乙型肝炎基因工程疫苗阻断乙型肝炎病毒母婴传播…

用转基因细胞分泌的乙型肝炎病毒表面抗原基因工程疫苗免疫母亲ＨＢｓＡｇ阳性的新生儿５０例，观察其阻断乙型肝炎病毒母婴传播的效果，随访１２个月，在３６例母亲为ＨＢｓＡｇ和ＨＢｅＡｇ均阳性的婴儿中仅１例为ＨＢｓＡｇ阳性，其余婴儿均有保护性抗体，预防保护率为９６．２％。     

套式PCR检测HBV与HCV垂直传播的比较研究

用套式ＰＣＲ法检测了２７例ＨＢｓＡｇ阳性母亲的新生儿脐带血ＨＢＶ ＤＮＡ，阳性３例（１１．１％），同时检测了７例抗－ＨＣＶ阳性母亲的新生儿脐带血ＨＣＶ ＲＮＡ及抗－ＨＣＶ，结果ＨＣＶ ＲＮＡ无１例阳性，抗－ＨＣＶ全部阳性，因此认为婴儿体内抗－ＨＣＶ是一种由母体被动获得的抗体，不能作为ＨＣＶ感染的客观指标。     

婴幼儿乙型肝炎病毒表面抗原阳性者的动态转归

对５４例婴幼儿时期斩乙型肝炎病毒表面抗原（ＨＢｓＡｇ）阳性者进行了动态研究。结果表明，婴幼儿ＨＢｓＡｇ阳性者的年阴转率很低，为１．０４％，ＨＢｓＡｇ阳性者年阴转率高低与母亲ＨＢｓＡｇ阳性者年阴转率高低与母亲ＨＢｓＡｇ是否阳性关系密切，与是否免疫关系为期母亲ＨＢｓＡｇ阳性者阴转率稍高。     

我国九个地区丁型肝炎病毒感染状况及其与乙型肝炎病毒复制指标关系的研究

对１５４５例各类乙型肝炎病毒表面抗原（ＨＢｓＡｇ）阳性肝病和无症状ＨＢｓＡｇ携带者的血清进行了乙型肝炎病毒（ＨＢＶ）与丁型肝炎病毒（ＨＤＶ）感染标记物的测定。结果表明，ＨＤＶ感染率为１３．０１％，其中ＨＤＡｇ和抗－ＨＤ阳性率分别为２．９１％和１０．０９％。而且在全国九个地区均有ＨＤＶ感染者存在，说明其分布是较为广泛的。同时还表现出，男性高于女性，慢性肝炎、重型肝炎及原发性肝癌高于急性肝炎和无症状ＨＢｓＡｇ携带者。提示ＨＢＶ与ＨＤＶ合并感染或重叠感染可能导致病情加重和感染的慢性化。本项研究结果还揭示，在ＨＢＶ与ＨＤＶ合并或重叠感染时，可能对ＨＢＶ的复制指标（ＨＢｅＡｇ·ＨＢＶＤＮＡ）有一定的抑制现象。     

应用重组质粒pAM－HBsAg在果蝇细胞中表达乙型肝炎病毒表面抗原及基因免疫的初步研究

应用果蝇（ＤＳ２）表达系统，构建了含有乙型肝炎病毒表面抗原（ＨＢｓＡｇ）、摄金蛋白启动子（ＭＴｎ－ｐｒｏｍｏｔｅｒ）的共表达质粒ｐＡＭ－ＨＢｓＡｇ，转染细胞，经克隆，存活细胞株的培养上清液经硫酸铵沉淀、氯化铯（ＣＳＣｌ）密度梯度离心沉淀，获得的抗原用酶联免疫吸附试验（ＥＬＩＳＡ）、放射免疫分析（ＲＩＡ）法检测抗体，免疫吸印法（Ｗｅｓｔｅｒｎｂｌｏｔ）和电泳凝胶银染显色证实分子量为２３０００和２７０００，免疫电镜观察显示表达产物为２２ｕｍ球型颗粒，通过重金属离子（ＣｕＳＯ４、ＺｎＳＯ４）的诱导可增加抗原的表达量。用共表达质粒ｐＡＭ－ＨＢｓＡｇ的ＤＮＡ，注射Ｂａｌｂ／Ｃ小鼠的股四头肌。经ＥＬＩＳＡ、ＲＩＡ检测抗体产生情况，结果免疫后的小鼠经硫酸锌喂养抗体高于普通喂养的小鼠。Ｓｏｕｔｈｅｒｎ杂交证实鼠肌肉细胞存在ＨＢｓＡｇ基因。小鼠免疫接种实验表明，ＤＳ２细胞表达的抗原与直接用ＤＮＡ含有ＨＢｓＡｇ的重组质粒）免疫小鼠均获抗ＨＢｓＡｇ的抗体。     

