abstract]
ABSTRACT: The pancreatic duct of anesthetized rabbits was cannulated and, in some animals, flow of pancreatic exocrine secretions was blocked by raising the cannula to a vertical position. Blockage for 3-7 h caused a rapid and significant rise in serum amylase activity and an increase in amylase activity within the pancreas. The concentration of lysosomal enzymes in the pancreas was not altered but they became redistributed among subcellular fractions and, as a result, an increased amount was recovered in the 1,000-g, 15-min pellet, which was enriched in zymogen granules. Immunofluorescence studies indicated that lysosomal enzymes become localized within organelles which, in size and distribution, resemble zymogen granules. They also contain digestive enzyme zymogens. Blockage of pancreatic secretions also caused lysosomal enzyme-containing organelles to become more fragile and subject to in vitro rupture. These changes noted after short-term pancreatic duct obstruction are remarkably similar to those previously noted to occur during the early stages of diet and secretagogue-induced experimental pancreatitis, observations that have suggested that colocalization of digestive enzyme zymogens and lysosomal hydrolases might result in intracellular digestive enzyme activation and be an important early event in the evolution of those forms of experimental acute pancreatitis.
Journal of Clinical Investigation 11/1989; 84(4):1260-6. · 15.39 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The effects of a potent cholecystokinin (CCK) receptor antagonist, L-364,718, on two forms of experimental acute pancreatitis in mice were evaluated. The antagonist prevented the hyperamylasemia, pancreatic edema, and acinar cell vacuolization that followed administration of a supramaximally stimulating dose of the cholecystokinin analogue cerulein. In contrast, the same dose of L-364,718 (1 mg/kg/6 h) and an even higher dose (10 mg/kg/6 h) failed to prevent the hyperamylasemia, acinar cell necrosis, and mortality that followed administration of a choline-deficient ethionine-supplemented diet. These observations are at variance with those previously reported to follow administration of the relatively weak cholecystokinin antagonist proglumide (Niederau C et al. J Clin Invest 1986;78:1056-63). The observations reported in this communication suggest that cholecystokinin does not play an important role in diet-induced pancreatitis and that CCK receptor antagonists are unlikely to be of benefit in the treatment of clinical acute pancreatitis.
Pancreas 02/1989; 4(6):739-43. · 2.39 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: A method of measuring amylase secretion from dispersed pancreatic acini is described that utilizes a 20-micron nylon mesh to separate cell-associated from secreted amylase. When compared with the standard centrifugation assay, the mesh technique permits easy measurement at multiple, closely spaced time points while reducing the number of samples generated and allowing rapid manipulation of the extracellular suspending medium.
Pancreas 02/1988; 3(5):508-11. · 2.39 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Rats infused with a dose of the secretagogue caerulein that is in excess of that which stimulates a maximal rate of pancreatic digestive enzyme secretion develop acute edematous pancreatitis. We have previously noted that infusion of this dose of caerulein (5 micrograms . kg-1 . h-1) induces the appearance of large heterogeneous vacuoles in acinar cell, blockade of exocytosis, and intracellular accumulation of digestive zymogens [O. Watanabe et al. Am. J. Physiol. 246 (Gastrointest. Liver Physiol. 9): G457-G467, 1984 and A. Saluja et al. Am. J. Physiol. 249 (Gastrointest. Liver Physiol. 12): G702-G710, 1985]. The current studies were performed to further elucidate these phenomena at the electron microscopic level of resolution and employed the techniques of pulse labeling, radioautography, and immunolocalization. Rats were infused with caerulein (5 micrograms . kg-1 . h-1) for 1 h, given a pulse of [3H]phenylalanine, and killed at selected times during the subsequent 5- to 180-min postpulse period during which caerulein infusion was continued. Transport from the endoplasmic reticulum to the Golgi cisternae was not altered by supramaximal stimulation, but transport through post-Golgi elements was altered. In particular, the maturation of condensing vacuoles into zymogen granules was found to be impaired. This led to the accumulation of partially condensed vacuoles and to the development of the large vacuoles containing newly synthesized digestive zymogens as well as the lysosomal hydrolase cathepsin D. The source of the latter could be impaired sorting of lysosomal and digestive enzymes and/or fusion of vacuoles with lysosomes. At the later times after pulse labeling, mature zymogen granules were also found to fuse with these large cathepsin D-containing vacuoles by a process analogous to crinophagy. Thus these studies indicate that the large heterogeneous vacuoles that appear during supramaximal secretagogue stimulation and that contain admixed digestive zymogens and lysosomal hydrolases arise by at least two mechanisms, impaired condensing vacuole maturation and crinophagy.
The American journal of physiology 11/1987; 253(4 Pt 1):G517-26.
