专题式教学法在感染病学教学中的实践与认识

专题式教学法是将过去以单病种授课的教学模式改为以疾病共性特征为专题而进行的讲座式教学的模式。与传统式教学方法相比具有很大的优越性，不仪能够系统地讲授相关疾病的基础知识，还能够培养学生的临床思维能力，既注重理论学习，也强调实际应用，是一种很适用于感染病学的教学方法。     

可回复性永生化逆转录病毒载体pLNCTIGIox的构建与表达

目的:为建立可回复性永生化肝细胞株,构建带有筛选基因NeoR、报道基因EGFP及重组位点loxP的可回复性永生化逆转录病毒载体.方法:将2.1kbSV40T亚克隆到pIRES2-EGFP的相应位点,再酶切下3.4kbSV40T-IRES-EGFP片段插入逆转录病毒载体pLNCX2的两个相同酶切位点之间,构成新载体pLNCTIG.最后,将线性化pLNCTIG和loxP双链片段混合连接.取连接产物转化高效感受态DH5α细胞.菌落PCR和酶切法鉴定阳性克隆.转染包装细胞PT67并在荧光显微镜下观察绿色荧光蛋白的表达.扩大培养稳定转染的细胞,免疫组化染色检测SV40T基因的表达.结果:随机挑取19个克隆,3个电泳分离出约1.1kb阳性条带.取上述阳性克隆进行酶切鉴定,结果仅出现一条约9.5kb的条带,证明该克隆为阳性重组体.转染PT67细胞24h后,荧光显微镜下可见散在多处绿色荧光.免疫组化染色显示细胞胞核呈阳性染色.结论:成功构建并表达可回复性永生化逆转录病毒载体pLNCTIGlox.     

人胚胎骨髓间充质干细胞在大鼠体内分化为肝细胞的研究

目的：观察人胚胎骨髓间充质干细胞(MSC)在受体大鼠体内的分化演变，研究人胚胎骨髓间充质干细胞在活体大鼠体内转化为人肝细胞的可行性。方法：将人胚胎MSC移植到肝损伤合并自身肝细胞再生抑制大鼠的脾脏。术后分时段取脾脏进行免疫组化染色，了解人白蛋白(ALB)、人角蛋白(CK8)、人CK18、人CK19的表达情况。结果：人胚胎MSC移植后10d大鼠脾脏的人CK8、人CK18、人CK19免疫组化染色可见阳性细胞；移植后15d大鼠脾脏组织内人ALB免疫组化染色可见阳性细胞。结论：人胚胎MSC能够在自身肝细胞再生抑制的肝损伤大鼠脾脏内分化为肝细胞。     

蛋白转导结构域介导乙型肝炎病毒聚合酶末端蛋白重链可变区抗体体外抑制病毒的复制

Objective To study a functional variable fragment of heavy chain(VH)antibody against the terminal protein(TP)region of hepatitis B virus(HBV)polymerase introduced by human immunodeficiency virus Tat protein transduction domain(TAT)and the inhibitive activity of TAT-VH on the replication of HBV in vitro.Methods The gene encoding TAT-VH was cloned into prokaryotic expression vector pET28a(+).Recombinant plasmid was transduced into E coli BL21(DE3)LysS,then the protein was expressed and purified.The purified TAT-VH fusion protein was added into HepG2.2.15 cell culture.The transduction efficiency was evaluated by indirect fluorescence assay(IFA).The cytotoxicity of TAT-VH was detected by Methabenzthiazuron(MTT)assay.HBV DNA level in HepG2.2.15 cell culture was measured using quantitative polymerase chain reaction(PCR).The data were analyzed by one-factor analysis of variance and t test.Results TAT-VH fusion protein was successfully expressed and purified.It was confirmed by IFA and MTT assay that TAT-VH was introduced into HepG2.2.15 cells and the cell growth was not affected.The level of HBV DNA in supernatant of HeDG2.2.15 cell culture with 5 000 nmol/L TAT-VH was(1.211±0.132)lg copy/mL,which was significantly lower than control group[(5.325±0.041)lg copy/mL,t=72.91,P＜0.05].Meanwhile,the level of intracellular HBV DNA was(3.521±0.411)lg copy/mL,which was significantly lower than control group[(8.532±0.132)lg copy/mL.t=28.41,P＜0.05].Conclusion The HBV replication is inhibited by anti-TP TAT-VH antibodies in vitro,which provides valuable experimemal basis for developing therapy of HBV infection with intracellular antibody.     

