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.
Acute pancreatitis (AP) is characterized by initial pancreatic injury resulting from the activation of digestive enzymes and, later, widespread inflammation to distant organs. The aim of this study was to study whether the time-course of inflammatory events during AP induced by bile-pancreatic duct obstruction (BPDO) varies after lowering the acinar enzyme content by L364,718 (0.1 mg/kg/day) administration over 7 days before inducing AP. Flow cytometric immunophenotyping was used to analyse the following at different AP stages: distribution of major circulating leucocyte subsets, activation state of circulating neutrophils and monocytes as reflected by CD11b expression and tumour necrosis factor-alpha (TNF-alpha) production and the contribution of T-cell-derived pro-(TNF-alpha) and anti-(IL-10) inflammatory mediators. TNF-alpha plasma levels and neutrophil infiltration in pancreas and lung were also measured. At early BPDO times, L364,718 treatment partially inhibited leukocytosis and increase in peripheral blood neutrophils and monocytes as well as TNF-alpha expression by monocytes. However, from 6 h onwards after BPDO, L364,718 treatment was unable to prevent either pancreatic and lung neutrophil infiltration or the release of TNF-alpha from activated monocytes. By its action on circulating lymphocytes, L364,718 treatment enhanced the severity of the inflammatory response induced by BPDO. Peripheral blood lymphocytes were recruited from earlier BPDO times, and 12 h after BPDO, T cells displayed a significantly higher reserve of TNF-alpha able to be released under stimulation but lower functional reserve of interleukin-10 (IL-10) than observed in untreated rats. It is concluded that lowering the acinar enzyme content through L364,718 treatment prevents earlier systemic immune events in BPDO-induced AP. However, at the point of maximal injury, the inflammatory response became pronounced, largely due to the role played by activated T lymphocytes.
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.
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