RESUMEN
The abundant stromal/desmoplastic reaction, a characteristic feature of a majority of pancreatic adenocarcinomas (PDAC), has only recently been receiving some attention regarding its possible role in the pathobiology of pancreatic cancer. It is now well established that the cells predominantly responsible for producing the collagenous stroma are pancreatic stellate cells (PSCs). In addition to extracellular matrix proteins, the stroma also exhibits cellular elements including, immune cells, endothelial cells and neural cells. Evidence is accumulating to indicate the presence of significant interactions between PSCs and cancer cells as well as between PSCs and other cell types in the stroma. The majority of research reports to date, using in vitro and in vivo approaches, suggest that these interactions facilitate local growth as well as distant metastasis of pancreatic cancer, although a recent study using animals depleted of myofibroblasts has raised some questions regarding the central role of myofibroblasts in cancer progression. Nonetheless, novel therapeutic strategies have been assessed, mainly in the pre-clinical setting, in a bid to interrupt stromal-tumour interactions and inhibit disease progression. The next important challenge is for the translation of such pre-clinical strategies to the clinical situation so as to improve the outcome of patients with pancreatic cancer.
Asunto(s)
Neoplasias Pancreáticas/patología , Microambiente Tumoral , Adenocarcinoma/patología , Animales , HumanosRESUMEN
Chronic pancreatitis is a necroinflammatory process characterized pathologically by acinar atrophy and fibrosis and clinically by abdominal pain, diabetes and maldigestion. In this review we summarize some of the recent advances in the understanding of the pathogenesis of pancreatitis and how they have shaped our current understanding of chronic pancreatitis. We pay particular attention to advances in the genetic basis of idiopathic, hereditary and tropical pancreatitis as well as research into the relationship between alcohol and the pancreas. We have also reviewed current practices with respect to diagnosis and management of chronic pancreatitis.
Asunto(s)
Pancreatitis Crónica/diagnóstico , Pancreatitis Crónica/metabolismo , Alcoholismo/complicaciones , Alcoholismo/metabolismo , Antioxidantes/uso terapéutico , Humanos , Páncreas/metabolismo , Páncreas/patología , Pancreatitis Crónica/etiología , Pancreatitis Crónica/terapiaRESUMEN
The infiltration of inflammatory cells into the pancreas is an early and central event in acute pancreatitis that promotes local injury and systemic complications of the disease. Recent research has yielded the important finding that resident cells of the pancreas (particularly acinar and pancreatic stellate cells) play a dynamic role in leukocyte attraction via secretion of chemokines and cytokines and expression of adhesion molecules. Significant progress has been made in recent years in our understanding of the role of leukocyte movement (adhesion to the blood vessel wall, transmigration through the blood vessel wall and infiltration into the parenchyma) in the pathophysiology of acute pancreatitis. This review discusses recent studies and describes the current state of knowledge in the field. It is clear that detailed elucidation of the numerous processes in the inflammatory cascade is an essential step towards the development of improved therapeutic strategies in acute pancreatitis. Studies to date suggest that combination therapy targeting different steps of the inflammatory cascade may be the treatment of choice for this disease.
Asunto(s)
Páncreas Exocrino/fisiología , Pancreatitis/inmunología , Enfermedad Aguda , Animales , Adhesión Celular/fisiología , Moléculas de Adhesión Celular/inmunología , Movimiento Celular/fisiología , Quimiocinas/inmunología , Quimiotaxis de Leucocito , Matriz Extracelular/fisiología , Humanos , Leucocitos/inmunología , Páncreas Exocrino/inmunología , Páncreas Exocrino/patología , Pancreatitis/patologíaRESUMEN
BACKGROUND AND AIMS: Activated pancreatic stellate cells (PSCs) are implicated in the production of alcohol induced pancreatic fibrosis. PSC activation is invariably associated with loss of cytoplasmic vitamin A (retinol) stores. Furthermore, retinol and ethanol are known to be metabolised by similar pathways. Our group and others have demonstrated that ethanol induced PSC activation is mediated by the mitogen activated protein kinase (MAPK) pathway but the specific role of retinol and its metabolites all-trans retinoic acid (ATRA) and 9-cis retinoic acid (9-RA) in PSC quiescence/activation, or its influence on ethanol induced PSC activation is not known. Therefore, the aims of this study were to (i) examine the effects of retinol, ATRA, and 9-RA on PSC activation; (ii) determine whether retinol, ATRA, and 9-RA influence MAPK signalling in PSCs; and (iii) assess the effect of retinol supplementation on PSCs activated by ethanol. METHODS: Cultured rat PSCs were incubated with retinol, ATRA, or 9-RA for varying time periods and assessed for: (i) proliferation; (ii) expression of alpha smooth muscle actin (alpha-SMA), collagen I, fibronectin, and laminin; and (iii) activation of MAPKs (extracellular regulated kinases 1 and 2, p38 kinase, and c-Jun N terminal kinase). The effect of retinol on PSCs treated with ethanol was also examined by incubating cells with ethanol in the presence or absence of retinol for five days, followed by assessment of alpha-SMA, collagen I, fibronectin, and laminin expression. RESULTS: Retinol, ATRA, and 9-RA significantly inhibited: (i) cell proliferation, (ii) expression of alpha-SMA, collagen I, fibronectin, and laminin, and (iii) activation of all three classes of MAPKs. Furthermore, retinol prevented ethanol induced PSC activation, as indicated by inhibition of the ethanol induced increase in alpha-SMA, collagen I, fibronectin, and laminin expression. CONCLUSIONS: Retinol and its metabolites ATRA and 9-RA induce quiescence in culture activated PSCs associated with a significant decrease in the activation of all three classes of MAPKs in PSCs. Ethanol induced PSC activation is prevented by retinol supplementation.
