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沙漠干热环境猪创伤失血性休克肾损伤氧化应激及Caspase-3变化研究

The changes of oxidative stress and caspase-3 in swine with traumatic hemorrhagic shock in dry-heat environment of desert

摘要:

目的 探讨沙漠干热环境下猪创伤失血性休克后继发性肾损伤和氧化应激及凋亡相关因子Caspase-3的变化研究.方法 48头长白仔猪随机(随机数字法)分为2组,建立常温下和干热环境下猪创伤失血性休克模型,常温环境失血性休克组(RTS),干热环境创伤失血性休克组(DHS),分别在常温和沙漠干热环境中暴露3 h(T0,n=6),T1(休克模型成功50 min,n=6)、T2(休克模型成功100 min,n=6)、T3(休克模型成功150 min,n=6),各组在相应的时间点收集血液检测血尿素氮(BUN)和肌酐(creatinine);收集尿液检测中性粒细胞明胶酶相关脂蛋白(NGAL);收集肾组织行形态学检查及肾小管评分,收集肾组织检测过氧化氢酶(CAT)、超氧化物歧化酶(SOD)及丙二醛(MDA),利用蛋白印迹法检测肾组织Caspase-3蛋白的表达情况.结果 在沙漠干热环境暴露3 h(T0组)与常温环境暴露组比较,肾损伤指标、抗氧化及Caspase-3均升高,但差异无统计学意义(P>0.05);DHS组从T1时间点开始, NGAL、CAT及SOD均明显升高,与常温休克组比较均差异有统计学意义(P<0.05或P<0.01),而尿素氮及肌酐在T2时间点差异有统计学意义(P<0.05).T3时间点,DHS组Caspase-3蛋白含量与RTS相比差异有统计学意义(P<0.01).NGAL与MDA的相关性分析,发现RTS组和DHS组NGAL的表达量与MDA的表达量均成正相关(r常温=0.935,r干热=0.858,P<0.01).与RTS组相比,光镜下发现,DHS组相继出现肾小囊扩张,内有变性脱落上皮细胞,近曲小管上皮脱落,间质水肿;电镜下发现,线粒体形态成多形性,内质网内褶增宽.结论 当干热环境合并创伤失血性休克时,干热环境可加重肾脏的损伤,可能是通过降低肾组织抗氧化酶含量和提高肾组织Caspase-3活性来促进肾组织细胞凋亡来实现.提示抗氧化应激及细胞凋亡可能是预防沙漠干热环境创伤失血性休克继发性肾损伤的重要途径.

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Objective To study the changes of oxidative stress and caspase-3 in swine with traumatic hemorrhagic shock in dry-heat environment of desert.Methods A total of 48 Landrace small swine were randomly(random number)divided into 2 groups(n=24 in each group), and then the traumatic hemorrhagic shock was established in room temperature environment and in dry-heat environmentin swine.Dry-heat environment traumatic hemorrhagic shock group (DHS), which was made in an artificial experiment cabin mimic the reality included swine exposed in the dry-heat environment of desert for 3 h (T0, n=6), T1 (50 min after shock modeling, n=6), T2 (100 min after shock modeling, n=6), T3 (150 min after shock modeling, n=6).At each interval, blood sample was collected to detect urea nitrogen (BUN) and creatinine, urine sample was collected to detect neutrophil gelatinase-associated lipoprotein (NGAL), kidney tissue samples were collected to evaluate renal morphological and tubular scores, as well as to detect catalase (CAT), superoxide dismutase (SOD) and malondialdehyde (MDA).Western blot was used to detect the level of caspase-3.Traumatic hemorrhagic shock group of room temperature environment (RTS) was established and variety of assays were carried out as same as those deteced in the dry-heat environment group.Results Compared with the room temperature environment exposed group,kidney damage index, antioxidant and caspase-3 were increased in desert dry-heat environment exposed for 3 h group, but there were no statistically significant difference(P> 0.05).And from T1 then on, the levels of NGAL, CAT and SOD in DHS groups were increased which were significant different from those in RTS group (P<0.05 or P<0.01).There were significant differences in BUN and creatinine at T2 between two groups(P<0.05).At T3, caspase-3 protein content in DHS group was significantly different from that in RTS group (P<0.01).Correlation analysis showed that the NGAL level was correlated with the levels to MDA (rRTS=0.935, rDHS =0.858, P<0.01) in RTS group and DHS group.Compared with RTS group, renal tissue under light microscope showed that Bowman appeared dilated with degeneration and exfoliated epithelial cells, proximal tubule epithelial shedding, and interstitial edema in DHS group.Electron microscope showed that mitochondria became pleomorphic, endoplasmic reticulum with fold broadening.Conclusions When traumatic hemorrhagic shock happened in the desert dry-heat environment, desert dry-heat environment can aggravate kidney damage, possibly by reducing the renal tissue antioxidant enzyme content and increase renal tissue caspase-3 activity to promote renal tissue apoptosis.Antioxidant stress and apoptosis may be an important role in the prevention of the secondary kidney injury induced by traumatic hemorrhagic shock in dry-heat environment.

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