用ELISA微板法检测乙型肝炎病毒核心抗原

以双抗体包被的抗体夹心法，用微板ＥＬＩＳＡ检测血清乙型肝炎病毒核心抗原（ＨＢＣＡｇ），确定双包被工作浓度ＭＣ－抗－ＨＢｃ（效价１０００）为０．０４μｌ／孔，ＭＣ－抗－ＨＢｓ（１ｍｇ／ｍｌ）为３～４μｌ／孔；最佳裂解剂及其工作浓度为７％ＮＰ－４０巯基乙醇溶液。分别用不同的酶标记抗体检测，均证明双包被具有特异性。加入抗－ＨＢｃ进行阻断试验，其阻断率为７９．３％。对８４４例ＨＢｓＡｇ阴性的血清及１１４例ＨＢＶ－ＤＮＡ探针阴性血清用本法进行ＨＢｃＡｇ检测，均为阴性。在临床应用上，本法的阳性率明显高于试管法的，与ＨＢＶ－ＤＮＡ探针的阳性符合率为９１．４％，并且特异性与ＨＢＶ－ＤＮＡ探针的一致。     

慢性肝炎和肝癌病人血清中乙型肝炎病毒DNA的检测

为了了解慢性肝炎和肝癌病人患者体内乙型肝炎病毒（ＨＢＶ）复制与ＨＢＶ血清标志之间的关系，用酶联免疫吸附实验（ＥＬＩＳＡ）、聚合酶链反应（ＰＣＲ）及斑点杂交方法对６１例慢性肝炎和４７例肝癌患者的ＨＢＶ表面抗原（ＨＢｓＡｇ）、相关ｅ抗原（ＨＢｅＡｇ）、表面抗体（抗－ＨＢｓ）、核心抗体（抗－ＨＢｃ）、相关ｅ抗体（抗－ＨＢｅ）进行了检测。结果表明：ＨＢＶＤＮＡ在ＨＢｓＡｇ、ＨＢｅＡｇ、／抗－ＨＢｃ阳性的慢性肝炎和肝癌患者血清中的检出率分别为９０．５０％和５０．００％；在ＨＢｓＡｇ／抗－ＨＢｅ、抗－ＨＢｃ阳性者的检出率分别为４５．４０％和７．１４％；在ＨＢｓＡｇ阳性、ＨＢｅＡｇ阴性／抗－ＨＢｅ阴性者中的检出率分别为６０．００％和４０．００％；ＨＢｓＡｇ阴性、／抗－ＨＢｃ阳性或／抗－ＨＢｅ阳性或／抗－ＨＢｓ阳性者中的检出率分别为２０．００％和２２．２２％；在血清学指标全阴性时，慢性肝炎和肝癌患者血清中ＨＢＶＤＮＡ的检出率均为０。实验提示：无论是肝炎或肝癌，在ＨＢｓＡｇ、ＨＢｅＡｇ同时阳性时，ＨＢＶ复制最为活跃；在单独ＨＢｓＡｇ阳性时，ＨＢＶ有一定程度的复制；ＨＢＶ复制在肝癌细胞中受到一定程度的抑制。     

乙型肝炎疫苗免疫持久性及预防效果的队列定人研究

报告了新生儿乙型肝炎疫苗免疫后１－９年乙型肝炎病毒表面抗原（ＨＢｓＡｇ）阳转率的定人随访结果。母亲ＨＢｓＡｇ阳性和阴性免疫后的ＨＢｓＡｇ阴性者，基ＨＢｓＡｇ的年阳转率分别为０．３６％和０．０３６％，前者是后者的１０倍。母亲ＨＢｓＡｇ阳性儿中１４例ＨＢｓＡｇ阳转，其中１０／１４（７１％）发生在免疫后２－３岁；１２／１４（８６％）变成慢性ＨＢｓＡｇ携带者。母亲ＨＢｓＡｇ阴性儿共发现３例ＨＢｓＡｇ阳转，     

