艾叶挥发油致急性肝损伤小鼠线粒体结构和功能的变化

Changes of structure and function of mitochondria in liver tissue of mice with acute liver injury induced by volatile oils from Artemisiae Argyi Folium

目的研究艾叶挥发油所致的急性肝损伤小鼠肝组织线粒体结构和功能的变化。方法水蒸气蒸馏法制备艾叶挥发油。健康成年小鼠,雌雄各半,随机分为5组:对照组(n=10),四氯化碳(CCl_4)组(n=10),低(n=10)、中(n=10)和高剂量(n=12)艾叶挥发油组(实验组)。对照组给予橄榄油,CCl_4组给予2.5g·kg^(-1)的CCl_4,低、中、高剂量艾叶挥发油组分别给予1.9,2.3,2.7 g·kg^(-1)的艾叶挥发油。各组均为一次性灌胃,容量为20 mL·kg^(-1)。6h后处死小鼠,检测谷草转氨酶(AST),分离肝组织线粒体,透射电镜观察肝组织线粒体形态,JC-1法检测线粒体膜电位,分光光度法检测线粒体肿胀敏感性,定磷法检测线粒体钠钾腺苷三磷酸酶(Na^+-K^+-ATPase)、钙腺苷三磷酸酶(Ca^(2+)-ATPase)和钙镁腺苷三磷酸酶(Ca^(2+)-Mg^(2+)-ATPase)活力,结果低、中、高剂量艾叶挥发油组血清AST水平分别为(224±56),(251±64),(316±101)U·L^(-1),对照组为(154±27)U·L^(-1),差异有统计学意义(P<0.05)。低、中、高剂量艾叶挥发油组肝组织线粒体膜电位分别为22.06±6.18,21.34±5.99,16.48±4.88,对照组为31.35±9.14(P<0.05)。低、中、高剂量艾叶挥发油组肿胀敏感性分别为(0.68±0.28)×10^(-2),(0.40±0.18)×10^(-2),(0.26±0.10)×10^(-2),对照组为(1.43±0.69)×10^(-2)(P<0.05)。低、中、高剂量艾叶挥发油组Na^+-K^+-ATPase活力分别为2.60±0.60,2.20±0.80,2.10±0.80,对照组为4.40±0.90(P<0.05)。低、中、高剂量艾叶挥发油组Ca^(2+)-ATPase活力分别为2.70±0.50,2.50±0.80,2.20±0.60,对照组为4.20±0.80(P<0.05)。低、中、高3个剂量艾叶挥发油组Ca^(2+)-2Mg^(2+)-ATPase活力分别为2.80±0.60,2.60±0.90,2.50±0.70,对照组为3.90±0.70(P<0.05)。各指标与对照组相比,差异均有统计学意义(P<0.05),且与剂量的变化趋势相关。结论艾叶挥发油可能通过诱发肝组织线粒体的结构和功能障碍而导致小鼠急性肝损伤。

Objective To explore the changes of structure and function of mitochondria in liver tissue of mice with acute liver injury induced by volatile oils from ArtemisiaeArgyi Folium（VOAAF）.Methods VOAAF was obtained by hydrodistillation extraction,specific pathogen free（SPF）Institute of Cancer Research（ICR） mice in half male and half female were randomized into five groups,including control group（n = 10）,model（carbon tetrachloride,CCl4） group（n = 10）,low dose（1.9 g · kg^-1,n = 10）,middle dose（2.3 g·kg^-1,n =10）,and high dose（2.7g · kg^-1,n = 12） VOAAF group（test group）.The biological samples were obtained in 6 h after gavaging equal volume（20 mL·kg^-1） drugs or solvent to mice at single dose.Aspartate transaminase（AST） in serum was detected.Morphological change of mitochondria in liver tissue was observed by transmission electron microscope.Change of mitochondrial membrane potential in liver tissue was examined by JC-1.Change of mitochondrial swelling sensibility in liver tissue was observed by spectrophotometer.Activity change of Na^＋-K^＋-ATPase,Ca^2＋-ATPase and Ca^2＋-Mg^2＋-ATPase in liver tissue were detected by phosphorus determination method.Results The levels of AST in control group and low,middle,high dose test group were（154 ±27）,（224 ±56）,（251 ±64）,（316 ± 101） U · L^-1（P〈0.05）.The mitochondria got swollen and hydropic in some degree,mitochondrial membrane potential were 31.35 ±9.14,22.06 ± 6.18,21.34 ±5.99,16.48 ±4.88（P〈0.05）.The mitochondrial swelling sensibility were（1.43 ± 0.69） × 10^-2,（0.68 ± 0.28） × 10^-2,（0.40× ± 0.18）× 10^-2,（0.26 ±0.10）×10^-2（P〈0.05）.The activity change of Na^＋-K^＋-ATPase were 4.40 ± 0.90,2.60 ± 0.60,2.20 ±0.80,2.10 ± 0.80（P〈0.05）.The Ca^2＋-ATPase were 4.20 ± 0.80,2.70 ± 0.50,2.50 ± 0.80,2.20 ±0.60（P〈0.05）.The Ca^2＋-Mg^2＋-ATPase were 3.90 ±0.70,2.80 ± 0.60,2.60 ± 0.90,2.50 ± 0.70（P〈0.05）.All changes were related to dose variation trend of volatile oils.Conclusion The disorder of the structure and function of mitochondria could be the mechanism of acute liver injury induced by VOAAF in mice.