高渗盐溶液减轻严重烫伤大鼠早期肺损伤的机制研究

Study on the mechanism of hypertonic salt solution alleviates lung injury of rats at the early stage of severe scald

目的探讨高渗盐溶液（HS）减轻严重烫伤大鼠早期肺损伤的机制。方法将32只雌性SD大鼠按随机数字表法分为假伤组、乳酸钠林格液（LR）组、HS200组（200 mmo/ HS组，1 L 200 mmo/ HS含LR 955 mL和10% NaCl 45 mL）和HS400组（400 mmo/ HS组，1 L 400 mmo/ HS含LR 846 mL和10% NaCl 154 mL），每组8只。将大鼠浸入98℃开水中12 s制备背部30%总体表面积（TBSA）Ⅲ度烫伤模型；假伤组大鼠背部置于37℃水浴中。3个药物干预组于制模后即刻分别经大鼠尾静脉泵入LR、200 mmo/ HS和400 mmo/ HS，按Parkland方案补液；假伤组不予液体复苏。各组大鼠于伤后8 h取腹主动脉血，采用酶联免疫吸附试验（ELISA）检测血清中白细胞介素（IL-6、IL-10、IL-17）的含量；处死大鼠取肺组织，应用紫外分光光度计检测肺组织中丙二醛（MDA）和超氧化物歧化酶（SOD）水平，采用蛋白质免疫印迹试验（Western Blot）检测肺组织中p38丝裂素活化蛋白激酶（p38MAPK）和细胞外信号调节激酶/（ERK/）表达，苏木素-伊红（HE）染色后光镜下观察肺组织病理学改变。结果与假伤组相比，LR组肺组织损伤明显，可见大量炎性细胞浸润、细胞水肿和肺间隔增厚；血清中IL-6、IL-10、IL-17含量以及肺组织MDA含量和p38MAPK、ERK/的磷酸化程度均明显升高，而SOD活性显著下降。与LR组相比，两个HS组肺损伤程度明显减轻，且血清中IL-6、IL-17含量以及肺组织MDA含量和p38MAPK、ERK/的磷酸化程度均明显降低，血清IL-10含量进一步升高，肺组织SOD活性明显增加，且均呈一定剂量依赖性，HS400组各指标与LR组比较差异均有统计学意义〔血IL-6（n/）：3.76±0.12比6.72±0.90，血IL-10（n/）：33.76±3.71比16.77±3.19，血IL-17（n/）：103.52±2.78比124.96±4.96，肺MDA（nmo/g）：5.59±0.24比7.09±0.39，肺SOD（/g）：226.7±3.9比172.7±3.4，肺磷酸化p38MAPK（p-p38MAPK/38MAPK：0.15±0.09比0.35±0.19，肺磷酸化ERK/（p-ERK//RK/：0.27±0.01比0.70±0.01，均P〈0.01〕。结论HS对严重烫伤大鼠肺损伤的保护作用优于LR，且呈一定剂量依赖性，其机制可能是HS能抑制肺组织p38MAPK和ERK/通路活化，提高抑炎因子水平，同时减少促炎因子释放，从而抑制肺组织过度炎症反应和氧化应激损伤。

ObjectiveTo explore the mechanism of hypertonic salt solution （HS） alleviates lung injury of rats at the early stage of severe scald.MethodsThirty-two female Sprague-Dawley （SD） rats were randomly assigned to sham group, lactated Ringer solution （LR） group, HS200 group （200 mmo/ HS group, 1 L 200 mmo/ HS contained 955 mL LR and 45 mL 10% NaCl） and HS400 group （400 mmo/ HS group, 1 L 400 mmo/ HS contained 846 mL LR and 154 mL 10% NaCl）, with 8 rats in each group. A 30% total body surface area （TBSA）Ⅲ degree scalded model was reproduced by scalded on the back with 98℃ boiling water for 12 seconds, whereas those in the sham group were exposed to 37℃ water without liquid resuscitation. Rats in the three drug intervention groups were resuscitated with LR, 200 mmo/ HS and 400 mmo/ HS by caudal vein according to the Parkland formula, respectively. All rats were sacrificed at 8 hours after scald injury to harvest abdominal aorta blood and lung tissues. Interleukins （IL-6, IL-10 and IL-17） in serum were determined by enzyme-linked immunosorbent assay （ELISA）. Samples from the lung tissue were used to measure malondialdehyde （MDA） and superoxide dismutase （SOD） levels by ultraviolet spectrophotometer. Expressions of p38 mitogen-activated protein kinase （p38MAPK） and extracellular regulated protein kinase / （ERK/） in the lung were determined by Western Blot. The lung tissue was stained with hematoxylin and eosin （HE）, and the pathological changes were observed with a light microscope.ResultsCompared with the sham group, the lung tissues in the LR group were damage obviously, which accompanied with more inflammatory cell infiltration, cell edema and pulmonary septum thickening, and the levels of IL-6, IL-10, IL-17 in serum and MDA content, the phosphorylation of p38MAPK and ERK/ in lung tissues were increased whereas the activity of SOD was decreased. Compared with the LR group, the lung injury was significantly alleviated, the levels of IL-6, IL-17 in serum and MDA content and the phosphorylation of p38MAPK and ERK/ were decreased, and the levels of IL-10 and SOD were increased in both HS groups with a dose-dependent manner. There were significant difference in above parameters between HS400 group and LR group [serum IL-6 （n/）： 3.76±0.12 vs. 6.72±0.90, serum IL-10 （n/）： 33.76±3.71 vs. 16.77±3.19, serum IL-17 （n/）： 103.52±2.78 vs. 124.96±4.96, lung MDA （nmo/g）： 5.59±0.24 vs. 7.09±0.39, lung SOD （/g）： 226.7±3.9 vs. 172.7±3.4, lung phosphorylation of p38MAPK （p-p38MAPK/38MAPK： 0.15±0.09 vs. 0.35±0.19, lung phosphorylation of ERK/ （p-ERK//RK/： 0.27±0.01 vs. 0.70±0.01, all P 〈 0.01].ConclusionHS protected against lung injury induced by severe burns in rats with a dose-dependent manner, and it was better than LR, and its possible mechanism was related with reducing the expression of p38MAPK and ERK/ pathway in lung tissue, increasing the level of anti-inflammatory cytokines and decreasing the release of pro-inflammatory cytokines, thus inhibiting excessive inflammation and oxidative stress injury in lung.