新生儿乙型肝炎疫苗免疫失败同孕妇血清乙型肝炎…

以ＨＢｓＡｇ，ＨＢｅＡｇ阳性孕妇作为研究对象，用病例－对照研究方法，比较婴儿乙型肝炎疫苗免疫失败和婴儿疫苗免疫成功两组孕妇分娩时血清乙型肝炎病毒ＨＢＶ－ＤＮＡ含量进一步用定群方法研究母亲ＨＢＶ－ＤＮＡ含量与其婴儿免疫后ＨＢｓＡｇ持续携带 关系，证实孕妇血清ＨＢＶ－ＤＮＡ高含量是婴儿免疫失败的主要原因。     

我国九个地区丁型肝炎病毒感染状况及其与乙型肝炎…

对１５４５例各类乙型肝炎病毒表面抗原（ＨＢｓＡｇ）阳性肝病和无症状ＨｓＡｇ携带者的血清进行了乙型肘炎病毒（ＨＢＶ）与丁型肝炎病毒（ＨＤＶ）感染标记物的测定。结果表明，ＨＤＶ感染率为１３．０１％，其中ＨＤＡｇ和抗－ＨＤ阳性率分别为２．９１％和１０．０９％。而且在全国九个地区均有ＨＤＶ感染者存在，说明其分布是较为广泛的。同时还表现出，男性高于女性，慢性肝炎、重型肝炎及原发性肝癌高于急性肝炎和无症状ＨＢ     

HBsAg和HBsAb双阳性乙肝患者血清中HBsAb确认方法的建立

目的 建立一种基于酶联免疫吸附实验（ ELISA）的HBsAb确认方法,验证HBsAg和HBsAb双阳性的乙肝患者血清中HBsAb阳性的真实性,剔除假阳性结果,避免错误诊断.方法收集60例电化学发光免疫分析法（ ECLIA）检测的HBsAg浓度在1000 COI以上的混合血清作为确认血清,将不同稀释度的确认血清与收集的HBsAb阳性混合血清中和,筛选并确定确认血清中HBsAg的最佳试验浓度.收集40例HBsAg和HBsAb双阳性的血清,与确认血清中和后采用ELISA检测COI的下降率,验证HBsAg和HBsAb双阳性标本中HBsAb阳性的真实性.结果 确认血清HBsAg浓度为2000 COI时对HBsAb的中和效果最好,ELISA确认法对40例HBsAg和HBsAb双阳性标本确认结果为37例真阳性和3例假阳性,与ECLIA法完全一致.结论 成功建立了HBsAg和HBsAb双阳性血清HBsAb的确认方法,该方法简单、准确且成本较低.     

山东省枣庄市乙型病毒性肝炎流行病学调查

目的　了解枣庄市人群中乙型肝炎的流行特征。方法　于 2 0 0 0年采用随机分层抽样 ,调查 312户家庭的 96 3人 ,以RIA法检测HBsAg、抗 HBs和抗 HBc。结果　HBsAg、抗 HBs、抗 HBc和HBV标化流行率分别为 7.0 8%、37.5 6 %、4 1.35 %和 4 4 .37%。HBsAg流行率男性高于女性 (P <0 .0 5 ) ,城区高于农村 (P <0 .0 1) ,在不同年龄及职业人群中差异无显著性 (P >0 .0 5 )。抗 HBs、抗 HBc和HBV感染率有随年龄增长而递增的趋势 (P <0 .0 1)。HBV总感染率男性高于女性 (P <0 .0 5 ) ,农村高于城市 (P <0 .0 5 )。结论　枣庄市人群HBV感染率较高 ,应积极采取预防和控制措施 ,减少发病。     

人外周血初始B细胞和记忆性B细胞亚群的特征和功能

B淋巴细胞是免疫系统的重要免疫成份,主要功能是介导体液免疫应答.在人外周血中,按照B淋巴细胞的发育阶段及功能的不同,可将B淋巴细胞分为初始成熟B细胞、记忆B细胞和浆细胞.记忆B细胞又可分为IgM记忆B细胞和类型转换的记忆B细胞.近年来的研究表明,B淋巴细胞的亚群远比人想象中的复杂,因此对人B细胞各亚群的起源、发育和功能进行更深入的研究,将有助于治疗自身免疫性疾病,在慢性感染性疾病的治疗过程中找到新的策略,并指导研发安全有效的疫苗.     

