摘要
目的:探讨玉竹提取物A对小鼠免疫性肝损伤的保护作用,探讨其发挥此作用的可能的机制。方法:①实验于2004-06/10在锦州医学院免疫实验室完成。选用健康雄性昆明小鼠50只,普通级。将50只健康雄性昆明小鼠随机分为5组:正常对照组、模型对照组、玉竹提取物A0.5g/kg组、玉竹提取物A1g/kg组、玉竹提取物A2g/kg组,每组10只。②玉竹提取物A是玉竹的干燥根茎经水醇法提取而得。③正常对照组和模型对照组小鼠腹腔注射生理盐水,玉竹提取物A0.5,1,2g/kg组小鼠分别腹腔注射相应浓度的玉竹提取物A,每次每只0.5mL,3次/d,连续注射4d。最后一次注射后8h,除正常对照组注射生理盐水外,其余各组均尾静脉注射20mg/kg的刀豆蛋白A0.5mL,制备免疫性肝损伤模型。④注射刀豆蛋白A8h后摘眼球取血低温保存,取肝待行光镜检查,以观察肝脏组织病理学改变(常规苏木精-伊红染色),取脾待行淋巴细胞增殖实验(采用四甲基偶氮唑盐法)和流式细胞术以检测T淋巴细胞亚群。各组小鼠血清谷丙转氨酶活性检测按购自南京建成生物研究所的谷丙转氨酶试剂盒说明进行。⑤组间计量资料差异比较采用方差分析。结果:昆明小鼠50只均进入结果分析。①肝脏组织病理学改变:模型对照组可见明显免疫性肝损伤病理改变。玉竹提取物A3个实验组小鼠肝损伤病理改变均不同程度的好于模型对照组。②玉竹提取物A对血清谷丙转氨酶活性的影响:模型对照组血清谷丙转氨酶活性明显高于正常对照组(t=13.8,P<0.01),玉竹提取物A3个实验组血清谷丙转氨酶活性低于模型对照组,尤以玉竹提取物A2g/kg组与模型对照组差异最明显(t=5.62,P<0.01),且存在剂量依赖关系。③玉竹提取物A对T淋巴细胞转化增值的影响:刀豆蛋白A刺激体外培养的各组脾淋巴细胞,模型对照组的刺激指数明显高于正常对照组(t=9.02,P<0.01),玉竹提取物A3个实验组的刺激指数明显低于模型对照组(t=6.99～11.06,P<0.01),且存在剂量依赖关系。④玉竹提取物A对脾组织T淋巴细胞亚群的影响:各组,CD4+T,CD8+T淋巴细胞的数量和比例差异不明显(P>0.05)。结论:①玉竹提取物A具有抑制肝细胞破坏及改善肝脏微循环的作用。②玉竹提取物A可以显著抑制T淋巴细胞的转化增殖,并呈剂量依赖关系,玉竹提取物A发挥肝保护作用的一个机制可能就是显著抑制T淋巴细胞的转化增殖,减少肝损伤细胞因子的释放,抑制活化增殖的T淋巴细胞对肝细胞的直接细胞毒作用。③在肝损伤的早期可能只是大量免疫细胞聚集于肝脏并且各种分泌及破坏功能增强,而非其数量明显增加。
AIM： To investigate the protective effect of extraction A of polygonatum odoratum （EA-PAOA） on the immune hepatic injury in mice and study its protective mechanisms.
METHODS：（1）The experiment was carried out in the immunological laboratory of Jinzhou Medical College from June to October 2004. Fifty Kunmlng mice （common grade） were selected and randomized into five groups with ten mice in each group： normal control group, model control group, test 1 （EA-PAOA 0.5 g/Kg） group, test 2 （EA-PAOA 1 g/Kg） group and test 3 （EA-PAOA 2 g/Kg） group. （2）The EA-PAOA was extracted from dry root and petiol with water-ethanol method （30 mL/L ethanol）. （3）Saline was injected into abdominal cavity of mice in the normal control group and the model control group. Mice in the test 1, test 2 and test 3 groups were given with the corresponding EA-PAOA through intraperitoneal injection, respectively, once 0.5 mL into every mice, 3 times per day, for 4 days. Eight hours after the last injection, the mice in the model control and three test groups, respectively, received intravenous injections of concanavalin A （20 mg/kg） 0.5 mL, except these in the normal control group were injected with saline. Then the immune liver injury mice were prepared. （4）Blood gained from the picked eyeball were collected at eight hours after intravenous injections of concanavalin A, and then kept in hypothermia. The liver was reserved for routine light microscopy so as to observe the pathological changes of liver tissue （hematoxylin-eosin staining）. The spleen was aimed to do lymphocyte proliferation experiment and T-cell proliferactive activity tests with MTT, The level of alanine aminotransferase was tested with Kit made from Nanjing Jiancheng Bioengineering Institute. （5））The analysis of variance was applied to the compare of computation data.
RESULTS： Totally 50 Kumming mice were involved in the result analysis. （1） Pathological change of liver tissue： Obvious immune hepatic injury appeared in the mice of the model control group. However, the three test groups had improved in different extent dramatically as compared with the model control group. （2）Effect of EA-PAOA on semm ALT： The level of serum ALT in the model control group was higher significantly than that in the normal control group （t=13.8, P 〈 0.01）. The level of serum ALT in the three test groups was lower than that in the model control group, and had most significant difference between the test 3 group and the model control group, especially （t=5.62,P 〈 0.01）, and existed dose-dependent relation. （3）Effect of EA-PAOA on conversion and proliferation of T-lymphocytes： The concanavalin A stimulated the lienal lymphocytes of every group cultured in vitro. The stimulate-index was higher in the model control group than that in the normal conti＇ol group dramatically （t=9.02,P 〈 0.01）. It was lower markedly in the three test groups than that in themodel control group （t=6.99-11.06,P 〈 0.01）, and had dose-dependent relation. （4）Effect of EA-PAOA on T-cell of lienal tissue： The number of CD： T and CD8^＋ T iymphocytes in spleen and the proportion were insignificant （P 〉 0.05）.
CONCLUSION： （1） The EA-PAOA can inhibit the liver injury and improve the liver microcirculation. （2）The EA-PAOA can obviously inhibit T-cell proliferactive activity and dose-dependent relation. One of anti-liver injury mechanisms is that the EA-PAOA can obviously inhibit T-cell proliferactive activity to decrease the level of cell factor, and then all the effects lead to inhibit the injury by the multiplation of T-cell. （3）In the early stage, the liver injury is only due to lots of immunocytes collecting in liver and its secretion and destroyed function increased dramatically, but the number does not increase markedly.
出处
《中国临床康复》
CSCD
北大核心
2006年第3期99-101,共3页
Chinese Journal of Clinical Rehabilitation
