Biomedical subjects

Stephen J Pandol

Publications and source records attributed to Stephen J Pandol.

34 records · Page 2Linked to original sources

PubMed2004

Neurohumoral control of exocrine pancreatic secretion.

PURPOSE OF REVIEW: Advancing knowledge about normal physiology of the exocrine pancreas is essential for investigations into the mechanisms of disorders of the pancreas. To this end, reports published during the past year give further insights into the complexity of the hormonal and neural interactions and processes involved in exocrine pancreatic physiology. RECENT FINDINGS: Key findings include demonstrations of different biologic effects of cholecystokinin peptides on pancreatic secretion, underlining the importance of measurement techniques to determine specific forms of peptide hormones involved in physiologic processes; investigations of the details of the central neural pathways involved in the vago-vagal reflex in pancreatic secretion; the demonstration of the essential role for intrapancreatic nerves in mediating meal-induced responses; and identification of a modulatory role for leptin in exocrine pancreatic secretion. SUMMARY: These findings should not only spur further investigations into mechanisms of normal physiology but also provide impetus for application of these findings to studies of pancreatic disorders.

Journal Article↗

PubMed2003

Ethanol differentially regulates NF-kappaB activation in pancreatic acinar cells through calcium and protein kinase C pathways.

Mechanisms of alcoholic pancreatitis remain unknown. Previously, we showed that ethanol feeding sensitizes rats to pancreatitis caused by CCK-8, at least in part, by augmenting activation of the proinflammatory transcription factor NF-kappaB. To elucidate the mechanism of sensitization, here we investigate the effect of ethanol on Ca(2+)- and PKC-mediated pathways of CCK-induced NF-kappaB activation using an in vitro system of rat pancreatic acini incubated with ethanol. Ethanol augmented CCK-8-induced activation of NF-kappaB, similar to our in vivo findings with ethanol-fed rats. In contrast, ethanol prevented NF-kappaB activation caused by thapsigargin, an agent that mobilizes intracellular Ca(2+) bypassing the receptor. Pharmacological analysis showed that NF-kappaB activation by thapsigargin but not by CCK-8 is mediated through the calcineurin pathway and that the inhibitory effect of ethanol on the thapsigargin-induced NF-kappaB activation could be through inhibiting this pathway. Ethanol augmented NF-kappaB activation induced by the phorbol ester PMA, a direct activator of PKC. Inhibitory analysis demonstrated that Ca(2+)-independent (novel and/or atypical) PKC isoforms are involved in NF-kappaB activation induced by both CCK-8 and PMA in cells treated and not treated with ethanol. The results indicate that ethanol differentially affects the Ca(2+)/calcineurin- and PKC-mediated pathways of NF-kappaB activation in pancreatic acinar cells. These effects may play a role in the ability of ethanol to sensitize pancreas to the inflammatory response and pancreatitis.

Animals↗

PubMed2003

Emerging concepts for the mechanism of alcoholic pancreatitis from experimental models.

The pathophysiologic mechanisms that underlie acute and chronic pancreatitis arising from alcohol abuse are poorly understood. The reasons for this state of knowledge result historically from a lack of models for experimental investigation. Ethanol feeding alone, even at high doses, has minimal and inconsistent effects on morphologic findings in the pancreas in experimental animals. This experience, plus the fact that alcohol abuse causes pancreatic pathology in only a minority of patients, suggest that ethanol acts to sensitize the pancreas to the deleterious effects of other stimuli. In this article, we discuss findings to support this concept of ethanol as a sensitizing agent and experimental models developed that can be used to investigate the effects of ethanol on the pathologic processes of pancreatitis. These pathologic processes include inflammation, cell death, intrapancreatic digestive enzyme activation, and fibrosis.

Animals↗

PubMed2003

Extracellular matrix proteins protect pancreatic cancer cells from death via mitochondrial and nonmitochondrial pathways.

