| BackgroundAcute pancreatitis is a kind of disease characterized by pancreatic interstitial edema, bleeding, acinous cell vacuolar degeneration, necrosis and inflammatory cells infiltration. Most acute pancreatitis accompany with whole body inflammatory reaction, about 20%patients develop to severe acute pancreatitis (SAP), which is 30%-50%mortality rate. With the changing of life habits and food structure, acute pancreatitis (including SAP) becomes more and more common. Therefore SAP has become a frequent and severe threaten disease to people.Acute pancreatitis-associated lung Injury (APALI) is the most frequent, earliest and severest complication of SAP. Pathogenesy of APALI is still not clear, recently most studying focused on the inflammatory cells and medium. In common APALI is considered to be a part of whole body inflammatory reaction, and because of the tissues specificity of lung it's easy to get hurt from inflammation. Accumulative evidences indicate that TNF-α, IL-1, IL-6 secreted by active mononuclear macrophage, polynuclear granulocytes in pancreas are very important factors to induce pancreatitis and APALI. The overexpression of pro inflammatory factor can aggravate experimental pancreatitis and APALI. So to decrease the inflammatory factors in lung is a hot point that people study how to treat APALI. Abundant animal experiments show that low the level of inflammatory factor in lung or whole body can treat APALI. But the real mechanism of increasing of inflammatory factor in lung during pancreatitis is still unknown. Only finding the real mechanism it can guide the treatment of APALI and decrease the mortality rate of SAP.Recently, JNK path has got more and more people's attention in the developing of acute pancreatitis. In acute pancreatitis animal models JNK path is activated and induce inflammatory factors release. JNK path is activated, which is an early incidence in the developing of acute pancreatitis. Using specific inhibitor of JNK path (SP600125) can decrease the expression of TNF-α, ICAM-1, IL-1βand so on, it can significantly improve the impairing of pancreas and treat the acute pancreatitis. The evidences show that JNK path plays a very important role during the development of acute pancreatitis. At the same time people have found the connection between APALI and acute pancreatitis. In vitro, studying show that JNK path plays an important role in the APALI animal models induced by nickel or LPS, and JNK path specific inhibitor can be used to treat APALI especially animal models induced by LPS. But it's still not clear the mechanism of JNK path in acute pancreatitis.The studying is divided into 2 parts. Part one is to observe the activation of JNK path in the lung of rat acute hemorrhagic necrotizing pancreatitis induced by bile salts and investigate the correlation between JNK path and APALI. At the same time SP600125 is used to confirm the relationship between JNK path and APALI. Part two is to investigate if tanshinone can treat APALI and the mechanism of it.Part one The mechanisms of JNK pathway in acute pancreatitis associated lung injury in rat AimThe aim of studying is to detect the mechanism of JNK path in the developing of acute pancreatitis, and primarily research the therapeutical effect of tanshinone in APALI.Methods72 SD rats were individed into 3 groups. Group S:sham operated group,24 rats, after opening abdomen duodenum and pancreas were lightly touched several times, than abdomen was closed again. Group SAP:To 1ml/kg dose retrograde cholangiopancreatography sterile injection of 5%sodium taurocholate, induced SD rat model of acute hemorrhagic necrotizing pancreatitis. Group SP600125:Acute hemorrhagic necrotizing pancreatitis in rats 1 hour before induction to the dose of 35mg/kg by intraperitoneal injection of SP600125. Every group was divided into 3h,6h,12h and 24h stages. We killed the rats at different stages and collected serum, pancreas and lung. The severity of pulmonary edema was expressed by wet/dry. TNF-a, IL-1βin lung and serum amylase were detected in serum. HE staining was used to value the injure of pancreas and lung. Spectrophotometer was used to detected myeloperoxidase in lung tissue and expression of ICAM-1 was tested by immunohistochemistry in lung. Western-blot was used to detect the expression of JNK and PJNK in lung tissue.ResultsIn Group S at 3hr,6hr,12hr and 24hr four different steps serum amylase were 501±85,512±91,518±94 and 532±86; pancreas pathology score was 0; lung pathology score was 0; lung wet/dry ratio were 2.29±0.10,2.22±0.23,2.17±0.11 and 2.31±0.13; the activity of MPO in lung was 1.33±0.56,1.56±0.43,1.51±0.2andl.47±0.48,IL-1βin lung were 