Asunto(s)
Páncreas/efectos de los fármacos , Vitamina A/farmacología , Alitretinoína , Animales , Proteínas de Ciclo Celular/metabolismo , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Fosfatasa 1 de Especificidad Dual , Activación Enzimática/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Etanol/antagonistas & inhibidores , Etanol/farmacología , Proteínas de la Matriz Extracelular/metabolismo , Fibrosis , Proteínas Inmediatas-Precoces/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Páncreas/citología , Páncreas/metabolismo , Páncreas/patología , Fosfoproteínas Fosfatasas/metabolismo , Proteína Fosfatasa 1 , Proteínas Tirosina Fosfatasas/metabolismo , Ratas , Receptores de Ácido Retinoico/metabolismo , Receptores X Retinoide/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Tretinoina/farmacología , Vanadatos/farmacologíaRESUMEN
Alcoholic pancreatitis is a major complication of alcohol abuse. Since only a minority of alcoholics develop pancreatitis, there has been a keen interest in identifying the factors that may confer individual susceptibility to the disease. Numerous possibilities have been evaluated including diet, drinking patterns and a range of inherited factors. However, at the present time, no susceptibility factor has been unequivocally identified. In contrast, considerable progress has been made with respect to the constant effects of alcohol on the pancreas. The molecular mechanisms of alcohol-induced pancreatic injury are being increasingly defined with an emphasis, in recent years, on the acinar cell itself as the principal site on ethanol-related damage. It has now been established that the acinar cell is capable of metabolizing alcohol and that the direct toxic effects of alcohol and/or its metabolites on acinar cells may predispose the gland to autodigestive injury in the presence of an appropriate triggering factor. A significant recent development relates to the characterization of pancreatic stellate cells, increasingly implicated in alcoholic pancreatic fibrosis. Here the current concepts regarding the mechanisms/pathways mediating alcohol-induced pancreatic injury are outlined.
Asunto(s)
Alcoholismo/complicaciones , Citocromo P-450 CYP2E1/genética , Estrés Oxidativo/fisiología , Pancreatitis Alcohólica/genética , Pancreatitis Crónica/genética , Animales , Biopsia con Aguja , Citocromo P-450 CYP2E1/metabolismo , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Relación Dosis-Respuesta a Droga , Activación Enzimática , Etanol/efectos adversos , Femenino , Regulación de la Expresión Génica , Humanos , Inmunohistoquímica , Masculino , Biología Molecular , Pancreatitis Alcohólica/enzimología , Pancreatitis Alcohólica/patología , Pancreatitis Crónica/enzimología , Pancreatitis Crónica/patología , Pronóstico , Ratas , Índice de Severidad de la EnfermedadRESUMEN
OBJECTIVES: Pancreatic cancer has a very poor prognosis, largely due to its propensity for early local and distant spread. Histopathologically, most pancreatic cancers are characterized by a prominent stromal/fibrous reaction in and around tumor tissue. The aims of this study were to determine whether (1) the cells responsible for the formation of the stromal reaction in human pancreatic cancers are activated pancreatic stellate cells (PSCs) and (2) an interaction exists between pancreatic cancer cells and PSCs that may facilitate local and distant invasion of tumor. METHODS: Serial sections of human pancreatic cancer tissue were stained for desmin and glial fibrillary acidic protein (stellate cell selective markers) and alpha-smooth muscle actin (alphaSMA), a marker of activated PSC activation, by immunohistochemistry, and for collagen using Sirius Red. Correlation between the extent of positive staining for collagen and alphaSMA was assessed by morphometry. The cellular source of collagen in stromal areas was identified using dual staining methodology, ie, immunostaining for alphaSMA and in situ hybridization for procollagen alpha1I mRNA. The possible interaction between pancreatic cancer cells and PSCs was assessed in vitro by exposing cultured rat PSCs to control medium or conditioned medium from 2 pancreatic cancer cell lines (PANC-1 and MiaPaCa-2) for 24 hours. PSC activation was assessed by cell proliferation and alphaSMA expression. RESULTS: Stromal areas of human pancreatic cancer stained strongly positive for the stellate cell selective markers desmin and GFAP (indicating the presence of PSCs), for alphaSMA (suggesting that the PSCs were in their activated state) and for collagen. Morphometric analysis demonstrated a close correlation (r = 0.77; P < 0.04; 8 paired sections) between the extent of PSC activation and collagen deposition. Procollagen mRNA expression was localized to alphaSMA-positive cells in stromal areas indicating that activated PSCs were the predominant source of collagen in stromal areas. Exposure of PSCs to pancreatic cancer cell secretions in vitro resulted in PSC activation as indicated by significantly increased cell proliferation and alphaSMA expression. CONCLUSIONS: Activated PSCs are present in the stromal reaction in pancreatic cancers and are responsible for the production of stromal collagen. PSC function is influenced by pancreatic cancer cells. Interactions between tumor cells and stromal cells (PSCs) may play an important role in the pathobiology of pancreatic cancer.