BACKGROUND AND AIMS: Pancreatic cancer is a very aggressive malignancy. Normal cells die through apoptosis when detached from extracellular matrix (ECM), but the role of ECM in cancer cell survival is poorly understood. Here, we determined the effects of ECM proteins on death responses and underlying signaling pathways in human pancreatic cancer cells. METHODS: We measured apoptosis and necrosis, caspase activation, and mitochondrial dysfunction in MIA PaCa-2 and PANC-1 pancreatic carcinoma cells both detached and attached to ECM proteins. RESULTS: Detachment of pancreatic cancer cells from ECM did not induce classic apoptosis, as it does in normal cells, but induced necrosis and apoptosis associated with secondary necrosis. It caused a pronounced mitochondrial depolarization and release of cytochrome c and Smac/DIABLO. However, as different from normal cells, cytochrome c release did not result in downstream caspase activation. Executioner caspases were activated in detached pancreatic cancer cells independent of cytochrome c. Laminin and fibronectin, but not collagen I, markedly increased pancreatic cancer cell survival by inhibiting both mitochondrial dysfunction (leading to inhibition of necrosis) and caspase activity (leading to decreased apoptotic DNA fragmentation). CONCLUSIONS: ECM proteins greatly protect pancreatic cancer cells from death by mechanisms different from those operating in normal cells. The results suggest ECM proteins and their receptors as potential targets for treatment of pancreatic cancer.

Adenocarcinoma↗

PubMed2003

Nonoxidative ethanol metabolites alter extracellular matrix protein content in rat pancreas.

BACKGROUND & AIMS: The mechanisms involved in ethanol-induced pancreas fibrosis are poorly understood. Here we show that fatty acid ethyl esters (FAEEs), nonoxidative ethanol metabolites, increase extracellular matrix (ECM) protein levels in pancreas. METHODS: Rat pancreatic acini were incubated for 1-4 hours with FAEEs or acetaldehyde. In another set of experiments, rats received an intravenous infusion of FAEEs for 6 hours. Collagens were assessed by a hydroxyproline assay. Laminin and fibronectin were analyzed by Western blotting. Gene expression of ECM proteins was measured by conventional and real-time reverse-transcription polymerase chain reaction (RT-PCR). Matrix metalloproteinase (MMP), plasmin, and urokinase-type plasminogen activator (uPA) activities were determined by zymography and fluorogenic assays. RESULTS: FAEEs increased collagen, laminin, and fibronectin levels in pancreatic acini without affecting messenger RNA (mRNA) expression for these proteins. Actinomycin D, a transcriptional inhibitor, did not block the increase in ECM proteins induced by FAEEs. FAEEs reduced the activity of the serine protease, plasmin, and that of the uPA. Consistent with these results, the serine protease inhibitor aprotinin reproduced the effects of FAEEs and prevented the further increase in ECM proteins induced by FAEEs. In vivo administration of FAEEs reduced plasmin and uPA activities and increased ECM protein levels in pancreas. Acetaldehyde had minor effects on ECM protein levels and did not affect plasmin activity. CONCLUSIONS: FAEEs increase ECM protein levels in pancreas. The results suggest that this effect is caused primarily by an inhibition in ECM degradation via serine proteases including the plasminogen system.

Acetaldehyde↗

PubMed2003

Neurohumoral control of exocrine pancreatic secretion.

Reports over the past year provide significant advances in our knowledge of the neurohumoral control of exocrine pancreatic secretion, especially related to human physiology. Major findings include those demonstrating that human pancreatic acinar cells do not respond to cholecystokinin stimulation and that a major circulating form of cholecystokinin is CCK-58. These findings establish that in humans, cholecystokinin causes pancreatic secretion via a neural circuit after interacting with neural sensory receptors in the mucosa of the intestine and that CCK-58 is the likely form of cholecystokinin that stimulates the neural pathways. Other findings demonstrate significant differences in the pancreatic secretory response in humans as a function of the type of nutrient delivered to the gut, especially the fact the elemental diets and medium-chain triglycerides cause much less stimulation of pancreatic secretion than do complex diets. Finally, convincing evidence demonstrating that pancreatic proteases cause inhibition of pancreatic secretion in humans has been presented. In addition to new insights into the neurohumoral control of pancreatic secretion, these findings provide information relevant to both the pathogenesis of pancreatic disorders and their treatment.

