Effect Of Rosiglitazone On Acute Pancreatitis Associated Lung Injury And Its Mechanisms

Part 1. Dose-response relationship of rosiglitazone administered via vein on acute pancreatitis in ratsObjective:The aim of this study is to investigate dose-response relationship of rosiglitazone administered via vein on acute pancreatitis in order to obtain the optimal dose of rosiglitazone for preventing sodium taurocholate-induced acute pancreatitis associated-lung injury.Methods:Forty-two male Wistar rats, weighing 200-250g, were randomly divided into seven groups (N=6):control group (CON group), acute pancreatitis model group(AP group), different dosage(0.3mg/kg,3mg/kg,6mg/kg and 10mg/kg) rosiglitazone pretreatment group (ROSI group) and drug control group (Drug-CON group) (dosage according to the effect of rosiglitazone). Prior to the experiment, rats were deprived of food for 12 hours, while drinking water was available ad libitum. Anaesthesia was conducted by intraperitoneal injection of 10% chloraldurat (0.3ml/ 100g) and rats underwent sterile laparotomy. The bile-pancreatic duct was cannulated through the duodenum and acute pancreatitis model was induced by a standardized pressure-controlled retrograde infusion 5% sodium taurocholate(0.1ml/100g) into the bile-pancreatic duct. The CON group was identical to the AP group except that the saline was injected instead of sodium taurocholate. The CON group and the AP group were received 10% dimethyl sulphoxide (DMSO) (0.2ml/100g) administered via femoral vein 30min prior to the operation. In the ROSI group, different concentration rosiglitazone were injected partes aequales respectivly instead of 10% DMSO. The drug control group was identical to the CON group except that partes aequales rosiglitazone was injected instead of 10% DMSO. Rats were killed at 12 hours after operation. Blood were obtained from the left ventricle of heart. The samples were centrifuged and the serum amylase level, alanine aminotransferase level were detected by automatic biochemistry analyzer. Continuous sections of the paraffin embedded tissue were taken for pathological examination with hematoxylin-eosin (HE) staining. Morphometric documentation for pancreatic and pulmonary sections under light microscope were evaluated. Results:Compared with the AP group,0.3mg/kg,3mg/kg dosage failed to improve the levels of amylase and alanine aminotransferase (P> 0.05), but attenuated severity of pancreatic injury (P<0.05).6mg/kg dosage had the best effect on improving the levels of amylase, alanine aminotransferase and pathologic score (P<0.05). The dosage of 10mg/kg also improve level of amylase and pathologic score (P<0.05), but failed to improve the level of alanine aminotransferase (P>0.05). There was no statistically significant between 6mg/kg dosage and 10mg/kg dosage of all the indexes. Compared with the CON group, rats of preteatment with rosiglitazone (6mg/kg) in the absence of sodium taurocholate revealed virtually the same serum amylaselevel, alanine aminotransferase level and pathologic score (P>0.05).Conclusion:Rosiglitazone (6mg/kg) administered intravenously was the safe, effective dosage for attenuating the severity of acute pancreatitis induced by retrograde infusion sodium taurocholate.Part 2. Effect of rosiglitazone on acute pancreatitis associated lung injury in ratsObjective:The aim of this study is to investigate the effect of rosiglitazone on acute pancreatitis associated lung injury in rats.Methods:Fifty-four male Wistar rats, weighing 200-250g, were randomly divided into three groups (N=18):control group (CON group), acute pancreatitis model group (AP group) and rosiglitazone pretreatment group (ROSI group). Prior to the experiment, rats were deprived of food for 12 hours, while drinking water was available ad libitum. Anaesthesia was conducted by intraperitoneal injection of 10% chloraldurat(0.3ml/100g) and rats underwent sterile laparotomy. The bile-pancreatic duct was cannulated through the duodenum and acute pancreatitis model was induced by a standardized pressure-controlled retrograde infusion 5% sodium taurocholate (0.1ml/100g) into the bile-pancreatic duct. The CON group was identical to the AP group except that the saline was injected instead of sodium taurocholate. The CON group and the AP group were received 10% dimethyl sulphoxide (DMSO) (0.2ml/ 100g) administered via femoral vein 30min prior to the operation. In