Asunto(s)
Neoplasias Pancreáticas/patología , Células del Estroma/patología , Actinas/análisis , Actinas/biosíntesis , Animales , Biomarcadores de Tumor/análisis , División Celular/efectos de los fármacos , Línea Celular Tumoral/metabolismo , Células Cultivadas/efectos de los fármacos , Colágeno/análisis , Medios de Cultivo Condicionados/farmacología , Desmina/análisis , Proteína Ácida Fibrilar de la Glía/análisis , Humanos , Invasividad Neoplásica , Proteínas de Neoplasias/análisis , Páncreas/citología , Neoplasias Pancreáticas/química , ARN Mensajero/análisis , Ratas , Células del Estroma/químicaRESUMEN
Pancreatic fibrosis, a characteristic histopathological feature of chronic pancreatitis, is no longer considered an epiphenomenon of chronic injury, but an active process that may be reversible in the early stages. The identification and characterization of pancreatic stellate cells (PSCs) in recent years has had a significant impact on research into pancreatic fibrogenesis. Accumulating evidence from both in vivo studies (using human pancreatic sections and experimental models of pancreatic fibrosis) and in vitro studies (using cultured pancreatic stellate cells) indicates a key role for activated PSCs in the fibrotic process. These cells are now known to be activated by ethanol and its metabolites and by several factors that are upregulated during pancreatic injury including growth factors, cytokines and oxidant stress. Based on this knowledge, potential antifibrotic strategies such as antioxidants and cytokine inhibition have been assessed in experimental models. Studies are also underway to characterise the signaling pathways/molecules responsible for mediating PSC activation, in order to identify potential therapeutic targets for the inhibition of PSC activation, thereby preventing or reversing the development of pancreatic fibrosis.
Asunto(s)
Páncreas/patología , Enfermedades Pancreáticas/patología , Animales , Células Cultivadas , Citocinas/metabolismo , Fibrosis/metabolismo , Fibrosis/patología , Sustancias de Crecimiento/metabolismo , Humanos , Estrés Oxidativo/fisiología , Páncreas/metabolismo , Enfermedades Pancreáticas/metabolismoRESUMEN
Metabolism of ethanol by acinar and other pancreatic cells and the consequent generation of toxic metabolites are postulated to play an important role in the development of alcohol-related acute and chronic pancreatic injury. Studies using cultured pancreatic acinar cells and isolated pancreatic acini have established that (i) the pancreas can metabolize ethanol via the oxidative pathway involving the enzymes alcohol dehydrogenase (ADH) and possibly cytochrome P4502E1 (although the role of the latter remains to be fully delineated) as well as the nonoxidative pathway [involving fatty acid ethyl ester (FAEE) synthases] and (ii) the oxidative pathway (which generates acetaldehyde) is quantitatively greater than the nonoxidative pathway, which yields FAEEs. Most recently, pancreatic stellate cells (PSCs) (implicated in pancreatic fibrogenesis) have been reported to exhibit ADH activity, suggesting that the capacity of the pancreas to metabolize ethanol may reside not only in parenchymal (acinar) cells but also in nonparenchymal cells. Polymorphisms/mutations of ethanol metabolizing enzymes have been examined to determine whether they may confer individual susceptibility to alcoholic pancreatitis. However, no association has been demonstrated between ADH and CYP2E1 polymorphisms and the predisposition to alcoholic pancreatitis. Other candidate factors that remain to be studied include polymorphisms of FAEE synthetic enzymes and proteins relevant to antioxidant pathways in the cell. Injury to the pancreas due to its capacity to metabolize ethanol may be mediated by direct effects of both acetaldehyde and FAEEs and by alterations induced within the cells during ethanol metabolism, such as changes in the intracellular redox state and the generation of oxidant stress.