Journal Article↗

PubMed2003

Prevention of metastatic pancreatic cancer growth in vivo by induction of apoptosis with genistein, a naturally occurring isoflavonoid.

INTRODUCTION: The critical need for novel therapeutic approaches to pancreatic cancer treatment is clear. Genistein, a naturally occurring isoflavonoid, is active against certain solid malignancies, but its effect on pancreatic cancer is unknown. AIMS: To investigate the bioactivity of genistein in experimental pancreatic cancer in vitro and in vivo. METHODOLOGY: The effect of intraperitoneal genistein administration on local tumor growth and metastatic disease was determined in an orthotopic nude mouse model. Apoptosis in tumor specimens was determined by the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) technique. In vitro, the effect of genistein on cell growth was assessed by cell count and MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) colorimetric assay. Apoptosis was determined in vitro by DNA laddering and annexin-V. Caspase-3 and nuclear factor-kappaB activity were measured following genistein treatment. RESULTS: In vivo, genistein significantly improved survival, almost completely inhibited metastasis, and increased apoptosis in an orthotopic model of pancreatic cancer. In vitro genistein treatment resulted in apoptosis in all pancreatic cancer cell lines tested, and this appeared to be mediated by activation of caspase-3. CONCLUSION: These findings suggest that the antimetastatic effect of genistein treatment in vivo is mediated by induction of apoptosis. Genistein may have a therapeutic benefit for patients with pancreatic cancer, in particular after surgery, to prevent recurrence of metastatic disease.

Animals↗

PubMed2003

Animal and in vitro models of alcoholic pancreatitis: role of cholecystokinin.

Although ethanol abuse is the major etiologic factor in the development of acute and chronic pancreatitis, the mechanisms of ethanol effects to cause pancreatitis are poorly understood. The major reason for the lack of progress is the relative lack of animal models that reproduce the deleterious effects of ethanol on the pancreas that are observed in human disease. We propose that the effect of ethanol on the pancreas is due to its ability to sensitize animals and humans to the potentially injurious effects of other stimuli. We have developed models of ethanol-induced acute and chronic pancreatitis in rats as well as pancreatic acinar cells in primary culture demonstrating that ethanol sensitizes the pancreas to the inflammatory, cell death, and fibrosing responses caused by cholecystokinin (CCK). Our results indicate that the ethanol-sensitized inflammatory response is the key or trigger event for the development of the other pathologic responses in both acute and chronic pancreatitis, such as cell death, intracellular digestive enzyme activation, and fibrosis. These findings suggest that experimental strategies designed to reveal the modulating effects of ethanol on the mechanisms underlying the inflammatory, cell death, and fibrosing responses stimulated by CCK will provide the key information needed to understand how ethanol abuse causes pancreatitis.

Animals↗

PubMed2003

Curcumin ameliorates ethanol and nonethanol experimental pancreatitis.

Treatments for pancreatitis are limited. Activation of transcription factor NF-kappaB, a key regulator of inflammatory molecule expression, is an early event in experimental pancreatitis and correlates with the inflammatory response. We report here that curcumin, a natural phytochemical known to inhibit NF-kappaB and activator protein (AP)-1, another important proinflammatory transcription factor, ameliorates pancreatitis in two rat models. In both cerulein pancreatitis and pancreatitis induced by a combination of ethanol diet and low-dose CCK, curcumin improved the severity of the disease as measured by a number of parameters (histology, serum amylase, pancreatic trypsin, and neutrophil infiltration). Curcumin markedly inhibited NF-kappaB and AP-1 activation, assessed by DNA binding and degradation of inhibitory IkappaB proteins, and the induction of mRNAs for cytokines IL-6 and TNF-alpha, the chemokine KC, and inducible nitric oxide synthase in pancreas. Curcumin also blocked CCK-induced NF-kappaB and AP-1 activation in isolated pancreatic acini. Our findings indicate that blocking key signals of the inflammatory response ameliorates pancreatitis in both ethanol and nonethanol models. They suggest that curcumin, which is currently in clinical trials for cancer prevention, may be useful for treatment of pancreatitis.