the ROSI group, rosiglitazone (6mg/kg) dissolved in 10% DMSO was injected by femoral vein 30 minutes piror to the operation. Rats were killed at 3,6 and 12 hours after operation. Blood were obtained from the left ventricle of heart. The samples were centrifuged and the serum amylase level was detected by automatic biochemistry analyzer. Continuous sections of the paraffin embedded tissue were taken for pathological examination with hematoxylin-eosin (HE) staining. Morphometric documentation for pancreatic and pulmonary sections under light microscope were evaluated. Lung myeloperoxidase (MPO) activity was detceted by chromatometry(reflect neutrophil infiltration). The left main bronchus was instilled with 3 mL saline at appropriate pressure, and bronchoalveolar lavage fluid (BALF) was collected for detecting protein content (reflect pulmonary microvascular permeability). The lung wet/dry ratio (W/D) was determined by calculating from the initial weight of the right lung middle lobe (wet weight) to its weight after desiccation at 70℃for 24 hours (dry weight) (reflect pulmonary edema extent).Results In the AP group, serum amylase level, lung MPO activity, W/D ratio, protein content of BALF, pancreatic and pulmonary pathologic score reached the peak level at 12 hours after operation(5353.0±728.2 U/L,1.12±0.14 U/g,3.00±0.14, 0.438±0.056,11.17±0.93 and 8.17±0.75 respectively). All the indexes were increased significantly at each time point in the AP group than in the CON group (P< 0.05). Compared with the AP group, pretreatment with rosiglitazone reduced serum amylase level, lung MPO activity, W/D weight ratio, protein content of BALF, pancreatic and pulmonary pathologic score at 6h and 12h(P<0.05).Conclusions:Rosiglitazone exerts the protective effect against acute pancreatitis associated lung injury by inhibition of lung MPO activity, reducing pulmonary edema and pulmonary microvascular permeability.Part 3. Effect mechanism of rosiglitazone on acute pancreatitis associated lung injury in ratsObjective:The aim of this study is to investigate the effect of rosiglitazone on adhesion molecules expression in acute pancreatitis associated lung injury in rats.Methods:Fifty-four male Wistar rats, weighing 200-250g, were randomly divided into three groups (N=18):control group (CON group), acute pancreatitis model group(AP group)and rosiglitazone pretreatment group(ROSI group). Prior to the experiment, rats were deprived of food for 12 hours, while drinking water was available ad libitum. Anaesthesia was conducted by intraperitoneal injection of 10% chloraldurat(0.3ml/100g) and rats underwent sterile laparotomy. The bile-pancreatic duct was cannulated through the duodenum and acute pancreatitis model was induced by a standardized pressure-controlled retrograde infusion 5% sodium taurocholate (0.1ml/100g) into the bile-pancreatic duct. The CON group was identical to the AP group except that the saline was injected instead of sodium taurocholate. The CON group and the AP group were received 10% dimethyl sulphoxide (DMSO) (0.2ml/ 100g) administered via femoral vein 30min prior to the operation. In the ROSI group, rosiglitazone (6mg/kg) dissolved in 10% DMSO was injected by femoral vein 30 minutes piror to the operation. Rats were killed at 3,6 and 12 hours after operation. In pulmonary tissue, nuclear factor-kappa B (NF-κB) p65 expression was assayed by immuno-histochemistry. Tumor necrosis factor-α(TNF-α), intercellular adhesion molecule-1 (ICAM-1) and P-selectin mRNA expression were detected by reverse transcriptase polymerase chain reaction (RT-PCR). ICAM-1 and P-selectin protein expression were studied using Western blot analysis.Results In the AP group, the peak of NF-κB score, TNF-a mRNA expression ICAM-1 mRNA expression, P-selectin mRNA expression of the lung tissue were at 12 hours (1.04±0.13, 0.57±0.03,1.53±0.08,0.51±0.09). All the indexes were increased significantly at each time point in the AP group than in the CON group (P< 0.05). Compared with the AP group, pretreatment with rosiglitazone decreased the NF-κB score and the expression of TNF-a mRNA at 6h and 12h(P< 0.05), down-regulation the expression of ICAM-1, P-selectin mRNA and protein at 12 hours (P<0.05).Conclusions:Rosiglitazone exerts the protective effect against acute pancreatitis associated lung injury by inhibition of NF-κB and TNF-α, down-regulation the expression of ICAM-1 and P-selectin.