Role of RNA secondary structure in emergence of compartment specific hepatitis B virus immune escape variants

AIM To investigate the role of subgenotype specific RNA secondary structure in the compartment specific selection of hepatitis B virus(HBV)immune escape mutations.METHODS This study was based on the analysis of the specific observation of HBV subgenotype A1 in the serum/plasma,while subgenotype A2 with G145R mutation in the peripheral blood leukocytes(PBLs).Genetic variability found among the two subgenotypes was used for prediction and comparison of the full length pregenomic RNA(pgRNA)secondary structure and base pairings.RNA secondary structures were predicted for 37℃using the Vienna RNA fold server,using default parameters.Visualization and detailed analysis was done using RNA shapes program.RESULTS In this analysis,using similar algorithm and conditions,entirely different pgRNA secondary structures for subgenotype A1 and subgenotype A2 were predicted,suggesting different base pairing patterns within the two subgenotypes of genotype A,specifically,in the HBV genetic region encoding the major hydrophilic loop.We observed that for subgenotype A1 specific pgRNA,nucleotide 358U base paired with 1738A and nucleotide 587G base paired with 607C.However in sharp contrast,in subgenotype A2 specific pgRNA,nucleotide 358U was opposite to nucleotide 588G,while 587G was opposite to 359U,hence precluding correct base pairing and thereby lesser stability of the stem structure.When the nucleotides at 358U and 587G were replaced with 358C and 587A respectively(as observed specifically in the PBL associated A2 sequences),these nucleotides base paired correctly with 588G and 359U,respectively.CONCLUSION The results of this study show that compartment specific mutations are associated with HBV subgenotype specific alterations in base pairing of the pgRNA,leading to compartment specific selection and preponderance of specific HBV subgenotype with unique mutational pattern.

八珍汤合化积丸加减联合核苷类药物治疗“正虚血瘀型”乙肝肝硬化继发脾功能亢进的效果

目的:探讨八珍汤合化积丸加减联合核苷类药物治疗“正虚血瘀型”乙肝肝硬化继发脾功能亢进的效果。方法:选取2020年6—12月在武威市凉州医院就诊的西医诊断为“乙型肝炎后肝硬化继发脾功能亢进”、中医辨证为“正虚血瘀型”的96例患者作为研究对象,根据随机数表法分为对照组和观察组,各48例。对照组给予替诺福韦酯,观察组给予替诺福韦酯联合八珍汤合化积丸,比较两组白细胞(WBC)、血小板(PLT),凝血酶原活动度(PTA)、脾脏厚度、门静脉内径变化、肝脏硬度值及临床疗效。结果:治疗前、治疗3个月,两组WBC、PLT、PTA水平及脾脏厚度、门静脉内径比较,差异无统计学意义(P>0.05);治疗6个月,两组WBC、PLT、PTA水平较治疗前升高,脾脏厚度、门静脉内径较治疗前降低,且观察组优于对照组,差异有统计学意义(P<0.05)。两组治疗前肝脏FibroScan弹性值比较,差异无统计学意义(P>0.05);观察组治疗6个月肝脏FibroScan值小于对照组,差异有统计学意义(P<0.05)。观察组总有效率为87.5%,高于对照组的62.5%,差异有统计学意义(P<0.05)。结论:八珍汤和化积丸加减联合核苷类药物治疗“正虚血瘀型”乙肝肝硬化继发脾功能亢进,不仅能改善患者的临床症状,还能提高WBC、PLT,改善PTA,在降低肝脏硬度值及脾脏厚度和门静脉内径方面效果显著。

Persistent occult hepatitis B virus infection:Experimental findings and clinical implications

Hepatitis B virus (HBV) is a highly pathogenic virus that causes chronic liver diseases in millions of people globally. In addition to a symptomatic, serologically evident infection, occult persistent HBV carriage has been identified since nucleic acid amplification assays of enhanced sensitivity became introduced for detection of hepadnaviral genomes and their replicative intermediates. Current evidence indicates that occult HBV infection is a common and long-term consequence of resolution of acute hepatitis B. This form of residual infection is termed as secondary occult infection (SOI). The data from the woodchuck model of HBV infection indicate that exposure to small amounts of hepadnavirus can also cause primary occult infection (POI) where virus genome, but no serological makers of exposure to virus, are detectable, and the liver may not be involved. However, virus replicates at low levels in the lymphatic system in both these forms. We briefly summarize the current understanding of the nature and characteristics of occult hepadnaviral persistence as well as of its documented and expected pathological consequences.