蛋白转导结构域介导乙型肝炎病毒聚合酶末端蛋白重链可变区抗体体外抑制病毒的复制

Objective To study a functional variable fragment of heavy chain(VH)antibody against the terminal protein(TP)region of hepatitis B virus(HBV)polymerase introduced by human immunodeficiency virus Tat protein transduction domain(TAT)and the inhibitive activity of TAT-VH on the replication of HBV in vitro.Methods The gene encoding TAT-VH was cloned into prokaryotic expression vector pET28a(+).Recombinant plasmid was transduced into E coli BL21(DE3)LysS,then the protein was expressed and purified.The purified TAT-VH fusion protein was added into HepG2.2.15 cell culture.The transduction efficiency was evaluated by indirect fluorescence assay(IFA).The cytotoxicity of TAT-VH was detected by Methabenzthiazuron(MTT)assay.HBV DNA level in HepG2.2.15 cell culture was measured using quantitative polymerase chain reaction(PCR).The data were analyzed by one-factor analysis of variance and t test.Results TAT-VH fusion protein was successfully expressed and purified.It was confirmed by IFA and MTT assay that TAT-VH was introduced into HepG2.2.15 cells and the cell growth was not affected.The level of HBV DNA in supernatant of HeDG2.2.15 cell culture with 5 000 nmol/L TAT-VH was(1.211±0.132)lg copy/mL,which was significantly lower than control group[(5.325±0.041)lg copy/mL,t=72.91,P＜0.05].Meanwhile,the level of intracellular HBV DNA was(3.521±0.411)lg copy/mL,which was significantly lower than control group[(8.532±0.132)lg copy/mL.t=28.41,P＜0.05].Conclusion The HBV replication is inhibited by anti-TP TAT-VH antibodies in vitro,which provides valuable experimemal basis for developing therapy of HBV infection with intracellular antibody.     

重型肝炎患者血浆内皮素变化的意义

重型肝炎患者血浆内皮素变化的意义陈耀凯Ｓｕｂｊｅｃｔｈｅａｄｉｎｇｓｈｅｐａｔｉｔｉｓ，ｖｉｒａｌ，ｈｕｍａｎ／ｂｌｏｏｄ；ｈｅｐａｔｉｔｉｓ，ｖｉｒａｌ，ｈｕｍａｎ／ｄｉａｇｎｏｓｉｓ；ｅｎｄｏｔｈｅｌｉｎ１／ｂｌｏｏｄ主题词肝炎，病毒性，人...     

干扰素对慢性乙型肝炎患者血清Ⅲ型前胶原的影响

用α干扰素治疗慢性乙型肝炎３１例，并检测了治疗前后血清Ⅲ型前胶原水平。结果：治疗线疗程结束后血清Ⅲ型胶胶原含量显著降低，而对照组无明显变化，表明干扰素可减轻慢性乙型肝炎患者内胶原增生的程度及活动性。     

慢性重型肝炎患者肝组织内胆管增生的免疫组织化学研究

在慢性重型肝炎患者的肝组织切片中,除肝细胞大面积坏死、纤维组织增生、炎症细胞浸润及不同程度的肝细胞再生等病理特征外,还常有明显的胆管增生现象,但这种胆管增生的本质及意义尚不清楚。新近提出肝组织内胆管增生与肝干细胞     

肝衰竭/重型肝炎的研究进展

本文对肝衰竭／重型肝炎的新进进行综述。在定义上倾向于将急性肝衰竭与慢性肝衰竭及脑病型与非脑病型分开；对新近提出的终末期肝病模型（MELD）预后判断公式进行评价；应用拉米夫定（LAM）治疗伴乙型肝炎病毒（HBV）复制的肝衰竭有效，但肝炎肝硬化患者因不当停药所致肝功能不全患者有所增多；LAM加乙型肝炎病毒免疫球蛋白（HBIg)预防肝移植后HBV感染有效，但其逸逸株有增多趋势；当前国际上将肝性脑病（HE）分为三种类型，A型为急性肝衰竭相关HE（ALFA－HE），B型为不伴有内在肝病的门体分离，C型指在慢性肝病／肝硬化基础上发生的HE。     

乏力、黄疸、发热、声嘶、呼吸困难

病史摘要患者 ,男 ,2 8岁 ,农民。因咽痛、乏力、纳差 1个月 ,巩膜黄染、畏寒、高热伴声嘶 12ｄ ,呼吸困难 3ｄ于 2 0 0 1年 3月14日 2 1∶30入院 ,住院 16ｈ后死亡。患者 1个月前受凉后感咽痛 ,经对症处理后好转 ,继而出现明显全身乏力、纳差、厌油、尿黄。入院前 15ｄ解黏液稀黄便 5～ 7次 /ｄ ,无腹痛及里急后重。入院前 12ｄ出现巩膜、皮肤黄染伴恶心、呕吐 ,畏寒 ,体温多在 38～ 4 0℃ ,咽痛加重 ,声音嘶哑。入院前 8ｄ感腹胀、尿少。入院前 7ｄ在外院诊断为“重型肝炎” ,给予“还原型谷胱甘肽、人血白蛋白、血浆、氧氟沙星、氨苄西林…