【作者】 范巧云；

【导师】 刘群红； 李朝品；

【作者基本信息】 安徽理工大学 ， 病原生物学， 2007， 硕士

【摘要】 目的观察NAPP体内外抗乙型肝炎病毒(HBV)的效果,为临床开发新的抗病毒药物提供理论依据。方法体外实验采用HepG2.2.15细胞株为模型,在细胞培养液中加入不同浓度的NAPP溶液进行混合培养,通过四甲基偶氮唑盐(MTT)比色分析法检测药物的细胞毒性,9天后应用酶联免疫吸附法(ELISA)、实时荧光定量PCR(FQ-PCR)技术检测上清液中的乙型肝炎病毒表面抗原(HBsAg)、乙型肝炎病毒e抗原(HBeAg)和乙型肝炎病毒DNA(HBV-DNA)的载量,作为药物抗HBV的观察指标,并与抗HBV药物拉米夫定进行对照研究,对NAPP体外抗HBV作用进行评价;体内实验采用先天感染鸭乙肝模型,分设模型对照组、拉米夫定组和NAPP组,各组按20mg/(kg·d)连续灌胃给药10d,检测用药前后鸭血清中乙肝病毒脱氧核糖核酸(DHBV-DNA)的动态变化,实验结束时取新鲜肝组织,HE染色观察肝脏组织病理学变化,对NAPP体内抗HBV的作用进行评价。结果体外实验,NAPP浓度在1mg/ml时,对细胞无明显毒性;NAPP在所稀释的各浓度下,对HepG2.2.15细胞分泌HBsAg、HBeAg均有一定的抑制作用,且抑制作用具有剂量依赖关系;NAPP对HBsAg、HBeAg的治疗指数(TI)分别大于188.7和64.5。体内实验,NAPP组在连续灌胃5天、10天时,DHBV-DNA滴度显著下降,与模型组比较差异有显著性意义(T5,P＜0.05;T10,P＜0.01),与拉米夫定组比较,差异无显著性意义(P＞0.05);病理检查发现NAPP组肝细胞点状坏死明显减轻,与模型对照组比较,差异有显著性意义(P＜0.05)。结论NAPP在体内外均具有抗乙肝病毒作用,并具有明显的抗炎、保肝作用。更多还原

【Abstract】 Objective To observe the inhibitory effect of NAPP on Hepatitis B virus both in vitro and in vivo. It may lead a theoretical foundation on developing of a new kind of anti-virus medicine.Methods The cell 2.2.15 line was taken as cell model. The different concentration of NAPP water solution were respectively added into cell culture fluid of 2.2.15 cell strain. The cell toxicity of NAPP was determined with MTT method and the inhibitory effect of NAPP on HBsAg and HBeAg secreted by 2.2.15 cells was evaluated on the ninth day with ELISA method. Real-time fluorescence quantitative PCR was used for detection of HBV-DNA in the culture medium. At same time, we took lamivudine as a control to evaluate the effect on anti-HBV. Duck model of congenital infection of HBV was used as animal model. The ducks were divided into three groups at random and the each group ducks was given the drug at the 20mg/(kg·d) for 10 successive days. The dynamic changes of serum duck HBV-DNA titer was detected by dot-hybridization test before treatment and after treatment. Fresh liver tissue ware sheared. The pathological changes of the duck liver were also examined by HE staining.Results In the cell model, NAPP had no obvious cell toxicity at the concentration of 1mg/ml. NAPP had obvious restrain effect on HBsAg and HBeAg at the different concentration and had a dose-dependent. The therapeutic index of NAPP was 188.7 for HBsAg and 64.5 for HBeAg. In the animal model, the optic density value of DHBV-DNA in ducks serum got down markedly after giving medication with NAPP for 5 days and 10 days, there were remarkable difference (T5, P<0.05; T10, P<0.01). Compared with lamivudine control group, there was no significant difference (P>0.05). Spotty of hepatocyte in NAPP group was improved more obviously than that in model group.Conclusion NAPP exerts a certain inhibitory effect on Hepatitis B Virus both in vitro and in vivo. NAPP can protect duck liver against inflammation.更多还原