Asunto(s)
Etanol/metabolismo , Pancreatitis Alcohólica/metabolismo , Alcohol Deshidrogenasa/metabolismo , Citocromo P-450 CYP2E1/metabolismo , Etanol/administración & dosificación , Humanos , Oxidación-Reducción , Páncreas/metabolismo , Páncreas/patología , Pancreatitis Alcohólica/inducido químicamenteRESUMEN
The pathogenesis of pancreatic fibrosis, a characteristic feature of alcohol-induced chronic pancreatitis, has received increasing attention over the past few years, largely due to the identification and characterization of stellate cells in the pancreas. These cells are morphologically similar to hepatic stellate cells, the principal effector cells in liver fibrosis. The role of pancreatic stellate cells (PSCs) in alcoholic pancreatic fibrosis has been studied using 2 approaches: (i) in vivo studies using pancreatic tissue from patients with alcohol-induced chronic pancreatitis and from animal models of experimental pancreatitis and (ii) in vitro studies using cultured PSCs. These studies indicate that PSCs are activated early in the course of pancreatic injury and are the predominant source of collagen in the fibrotic pancreas. Several factors that may be responsible for mediating PSC activation during chronic alcohol exposure have also been identified. From the findings to date, it may be speculated that the pathogenesis of alcoholic pancreatic fibrosis may involve 2 pathways: (i) a necroinflammatory pathway involving cytokine release and PSC activation and (ii) a nonnecroinflammatory pathway involving direct activation of PSCs by ethanol via its metabolism to acetaldehyde and the generation of oxidant stress.
Asunto(s)
Páncreas/patología , Pancreatitis Alcohólica/patología , Actinas/biosíntesis , Animales , Citocinas/biosíntesis , Modelos Animales de Enfermedad , Etanol/farmacología , Proteínas de la Matriz Extracelular/biosíntesis , Fibrosis , Humanos , Ratones , Modelos Biológicos , Músculo Liso/química , Páncreas/efectos de los fármacos , Páncreas/metabolismo , RatasRESUMEN
BACKGROUND: Pancreatic fibrosis is a characteristic feature of alcoholic chronic pancreatitis. Recent studies suggest that activated pancreatic stellate cells (PSCs) are the major cell-type involved in pancreatic fibrogenesis. Cultured PSCs become activated when exposed to ethanol or its metabolite acetaldehyde (as indicated by increased alpha-smooth muscle actin [alpha-SMA] expression and increased collagen synthesis). However the intracellular signaling mechanisms responsible for ethanol- or acetaldehyde-induced PSC activation remain to be fully elucidated. One of the major signaling pathways known to regulate protein synthesis in mammalian cells is the mitogen-activated protein kinase (MARK) pathway. AIMS: To examine the effects of ethanol and acetaldehyde on the MAPK pathway (by assessing the activities of the 3 major subfamilies (extracellular-regulated kinases 1 and 2 [ERK 1/2], JNK and p38 kinase) in PSCs and to examine the role of p38 kinase in mediating the ethanol- and acetaldehyde-induced increase in alpha-SMA expression in activated rat PSCs. METHODS: Rat PSCs were incubated with ethanol (50 mM) or acetaldehyde (200 microM) for 15 min, 30 min, 60 min, and 24 h; and activities of ERK 1/2, JNK, and p38 kinase were assessed in cell lysates using kinase assays and Western blot. In addition, rat PSCs were treated with the specific p38 MAPK inhibitor SB203580 in the presence or absence of ethanol or acetaldehyde for 24h, and activation of the downstream protein kinase MAPKAP kinase-2 (an indicator of p38 MAPK activity) was assessed by Western blot. Specific inhibitors were also used to inhibit the activity of ERK 1/2 and JNK. Following inhibition of the above signaling pathways, alpha-SMA expression by PSCs was assessed by Western blot. RESULTS: Ethanol and acetaldehyde increased the activation of all 3 subfamilies (ERK 1/2, JNK and p38 kinase) of the MAPK pathway in PSCs. Treatment of PSCs with SB203580 abolished the ethanol- and acetaldehyde-induced increase in p38 MAPK activity and also prevented the induction of alpha-SMA expression in PSCs. However, inhibition of ERK 1/2 and JNK had no effect on ethanoland acetaldehyde-induced alpha-SMA expression in PSCs. CONCLUSIONS: (1) The MAP kinase pathway is induced in PSCs after exposure to ethanol or acetaldehyde and this induction is sustained for at least 24h. (2) The p38 MAPK pathway mediates the activation (as indicated by increased alpha-SMA expression) of PSCs by ethanol or acetaldehyde.
