Relationship between HBV viremia level of pregnant women and intrauterine infection:neated PCR for detection of HBV DNA

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血红素加氧酶-1表达对HBV复制的反向调控作用

目的评价血红素加氧酶-1（HO-1）对HBV复制的调控作用。方法采用分子克隆的方法分别构建人H（）_1真核表达载体——pH0和HO-1RNA干扰质粒——pHi，同HBV可复制性克隆pHBV1．3体外共转染人肝癌细胞系Huh-7，检测HBV抗原分泌情况和HBV相关mRNA含量。利用HBV急性感染小鼠模型，腹腔注射钴原卟啉（CoPP）IX和锌原卟啉（ZnPP）IX分别诱导和抑制HO-1表达，检测血清HO～1水平、HBV效价，免疫组织化学染色观察HBcAg在肝细胞内表达情况。分别在细胞水平和动物体内水平，评价HO-1表达对HBV复制的调控作用。结果同空载体共转染细胞相比，pHO共转染后HO-1的分泌水平明显上调（P〈0．01），HBsAg／HBeAg的分泌受到抑制（均P〈0．01）；pHi共转染后HO-1的分泌水平下降（P〈0．01），而HBsAg／HBeAg分泌增加（P均〈0．01）；但各组HBV相关RNA水平无明显差异。CoPPIX注射后诱导小鼠血清H0—1高表达（P％0．01），ZnPPIX则有效抑制了HO-1表达（P％0．01）。同对照组相比，CoPPIX组小鼠血清HBVDNA含量降低，肝脏内HBcAg阳性染色信号也随之明显减弱（P均〈0．01）；而ZnPPIX组小鼠血清HBVDNA含量增加，肝脏内HBcAg表达明显增强（均P〈0．01）。结论上调HO-1表达可有效抑制HBV复制，而下调其表达有利于HBV复制，且其可能是在转录后环节上发挥抗HBV作用。     

Clinical evaluation (phase I) of a combination of two human monoclonal antibodies to HBV: safety and antiviral properties

Treatment of chronic hepatitis B virus (HBV) infection with interferon alfa and lamivudine is characterized by lack of viral clearance, loss of response, or emergence of drug-resistant mutants. Thus, new and multiple drug approaches are needed. We have developed two fully human monoclonal antibodies, directed against different epitopes of hepatitis B surface antigen (HBsAg) that bind to all major HBV subtypes. A phase I clinical study was conducted to evaluate the safety, tolerability, and efficacy of a mixture of these two monoclonal antibodies, HBV-AB(XTL). A total of 27 chronic HBV patients were enrolled. In part A of the study 15 patients in 5 cohorts received a single intravenous infusion of antibodies with doses ranging from 0.26 mg (260 IU) to 40 mg (40,000 IU). All patients completed 16 weeks of follow-up. In the second part of the study (part B), 12 patients in 4 cohorts received 4 weekly infusions of 10, 20, 40, or 80 mg each of HBV-AB(XTL) and were followed for 4 additional weeks. Administration of antibodies was well tolerated. Patients administered doses at an Ab:Ag molar ratio of 1:2 to 1:20 showed a rapid and significant decrease in HBsAg to undetectable levels, with a corresponding reduction of HBV-DNA levels. In part B, HBV-AB(XTL) induced a significant reduction in both HBsAg and HBV-DNA levels repeatedly after administration. In conclusion, these data suggest that HBV-AB(XTL) binds HBV particles and reduces serum viral titers and HBsAg levels. HBV-AB(XTL) could be combined with other monotherapies that are currently used to treat HBV carriers.     