Pathogenesis of occult chronic hepatitis B virus infection

Occult hepatitis B infection(OBI) is characterized by hepatitis B virus(HBV) DNA in serum in the absence of hepatitis B surface antigen(HBsAg) presenting HBsAg-negative and anti-HBc positive serological patterns.Occult HBV status is associated in some cases with mutant viruses undetectable by HBsAg assays;but more frequently it is due to a strong suppression of viral replication and gene expression.OBI is an entity with world-wide diffusion.The failure to detect HBsAg,despite the persistence of the viral DNA,is due in most cases to the strong suppression of viral replication and gene expression that characterizes this"occult"HBV infection;although the mechanisms responsible for suppression of HBV are not well understood.The majority of OBI cases are secondary to overt HBV infection and represent a residual low viremia level suppressed by a strong immune response together with histological derangements which occurred during acute or chronic HBV infection.Much evidence suggests that it can favour the progression of liver fibrosis and the development of hepatocellular carcinoma.

Immunogenicity and reactogenicity of a recombinant hepatitis B vaccine in subjects over age of forty years and response of a booster dose among nonresponders

AIM:The study was initiated to evaluate the reactogenicity and immunogenicity of a recombinant hepatitis B vaccine in age group >40 years and to study the response of a single booster dose in primary non-responders to the hepatitis B vaccination. METHODS:A total of 102 volunteers without markers of hepatitis B infection (negative for HBsAg,anti-HBc antibody, HBeAg and anti-HBs antibody) received 20μg of recombinant HB vaccine intramuscularly at 0,1,and 6 months.Anti HBs titers were evaluated by a quantitative Elisa kit at 90 and 210 days.A booster dose of 20μg HB vaccine was given after 6 months of the 3^(rd) vaccine dose to the 15 non- responders and anti-HBs titers were measured after i month. RESULTS:Seroprotection (anti-HBs GMT^3 10 IU/L) was achieved in 85.3 % (87/102) volunteers.The mean GMT titers of the vaccine responders was 136.1 IU/L.Of the seroprotected individuals,there were 32.4% (33/102) hyporesponders (anti- HBs titers <10-99 mIU/ml) and 52.9% (54/102) were responders (anti-HBs titers >100 IU/L).All the non-responders (15/15) responded to a single dose of the booster dose of recombinant HB vaccine and their mean anti-HBs antibody titers were more than 100.5 mIU/ml after the booster dose. CONCLUSION:Recombinant hepatitis B vaccine offers good seroprotection in the age group >40 years and has a good safety profile.A single booster dose after 6 months in primary non-responders leads to good seroprotective anti-HBs antibody titers.However,larger population based studies are needed to evaluate the role of a booster dose in selected group of non-responders and whether such an approach will be cost effective.

Percutaneous liver biopsy： retrospective study of primary and secondary hepatic lymphoma in twenty-one patients

BACKGROUND： Hepatic lymphoma （HL） is categorized as primary and secondary hepatic lymphoma （PHL and SHL）. This disorder can present as hepatic mass or mass-like lesion. Chemotherapy often is the first line treatment for patients with HL. Thus, an accurate pre-management histological diagnosis is essential to potentially improve clinical outcomes. The present study was to explore the prevalence of HL in ultrasound guided liver biopsies for hepatic mass or mass-like lesions, to investigate HL associated clinicopathological features, to raise the awareness of early recognition and proper diagnosis of this entity, and to assess specimen adequacy in needle core biopsy. METHODS： Twenty-one cases of HL were enrolled. Clinical and pathological characteristics were evaluated, quality of biopsies was assessed and pertinent literature was reviewed. RESULTS： HL was diagnosed in 0.94% of 2242 liver biopsy cases with ambiguous clinical presentation, laboratory tests and image studies. There were two cases of PHL （0.09%）, and nineteen cases of SHL （0.85%）. Histopathologically, diffuse large B-cell lymphoma was the most common type, followed by B-cell lymphoma not otherwise specified, T-ceU lymphoma, Hodgkin＇s lymphoma, and B-cell chronic lymphocytic leukemia/small lymphocytic lymphoma. Additionally, three lym- phocytic infiltration patterns were documented microscopically. The nodular infiltration was the most common type. CONCLUSIONS： HL is a rare entity and histopathology along with ancillary tests remains the only way to make the diagnosis.Clinicians＇ awareness of this entity and early liver biopsy are essential in patient management.

