解毒饮体外抗乙肝病毒及体内抗肝损伤的实验分析-experimental analysis of jiedu decoction on anti-hepatitis b virus in vitro and anti-liver injury in vivo.docxVIP

中 文 摘 要目 的：探讨中药复方解毒饮保肝护肝及抗乙型肝炎病毒的作用，为中 医药复方治疗慢性乙型肝炎提供理论和实验依据。方 法：体外实验应用血清药理学方法制备大鼠药物血清及正常血清， 将不同血清作用于乙型肝炎病毒基因组转染的 HepG2.2.15 细胞，采用 MTT 法检测药物血清细胞毒性，ELISA 法测定细胞上清液中 HBsAg、 HBeAg 的滴度，FQ-PCR 法检测上清液中 HBV-DNA 的含量；体内实验 中采用 CCL4 灌胃制备小鼠慢性肝损伤模型，每日解毒饮灌胃给药，并以 水飞蓟素（利加隆）作为对照，采用全自动生化分析仪检测血清总胆红 素（TBIL）、丙氨酸氨基转氨酶（ALT）、门冬氨酸氨基转移酶（AST） 水平，光镜下观察肝脏组织形态学变化，并按试剂盒方法检测肝组织匀 浆中 MDA、SOD 的含量。结果：体外实验中解毒饮药物血清对 HepG2.2.15 细胞的破坏百分率极 低，能明显抑制细胞上清液中 HBsAg、HBeAg 的分泌及 HBV-DNA 的含 量，抑制作用呈量效关系；体内实验中模型组小鼠血清 TBIL、ALT 及 AST 显著升高，肝组织匀浆中 MDA 含量明显升高，SOD 活性明显下降， 光镜下肝小叶结构紊乱，肝细胞弥漫疏松变性，可见多灶状坏死。解毒 饮组 TBIL、ALT 及 AST 水平明显降低，肝匀浆中 MDA 含量明显减少， SOD 活性显著升高，光镜下病变轻微，肝细胞弥漫轻度疏松变性，未见 坏死。结 论：中药复方解毒饮能保肝降酶，保护肝细胞，改善肝脏组织的损 伤，对慢性肝损伤有明显的保护作用；解毒饮药物血清对 HepG2.2.15 细 胞毒性很低，安全性好；解毒饮药物血清能有效抑制细胞 HBsAg、HBeAg 的分泌及 HBV-DNA 的合成，具有直接抗 HBV 的作用。关 键 词： 解毒饮；抗乙型肝炎病毒；HepG2.2.15 细胞；血清药理学； 慢性肝损伤Experimental Study of Jieduyin Decoction on Anti-hepatitis B virus in vitro and Anti-liver injury in vivoABSTRACTObjectives:To provide theoretical and experimental evidences for treating chronic hepatitis B virus with compound Jieduyin decoction.Methods:Human HepG2.2.15 cell contaminated by hepatic B virus genome was set up as the cell model. The control group was given distilled water twice daily while the treatment group with Jieduyin decoction. Then, the serum was given to HepG2.2.15 cell after having processed into herbal solution. The cell toxicity of herbal serum was determined by MTT method. The cell upper clear parts of cell culture were collected to determine the titer of HBsAg and HBeAg by ELISA method as well as the content of HBV-DNA by FQ-PCR method.The mouses were made into models of hepatic injury by gavaging CCL4. Then 60 healthy Kunming mouses were randomly divided into the control group, the model group, the Jieduyin group and the Lijialong group. 24 hours later, the serum extracted from the blood in eyeball to test the level of TBIL, ALT, and AST by ACA machine, and observe the morphologic changes of liver tissues under light microscope as well as