Animals↗

PubMed2002

Role of S-adenosylmethionine in two experimental models of pancreatitis.

Severe necrotizing pancreatitis occurs in young female mice fed a choline-deficient and ethionine-supplemented (CDE) diet. Although the mechanism of the pancreatitis is unknown, one consequence of this diet is depletion of hepatic S-adenosylmethionine (SAM). SAM formation is catalyzed by methionine adenosyltransferases (MATs), which are encoded by liver-specific (MAT1A) and non-liver-specific (MAT2A) genes. In this work, we examined changes in pancreatic SAM homeostasis in mice receiving the CDE diet and the effect of SAM treatment. We found that both MAT forms are expressed in normal pancreas and pancreatic acini. After 48 h of the CDE diet, SAM levels decreased 50% and MAT1A-encoded protein disappeared via post-translational mechanisms, whereas MAT2A-encoded protein increased via pretranslational mechanisms. CDE-fed mice exhibited extensive necrosis, edema, and acute pancreatic inflammatory infiltration, which were prevented by SAM treatment. However, old female mice consuming the CDE diet that do not develop pancreatitis showed a similar fall in pancreatic SAM level. SAM was also protective in cerulein-induced pancreatitis in the rat, but the protection was limited. Although the pancreatic SAM level fell by more than 80% in the MAT1A knockout mice, no pancreatitis developed. This study thus provides several novel findings. First, the so-called liver-specific MAT1A is highly expressed in the normal pancreas and pancreatic acini. Second, the CDE diet causes dramatic changes in the expression of MAT isozymes by different mechanisms. Third, in contrast to the situation in the liver, where absence of MAT1A and decreased hepatic SAM level can lead to spontaneous tissue injury, in the pancreas the roles of SAM and MAT1A appear more complex and remain to be defined.

Administration, Oral↗

PubMed2002

Expression of cytokine and chemokine mRNA and secretion of tumor necrosis factor-alpha by gallbladder epithelial cells: response to bacterial lipopolysaccharides.

BACKGROUND: In addition to immune cells, many other cell types are known to produce cytokines. Cultured normal mouse gallbladder epithelial cells, used as a model system for gallbladder epithelium, were examined for their ability to express the mRNA of various cytokines and chemokines in response to bacterial lipopolysaccharide. The synthesis and secretion of the tumor necrosis factor-alpha (TNF-alpha) protein by these cells was also measured. RESULTS: Untreated mouse gallbladder cells expressed mRNA for TNF-alpha, RANTES, and macrophage inflammatory protein-2 (MIP-2). Upon treatment with lipopolysaccharide, these cells now produced mRNA for Interleukin-1beta (IL-1beta), IL-6, monocyte chemoattractant protein-1 (MCP-1), and showed increased expression of TNF-alpha and MIP-2 mRNA. Untreated mouse gallbladder cells did not synthesize TNF-alpha protein; however, they did synthesize and secrete TNF-alpha upon treatment with lipopolysaccharide. METHODS: Cells were treated with lipopolysaccharides from 3 strains of bacteria. Qualitative and semi-quantitative RT-PCR, using cytokine or chemokine-specific primers, was used to measure mRNA levels of TNFalpha, IL-1beta, IL-6, IL-10, KC, RANTES, MCP-1, and MIP-2. TNF-alpha protein was measured by immunoassays. CONCLUSION: This research demonstrates that gallbladder epithelial cells in response to lipopolysaccharide exposure can alter their cytokine and chemokine RNA expression pattern and can synthesize and secrete TNFalpha protein. This suggests a mechanism whereby gallbladder epithelial cells in vivo may mediate gallbladder secretory function, inflammation and diseases in an autocrine/paracrine fashion by producing and secreting cytokines and/or chemokines during sepsis.