Hepatitis C virus NS3/4A with sequence variation at amino-terminus has different serine protease activities and inhibitory activities on IFN-β induction and p53-dependent transcriptional activation

Objective： To construct the point mutation plasmids expressing HCV NS3/4A with different secondary structures at the N-terminus, and to analyze their serine protease activities. Methods： The point mutation plasmid constructs were generated by using the QuickChange site-directed mutagenesis kit with the backbone of M-H05-5 （AI-1）, and were named as subgroup A1-2, A2-1, A2-2, BI-1, B1-2, B2-1, and B2-2 respectively. The transient expression of the constructs was investigated by immunofluorescence assay and Western blot analysis. The difference in in cis and in trans NS3 serine protease activity between each subgroup was determined by Western blot analysis. Luciferase reporter assay was used to observe the inhibitory effects of the constructs on RIG-I induced IFN-β promoter activity and on p53-dependent transcriptional activation. Results： The point mutation plasmid constructs were verified for the correct sequence by DNA sequencing. The immunofluorescence assay revealed 4 subcellular localization patterns of NS3, including dot-like staining, diffuse staining, doughnut-like staining, and rod-shape staining. Western blot analysis indicated that the incomplete cleavage of NS3/4A appeared in subgroups A2-1 and B2-1, indicating that the in cis NS3 serine protease activities of subgroup A2-1 and B2-1 were weaker when compared with the other subgroups. By using NS5A/SBAC as a substrate for NS3/4A serine protease, it was also found that the in trans NS3 serine protease activities of subgroup A2-1 and B2-1 were also weaker compared the other subgroups. Differences in inhibitory effects of HCV NS3 on RIG-I induced IFN-β promoter activity and on p53-dependent transcriptional activation were also observed between subgroup A2-1, B2-1 and the other subgroups. Conclusion： The results suggest that subgroup A2-1 and B2-1 has weaker serine protease activities and weaker inhibitory activities on host cell functions than the other subgroups, which might be explained by the different secondary structure of the 120-aa sequence at N-terminus of NS3.

Strategies in secondary biliary fibrosis

The cellular and molecular mechanisms that mediate hepatic fibrosis have provided a framework of different therapeutic foci to prevent, delay or in such case revert fibrosis and cirrhosis. The fundamental event in development of hepatic fibrosis caused by secondary biliary cholestasis is based on activation of the hepatic stellar cells (HSC) which the primary function is forming fibrosis. The activated HSC cells transform into myofibroblasts with capacity to produce alpha smooth muscle actin (α-SMA). As a result, the HSC activates proliferation of the cholangiocytes and epithelial cells whose functions represent important anti-fibrotic objectives. Some strategies are described as targeting against molecule involved in fibrosis production;and some medications with anti-fibrotic functions that are actually available in the medical arsenal have been tested in experimental animal models and in few clinical studies, and their components act in relation to the fibrotic cascade. In the end, the treatment strategies for hepatic fibrosis can vary on an individual basis depending on the etiology, the risk of fibrosis progression and the predominant pathogenic medium, which indicates that a multi-factorial approach could be necessary. Orthotopic liver transplant continues being the last final alternative for hepatic insufficiency from any cause;however, in no way does it supersede healthy natural liver in survival and adequate function. The investigative arsenal continues to develop rapidly, giving rise to other possible objectives in pre-clinical studies of conceptual trials, such as the utilization of molecular, cellular, drugs therapy and Chinese herbs. Despite being aforementioned, there are no existing ideal alternatives that completely reverse fibrosis in humans. Future usefulness of the majority of management alternatives seems probable and could be feasible.