流体动力学法乙型肝炎病毒感染动物模型的建立

目的:建立一种简便、有效、稳定的乙型肝炎病毒(HBV)感染的动物模型,观察该模型动物体内不同时间点各种HBV标志物的表达情况．方法:以流体动力学法尾静脉注射BALB／c 小鼠pcDNA3．1-HBV,1 wk内检测各种HBV 标志物,时间分辨免疫荧光分析法(IFMA)检测血清中HBsAg,HBeAg,抗HBs,抗HBe,抗 HBc,荧光定量PCR法(FQ-PCR)检测血清HBV DNA,免疫组织化学法检测肝组织HBsAg和 HBcAg．结果:成功建立一种急性HBV感染动物模型, 第1天血清中HBsAg表达达高峰,后逐渐降低, HBeAg表达量少,分别在第4、5、7天检测到抗HBc,抗HBe,抗HBs,d1 HBV DNA滴度亦达高峰,后渐下降,免疫组织化学示HBsAg呈胞质内弥漫性分布,HBcAg亦主要为胞质型分布．结论:以流体动力学法建立的HBV模型是一种新型有效的HBV感染动物模型,能稳定较高水平表达大部分HBV标志物．     

高压水动力法建立表面抗原表达缺失的乙型肝炎转染小鼠模型及初步评估

目的探讨HBsAg在小鼠体内模型中对宿主免疫应答调节及病毒感染慢性化中的作用。方法在已有高压水动力法转染pAAV-HBV1.2质粒建立乙肝小鼠模型的基础上,通过点突变技术引入终止密码子至pAAV-HBV1.2质粒上的HBsAg读码框,构建HBsAg无法转录的突变质粒,再利用该质粒构建HBsAg表达缺失的乙肝小鼠模型。通过比较原始质粒建立的乙肝模型小鼠,检测分析外周血中HBV抗原与DNA等病毒感染指标,比较肝脏病变和免疫细胞浸润情况。结果成功构建了HBsAg缺失的乙肝模型小鼠:在乙肝模型小鼠和HBsAg缺失突变的模型小鼠外周血中均能检测到4 COI/mL水平的HBeAg,两组小鼠没有差异,但只有乙肝小鼠体内能检测到102 COI/mL以上的HBsAg和103至106拷贝/mL的HBV DNA,HBsAg缺失的小鼠中无法检测到HBsAg和HBV DNA;动态监测显示HBeAg在乙肝和HBsAg缺失突变的两组小鼠中的阳性率均逐步降低,在HBsAg缺失突变组中降低的更明显,第8周时仅有20%的小鼠为HBeAg阳性,而此时未突变乙肝组的阳性率为60%;免疫组织化学结果显示:在阴性对照小鼠的肝脏结构较完整,未见聚集的浸润免疫细胞,而乙肝和HBsAg缺失的乙肝小鼠肝小叶结构均松散,乙肝小鼠肝脏中有聚集的浸润性免疫细胞,HBsAg缺失的小鼠肝脏缺少免疫细胞的聚集,却存在严重的肝细胞毛玻璃样变性。结论成功构建了HBsAg缺失的乙肝转染小鼠模型;并发现HBsAg在HBV完整病毒颗粒包装和出胞,以及肝脏病变和免疫细胞浸润过程中发挥了重要的作用。     

Replication and gene expression of mutant hepatitis B virus in a transgenic mouse containing the complete viral genome with mutant s gene

