氢饱和生理盐水对大鼠重症急性胰腺炎肺损伤保护作用及对JNK信号通路调控的实验研究

发布时间：2018-04-25 10:44

本文选题：重症急性胰腺炎 + 肺损伤　；参考：《泸州医学院》2014年硕士论文

【摘要】：目的：探讨静脉注射氢饱和生理盐水对牛磺胆酸钠诱导的大鼠重症急性胰腺炎相关性肺损伤是否具有保护作用并从JNK信号通路的调控探讨其作用机制。方法：1.氢饱和生理盐水的制备：利用氢气发生器生产高压氢气，再将高压氢气通入氯化钠溶液中达到饱和浓度即可，现制现用。2.大鼠重症急性胰腺炎相关性肺损伤模型的建立及处理：将54只健康SD雄性大鼠随机分成假手术组（Sham组）、模型组（SAP+NS组）和氢水处理组（SAP+H2组），各组再分成6h、12h、24h三个时间点，每个时间点6只大鼠。Sham组大鼠开腹翻动胰腺数次后随即关腹，不作其他处理，SAP+NS组和SAP+H2组以5%牛磺胆酸钠（1ml/kg）经胆胰管开口逆行注入（0.1ml/min）建立模型，在建模成功后1h经尾静脉分别注射等量的生理盐水或氢饱和生理盐水(5ml/kg)。各组分别在6h、12h、24h三个时间点处死大鼠，收集血清、肺组织及胰腺组织。用酶联免疫吸附试验(ELISA)法检测血清TNF-α、IL-1β含量；分光光度计法测定肺组织中髓过氧化物酶（MPO）的活性；荧光定量PCR法检测肺组织中TNF-α-mRNA、IL-1β-mRNA的表达；Western blot法检测肺组织中P-JNK蛋白的表达；用肺组织湿干重比,反映肺脏水肿程度；并对胰腺组织、肺组织进行HE染色病理学检查。以上所得数据采用均数±标准差（X—±S）表示，，运用SPSS17.0统计软件进行统计学分析；样本均数的比较采用方差分析；两者的相关性采用直线相关和回归分析，P＜0.05，则差异有统计学意义。结果：（1）SAP+NS组和SAP+H2组血清中TNF-α含量、肺组织中TNF-αmRNA表达水平、肺组织中P-JNK蛋白的表达、肺组织湿干重比，在6h、12h、24h各个时间点均高于Sham组（P0.05），SAP+H2组与SAP+NS组比较，SAP+H2组在各个时间点均低于SAP+H2组（TNF-α含量：204.1±8.5VS215.3±6.0，P0.05；122.4±10.3VS263.2±7.4，P0.05；84.1±8.6VS288.0±5.6，P0.05。TNF-α mRNA表达水平：2.81±0.09VS3.94±0.20，P0.05；2.51±0.11VS4.40±0.24，P0.05；1.77±0.06VS6.00±0.37，P0.05。P-JNK蛋白的表达：11.29±0.01VS11.77±0.01，P0.05；10.59±0.02VS12.73±0.01，P0.05；9.43±0.01VS14.12±0.01，P0.05。肺组织湿干重比：3.7±0.2VS4.2±0.2，P0.05；3.3±0.3VS4.9±0.2，P0.05；3.2±0.2VS4.6±0.3，P0.05。）。（2）SAP+NS组和SAP+H2组血清中IL-1β含量、胰腺和肺组织病理评分、肺组织中MPO活性、肺组织中IL-1β mRNA表达水平，在6h、12h、24h各个时间点均高于Sham组（P0.05），SAP+H2组与SAP+NS组比较，6h时无统计学差异，在12h、24h时SAP+H2组均低于SAP+NS组（IL-1β含量：80.3±8.3VS215.4±10.4，P0.05；59.3±8.2VS254.3±8.6，P0.05。胰腺和肺组织病理评分：7.5±1.1VS9.5±0.4，P0.05；7.3±0.4VS9.8±0.3，P0.05。4.6±0.2VS5.1±0.2，P0.05；3.8±0.3VS6.3±0.4，P0.05。MPO活性：3.3±0.2VS4.7±0.2，P0.05；3.1±0.1VS4.9±0.2，P0.05。IL-1β mRNA表达：1.77±0.16VS1.97±0.11，P0.05；1.29±0.03VS2.92±0.26，P0.05）。（3）Person相关性分析表明肺组织中P-JNK蛋白的表达与肺组织损伤程度存在相关性。结论：1.经尾静脉注射氢饱和生理盐水对APALI有一定治疗保护作用。2.氢饱和生理盐水可能是通过其选择性抗氧化作用抑制JNK细胞信号通路的激活来实现对APALI的保护治疗作用。
[Abstract]:Objective: To explore the protective effect of intravenous hydrogen saturated saline on severe acute pancreatitis associated lung injury induced by sodium taurocholate in rats and to explore its mechanism from the regulation of JNK signaling pathway. Methods: 1. hydrogen saturated saline was prepared by hydrogen generator to produce high pressure hydrogen and then high pressure hydrogen. The establishment and treatment of the present model of severe acute pancreatitis associated lung injury in.2. rats was established and treated with saturated concentration of Sodium Chloride Solution. 54 healthy SD male rats were randomly divided into sham operation group (group Sham), model group (group SAP+NS) and hydrogen water treatment group (SAP+ H2 group), each group was then divided into 6h, 12h, 24h three time points, each time. The rats in the 6 rats of the 6 rats were operated on the pancreas to turn off the pancreas for several times and no other treatment was done. The SAP+NS group and the SAP+H2 group were injected with 5% sodium taurocholate (1ml/kg) through the biliary pancreatic duct