Asunto(s)
Acetaldehído/farmacología , Páncreas/efectos de los fármacos , Actinas/biosíntesis , Animales , Células Cultivadas , Activación Enzimática/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Imidazoles/farmacología , Proteínas Quinasas JNK Activadas por Mitógenos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Músculo Liso/química , Páncreas/citología , Páncreas/metabolismo , Inhibidores de Proteínas Quinasas , Proteínas Quinasas/metabolismo , Piridinas/farmacología , Ratas , Factores de Tiempo , Proteínas Quinasas p38 Activadas por MitógenosRESUMEN
Alcoholic pancreatitis is a major complication of alcohol abuse. Until recently, it was generally accepted that alcoholic pancreatitis was a chronic disease from the outset. However, evidence is now emerging in support of the 'necrosis-fibrosis' hypothesis that alcoholic pancreatitis begins as an acute process and that repeated episodes of acute injury lead to the changes of chronic pancreatitis (acinar atrophy and fibrosis) resulting in exocrine and endocrine dysfunction. The treatment of acute pancreatitis follows the regimen of bed rest, nasogastric suction, analgesia and intravenous support. The role of additional therapeutic measures such as prophylactic antibiotics, antioxidants and enteral nutrition in severe cases has not yet been precisely defined. The treatment of chronic pancreatitis involves attention to its three cardinal features: pain, maldigestion and diabetes. With respect to the pathogenesis of alcoholic pancreatitis, the focus of research over the past 30 years has shifted from the sphincter of Oddi and ductular abnormalities to the acinar cell itself. It has now been established that the acinar cell is capable of metabolizing alcohol and that direct toxic effects of alcohol and/or its metabolites on acinar cells may predispose the gland to injury in the presence of an appropriate trigger factor. A significant recent development relates to the characterization of pancreatic stellate cells, increasingly implicated in alcoholic pancreatic fibrosis. This chapter summarizes the natural history, clinical features, current trends in treatment as well as recent advances in our understanding of the pathogenesis of alcoholic pancreatitis.
Asunto(s)
Alcoholismo/complicaciones , Etanol/toxicidad , Pancreatitis Alcohólica/etiología , Pancreatitis Alcohólica/terapia , Enfermedad Aguda , Animales , Enfermedad Crónica , Diabetes Mellitus/etiología , Diabetes Mellitus/terapia , Fibrosis , Predisposición Genética a la Enfermedad , Humanos , Mediadores de Inflamación/metabolismo , Síndromes de Malabsorción/etiología , Síndromes de Malabsorción/terapia , Dolor/etiología , Manejo del Dolor , Páncreas/citología , Páncreas/efectos de los fármacos , Páncreas/enzimología , Páncreas/patología , Pancreatitis Alcohólica/diagnóstico , Pancreatitis Alcohólica/patologíaRESUMEN
BACKGROUND: Pancreatic stellate cells (PSCs), implicated as key mediators of pancreatic fibrogenesis, are found in increased numbers in areas of pancreatic injury. This increase in PSC number may be due to increased local proliferation and/or migration of these cells from adjacent areas. The ability of PSCs to proliferate has been well established but their potential for migration has not been examined. AIMS: Therefore, the aims of this study were to determine whether cultured rat PSCs have the capacity to migrate and, if so, to characterise this migratory capacity with respect to the influence of basement membrane components and the effect of platelet derived growth factor (PDGF, a known stimulant for migration of other cell types). METHODS: Migration of freshly isolated (quiescent) and culture activated (passaged) rat PSCs was assessed across uncoated or Matrigel (a basement membrane-like substance) coated porous membranes (pore size 8 micro m) in the presence or absence of PDGF (10 and 20 ng/ml) in the culture medium. A checkerboard assay was performed to assess whether the effect of PDGF on PSC migration was chemotactic or chemokinetic. RESULTS: Cell migration was observed with both freshly isolated and passaged PSCs. However, compared with passaged (culture activated) cells, migration of freshly isolated cells was delayed, occurring only at or after 48 hours of incubation when the cells displayed an activated phenotype. PSC migration through Matrigel coated membranes was delayed but not prevented by basement membrane components. PSC migration was increased by PDGF and this effect was predominantly chemotactic (that is, in the direction of a positive concentration gradient). CONCLUSIONS: (i) PSCs have the capacity to migrate. (ii) Activation of PSCs appears to be a prerequisite for migration. (iii) PDGF stimulates PSC migration and this effect is predominantly chemotactic. IMPLICATION: Chemotactic factors released during pancreatic injury may stimulate the migration of PSCs through surrounding basement membrane towards affected areas of the gland.