AIM: To establish the transgenic mouse line harbouring complete hepatitis B virus (HBV) genome with mutant s gene (adr subtype). METHODS: Transgenic mice were generated by microinjecting HBV genome into fertilized eggs. Integration, expression, replication of HBV gene and histological changes in transgenic mice were estimated by genomic DNA PCR, serum DNA PCR, Southern blot, ELISA, HE staining, immunohistochemistry and transmission electron microscopy. Transgenic mice with HBsAg positive in serum were bred and analyzed. RESULTS: A total of 288 eggs survived from microinjections were transplanted into the oviducts of 13 pseudopregnant mice and 49 pups were produced. Twenty-six mice were identified to have the integrated HBV gene. Serum HBsAg and HBeAg were detected in 2 of 43 mice. HBsAg and HBcAg in cytoplasm or nuclei of hepatocytes were detected in 10 mice. Founders with HBsAg in serum were named lineages G145R-15 and G145R-18. Of the 16 F1 offsprings generated by G145R-15 founder, 12 were positive for HBV genome with PCR, 10 were positive for HBsAg and HBcAg with immunohistochemistry and 7 were positive for HBsAg and HBeAg with ELISA. Only 1 of 8 F1 offsprings generated by G145R-18 founder was survived and it was detected positive for HBV genome, HBsAg, HBcAg and HBeAg. Both of the two lineages had some pathological characteristics of mild chronic hepatitis B in the liver, such as swelling of hepatocytes and focal hepatocellular necrosis and parenchymal lymphomononuclear cell infiltrate. CONCLUSION: Transgenic mice harbouring HBV with mutant s gene can be generated. The HBV genes are integrated in the transgenic mice genome and can be expressed, replicated, packaged and excreted. HBV DNA can be stably transmitted in the transgenic mice.     

商陆抗病毒蛋白在急性人乙型肝炎病毒感染小鼠体内的抑制作用

目的 探讨商陆抗病毒蛋白(PAP)在急性HBV感染小鼠模型体内抗HBV的效果.方法 通过尾静脉注射具有复制能力的HBV真核表达质粒来建立小鼠急性HBV感染模型.根据注射后第1天小鼠血清HBsAg和HBeAg水平,从35只小鼠中选出24只进行配对,分为PAP治疗组(腹腔注射0.25 mg/kg PAP)和对照组.于PAP注射前、注射后第1、3、5、7天,时间分辨免疫荧光分析法检测小鼠血清HBsAg、HBeAg和抗-HBc水平,荧光定量PCR检测小鼠血清HBV DNA水平.注射后第7天HE染色检测肝组织病理变化,免疫组织化学检测肝组织HBsAg、HBeAg表达.采用配对t检验进行统计学分析.结果 与对照组相比,在PAP腹腔注射后第1、3、5天对HBsAg的抑制率分别为23%(t=116.3,P<0.05)、47%(t=38.2,P<0.05)、68%(t=23.7,P<0.05),对HBeAg的抑制率分别为36%(t=34.2,P<0.05)、55%(t=61.6,P<0.05)、83%(t=98.8,P<0.05),对HBV DNA的抑制率分别为70.7%(t=6.6,P<0.05)、86.9%(t=5.9,P<0.05)、95.2%(t=36.6,P<0.05)、95.3%(t=19.7,P<0.05).结论 0.25 mg/kg剂量的PAP在急性HBV感染小鼠模型体内对血清和肝组织的HBsAg、HBeAg的表达及HBV DNA的复制均有明显的抑制效果.     

日本血吸虫重叠 HBV 感染小鼠模型的建立

目的：建立日本血吸虫重叠 HBV 感染小鼠模型,观察其生物学表现。方法日本血吸虫尾蚴感染6～8周龄雌性 BALB／cJ 小鼠,8周后,通过高压尾静脉注射 B 基因型 HBV 可复制性克隆,分别在注射后第3、7天,检测血清HBV DNA、HBsAg、HBeAg、ALT、透明质酸（HA）水平;检测 HBcAg 在肝细胞内表达情况和虫卵肉芽肿形成情况。比较重叠感染相关生物学指标和单独感染组的不同。结果重叠感染组小鼠肝组织内可见 HBcAg 在虫卵肉芽肿周围肝细胞内表达;可以正常分泌 HBsAg 和 HBeAg 以及子代病毒;其血清 ALT 水平低于 HBV 感染组,高于血吸虫感染组;HA 含量相对明显升高。结论成功建立了日本血吸虫重叠 HBV 感染小鼠模型,为后续研究此类重叠感染提供了良好的小动物实验平台。     