Autoimmune hepatitis in a patient with immunoglobulin A nephropathy:A case report

BACKGROUND Immunoglobulin A nephropathy(IgAN)is the most commonly encountered glomerular disease in Asian countries.It has a broad clinical presentation,and it is frequently associated with other conditions.Chronic liver disease is well recognized as the leading cause of secondary IgAN.However,cases of IgAN associated with autoimmune hepatitis(AIH)have seldom been reported.CASE SUMMARY A 63-year-old Korean woman was admitted to Pusan National University Hospital for an evaluation of abdominal pain and elevated liver enzymes.Two weeks prior,she had presented to our hospital with proteinuria of approximately 1350 mg/d and hematuria and was diagnosed with IgAN.Autoimmune profiles were highly positive for antinuclear antibodies,and symptoms related to portal hypertension including ascites and peripheral edema were present.A diagnosis of AIH was made according to the simplified scoring system of the International Autoimmune Hepatitis Group.Despite immunosuppression with prednisolone and azathioprine,rapid deterioration of liver function led to end-stage liver disease.After a living-donor liver transplantation,liver function gradually improved,and she had maintained stable liver and kidney function at the six months follow-up.CONCLUSION Cases of secondary IgAN with chronic liver disease have been frequently reported in the literature but are rarely associated with AIH.We encountered an IgAN patient with concurrent progressive liver failure due to AIH.

基于“肝藏血、血舍魂”理论探讨慢性乙型肝炎继发抑郁的病机及治疗

慢性乙型肝炎病情缠绵难愈,部分患者容易继发抑郁症,使病情更加复杂。基于“肝藏血、血舍魂”理论在情志疾病中的作用及其病理表现,慢性乙型肝炎继发抑郁症的核心病机为肝血亏虚,血不养魂,最终致肝魂失调。临床诊治慢性乙型肝炎患者时,辨证加用养肝安魂、补肾安魂、健脾安魂以及重镇安魂之品,可以防治抑郁症,从而起到“未病先防”“既病防变”的作用。

HBV母婴传播阻断效果及影响因素分析

目的评价应用不同剂量乙型肝炎疫苗(Hep B)联合乙型肝炎免疫球蛋白(HBIG)阻断HBV母婴传播的效果及HBV母婴传播的影响因素。方法选取2012年7月-2015年2月在吉林大学第一医院进行HBV母婴阻断的785对乙型肝炎孕妇及其新生儿作为研究对象,根据孕妇产前HBV血清学标志物检查结果,对HBs Ag单阳性、HBs Ag与HBe Ag双阳性母亲的新生儿制订不同剂量阻断方案,在出生后2 h内分别注射10μg或20μg Hep B联合100 IU HBIG,并于1、6月龄继续接种Hep B。对入组孕妇及婴儿进行长期随访,采集婴儿7和12月龄静脉血检测HBV血清学标志物。计数资料组间比较采用χ2检验或Fisher确切概率法,组间HBV DVA的比较经对数转换后采用Mann-Whitney U秩和检验。探讨HBV相关因素采用非条件Logistic回归分析,并做影响HBV母婴传播的多因素分析。结果 785例新生儿中,HBs Ag阳性但HBe Ag阴性的529名孕妇所生的新生婴儿无一感染,阻断成功率为100%;256例HBs Ag与HBe Ag双阳性孕妇所生新生儿中,有14例婴儿感染,阻断成功率为94.53%。母亲HBe Ag阳性、HBV DNA>108IU/ml与阻断失败密切相关(P<0.001)。本研究未发现母亲的分娩方式、新生儿喂养方式等因素与阻断失败有关。探讨干预依从性时发现,首针疫苗未得到及时足量注射的新生儿更易发生感染。进一步Logistic回归分析发现,母亲HBV DNA>108IU/ml、首针疫苗未及时或未足量接种及男性婴儿更易发生HBV母婴传播。结论 HBs Ag阳性母亲的新生儿经过联合免疫后,可有效提高HBV母婴阻断成功率。母亲的感染状态、病毒载量和首针疫苗注射情况是影响HBV母婴传播阻断成功与否的决定性因素。

