丹参改善烧伤大鼠肠黏膜上皮细胞线粒体功能的亚细胞水平观察

背景严重烧伤时缺血、缺氧、再灌注造成的氧自由基损伤及感染会对肠黏膜造成损伤.目的从亚细胞水平观察丹参能否改善大鼠烧伤后肠黏膜上皮细胞线粒体呼吸功能及减少氧自由基的产生,从而保护肠黏膜.设计单一样本,随机对照动物实验.单位解放军第三医院烧伤科.材料实验于2001-12/2002-02在解放军第三医院外科研究所完成.选用清洁级SD大鼠96只,随机数字表法分为3组,正常对照组24只,烧伤模型组和丹参治疗组各36只.1 mL丹参注射液含丹参生药1.5 g.方法①烧伤模型组和丹参治疗组大鼠建立烧伤模型,造成20%总体表面积Ⅲ度烫伤.②正常对照组不致伤.③丹参治疗组造模后立即经股静脉缓慢注射丹参注射液1 mL/kg,正常对照组和烧伤模型组注射等量生理盐水.④烧伤模型组、丹参治疗组分别于造模后1,2,6 h各处死12只大鼠,正常对照组分别于对应时间点各处死8只.⑤采集小肠标本,检测肠黏膜上皮细胞线粒体细胞色素aa3、细胞色素C、能荷水平和超氧化物歧化酶的活力.主要观察指标造模后1,2,6 h各组肠黏膜上皮细胞线粒体内细胞色素aa3、细胞色素C、能荷水平及超氧化物歧化酶活力的变化.结果实验选用96只SD大鼠,均进入结果分析.①造模后不同时间点各组肠黏膜上皮细胞线粒体内细胞色素aa3水平的测定与正常对照组比较,烧伤模型组、丹参治疗组造模后2 h内肠黏膜上皮细胞线粒体内细胞色素aa3水平均无明显变化(P均＞0.05),造模后6 h均明显低于正常对照组(P均＜0.05),且丹参治疗组显著高于烧伤模型组(3.16±0.13),(2.56±0.15)μkat/g,P＜0.05].②造模后不同时间点各组肠黏膜上皮细胞线粒体内细胞色素C、能荷水平、超氧化物歧化酶活力的测定与正常对照组比较,烧伤模型组在造模后1,2,6 h肠黏膜上皮细胞线粒体内细胞色素C、能荷水平、超氧化物歧化酶活力均明显下降(P＜0.05或0.01);而丹参治疗组在各时间点以上指标均显著高于烧伤模型组(P＜0.05或0.01).结论丹参能显著提高烧伤大鼠肠黏膜上皮细胞线粒体细胞色素aa3、细胞色素C及能荷水平,明显减少氧自由基的产生,从而改善大鼠烧伤后肠黏膜上皮细胞线粒体的呼吸功能.

Observation of sub-cell level of the effect of danshen on mitochondrial function of epithelial cells in intestinal mucosa of burnt rats

BACKGROUND: Intestinal mucosa was damaged by ischemia and hypoxia during severe burn, and injury and infection of oxygen-derived free radicals during reperfusion.OBJECTIVE: To study on the basis of sub-cell level whether danshen can improve respiratory function of mitochondrion of epithelial cell in intestinal mucosa and decrease the production of oxygen-derived free radicals so as to protect intestinal mucosa of burnt rats.DESIGN: Randomized controlled animal study with single sample.SETTING: Department of Burn, the Third Hospital of Chinese PLA.MATERIALS: The experiment was conducted at the Surgery Institute of the Third Hospital of Chinese PLA from December 2001 to February 2002.Totally 96 SD rats of clean grade were randomly divided into normal control group (n=24), burning model group (n=36) and danshen treatment group (n=36). 1 mL danshen parenteral solution had 1.5 g raw materials.METHODS: ① Rats in the burning model group and danshen treatment group were used to establish grade Ⅲ burning models with 20% scald of total body surface. ② Rats in the normal control group were not injured.③ After modeling, 1 mL/kg danshen parenteral solution was slowing injected into rats in the danshen treatment group through femoral vein, but saline was injected slowly into rats in the normal control group and burning model group. ④ Twelve rats from each of the burning mgdel group and the danshen treatment group were sacrificed at 1, 2 and 6 hours after modeling, and 8 rats in the normal control group were sacrificed at relevant time points respectively. ⑤ Samples of small intestine were collected to measure cytochrome aa3, cytochrome C, level of energy charge and activity of superoxide dismutase of mitochondrion of epithelial cells in intestinal mucosa.MAIN OUTCOME MEASURES: Cytochrome aa3, cytochrome C, level of energy charge and activity of superoxide dismutase of mitochondrion of epithelial cells in intestinal mucosa 1, 2 and 6 hours after modeling.RESULTS: Totally 96 SD rats entered the final analysis. ① Level of cytochrome aa3 of mitochondrion of epithelial cells in intestinal mucosa at various time points after modeling: Levels of cytochrome aa3 did not changed obviously in the burning model group and danshen treatment group 2 hours after modeling as compared with that in the normal control group (P＞ 0.05). Six hours after burning, levels in the burning model group were obviously lower than those in the normal control group (P ＜ 0.05), but those in the danshen treatment group were obviously higher than those in the burning model group (3.16±0.13), (2.5640.15) μkat/g, P ＜ 0.05]. ②Measurements of cytochrome C, level of energy charge and activity of superoxide dismutase of mitochondrion of epithelial cells in intestinal mucosa at various time points after modeling: As compared with those in the normal control group, measurements were decreased obviously in the burning model group 1, 2 and 6 hours after modeling (P ＜ 0.05 or 0.01), but those in the danshen treatment group were obviously higher than those in the burning model group (P ＜ 0.05 or 0.01).CONCLUSION: Danshen can raise the levels of cytochrome aa3, cytochrome C, energy production and SOD, and also reduce the production of oxygen-derived free radicals so as to improve respiratory function of mitochondria of epithelial cells in intestinal mucosa of rats.