Inhibition of hepatitis B virus replication by APOBEC3G in vitro and in vivo

AIM: To investigate the effect of APOBEC3G mediated antiviral activity against hepatitis B virus (HBV) in cell cultures and replication competent HBV vector-based mouse model. METHODS: The mammalian hepatoma cells Huh7 and HepG2 were cotransfected with various amounts of CMV-driven expression vector encoding APOBEC3G and replication competent 1.3 fold over-length HBV. Levels of HBsAg and HBeAg in the media of the transfected cells were determined by ELISA. The expression of HBcAg in transfected cells was detected by western blot. HBV DNA and RNA from intracellular core particles were examined by Northern and Southern blot analyses. To assess activity of the APOBEC3G in vivo, an HBV vector-based model was used in which APOBEC3G and the HBV vector were co-delivered via high-volume tail vein injection. Levels of HBsAg and HBV DNA in the sera of mice as well as HBV core-associated RNA in the liver of mice were determined by ELISA and quantitative PCR analysis respectively. RESULTS: There was a dose dependent decrease in the levels of intracellular core-associated HBV DNA and extracellular production of HBsAg and HBeAg. The levels of intracellular core-associated viral RNA also decreased, but the expression of HBcAg in transfected cells showed almost no change. Consistent with In vitro results, levels of HBsAg in the sera of mice were dramatically decreased. More than 1.5 log 10 decrease in levels of serum HBV DNA and liver HBV RNA were observed in the APOBEC3G-treated groups compared with the control groups. CONCLUSION: These findings indicate that APOBEC3G could suppress HBV replication and antigen expression both in vivo and in vitro, promising an advance in treatment of HBV infection.     

Inhibitory effect of emodin and Astragalus polysaccharide on the replication of HBV

AIM: To evaluate the anti-viral effect of emodin plus Astragalus polysaccharide (APS) in hepatitis B virus (HBV) transgenic mice.METHODS: Sixty HBV transgenic mice (HBV TGM) whose weight varied between 18 and 24 g were randomly divided into 3 groups, with 20 mice in each group. Group A was the normal control, where the mice were treated with physiological saline; group B was the positive control where the mice were treated with lamivudine solution (100 mL/kg per day). Group C was the experimental group where the mice were treated with physiological saline containing emodin and APS (57.59 mg/kg per day and 287.95 mg/kg per day, respectively). The mice were treated daily for 3 wk.After 1 wk recovery time, the mice were sacrificed and serum as well as liver tissues were collected for ELISA and histological examination.RESULTS: After 21 d treatment, HBV DNA levels in group B and group C significantly declined when compared with group A ( P < 0.05). However, a significant increase in HBV DNA content was observed in group B, whereas this phenomenon was not observed in group C. A reduction in the contents of HBsAg,HBeAg and HBcAg in the mice from group B and C was observed when compared with group A.CONCLUSION: Emodin and APS have a weak but persistent inhibitory effect on HBV replication in vivo,which may function as a supplementary modality in the treatment of hepatitis B infection.     

Intrahepatic distribution of hepatitis B virus antigens in patients with and without hepatocellular carcinoma

