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1.
Biochim Biophys Acta ; 1639(3): 177-84, 2003 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-14636949

RESUMO

Although oxygen free radicals (OFR) are considered to be one of the pathophysiological mechanisms involved in acute pancreatitis (AP), the contribution of acinar cells to their production is not well established. The aim of the present study was to determine the effect of N-acetylcysteine (NAC) in the course of AP induced by pancreatic duct obstruction (PDO) in rats, directly analysing by flow cytometry the quantity of OFR generated in acinar cells. NAC (50 mg/kg) was administered 1 h before and 1 h after PDO. Measurements by flow cytometry of OFR generated in acinar cells were taken at different PDO times over 24 h, using dihydrorhodamine-123 as fluorescent dye. Histological studies of pancreas and measurements of neutrophil infiltration in the pancreas, pancreatic glutathione (GSH), malondialdehyde (MDA) levels, plasma amylase activity and hemoconcentration were carried out in order to assess the severity of AP at different stages. NAC effectively blunted GSH depletion at early AP stages and prevented OFR generation found in acinar cells as a consequence of AP induced by PDO. This attenuation of the redox state impairment reduced cellular oxidative damage, as reflected by less severe pancreatic lesions, normal pancreatic MDA levels, as well as diminished neutrophil infiltration in pancreas. Hyperamylasemia and hemoconcentration following AP induction were ameliorated by NAC administration at early stages, when oxidative stress seems to be critical in the development of pancreatitis. In conclusion, NAC reinforces the antioxidant defences in acinar cells, preventing OFR generation therefore attenuating oxidative damage and subsequently reducing the severity of PDO-induced AP at early stages of the disease.


Assuntos
Acetilcisteína/uso terapêutico , Sequestradores de Radicais Livres/uso terapêutico , Ductos Pancreáticos/efeitos dos fármacos , Pancreatite/tratamento farmacológico , Doença Aguda , Amilases/sangue , Animais , Modelos Animais de Doenças , Radicais Livres/metabolismo , Masculino , Pâncreas/efeitos dos fármacos , Pâncreas/patologia , Pâncreas/ultraestrutura , Ratos , Ratos Wistar
2.
Cell Prolif ; 36(5): 279-89, 2003 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-14521521

RESUMO

Oxygen free radicals (OFR) are produced in the course of acute pancreatitis (AP). In addition to injurious oxidative effects, they are also involved in the regulation of cell growth. The aim of the present study was to examine the relationship between the effectiveness of N-acetyl-l-cysteine (NAC) to prevent the generation of OFR and the changes in the cell-cycle pattern of acinar cells in the course of AP induced in rats by pancreatic duct obstruction (PDO). NAC (50 mg/kg) was administered 1 h before and 1 h after PDO. Flow-cytometric measurement of OFR generation in acinar cells was carried out using dihydrorhodamine as fluorescent dye. Plasma amylase activity, pancreatic glutathione (GSH) content and TNF-alpha plasma levels were also measured. The distribution of acinar cells throughout the different cell-cycle phases was analysed at different AP stages by flow cytometry using propidium iodide staining. NAC administration reduced the depletion of pancreatic GSH content and prevented OFR generation in acinar cells of rats with PDO-induced acute pancreatitis. As a result, AP became less severe as reflected by the significant improvement of hyper-amylasaemia and maintenance of plasma TNF-alpha levels at values not significantly different from controls were found. NAC administration inhibited progression of cell-cycle phases, maintaining acinar cells in quiescent state at early PDO times. The protection from oxidative damage by NAC treatment during early AP, allows the pancreatic cell to enter S-phase actively at later stages, thereby allowing acinar cells to proliferate and preventing the pancreatic atrophy provoked by PDO-induced AP. The results provide evidence that OFR play a critical role in the progression of acinar cell-cycle phases. Prevention of OFR generation of acinar cells in rats with PDO-induced AP through NAC treatment, not only protects pancreas from oxidative damage but also promotes beneficial changes in the cell cycle progression which reduce the risk of pancreatic atrophy.


Assuntos
Acetilcisteína/farmacologia , Ciclo Celular/efeitos dos fármacos , Radicais Livres/metabolismo , Pancreatite/patologia , Doença Aguda , Amilases/sangue , Animais , Citometria de Fluxo , Masculino , Estresse Oxidativo/efeitos dos fármacos , Pancreatite/metabolismo , Ratos , Ratos Wistar
3.
Digestion ; 68(1): 34-40, 2003.
Artigo em Inglês | MEDLINE | ID: mdl-12949437