肝动脉加门静脉灌注化疗在结直肠癌肝转移中的应用

目的 评价肝动脉加门静脉灌注化疗药物治疗结直肠癌肝转移的临床效果。方法 将 4 8例结直肠癌肝转移患者随机分成 3组 :肝动脉加门静脉灌注化疗 (Ⅰ组 ) 17例 ,肝动脉灌注化疗 (Ⅱ组 ) 16例 ,门静脉灌注化疗 (Ⅲ组 ) 15例。术后 2周开始化疗 ,化疗方案为氟尿嘧啶 (5 FU) 5 0 0mg/m2 +丝裂霉素 (MMC) 4mg/m2 +表阿霉素 (EPI) 6 0mg/m2 ,每周 1次 ,2～ 3次为 1个疗程。 3组化疗方案的剂量、频次均相同。结果 肝转移灶治疗有效率 (CR +PR) :Ⅰ组为 76 5 % ,Ⅱ组为 6 2 5 % ,Ⅲ组为 4 6 7% ,Ⅰ组与Ⅱ、Ⅲ组比较差异有统计学意义 (P <0 0 1) ;患者 0 5、1、2年生存率Ⅰ组为 10 0 %、82 4 %和 5 2 9% ,Ⅱ组为 87 5 %、6 2 5 %和 4 3 7% ,Ⅲ组为93 3%、6 0 %和 33 3% ,Ⅰ组与Ⅱ、Ⅲ组比较差异有统计学意义 (P <0 0 1)。 3组患者均无严重毒副反应及并发症出现。结论 肝动脉加门静脉灌注化疗可使局部药物达较高浓度 ,分布更均匀 ,是治疗和预防结直肠癌肝转移的有效方法 ,其疗效明显优于单纯肝动脉灌注化疗或门静脉灌注化疗。

天津市1992—2010年新生儿乙型肝炎疫苗免疫策略成本效果分析

目的探讨天津市1992—2010年新生儿乙型肝炎(乙肝)疫苗免疫策略的效果和经济效益。方法以天津市疾病预防控制中心历年计划免疫报告资料和血清学监测结果为基础,借助社区大规模乙肝及相关疾病监测,以乙肝及相关疾病患者为经济负担调查对象,运用成本效益分析方法进行综合评价。结果现行乙肝疫苗免疫接种的策略下,天津市20岁以下人群报告发病率由14.14/10万(1992年)下降到9.46/10万(2010年),下降幅度达33.00%。1992—2010年共减少HBsAg携带者98 984例,慢性乙肝17 830例,乙肝肝硬化5310例,肝癌1606例。共产出效益473.51亿元,净效益473.12亿元,效益成本比为1213∶1。结论天津市实施儿童乙肝疫苗免疫策略可获得巨大经济效益和社会效益。

腹腔镜与手助腹腔镜脾切除术的临床应用

目的探讨腹腔镜与手助式腹腔镜脾切除术的方法、安全性和有效性。方法需行脾切除术的患者26例,包括肝硬化继发性脾功能亢进13例,原发性血小板减少性紫癜6例,外伤性脾破裂4例,假性脾囊肿1例,遗传性红细胞增多症2例。脾亢患者中有8例接受手助式腹腔镜脾切除术,8例中有5例同时行贲门周围血管离断术,4例脾外伤行手助式腹腔镜脾切除术,其余14例患者均采用腹腔镜脾切除术。结果除1例因大出血而中转开腹,余25例顺利完成手术,术中出血量约200～400mL,平均手术时间约为2.5～4.0h,平均术后5～9d出院,无手术并发症。结论腹腔镜及手助式腹腔镜脾切除术安全可行,创伤小,恢复快,值得临床推广。

拉米夫定治疗慢性乙型肝炎疗效和基因变异的研究(英文)

目的就拉米夫定治疗慢性乙型肝炎(C H B)1a的疗效、H BV D N A P基因变异特点及编码的蛋白质的二级结构加以研究。方法提取H BV D N A后,经半巢式聚合酶链反应(PC R)扩增,PC R产物自动测序。总结拉米夫定的疗效、P基因变异出现的时间,用SAS软件分析影响疗效的因素。用D N A ST AR软件的C LU STA L V方法对H BV D N A P基因片段的核苷酸和氨基酸差异、变异类型进行分析。用D N Aclub、D N Asis软件分析出现P基因变异前后所编码的蛋白质的二级结构的差异。结果按照ALT水平分组,各组间促进A LT正常化没有明显差异。在治疗后6个月血清标本中未发现Y M D D变异,在治疗后12个月血清标本中发现2例Y M D D变异。在对照组中发现了1例L528M变异;在治疗后12个月发现了2例Y V D D变异,还发现了2例只有L528M的变异。在发生变异后,Y M D D的转角结构变为折叠,L528M的转角结构没变。结论1a的拉米夫定治疗可以有效抑制H BV D N A的复制。Y M D D变异发生在拉米夫定治疗6个月之后,L528M变异可单独在体内存在,未接受抗病毒治疗的慢性H BV携带者可发生L528M变异。发生Y M D D变异的P基因编码的蛋白质二级结构由转角变为折叠。L528M变异不影响蛋白质二级结构。