AIM: To study the intrahepatic expression of hepatitis B surface antigen(HBs Ag) and hepatitis B core antigen(HBc Ag) in chronic hepatitis B patients with and without hepatocellular carcinoma. METHODS: A total of 33 chronic hepatitis B patients(mean age of 40.3 ± 2.5 years), comprising of 14 HBe Ag positive and 19 HBe Ag negative patients; and 13 patients with hepatitis B virus related hepatocellular carcinoma(mean age of 49.6 ± 4.7 years), were included in our study. Immunohistochemical staining for HBc Ag and HBs Ag was done using standard streptavidin-biotin-immunoperoxidase technique on paraffin-embedded liver biopsies. The HBc Agand HBs Ag staining distributions and patterns were described according to a modified classification system. RESULTS: Compared to the HBe Ag negative patients, the HBe Ag positive patients were younger, had higher mean HBV DNA and alanine transaminases levels. All the HBe Ag positive patients had intrahepatic HBc Ag staining; predominantly with "diffuse" distribution(79%) and "mixed cytoplasmic/nuclear " pattern(79%). In comparison, only 5% of the HBe Ag-negative patients had intrahepatic HBc Ag staining. However, the intrahepatic HBs Ag staining has wider distribution among the HBe Ag negative patients, namely; majority of the HBe Ag negative cases had "patchy" HBs Ag distribution compared to "rare" distribution among the HBe Ag positive cases. All but one patient with HCC were HBe Ag negative with either undetectable HBV DNA or very low level of viremia. Intrahepatic HBc Ag and HBs Ag were seen in 13(100%) and 10(77%) of the HCC patients respectively. Interestingly, among the 9 HCC patients on anti-viral therapy with suppressed HBV DNA, HBc Ag and HBs Ag were detected in tumor tissues but not the adjacent liver in 4(44%) and 1(11%) patient respectively. CONCLUSION: Isolated intrahepatic HBc Ag and HBs Ag can be present in tumors of patients with suppressed HBV DNA on antiviral therapy; that may predispose them to cancer development.     

乙型肝炎病毒复制调控元件对HBV DNA疫苗诱导的免疫应答

目的研究乙型肝炎病毒（HBV）复制调控元件增强子Ⅰ（ENHⅠ）及前S2（Pre-S2）抗原基因对HBV DNA疫苗诱导的免疫应答的影响。方法采用常规聚合酶链反应（PCR）从HBV adr亚型全基因DNA序列中分别扩增HBsAg、PreS2-HBsAg、HBsAg-ENHI和PreS2-HBsAg-ENHⅠ基因片段，重组到VR1012载体中，构建4种HBV DNA疫苗，转染HepG2细胞并免疫Balb／C小鼠。通过细胞免疫化学、酶联免疫分析（ELISA）、酶联免疫斑点试验（ELISPOT）等方法检测其在HepG2细胞内的表达及小鼠的体液及细胞免疫应答。结果转染的HepG2细胞表达相应的目的蛋白．ENHⅠ及Pre-S2抗原基因均可增强HBV DNA疫苗转染HepG2细胞表达HBsAg；免疫接种小鼠后第2周产生抗-HBs及HBsAg特异性细胞毒T淋巴细胞（CTL），Pre—S2抗原基因可增强HBV DNA疫苗免疫Balb／C小鼠诱导的抗-HBs及HBsAg特异性CTL的产生，ENHⅠ基因对免疫应答无影响。结论ENHI及Pre—s2抗原基因均可增强HBVDNA疫苗转染HepG2细胞表达HBsAg．Pre-S2抗原基因可增强HBVDNA疫苗免疫Balb／C小鼠引起的免疫应答。     