Asunto(s)
Movimiento Celular/fisiología , Páncreas/citología , Animales , Membrana Basal/metabolismo , Materiales Biocompatibles/farmacología , División Celular/fisiología , Movimiento Celular/efectos de los fármacos , Células Cultivadas/fisiología , Quimiotaxis/fisiología , Colágeno/farmacología , Combinación de Medicamentos , Laminina/farmacología , Factor de Crecimiento Derivado de Plaquetas/farmacología , Proteoglicanos/farmacología , RatasRESUMEN
BACKGROUND: Pancreatic fibrosis is a characteristic feature of chronic pancreatic injury and is thought to result from a change in the balance between synthesis and degradation of extracellular matrix (ECM) proteins. Recent studies suggest that activated pancreatic stellate cells (PSCs) play a central role in pancreatic fibrogenesis via increased synthesis of ECM proteins. However, the role of these cells in ECM protein degradation has not been fully elucidated. AIMS: To determine: (i) whether PSCs secrete matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) and, if so (ii) whether MMP and TIMP secretion by PSCs is altered in response to known PSC activating factors such as tumour necrosis factor alpha (TNF-alpha), transforming growth factor beta1 (TGF-beta1), interleukin 6 (IL-6), ethanol, and acetaldehyde. METHODS: Cultured rat PSCs (n=3-5 separate cell preparations) were incubated at 37 degrees C for 24 hours with serum free culture medium containing TNF-alpha (5-25 U/ml), TGF-beta1 (0.5-1 ng/ml), IL-6 (0.001-10 ng/ml), ethanol (10-50 mM), or acetaldehyde (150-200 micro M), or no additions (controls). Medium from control cells was examined for the presence of MMPs by zymography using a 10% polyacrylamide-0.1% gelatin gel. Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to examine gene expression of MMP9 and the tissue inhibitors of metalloproteinases TIMP1 and TIMP2. Western blotting was used to identify a specific MMP, MMP2 (a gelatinase that digests basement membrane collagen and the dominant MMP observed on zymography) and a specific TIMP, TIMP2. Reverse zymography was used to examine functional TIMPs in PSC secretions. The effect of TNF-alpha, TGF-beta1, and IL-6 on MMP2 secretion was assessed by densitometry of western blots. The effect of ethanol and acetaldehyde on MMP2 and TIMP2 secretion was also assessed by this method. RESULTS: Zymography revealed that PSCs secrete a number of MMPs including proteinases with molecular weights consistent with MMP2, MMP9, and MMP13. RT-PCR demonstrated the presence of mRNA for metalloproteinase inhibitors TIMP1 and TIMP2 in PSCs while reverse zymography revealed the presence of functional TIMP2 in PSC secretions. MMP2 secretion by PSCs was significantly increased by TGF-beta1 and IL-6, but was not affected by TNF-alpha. Ethanol and acetaldehyde induced secretion of both MMP2 and TIMP2 by PSCs. CONCLUSIONS: Pancreatic stellate cells have the capacity to synthesise a number of matrix metalloproteinases, including MMP2, MMP9, and MMP13 and their inhibitors TIMP1 and TIMP2. MMP2 secretion by PSCs is significantly increased on exposure to the proinflammatory cytokines TGF-beta1 and IL-6. Both ethanol and its metabolite acetaldehyde increase MMP2 as well as TIMP2 secretion by PSCs. IMPLICATION: The role of pancreatic stellate cells in extracellular matrix formation and fibrogenesis may be related to their capacity to regulate the degradation as well as the synthesis of extracellular matrix proteins.
Asunto(s)
Metaloproteinasas de la Matriz/biosíntesis , Páncreas/enzimología , Inhibidores Tisulares de Metaloproteinasas/biosíntesis , Acetaldehído/farmacología , Animales , Western Blotting/métodos , Células Cultivadas , Electroforesis en Gel de Poliacrilamida/métodos , Etanol/farmacología , Interleucina-6/farmacología , Metaloproteinasa 2 de la Matriz/análisis , Metaloproteinasa 2 de la Matriz/biosíntesis , Metaloproteinasa 9 de la Matriz/análisis , Metaloproteinasa 9 de la Matriz/biosíntesis , Metaloproteinasas de la Matriz/análisis , Páncreas/citología , Páncreas/efectos de los fármacos , ARN Mensajero/análisis , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Inhibidor Tisular de Metaloproteinasa-1/análisis , Inhibidor Tisular de Metaloproteinasa-1/biosíntesis , Inhibidor Tisular de Metaloproteinasa-2/análisis , Inhibidor Tisular de Metaloproteinasa-2/biosíntesis , Factor de Crecimiento Transformador beta/farmacología , Factor de Crecimiento Transformador beta1 , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
This article represents the proceedings of a workshop at the 2000 ISBRA Meeting in Yokohama, Japan. The presentations were (1) Phenotypic alteration of myofibroblast during ethanol-induced pancreatic injury: its relation to bFGF, by Masahiko Nakamura, Kanji Tsuchimoto, and Hiromasa Ishii; (2) Activation of pancreatic stellate cells in pancreatic fibrosis, by Paul S. Haber, Gregory W. Keogh, Minoti V. Apte, Corey S. Moran, Nancy L. Stewart, Darrell H.G. Crawford, Romano C. Pirola, Geoffrey W. McCaughan, Grant A. Ramm, and Jeremy S. Wilson; (3) Pancreatic blood flow and pancreatic enzyme secretion on acute ethanol infusion in anesthetized RAT, by H. Nishino, M. Kohno, R. Aizawa, and N. Tajima; (4) Genotype difference of alcohol-metabolizing enzymes in relation to chronic alcoholic pancreatitis between the alcoholic in the National Institute on Alcoholism and patients in other general hospitals in Japan, by K. Maruyama, H. Takahashi, S. Matsushita, K. Okuyama, A. Yokoyama, Y. Nakamura, K. Shirakura, and H. Ishii; and (5) Alcohol consumption and incidence of type 2 diabetes, by Katherine M. Conigrave, B. Frank Hu, Carlos A. Camargo Jr, Meir J. Stampfer, Walter C. Willett, and Eric B. Rimm.
