Protective Effect Of Ethyl Pyruvate On Acute Necrotizing Pancreatitis-associated Acute Lung Injury In Rats

Acute lung injury (ALI) is a common complication of severe acute pancreatitis(SAP). Approximately one third of patients will develop acute lung injury and acute respiratory distress syndrome (ARDS), which account for 60% of all deaths within the first week. The mechanisms that initiate progression to lung injury in pancreatitis remain ill defined. Ventilation is the principle therapeutic maneuver, although recent interest has focused on suppressing the overly exuberant inflammatory response. Attempts to influence the interplay between proinflammatory and anti-inflammatory cytokines may offer the best prospect of diminishing pulmonary involvement. Animal models of pancreatitis have a propensity for responding to manipulation of cytokine pathways, but, lamentably, this response rarely translates into a demonstrable clinical effect in human trials and the pharmacodynamics or toxicity of these agents may limit their value in critically ill patients with multiple organ dysfunction syndrome (MODS). Furthermore, most of these agents aim to target on early inflammatory mediator so have short therapeutic window.Recent studies showed that being as an antoxidant and free radical scavenger, ethyl pyruvate can also regulated the releasing of early and late proinflammatory cytokines such as TNF-α, high mobility group box 1 protein (HMGB1). Treatment with ethyl pyruvate solution can improve outcome in a variety of animal models of critical illness, such as hemorrhagic shock, sepsis and ischemia, and reperfusion injury.Basis upon the above investigation, we established steady and repeatable easily model of acute lung injury induced by acute necrotizing pancreatitis in rats and explore the pathogenesis of the experimental ALL And then we observed the effect of delayed treatment with ethyl pyruvate on ALI induced by ANP and explore its possible mechanisms. HMGB1 has been demonstrated to be a novel proinflammatory mediator in sepsis and endotoxemia, we also aimed to investigate the role of HMGB1 in the experimental ALI.MethodsSeventy-two male Wistar rats were randomly divided into three groups: sham operation group(SO group), ANP group and EP treatment group. ANP model was established by retrograde injection of 5% sodium taurocholate into biliopancreatic duct. While the SO group received laparotomy and smooth the pancreas tissue some times only. Then the three groups rats were divided into 6 hour group, 12 hour group, 24 hour group according to the being killed time after model was induced. The rats of EP treatment group were injected with ethyl pyruvate solution (28mM) intravenously at 2h, 8h, 14h, 22h after ANP model was induced, each of dose is 40mg/kg. The serum levels of TNF-αand IL-6 were quantitated using ELISA kits. The lung tissue level of peroxidization (MDA) and superoxide dismutase (SOD) were also quantitated using colorimetric method. The HMGB1 mRNA level of lungs was detected by semi quantitative RT-PCR. The severity of Pancreatitis was determined by serum amylase, amounts of ascites and local pathologic lesions (gross and histopathologic scoring), the lung injury was measured by PaO₂, tissue water content, myeloperoxidase (MPO) activity of lung tissue, and histopathologic scoring.Results1. After model induced, serum level of AMY, the amount of the ascites and pathology scores of pancreas and lungs in ANP group kept rising continuously. Pancreas were observed edema, hemorrhage, necrosis spots; Massive alveolar edema, hemorrhage, and inflammatory cell infiltration were observed in lungs. The more longer the time, the more serious the damage. 2. The lung tissue level of MPO increased significantly after the ANP model induced. Contrasted with ANP group, the lung tissue level of MPO in treatment group rats were significantly lower at each time (P<0.05).3. The serum levels of TNF-αand IL-6 were quickly increased after ANP model induced, and reached peak at 6h, but decreased at 12h, whereas in treatment group, the serum levels of TNF-αand IL-6 were lower than ANP group (P<0.05).4. The HMGB1 mRNA level of lung tissue were increased significantly at 12h, and maintained to 24h after ANP model induced; whereas in treatment group, the HMGB1 mRNA level of lungs were lower than ANP group (P<0.05).5. The lung tissue level of MDA manifested a steady significant increase with time in ANP groups, while the level of SOD decreased gradually. In treatment group, the level of MDA was significantly lower at 6,12h (P<0.05) and the level of SOD was significantly higher at each time than ANP group (P<0.05).6. The histological injury of pancreas and lungs was milder than ANP group. PaO₂ were significantly higher in treatment group than ANP group (P<0.05).Conclusions1. This model is steady and repeatable, and is suitable in the research of acute pancreatitis-associated lung injury. This ANP model rats have lung injury in early phases, histologically manifested inflammatory cell infiltration, pulmonary edema, hemorrhage with respiratory dysfunction.2. After this ANP model induced, active pancreatic enzyme released and activated macrophages released lots of damaged inflammatory mediators such as cytokines into the systemic circulation which can result in accumulation and activation of inflammatory cells such as neutrophils within the pulmonary interstitial and alveolar spaces. Then noxious substance such as proinflammatory cytokines (e.g., TNF-α, HMGB1) and oxygen free radical was released and lead to lung injury.3. After this ANP model induced, the delayed kinetics of HMGB1 expression suggestted that it can be targeted in a wider therapeutic window in ANPALI than is provided by inhibitors that target the early cytokine response (e.g., anti-TNF or anti-IL-1).4. Ethyl pyruvate appears to have important anti-inflammatory and antioxidant properties so that it can lesson the severity of ANPALI. EP also has a wider therapeutic window in ANPALI than other inhibitors that target the early cytokine response (e.g., anti-TNF or anti-IL-1). Collectively, these data support the view that ethyl pyruvate warrants further evaluation as a therapeutic agent for APALI.