人外周血初始B细胞和记忆性B细胞亚群的特征和功能

B淋巴细胞是免疫系统的重要免疫成份,主要功能是介导体液免疫应答.在人外周血中,按照B淋巴细胞的发育阶段及功能的不同,可将B淋巴细胞分为初始成熟B细胞、记忆B细胞和浆细胞.记忆B细胞又可分为IgM记忆B细胞和类型转换的记忆B细胞.近年来的研究表明,B淋巴细胞的亚群远比人想象中的复杂,因此对人B细胞各亚群的起源、发育和功能进行更深入的研究,将有助于治疗自身免疫性疾病,在慢性感染性疾病的治疗过程中找到新的策略,并指导研发安全有效的疫苗.     

Molecular epidemiology of hepatitis B virus in Spain: identification of viral genotypes and prediction of antigenic subtypes by limited sequencing

The hepatitis B virus (HBV) genotypes were studied by a line probe assay (LiPA) and by direct sequencing of a 339 nucleotide fragment from the S region of the viral genome in samples from 269 carriers living in Spain, either native to Spain (231) or immigrants from Africa, Asia, and Eastern Europe (38). The sequences were also used to predict the HBV surface antigen (HBsAg) subtype on the basis of the amino acids specified at selected positions of the HBsAg molecule. Agreement between the two genotyping methods was found in most cases (98.1%) and a HBV genotype could be assigned to all samples. The viral groups D/ayw2 (30.1%), D/ayw3 (28.6%), and A/adw2 (21.2%) were prevalent, with an additional participation of the groups D/ayw4 (4.8%), F/adw4q- (1.9%), A/ayw1 (1.9%), and D/adw3 (0.7%), all of them present among the autochthonous carriers. Strains from genotypes B and C were found exclusively among Chinese immigrants. Genotype E strains were found in immigrants from Central Africa and in one patient native of Spain. Point mutations leading to amino acid changes of residues involved in the expression of the HBsAg subtype determinants were found in 12 samples (4.5%). Some mutations would predict the putative novel genotype-subtype associations A/adw4q+, A/ayr, D/ayr, and E/ayw1, while others would suggest the loss of subtype-specific determinants. The finding of HBV strains characteristic for Africa among the autochthonous carriers confirms the emergence of African HBV strains in Spain.     

Analysis of the entire nucleotide sequence of hepatitis B causing consecutive cases of fatal fulminant hepatitis in Miyagi Prefecture Japan

We encountered five consecutive patients with fulminant hepatitis induced by acute hepatitis B virus (HBV) infection in 2000--2001 in Japan. They had not had previous contact each other, and were referred to us from different hospitals. Although a 69-year-old woman could be rescued by intensive internal treatment, the four patients died. We analyzed the partial (nt 278-646) and entire nucleotide sequences of the HBV obtained from them, and their divergences were 0-0.3% and 0-0.2%, respectively. The results suggested that they had been infected with the same HBV isolates. The isolates belonged to genotype B and subgenotype B2 on the phylogenetic tree analysis (AB302942-AB302946). As for the nucleotides sequences of them, previously reported mutations of G1896A, A1762T, and G1764A were present. Amino acid analysis revealed that previously reported Ile97Leu and Pro130Non-Pro in the core region and Trp28Stop in the precore region were present. As for the entire nucleotide sequences among B2, AB302942 showed low divergences with AF121245 and AB073834 (1.7%), and X97850 from patients with fulminant hepatitis (3.2%). We compared the two consensus nucleotides derived from AB302942 and X97850 (fulminant hepatitis) versus AY121245 and AB073834 (non-fulminant hepatitis), which revealed a difference in nt 1,504 located in the P and X region. Nucleotide 1,504 was C for isolates from fulminant hepatitis and G for non-fulminant hepatitis, and it was recognized among most of the isolates belonging to B2 registered on GenBank. Further studies could disclose the mechanism of severe inflammation of liver that finally leads to fulminant hepatitis.     

High levels of viral replication during acute hepatitis B infection predict progression to chronicity

To assess the pattern of development of serologic markers during acute hepatitis B, levels of HBsAg, HBeAg, and hepatitis B virus (HBV) DNA were assayed in stored serum samples obtained sequentially from 12 subjects infected with HBV during experimental studies conducted in the 1950s. Six patients developed acute self-limited hepatitis, three developed chronic hepatitis, and three had an asymptomatic infection without HBsAg. HBsAg was the first serologic marker detected (mean = 52 days after exposure), followed by HBeAg (62 days) and HBV DNA (72 days). Peak HBsAg levels occurred before onset of symptoms and correlated with peak titers of HBeAg and HBV DNA. Patients who developed chronic hepatitis had higher peak levels of viral markers than those with self-limited disease: HBsAg (30 versus 5.4 μg/ml), HBeAg (1:2,000 versus 1:60 titer) and HBV DNA (3,192 versus 444 pg/ml). Thus, chronic HBV infection is characterized by high levels of viral replication appearing early during the acute phase of infection. © 1994 Wiley-Liss, Inc.