Asunto(s)
Depresores del Sistema Nervioso Central/farmacología , Etanol/farmacología , Factor 2 de Crecimiento de Fibroblastos/efectos de los fármacos , Páncreas/efectos de los fármacos , Enfermedades Pancreáticas/metabolismo , Consumo de Bebidas Alcohólicas/metabolismo , Alcoholismo/complicaciones , Alcoholismo/metabolismo , Animales , Diabetes Mellitus Tipo 2/etiología , Diabetes Mellitus Tipo 2/metabolismo , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Humanos , Masculino , Páncreas/irrigación sanguínea , Páncreas/metabolismo , Enfermedades Pancreáticas/etiología , Pancreatitis Alcohólica/etiología , Pancreatitis Alcohólica/metabolismo , Ratas , Ratas Sprague-Dawley , Ratas WistarRESUMEN
BACKGROUND & AIMS: Activated pancreatic stellate cells have recently been implicated in pancreatic fibrogenesis. This study examined the role of pancreatic stellate cells in alcoholic pancreatic fibrosis by determining whether these cells are activated by ethanol itself and, if so, whether such activation is caused by the metabolism of ethanol to acetaldehyde and/or the generation of oxidant stress within the cells. METHODS: Cultured rat pancreatic stellate cells were incubated with ethanol or acetaldehyde. Activation was assessed by cell proliferation, alpha-smooth muscle actin expression, and collagen synthesis. Alcohol dehydrogenase (ADH) activity in stellate cells and the influence of the ADH inhibitor 4-methylpyrazole (4MP) on the response of these cells to ethanol was assessed. Malondialdehyde levels were determined as an indicator of lipid peroxidation. The effect of the antioxidant vitamin E on the response of stellate cells to ethanol or acetaldehyde was also examined. RESULTS: Exposure to ethanol or acetaldehyde led to cell activation and intracellular lipid peroxidation. These changes were prevented by the antioxidant vitamin E. Stellate cells exhibited ethanol-inducible ADH activity. Inhibition of ADH by 4MP prevented ethanol-induced cell activation. CONCLUSIONS: Pancreatic stellate cells are activated on exposure to ethanol. This effect of ethanol is most likely mediated by its metabolism (via ADH) to acetaldehyde and the generation of oxidant stress within the cells.
Asunto(s)
Etanol/farmacología , Páncreas/efectos de los fármacos , Páncreas/patología , Acetaldehído/farmacología , Actinas/metabolismo , Animales , Ácido Ascórbico/farmacología , División Celular/efectos de los fármacos , Células Cultivadas , Colágeno/biosíntesis , Combinación de Medicamentos , Etanol/metabolismo , Compuestos Férricos/farmacología , Fibrosis , Peróxidos Lipídicos/metabolismo , Malondialdehído/metabolismo , Músculo Liso/metabolismo , Estrés Oxidativo , Páncreas/metabolismo , RatasRESUMEN
The mechanisms of pancreatic fibrosis are poorly understood. In the liver, stellate cells play an important role in fibrogenesis. Similar cells have recently been isolated from the pancreas and are termed pancreatic stellate cells. The aim of this study was to determine whether pancreatic stellate cell activation occurs during experimental and human pancreatic fibrosis. Pancreatic fibrosis was induced in rats (n = 24) by infusion of trinitrobenzene sulfonic acid (TNBS) into the pancreatic duct. Surgical specimens were obtained from patients with chronic pancreatitis (n = 6). Pancreatic fibrosis was assessed using the Sirius Red stain and immunohistochemistry for collagen type I. Pancreatic stellate cell activation was assessed by staining for alpha-smooth muscle actin (alphaSMA), desmin, and platelet-derived growth factor receptor type beta (PDGFRbeta). The relationship of fibrosis to stellate cell activation was studied by staining of serial sections for alphaSMA, desmin, PDGFRbeta, and collagen, and by dual-staining for alphaSMA plus either Sirius Red or in situ hybridization for procollagen alpha(1) (I) mRNA. The cellular source of TGFbeta was examined by immunohistochemistry. The histological appearances in the TNBS model resembled those found in human chronic pancreatitis. Areas of pancreatic fibrosis stained positively for Sirius Red and collagen type I. Sirius Red staining was associated with alphaSMA-positive cells. alphaSMA staining colocalized with procollagen alpha(1) (I) mRNA expression. In the rat model, desmin staining was associated with PDGFRbeta in areas of fibrosis. TGFbeta was maximal in acinar cells adjacent to areas of fibrosis and spindle cells within fibrotic bands. Pancreatic stellate cell activation is associated with fibrosis in both human pancreas and in an animal model. These cells appear to play an important role in pancreatic fibrogenesis.