RESUMO

AIM: To analyze the capability of N-acetylcysteine (NAC) to prevent major intra-acinar pathogenic mechanisms involved in the development of acute pancreatitis (AP). METHODS: AP was induced by pancreatic duct obstruction (PDO) in rats. Some animals received NAC (50 mg/kg) 1 h before and 1 h after PDO. During a 24-hour period of PDO, plasma amylase activity and pancreatic glutathione and malondialdehyde levels were measured. Cytosolic Ca(2+) levels and enzyme (amylase and trypsinogen) load in acinar cells were also analyzed by flow cytometry, and histological analysis of the pancreas was performed by electron microscopy. RESULTS: NAC avoided glutathione depletion at early AP stages, thereby preventing pancreatic oxidative damage, as reflected by normal malondialdehyde levels. By limiting oxidative stress, NAC treatment effectively prevented the impairment of Ca(2+) homeostasis found in acinar cells from early AP onwards, thus protecting the pancreas from damage. In addition, lower quantities of digestive enzymes were accumulated within acinar cells. This finding, together with the significantly lower hyperamylasemia observed in these animals, suggests that NAC treatment palliates the exocytosis blockade induced by PDO. CONCLUSION: By preventing oxidative stress at early AP stages, NAC administration prevents other pathological mechanisms of AP from being developed inside acinar cells, thus palliating the severity of disease.


Assuntos
Acetilcisteína/farmacologia , Sequestradores de Radicais Livres/farmacologia , Pâncreas/patologia , Pancreatite/patologia , Doença Aguda , Animais , Masculino , Microscopia Eletrônica , Estresse Oxidativo , Pâncreas/metabolismo , Pancreatite/prevenção & controle , Ratos , Ratos Wistar
4.
Biochim Biophys Acta ; 1588(2): 159-64, 2002 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-12385780

RESUMO

The time-course of oxygen free radicals (OFR) generation within acinar cells was studied at different stages of acute pancreatitis (AP) induced in rats by duct obstruction (PDO) for 48 h by flow cytometry, using dihydrorhodamine-123 (DHR) as fluorescent dye. Parallel measurements of the most common markers of oxidative stress such as glutathione (GSH) depletion and malondialdehyde (MDA) levels in pancreas were also performed. OFR production significantly increased within acinar cells at early stages of AP, concomitant with a marked depletion in pancreatic GSH. Lipid peroxidation was significantly enhanced 6 h after PDO, suggesting that the antioxidant defence system of the cell is overwhelmed by OFR production. Both MDA and OFR production in acinar cells decreased to normal values at late AP stages, thus allowing the recovery of pancreatic GSH levels 48 h after PDO. Among the two types of acinar cells differentiated by flow cytometry, R1 and R2, it was the R2 population that showed higher values of DHR dye. However, no differences between the two cell types were found regarding the amount of OFR generation. Our results demonstrate that individual acinar cells significantly contribute to produce large amounts of OFR at early stages of AP. The two existing populations of acinar cells displayed similar behaviour regarding oxidative stress over the course of the disease.


Assuntos
Ductos Pancreáticos/fisiopatologia , Pancreatite/fisiopatologia , Doença Aguda , Amilases/metabolismo , Análise de Variância , Animais , Separação Celular , Citometria de Fluxo , Glutationa/metabolismo , Hematócrito , Masculino , Malondialdeído/metabolismo , Estresse Oxidativo , Ductos Pancreáticos/citologia , Pancreatite/sangue , Pancreatite/metabolismo , Ratos , Ratos Wistar
5.
Biochim Biophys Acta ; 1509(1-2): 292-8, 2000 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-11118540

RESUMO

Flow-cytometric studies of lectin binding to individual acinar cells have been carried out in order to analyse the distribution of membrane glycoconjugates in cells from different areas of the pancreas: duodenal lobule (head) and splenic lobule (body and tail). The following fluoresceinated lectins were used: wheat germ agglutinin (WGA), Tetragonolobus purpureus agglutinin (TP) and concanavalin A (Con A), which specifically bind to N-acetyl D-glucosamine and sialic acid, L-fucose and D-mannose, respectively. In both pancreatic areas, two cell populations (R1 and R2) were identified according to the forward scatter (size). On the basis of their glycoconjugate pattern, R1 cells displayed higher density of WGA and TP receptors than R2 cells throughout the pancreas. Although no difference in size was found between the cells from duodenal and splenic lobules, N-acetyl D-glucosamine and/or sialic acid and L-fucose residues were more abundant in plasma membrane cell glycoconjugates from the duodenal lobule. The results provide evidence for biochemical heterogeneity among individual pancreatic cells according to the distribution of plasma membrane glycoconjugates.


Assuntos
Membrana Celular/metabolismo , Glicoconjugados/metabolismo , Pâncreas/metabolismo , Animais , Citometria de Fluxo , Fluoresceína-5-Isotiocianato/química , Glicoconjugados/química , Lectinas/química , Masculino , Pâncreas/citologia , Pâncreas/embriologia , Ratos , Ratos Wistar
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