Animals↗

PubMed2002

Cholecystokinin induces caspase activation and mitochondrial dysfunction in pancreatic acinar cells. Roles in cell injury processes of pancreatitis.

Apoptosis and necrosis are critical parameters of pancreatitis, the mechanisms of which remain unknown. Many characteristics of pancreatitis can be studied in vitro in pancreatic acini treated with high doses of cholecystokinin (CCK). We show here that CCK stimulates apoptosis and death signaling pathways in rat pancreatic acinar cells, including caspase activation, cytochrome c release, and mitochondrial depolarization. The mitochondrial dysfunction is mediated by upstream caspases (possibly caspase-8) and, in turn, leads to activation of caspase-3. CCK causes mitochondrial alterations through both permeability transition pore-dependent (cytochrome c release) and permeability transition pore-independent (mitochondrial depolarization) mechanisms. Caspase activation and mitochondrial alterations also occur in untreated pancreatic acinar cells; however, the underlying mechanisms are different. In particular, caspases protect untreated acinar cells from mitochondrial damage. We found that caspases not only mediate apoptosis but also regulate other parameters of CCK-induced acinar cell injury that are characteristic of pancreatitis; in particular, caspases negatively regulate necrosis and trypsin activation in acinar cells. The results suggest that the observed signaling pathways regulate parenchymal cell injury and death in CCK-induced pancreatitis. Protection against necrosis and trypsin activation by caspases can explain why the severity of pancreatitis in experimental models correlates inversely with the extent of apoptosis.

Amylases↗

PubMed2002

Food-derived polyphenols inhibit pancreatic cancer growth through mitochondrial cytochrome C release and apoptosis.

There is increasing evidence that food-derived polyphenols have a beneficial effect for cancers. Our purpose was to determine the effect and mechanism of action of these compounds on pancreatic cancer. We measured effects of quercetin on pancreatic cancer in a nude mouse model. We also investigated the effects of quercetin, rutin, trans-resveratrol and genistein on apoptosis and underlying signaling in pancreatic carcinoma cells in vitro. Quercetin decreased primary tumor growth, increased apoptosis and prevented metastasis in a model of pancreatic cancer. In vitro quercetin and trans-resveratrol, but not rutin, markedly enhanced apoptosis, causing mitochondrial depolarization and cytochrome c release followed by caspase-3 activation. In addition, the effect of a combination of quercetin and trans-resveratrol on mitochondrial cytochrome c release and caspase-3 activity was greater than the expected additive response. The inhibition of mitochondrial permeability transition prevented cytochrome c release, caspase-3 activation and apoptosis caused by polyphenols. Nuclear factor-kappa B activity was inhibited by quercetin and trans-resveratrol, but not genistein, indicating that this transcription factor is not the only mediator of the polyphenols' effects on apoptosis. The results suggest that food-derived polyphenols inhibit pancreatic cancer growth and prevent metastasis by inducing mitochondrial dysfunction, resulting in cytochrome c release, caspase activation and apoptosis.

Animals↗

PubMed2002

Ethanol metabolism and transcription factor activation in pancreatic acinar cells in rats.