慢性乙型肝炎患者肝组织HBV cccDNA、总HBV DNA与血清HBV DNA的相关性及其与临床的关系

目的探讨慢性乙型肝炎患者肝组织HBV共价闭合环状DNA（cccDNA）、肝组织总HBV DNA（HBV tDNA）与血清HBV DNA之间的相关性及其与临床的关系。方法 78例慢性乙型肝炎患者入选本研究。肝组织β- globinDNA、HBV cccDNA和HBV tDNA采用实时荧光定量PCR方法检测,平均每个肝细胞HBV cccDNA和HBV tDNA含量（拷贝/cell）=HBV cccDNA（实测值）/β-globin DNA（实测值）和HBV tDNA（实测值）/β3-globin DNA（实测值）,肝组织HBV cccDNA和HBV tDNA含量的计算单位定义为log₁₀拷贝/10⁶cell;采用实时荧光定量PCR、ELISA法检测血清HBVDNA和HBV标志物;采用免疫组织化学方法检测肝细胞中HBsAg和HBcAg的表达。统计分析采用pearson相关分析及t检验。结果 （1）肝组织HBV cccDNA与HBV tDNA定量呈正相关（r=0.696,P<0.001）;肝组织HBV cccDNA与血清HBV DNA定量呈正相关（r=0.304,P<0.01）;肝组织HBV tDNA与血清HBV DNA定量呈正相关（r=0.341,P<0.01）;（2）肝细胞内HBcAg定性检测阳性患者的血清HBV DNA定量明显高于阴性患者,且差异有统计学意义（P<0.05）;肝细胞内HBcAg定性检测阳性患者与阴性患者的肝组织HBV cccDNA和HBV tDNA定量差异均无统计学意义和（P均>0.05）;（3）肝细胞内HBsAg定性检测阳性患者的血清HBV DNA定量明显高于阴性患者,且差异有统计学意义（P<0.05）;肝细胞内HBsAg定性检测阳性患者与阴性患者的肝组织HBV cccDNA和HBV tDNA定量差异均无统计学意义（P>0.05）;（4）HBeAg（+）/抗-HBe（-）患者血清HBV DNA定量明显高于HBeAg（-）/抗-HBe（+）患者,且差异有统计学意义（P<0.05）;HBeAg（+）/抗-HBe（-）患者肝组织HBV cccDNA和HBV tDNA定量与HBeAg（-）/抗-HBe（+）患者比较差异均无统计学意义（均P>0.05）;（5）肝组织HBV cccDNA、HBV tDNA以及血清HBV DNA三者与肝脏炎症活动度及纤维化程度均无显著相关性（P>0.05）。结论 （1）血清HBV DNA定量结果并不一定能完全反映患者肝组织中HBV cccDNA和HBV tDNA含量,尤其在血清HBV DNA<500拷贝/mL时,肝组织中仍存在HBV cccDNA和HBV tDNA,且含量大小不等。（2）肝细胞内HBcAg定性检测阳性或者HBsAg定性检测阳性患者的血清HBV DNA定量均明显高于阴性患者;而两者的肝组织HBV cccDNA和HBV tDNA定量均没有显著差异;（3）HBeAg（+）/抗-HBe（-）患者血清HBV DNA定量明显高于HBeAg（-）/抗-HBe（+）患者,而两者的肝组织HBV cccDNA和HBV tDNA均没有显著差异;（4）肝组织HBV cccDNA、HBV tDNA及血清HBV DNA与肝脏炎症活动度和纤维化程度均无显著相关性。     

几种常见乙肝模式患者乙肝病毒前S1抗原检测的临床价值

目的：探讨乙肝前S1抗原（pre-S1）在几种常见乙肝模式患者鉴别诊断中的临床价值。方法：对353例乙肝两对半不同模式的患者血清及50例全阴对照组血清，同步进行pre-S1及HBV-DNA检测。结果：在HB—sAg（＋）、HBeAg（＋）、HBcAb（＋）模式组，pre—S1检出率为85．7％，HBV-DNA检出率为87．8％；在HBsAg（＋）、HBcAb（＋）、HBeAb（＋）模式组，pre-S1检出率为53．9％，HBV-DNA检出率为49．4％；在HBsAg（＋）、HBeAg（＋）模式组，pre—S1检出率为78．3％，HBV—DNA检出率为80．0％；在HBsAg（＋）、HBcAb（＋）模式组，pro-S1检出率为28．6％，HBV—DNA检出率为24．5％；在HBsAg（＋）、HBeAg（＋）、HBeAb（＋）模式组，pre-S1检出率为16．7％，HBV—DNA检出率为0．0％；在其他模式组，pre-S1和HBV-DNA检出率均为0．0％。结论：乙肝前S1抗原可作为HBV在体内复制的可靠标志。     