80.3±21.2,92.3±23.5,87.3±19.4 and 89.7±22.1; TNF-αin lung were70.6±21.2,67.6±19.8,68.4±19.5 and 74.5±22.6; ICAM-1 score in lung were 0.11±0.01,0.17±0.02,0.14±0.02 and 0.16±0.09 the relative optical density of PJNK was 0.65±0.12. In Group SAP at 3hr,6hr,12hr and 24hr four different steps serum amylase were 2506±215,3701±264,4101±411 and 3212±295; pancreas pathology score was 8.24±0.63,10.65±1.02,14.00±0.96 and 14.09±0.75; lung pathology score was 3.0±0.4,4.7±0.7,7.5±0.3 and 6.5±0.7; lung wet/dry ratio were 3.52±0.42,3.96±0.15,4.53±0.08 and 4.26±0.12; the activity of MPO in lung was 5.64±0.76,6.44±0.77,7.19±0.83 and 7.36±0.71IL-1βin lung were 211.2±19.3,315.4±34.2,399.2±43.2 and 353.4±38.3; TNF-αin lung were 348.5±42.8,433.5±63.4,394.3±56.3 and 365.3±39.9; ICAM-1 score in lung were 0.81±0.22,1.04±0.27,1.44±0.32 and 1.22±0.26; the relative optical density of PJNK was 1.03±0.22,1.24±0.17,1.38±0.21 and 1.28±0.24. In SAP group serum amylase, pancreas pathology score and lung pathology score were significantly higher than those in control group (P<0.05). We successfully set up the sever acute pancreatitis model in rats. Though JNK protein didn't increased in 4 time steps, PJNK in SAP group significantly increased. In Group SP600125 at 3hr,6hr,12hr and 24hr four different lung pathology score were 2.1±0.3,3.2±0.2,6.0±0.7 and 4.0±0.3; lung wet/dry ratio were 3.08±0.09,3.54±0.22,3.35±0.16 and 3.01±0.38; the activity of MPO in lung was 3.36±0.36,4.27±0.52,5.16±0.26and 5.46±0.61,IL-1βin lung were 154.3±11.2,221.4±21.5,289.4±23.4 and 267.7±15.1; TNF-a in lung were 243.8±32.3,317.6±22.8,272.3±20.5 and 225.5±32.6;ICAM-1 score in lung were 0.59±0.18,0.77±0.20,0.84±0.23,0.93±0.31; the relative optical density of PJNK were 0.69±0.14,0.58±0.22,0.85±0.17,0.18±0.05. To compare with SAP group, lung dry/wet ratio, MPO activity, IL-1β, TGF-αand ICAM-1 in SP60025 group significantly decreased. Though JNK protein didn't changed, PJNK protein significantly decreased and lung pathology score improved greatly (P<0.05). We found a significant correlation between the expression of PJNK and lung pathology score (P<0.05). Conclusion:We successful established acute pancreatitis associated lung injure in rat by retrograde pancreatic duct injection of 5%sodium taurocholate.JNK is a very important signal pathway in acute pancreatitis associated lung injure in rat. The activation of JNK is an early incident in the development of SAP. SP600125 can inhibit the activation of JNK, SP600125 can treat the acute pancreatitis associated lung injure and decrease the expression of inflammatory factor.Part two:Mechanisms of and effect of tashinone-ⅡA in acute pancreatitis associated lung injury in ratsAimThe aim of this part to investigate if tanshinone can treat APALI and the mechanism of it.Methods72 SD rats were individed into 3 groups. Group S:sham operated group,24 rats, after opening abdomen duodenum and pancreas were lightly touched several times, than abdomen was closed again. Group SAP:To lml/kg dose retrograde cholangiopancreatography sterile injection of 5%sodium taurocholate, induced SD rat model of acute hemorrhagic necrotizing pancreatitis.. Group tanshinoneⅡA:Acute hemorrhagic necrotizing pancreatitis in rats 30 minutes before induction to the dose of 20mg/kg by intraperitoneal injection of tanshinone IIA. Every group was divided into 3h,6h,12h and 24h stages. We killed the rats at different stages and collected serum, pancreas and lung. The severity of pulmonary edema was expressed by wet/dry. TNF-a, IL-1βin lung and serum amylase were detected in serum. HE staining was used to value the injury of pancreas and lung. Spectrophotometer was used to detected myeloperoxidase in lung tissue and expression of ICAM-1 was tested by immunohistochemistry in lung. Western-blot was used to detect the expression of JNK and PJNK in lung tissue.In Group tanshinone IIA at 3hr,6hr,12hr and 24hr four different steps lung wet/dry ratio were 3.17±0.29,3.24±0.22,3.49±0.26 and 3.21±0.38; the activity of MPO in lung was 20.13±0.39,19.65±1.26 and 24.84±0.71; TNF-αin lung were 253.8±27.3,337.6±22.3,312.3±22.5 and 311.5±32.6; ICAM-1 score in lung were0.63±0.15,0.82±0.19,1.01±0.23 and 0.96±0.31; lung pathology score were 2.3±0.3,3.5±0.2,6.3±0.7 and 5.1±0.3; the relative optical density of PJNK were 1.15±0.08,1.05±0.23,1.34±0.18,0.74±0.16. To compare with SAP group, the expression of PJNK, lung wet/dry ratio, MPO activity, IL-1β, TGF-βand ICAM-1 in lung were significantly decreased, and lung pathology score was also improved (P<0.05).Conclusion:Tanshinone IIA can block the expression of PJNK in lung of SAP rat, which could decrease injure of lung in SAP rat.
|
PDF Full Text Request