Asunto(s)
Páncreas/metabolismo , Pancreatitis Alcohólica/metabolismo , Actinas/metabolismo , Animales , Enfermedad Crónica , Colágeno/metabolismo , Desmina/metabolismo , Modelos Animales de Enfermedad , Fibrosis/inducido químicamente , Fibrosis/metabolismo , Fibrosis/patología , Proteína Ácida Fibrilar de la Glía/metabolismo , Humanos , Inmunohistoquímica , Masculino , Páncreas/efectos de los fármacos , Páncreas/patología , Pancreatitis Alcohólica/patología , Procolágeno/biosíntesis , ARN Mensajero/biosíntesis , Ratas , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/metabolismo , Factor de Crecimiento Transformador beta/metabolismo , Ácido TrinitrobencenosulfónicoRESUMEN
Chronic pancreatitis is characterized by progressive and irreversible loss of pancreatic exocrine and endocrine function. In the majority of cases, particularly in Western populations, the disease is associated with alcohol abuse. The major complications of chronic pancreatitis include abdominal pain, malabsorption, and diabetes. Of these, pain is the most difficult to treat and is therefore the most frustrating symptom for both the patient and the physician. While analgesics form the cornerstone of pain therapy, a number of other treatment modalities (inhibition of pancreatic secretion, antioxidants, and surgery) have also been described. Unfortunately, the efficacy of these modalities is difficult to assess, principally because of the lack of properly controlled clinical trials. Replacement of pancreatic enzymes (particularly lipase) in the gut is the mainstay of treatment for malabsorption; the recent discovery of a bacterial lipase (with high lipolytic activity and resistance to degradation in gastric and duodenal juice) represents an important advance that may significantly increase the efficacy of enzyme replacement therapy by replacing the easily degradable porcine lipase found in existing enzyme preparations. Diabetes secondary to chronic pancreatitis is difficult to control and its course is often complicated by hypoglycaemic attacks. Therefore, it is essential that caution is exercised when treating this condition with insulin. This paper reviews recent research and prevailing concepts regarding the three major complications of chronic pancreatitis noted above. A comprehensive discussion of current opinion on clinical issues relating to the other known complications of chronic pancreatitis such as pseudocysts, venous thromboses, biliary and duodenal obstruction, biliary cirrhosis, and pancreatic cancer is also presented.
Asunto(s)
Pancreatitis/complicaciones , Pancreatitis/terapia , Analgésicos/uso terapéutico , Enfermedad Crónica , Terapia Combinada , Femenino , Humanos , Masculino , Pancreatectomía/métodos , Pancreatitis/diagnóstico , PronósticoRESUMEN
BACKGROUND: The observation that only a minority of alcoholics develops clinical pancreatic disease has led to a search for a predisposing factor to the disease. One possible predisposing factor is mutation of the cystic fibrosis transmembrane conductance regulator (CFTR) gene as cystic fibrosis leads to pancreatic injury. We have recently demonstrated that 15 common CFTR mutations are not found in patients with alcoholic pancreatitis. Another common polymorphism of the CFTR gene has recently been implicated in the pathogenesis of idiopathic chronic pancreatitis, the 5T variant of the variable length polythymidine tract in intron 8 (the normal genotypes are 7T and 9T). The 5T variant inhibits transcription of exon 9 resulting in a CFTR protein lacking chloride channel activity. The aim of this study was to determine whether the 5T variant is associated with alcoholic pancreatitis. METHODS: Fifty-two patients with alcoholic pancreatitis were identified using standardized diagnostic criteria. Fifty alcoholics without pancreatitis were also studied as controls. Genomic DNA was extracted from peripheral blood leukocytes and the polythymidine tract of intron 8 was amplified by nested polymerase chain reaction using established primers. The polymerase chain reaction products were digested with MseI, separated by electrophoresis on 15% polyacrylamide gels and genotypes assigned by comparison with known positive controls. RESULTS: The 5T allele we found in only two patients with alcoholic pancreatitis (3.9% of th index group; 95% confidence intervals 0-10%) and in seven alco holic controls. Allele frequencies for 5T, 7T, and 9T in patients with alcoholic pancreatitis were 1.9%, 85.6%, and 12.5%, respectively These did not differ from the allele frequencies in alcoholic controls (7%, 79%, and 14% for 5T, 7T, and 9T, respectively). CONCLUSION: The 5T allele was not associated with alcoholic pancreatitis. Individual susceptibility to this disease remains unexplained.