北京市顺义区中学生病毒性肝炎健康教育效果评价

目的评价北京市顺义区中学生病毒性肝炎健康促进项目效果,为防控政策的制定提供科学依据。方法对全区中学生开展病毒性肝炎健康课堂、知识竞赛等健康促进项目,采用分阶段整群抽样方法抽取10所中学30个班级1800名中学生进行健康教育前后的问卷调查,评价项目的实施效果。结果共调查中学生1504名,教育前病毒性肝炎相关知识知晓率平均为44.68%,高中组学生知晓率高于初中组;教育后,平均知晓率上升到77.19%;表示会与病人保持交往的比率由教育前的80.07%上升到教育后的82.51%;认为接种乙肝疫苗能有效预防乙型肝炎的比率由教育前的80.58%上升到教育后的93.62%。结论在中学生中实施病毒性肝炎健康促进项目,能有效提高中学生病毒性肝炎防控知识和有关信念。

肝细胞向胆管上皮细胞转分化在继发性胆汁淤积性大鼠肝纤维化形成中的作用及黄芪汤组分的干预效应

目的探讨肝细胞向胆管上皮细胞转分化在继发性胆汁淤积性大鼠肝纤维化形成中的作用及黄芪汤组分抑制肝纤维化的效应机制。方法 75只SD雄性大鼠,采用胆总管结扎(bile duct ligation,BDL)制备继发性胆汁淤积性肝纤维化模型,假手术组(Sham)仅作胆总管分离,不作胆总管结扎。大鼠BDL术后1周随机分为对照组(M)与干预组(Y组,经灌胃给予黄芪汤组分4周),Sham组于术后1周末随机抽取5只大鼠处死取材,M组分别于术后1、2、3、4周末随机抽取5只大鼠做动态观察,余下各组大鼠均于术后第5周末处死取材。观测肝脏组织学及羟脯氨酸(Hyp)含量;激光共聚焦显微镜观察肝组织胆管上皮细胞标志物角蛋白-7(CK7)与肝细胞特异性抗原(HepPar)共定位,Western blot检测CK7、HepPar的蛋白表达,IPP软件分析肝组织天狼猩红胶原染色。结果随着造模时间推移,M组大鼠肝组织肝细胞Hep Par阳性细胞逐渐减少(Sham>M1周>M2周>M3周>M4周>M5周),胆管上皮细胞(CK7阳性细胞)及纤维化程度、Hyp含量、CK7蛋白表达均逐渐增加(Sham<M1周<M2周<M3周<M4周<M5周),CK7/Hep Par共定位细胞于术后1周即见增加,术后3周达到峰值,然后渐趋减少,肝组织Hep par蛋白表达量与CK7蛋白表达量呈显著负相关,Hep Par阳性细胞表达量与CK7阳性细胞表达量、胶原沉积程度均呈显著负相关,而CK7阳性细胞表达量与胶原沉积程度呈显著正相关;与M组比较,Y组大鼠死亡率、CK7阳性细胞、纤维化程度、肝组织Hyp含量、CK7蛋白表达量均显著下降(P<0.01),Hep Par阳性细胞及Hep Par蛋白表达含量显著增加。结论肝细胞向胆管上皮细胞转分化可能是继发性胆汁淤积性大鼠肝纤维化形成过程中的关键病理环节,抑制肝细胞向胆管上皮细胞转分化可能是黄芪汤组分有效干预继发性胆汁淤积性肝纤维化的主要效应机制之一。