1 This article is a US Government work and, as such, is in the public domain in the United States of America.

     

The Effect of Age on the B-Cell Repertoire

The antibody repertoire changes with age. This change reflects, in part, the age-associated impairment in the production of a diverse population of naive B cells in the bone marrow and, in part, by the decreased diversification of B cells in the germinal center where affinity maturation and isotype switching takes place. B cell number is strictly regulated and despite the decreased output of B cells by the bone marrow does not decline during aging. Self-renewal of peripheral B cells is sufficient to assure the stability of peripheral B cell number. However, when B cell production is stressed as, for example, following drug-induced lymphopenia, the rate of recovery of B cell number as well as of B cell diversity is compromised in old compared to young mice. Finally, aging is associated with the appearance of B cell clonal expansions which not only limit the diversity of the B cell repertoire but very likely give rise to monoclonal serum immunoglobulins and B cell neoplasms.     

Low-dose UVB radiation perturbs the functional expression of B7.1 and B7.2 co-stimulatory molecules on human Langerhans cells

In previous studies, we have shown that ultraviolet (UV) B radiation perturbs the APC function of Langerhans cells (LC) by interfering with as-yet unidentified co-stimulatory signals. Recently, B7.1 and B7.2 on APC were shown to deliver important co-stimulatory signals through interaction with their counterreceptors CD28 and CTLA-4 on T cells. To determine whether UVB affects the functional expression of B7.1 or B7.2 on LC, B7.1 and B7.2 expression was studied on human LC by multiparameter flow cytometry. Little, if any, B7.1 or B7.2 was detected on LC freshly isolated from skin. However, following 48 h of tissue culture, expression of both B7.1 and B7.2 were markedly up-regulated. To test whether these molecules were functional, primary mixed epidermal cell leukocyte reactions (MECLR) were performed. Blocking monoclonal antibody (mAb) to B7.1 or B7.2 both inhibited the MECLR, with anti-B7.2 being much more effective than anti-B7.1. UVB radiation dose-dependently (100–200 J/m²) suppressed the culture-induced up-regulation of B7.1 and B7.2 on LC. Since LC exposed to the same UVB flux (UVB-LC) failed to stimulate alloreactive T cells in a MECLR, we questioned whether this was related to their inability to provide B7 co-stimulation. Indeed, when effective B7-CD28 signaling was ascertained by adding submitogenic doses of exogenous anti-CD28 mAb to UVB-LC, the proliferative response of alloreactive T cells was restored. We conclude that the suppressive effects of low-dose UVB radiation on the APC function of LC are, at least in part, due to an inhibition of functional B7.1 and B7.2 expression.     

Novel HLA-B alleles, B*8201, B*3515 and B*5106, add to the complexity of serologic identification of HLA types

Three class I alleles, B*8201, B*3515 and B*5106, have been described using DNA and cDNA sequencing. The B*8201 allele is most structurally related to B*5602, differing from it by 14 nucleotide substitutions resulting in 5 amino acid differences. The other two alleles, B*3515 and B*5106, differ from their most closely related HLA-B alleles by 2–3 nucleotide substitutions resulting in 1–2 amino acid substitutions, respectively. The majority of nucleotide substitutions marking these new alleles are observed in other HLA-B alleles suggesting that gene conversion and/or reciprocal recombination have created this diversity. All of the amino acid substitutions are predicted to alter the antigen binding site of the HLA-B molecule. The newly defined HLA-B allelic products were originally defined by their unusual serologic reactivity patterns. The B*8201 allelic product is serologically typed as a B "blank" or as a variant of B22 or B45. These patterns and the serologic reactivity of the other newly described allelic products are consistent with the protein sequence homology among specific HLA-B molecules. While serology remains a powerful tool for detecting HLA diversity, alleles generated by events resulting in the sharing of HLA sequence polymorphisms among alleles at a locus will continue to create complexity in the interpretation of typing results.