乙型肝炎病毒全基因重组腺病毒感染免疫研究

目的 用腺病毒载体将乙型肝炎病毒(HBV)基因组导入小鼠体内,研究HBV表达产物诱发的机体免疫反应.方法 用AdEasy系统构建含HBV基因组的重组腺病毒AdHBV-WT,感染HepG2细胞后,分别用ELISA和western blot检测细胞培养上清液和细胞裂解物中的HBsAg、HBeAg和HBcAg.AdHBV-WT经在293细胞中扩增、纯化和浓缩后,经滴鼻途径感染BALB/C小鼠,检测体液和细胞免疫反应.结果 重组腺病毒AdHBV-WT感染HepG2细胞后,有效地表达出HBV的各种抗原.重组腺病毒感染小鼠可诱导抗-HBc的产生.3H-TdR摄入实验表明,AdHBV-WT可诱发小鼠产生HBcAg特异性的T细胞应答.结论 HBV全基因重组腺病毒在体外可表达HBV的各种抗原,并可诱发小鼠产生HBcAg特异性的体液免疫和细胞免疫.     

复方六月雪体外抗HBV的实验研究

目的观察复方六月雪(CLYX)体外抗乙型肝炎病毒(HBV)的作用。方法在最大无毒浓度(TC0)基础上观察不同浓度药物作用于HepG2.2.15细胞,分别在第4天和8天收集细胞培养上清液,采用实时荧光定量PCR法检测上清液HBV DNA的含量,采用ELISA法测定上清液HBsAg和HBeAg的滴度。结果无毒浓度的复方六月雪在HepG2.2.15细胞培养中可有效地抑制细胞HBV DNA的复制及HBsAg和HBeAg的分泌。结论CLYX在体外有显著的抗HBV的作用。     

乙型肝炎病毒转基因小鼠用于研究治疗乙型肝炎药物初探

目的：建立乙型肝炎病毒（HBV）转基因小鼠模型,探讨该模型能否用于抗HBV药物的验证,筛选,方法：原核显微注射法制备HBV转基因小鼠,用巢式PCR,Southern杂交,免疫组化学,ELISA等检测HBV基因的整合与表达,筛选60只血清HBVDNA,HBsAg阳性的小鼠,随机分为6组,3组为实验组,分别给予拉米夫定灌胃,胸腺肽腹腔注射,HBVS基因DNA疫苗,多点肌肉注射,另3组分别为生理盐水对照组,结果：显微注射产生的HBV转基因小鼠中有HBV的复制和表达,拉米夫定,胸腺肽,DNA疫苗用于转基因小鼠后,均能使血清HBVDNA阴转,以拉米夫定阴转率最高,并可看出作用开始,持续时间及反跳情况,结论：拉米夫定,胸腺肽,DNA疫苗可抑制转基因小鼠HBV的复制,HBV转基因小鼠有可能成为抗HBV药物验证,筛选的有力工具。     

急性乙型肝炎病毒感染小鼠模型的建立

目的采用尾静脉液压法建立小鼠急性HBV感染的动物模型。方法以液压法将具有复制能力的HBV质粒pAAV-HBV1.2通过尾静脉注射到免疫功能正常的BALB/c小鼠体内,注射后第1、2、4、6、8d,分别采用改良赖氏法、时间分辨免疫荧光法、实时荧光定量PCR检测小鼠血清中谷丙转氨酶(ALT)、HBsAg、HBeAg、抗HBs、抗HBe、HBV DNA的水平,免疫组化检测肝组织HBsAg、HBcAg的表达。结果16只小鼠注射pAAV-HBV1.2后,有14只(85.7%)小鼠在注射后第1d血清中可检测到HBsAg,小鼠血清中HBsAg和HBeAg水平在第1d达高峰,之后逐渐下降,第8d均未能检测到。小鼠血清中HBV DNA在第2d达高峰,之后仍维持在较高水平,至第8d时为1.9×104copies/mL。至第8d肝组织中可见约5%的HBcAg阳性肝细胞和2%的HBsAg阳性肝细胞。结论采用尾静脉液压法成功的建立了小鼠急性HBV感染的动物模型。