59例结直肠癌肝转移的外科治疗

背景与目的:结直肠癌(colorectalcancer,CRC)是最常见的恶性肿瘤之一,肝转移是结直肠癌治疗失败的最主要原因。结直肠癌肝转移若未经治疗,其中位生存期仅为6个月左右,手术切除是延长患者生存期的重要手段。本研究目的是探讨结直肠癌肝转移的外科治疗效果及影响预后的因素。方法:回顾性分析1987年1月～1998年12月行结直肠癌肝转移瘤切除59例患者资料。其中不规则性肝切除45例,规则性肝切除14例。术后发生并发症者4例(6.8%),未出现手术死亡。采用Kaplan鄄Meier法进行生存分析,log鄄rank检验进行统计学比较。应用Cox比例风险模型进行多因素分析。结果:全组总的1、3、5年生存率分别为91.4%、34.8%及21.9%。同时性肝转移患者的生存率显著高于异时性肝转移患者(P<0.01),肝转移瘤最大直径小于等于5.0cm患者的生存率高于大于5.0cm患者。肝转移发生时间与肿瘤最大直径大于5.0cm是影响预后的主要因素,而原发肿瘤淋巴结转移状况、术前癌胚抗原(CEA)水平及转移瘤数量对预后没有显著影响。结论:结直肠癌肝转移的外科治疗可使患者获得长期生存。通过加强随诊,早期发现肝转移瘤,可以提高手术切除率。

拉米夫定对乙型肝炎病毒DNA复制及基因变异的影响

目的:研究拉米夫定治疗慢性乙型肝炎(CHB)的疗效、乙型肝炎病毒(HBV)DNA P基因变异特点及编码蛋白质的二级结构。方法:提取HBV DNA后,半巢式聚合酶链反应(PCR)扩增,PCR产物自动测序。总结拉米夫定的疗效、P基因变异出现的时间,用SAS软件分析影响疗效的因素。用DNA STAR软件的CLUSTAL V方法对HBV DNA P基因片段的核苷酸和氨基酸差异、变异类型进行分析。用DNAc lub,DNAsis软件分析出现P基因变异前后所编码的蛋白质的二级结构的差异。结果:在治疗后6个月血清标本中未发现YMDD变异,在治疗后12个月血清标本中发现2例YMDD变异。在对照组中发现了1例L528M变异;在治疗后12个月发现了2例YVDD变异,2例只有L528M的变异。在发生变异后,YMDD的转角结构变为折叠,L528M的转角结构没变。结论:1年的拉米夫定治疗可以有效抑制HBV DNA的复制。YMDD变异发生在拉米夫定治疗6个月之后,L528M变异可单独在体内存在,未接受抗病毒治疗的慢性HBV携带者可发生L528M变异。发生YMDD变异的P基因编码的蛋白质二级结构由转角变为折叠。L528M变异不影响蛋白质二级结构。

丙型肝炎病毒F蛋白缺失对病毒复制及感染性的影响

探索了F蛋白缺失及核心蛋白(Core)二级结构改变对丙型肝炎病毒(HCV)复制和感染性的影响.利用定点突变方法,将J6JFH1的核心基因引进5个终止密码子以中断F蛋白的表达,从而获得F蛋白缺失的病毒复制子J6JFH1/ΔF.体外制备RNA转录体,并电穿孔转染Huh7.5.1细胞,采用免疫荧光、实时荧光定量PCR方法以及病毒感染等方法,观察F蛋白缺失对病毒复制、蛋白质表达及转染细胞上清感染性病毒颗粒产生的影响.在此基础上,构建5个单一突变病毒体,对HCV核心蛋白进行二级结构分析,观察核心蛋白二级结构对HCV复制和翻译的影响.结果显示,转染48 h后,J6JFH1/ΔF与野生型J6JFH1相比,J6JFH1/ΔF转染阳性细胞数明显降低,细胞内HCV RNA水平降低约95%,J6JFH1/ΔF转染后不同时间点细胞上清中HCV RNA拷贝数和病毒颗粒也明显降低.5个单一突变体不影响核心基因二级结构,病毒在细胞内复制和感染性与野生型水平一致.J6JFH1/ΔF所产生的改变可能是由于5处突变导致核心基因二级结构改变而造成的.结果说明,HCV F蛋白缺失不影响病毒的复制翻译及病毒颗粒的包装释放,核心蛋白二级结构的改变对病毒复制和翻译则产生较大影响.