BACKGROUND & AIMS: Ethanol metabolism by pancreatic acinar cells and the role of its metabolites in ethanol toxicity to the pancreas remain largely unknown. Here, we characterize ethanol metabolism in pancreatic acinar cells and determine the effects of ethanol metabolites on nuclear factor kappa B (NF-kappa B) and activator protein (AP)-1, transcription factors that are activated in pancreatitis and mediate expression of inflammatory molecules critical for this disease. METHODS: We measured activities of fatty acid ethyl ester (FAEE) synthase and alcohol dehydrogenase (ADH), as well as accumulation of ethanol metabolites. We measured the effects of ethanol and its metabolites on NF-kappa B and AP-1 activation by using a gel shift assay. RESULTS: Pancreas metabolizes ethanol via both oxidative and nonoxidative pathways. Acinar cells are the main source of ethanol metabolism in the pancreas. Compared with the liver, FAEE synthase activity in the pancreas is greater, whereas that of ADH is much less. FAEEs activated NF-kappa B and AP-1, whereas acetaldehyde inhibited NF-kappa B activation. Ethanol decreased NF-kappa B binding activity in acinar cells, which was potentiated by cyanamide. CONCLUSION: Oxidative and nonoxidative ethanol metabolites regulate transcription factors differently in pancreatic acinar cells. Ethanol may regulate NF-kappa B and AP-1 positively or negatively, depending on which metabolic pathway's effect predominates. These regulatory mechanisms may play a role in ethanol toxicity to the pancreas.

Acetaldehyde↗

PubMed2002

Neutrophils and NADPH oxidase mediate intrapancreatic trypsin activation in murine experimental acute pancreatitis.

BACKGROUND & AIMS: Intrapancreatic activation of digestive enzymes is a key event in the parenchymal cell injury of pancreatitis. We hypothesized that neutrophils recruited to the pancreas during pancreatitis may contribute to such activation. METHODS: To cause experimental pancreatitis, rats and mice were treated with high doses of cerulein. Activation of the digestive enzyme, trypsin, was measured in pancreatic homogenates using a fluorogenic assay and localized immunocytochemically with antibody to trypsin-activation peptide (TAP). RESULTS: Compared with controls, rats depleted of neutrophils with antineutrophil serum exhibited a marked attenuation in intrapancreatic trypsin activation and acinar cell TAP labeling induced by high-dose cerulein. To examine the mechanism, mice deficient in either nicontinamide adenine dinucleotide phosphate (NADPH) oxidase, or myeloperoxidase (MPO) were studied for trypsin activation. Mice deficient in NADPH oxidase exhibited attenuation of the cerulein-induced trypsin activation, but those deficient in MPO did not. Using measurements of Western blot analysis, generation of reactive oxygen species, and immunocytochemistry, we demonstrated the NADPH oxidase activity is in neutrophils and not pancreatic acinar tissue. CONCLUSIONS: The results demonstrate a novel role for neutrophils infiltrating the pancreas in pathologic activation of digestive enzymes in acute pancreatitis and indicate that this effect is mediated by products of NADPH oxidase.

Animals↗

PubMed2002

Caspase 8-mediated cleavage of plectin precedes F-actin breakdown in acinar cells during pancreatitis.

Pancreatic acinar cells depend on the integrity of the cytoskeleton for regulated secretion. Stimulation of isolated rat pancreatic acini with the secretagogue CCK serves as a model for human acute edematous pancreatitis. It induces the breakdown of the actin filament system (F-actin) with the consecutive inhibition of secretion and premature activation of digestive enzymes. However, the mechanisms that regulate F-actin breakdown are largely unknown. Plectin is a versatile cytolinker protein regulating F-actin dynamics in fibroblasts. It was recently demonstrated that plectin is a substrate of caspase 8. In pancreatic acinar cells, plectin strongly colocalizes with apical and basolateral F-actin. Supramaximal secretory stimulation of acini with CCK leads to a rapid redistribution and activation of caspase 8, followed by degradation of plectin that in turn precedes the F-actin breakdown. Inhibition of caspase 8 before CCK hyperstimulation prevents plectin cleavage, stabilizes F-actin morphology, and reverses the inhibition of secretion. Thus we propose that the caspase 8-mediated degradation of plectin represents a critical biochemical event during CCK-induced secretory blockade and cell injury.

Actins↗