opening retrograde injection (0.1ml/min) to establish the model. After the successful modeling, 1H was injected with equal amount of normal saline or hydrogen saturated saline (5ml/kg). The rats were killed at the three time points of 6h, 12h and 24h. The serum, lung and pancreas tissues were collected. The serum TNF- alpha and IL-1 beta content was detected by enzyme linked immunosorbent assay (ELISA); the activity of myeloperoxidase (MPO) in lung tissue was measured by spectrophotometer; the expression of TNF- alpha -mRNA, IL-1 beta -mRNA in lung tissue was detected by the fluorescence quantitative PCR method. Ern blot method was used to detect the expression of P-JNK protein in lung tissue, the ratio of wet dry weight to lung tissue was used to reflect the degree of pulmonary edema, and the pathological examination of pancreas tissue and lung tissue was carried out by HE staining. The above data were expressed with mean mean standard deviation (X - S) and SPSS17.0 statistical software was used for statistical analysis. The comparison of the average number of samples was used. The correlation was linear correlation and regression analysis. The difference was statistically significant in P < 0.05. Results: (1) the content of TNF- alpha in serum of SAP+NS and SAP+H2 group, the level of TNF- alpha mRNA in lung tissue, the expression of P-JNK protein in lung tissue, the ratio of wet dry weight in lung tissue, and higher than Sham group (P0.05) at all time points in 6h, 12h and 24h. In group AP+H2 and group SAP+NS, group SAP+H2 was lower than group SAP+H2 at all time points (TNF- alpha content: 204.1 + 8.5VS215.3 +, P0.05; 122.4 + 10.3VS263.2 + 7.4, P0.05; 84.1 + 8.6VS288.0 + 5.6, P0.05.TNF- alpha mRNA expression level: 2.81 + + 0.20, 2.51 + + 0.24, 1.77 + 0.37 The expression of NK protein: 11.29 + 0.01VS11.77 + 0.01, P0.05; 10.59 + 0.02VS12.73 + 0.01, P0.05; 9.43 + 0.01VS14.12 + 0.01, P0.05. lung tissue wet dry weight ratio: 3.7 + 0.2VS4.2 + 0.2, P0.05; 3.3 + 0.3VS4.9 + 0.2, P0.05; 3.2 + 0.2VS4.6 + 0.3. The activity of MPO in lung tissue and the expression level of IL-1 beta mRNA in lung tissue were higher than those in group Sham (P0.05) at all time points in 6h, 12h and 24h. There was no statistical difference between SAP+H2 group and SAP+NS group. Score: 7.5 + 1.1VS9.5 + 0.4, P0.05; 7.3 + 0.4VS9.8 + 0.3, P0.05.4.6 + 0.2VS5.1 + 0.2, P0.05; 3.8 + 0.3VS6.3 + 0.4, P0.05.MPO activity: 3.3 + 0.2VS4.7 + 0.2, P0.05; 3.1 + 0.1VS4.9 + 0.2. There is a correlation between the expression of P-JNK protein in the fabric and the degree of lung tissue damage. Conclusion: 1. the hydrogen saturated saline injected into the tail vein has certain protective effect on APALI..2. hydrogen saturated saline may be the protective and therapeutic effect of the JNK cell signaling pathway by its selective antioxidant activity.

【学位授予单位】：泸州医学院
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R576

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