Randomised controlled trial of lymphoblastoid interferon alfa in Europid men with chronic hepatitis B virus infection.

OBJECTIVE--To confirm the findings of pilot studies that interferon alfa is an effective treatment of Europid men with chronic hepatitis B virus infection. DESIGN--Randomised controlled trial of three months treatment with interferon alfa followed by 12 months of observation. SETTING--Outpatient clinic of a tertiary referral centre. PATIENTS--37 Treated men (six anti-HIV positive) and 34 untreated men (nine anti-HIV positive) who met the criteria for the trial. Four controls failed to complete follow up. INTERVENTIONS--The treated group received subcutaneous injections of 5-10 MU interferon alfa/m2 daily for five days, then 10 MU/m2 thrice weekly for 11 weeks. Follow up continued at monthly intervals for 12 months. Untreated controls were monitored over the same period. MAIN OUTCOME MEASURE--Hepatitis B e antigen and hepatitis B virus DNA state after 15 months of observation. RESULTS--12 Of the 37 treated patients cleared hepatitis B e antigen and hepatitis B virus DNA, whereas only one of 30 untreated controls seroconverted over the same period--an increased response rate of 29% (95% confidence interval 13% to 45%). The life table estimate of response at 15 months was 35% in treated patients, an increase of 32% above controls (95% confidence interval 16% to 48%). The response rates in groups by predictive pretreatment variables were 12 of 31 anti-HIV negative patients (excess response 34%; 95% confidence interval 14% to 54%), 12 of 26 with chronic active hepatitis before treatment (excess response 46%; 27% to 65%), and 12 of 21 with a pretreatment serum aspartate aminotransferase activity greater than 70 IU/l (excess response 46%; 16% to 76%). The combination of these factors predicted response with a sensitivity of 100% and a specificity of 80%. Four of the 12 responders, who had all been infected for less than two years, also lost hepatitis B surface antigen. Treatment was well tolerated. CONCLUSIONS--Interferon alfa is effective in the treatment of a proportion of Europid men with chronic hepatitis B virus infection, who might be identified before treatment. Additional strategies are required to improve the rate of response.     

Human and mouse LSP1 genes code for highly conserved phosphoproteins

With use of the mouse LSP1 cDNA we isolated a human homologue of the mouse LSP1 gene from a human CTL cDNA library. The predicted protein sequence of human LSP1 is compared with the predicted mouse LSP1 protein sequence and regions of homology are identified in order to predict structural features of the LSP1 protein that might be important for its function. Both the human and mouse LSP1 proteins consist of two domains, an N-terminal acidic domain and a C-terminal basic domain. The C-terminal domains of the mouse and human LSP1 proteins are highly conserved and include several conserved, putative serine/threonine phosphorylation sites. Immunoprecipitation of LSP1 protein from 32P-orthophosphate-loaded cells show that both the mouse and human LSP1 proteins are phosphoproteins. The sequences of the putative Ca2(+)-binding sites present in the N-terminal domain of the mouse LSP1 protein are not conserved in the human LSP1 protein; however, a different Ca2(+)-binding site may exist in the human protein, indicating a functional conservation rather than a strict sequence conservation of the two proteins. The expression of the human LSP1 gene follows the same pattern as the expression of the mouse LSP1 gene. Southern analysis of human genomic DNA shows multiple LSP1-related fragments of varying intensity in contrast to the simple pattern found after similar analysis of mouse genomic DNA. By using different parts of the human LSP1 cDNA as a probe, we show that most of these multiple bands contain sequences homologous to the conserved C-terminal region of the LSP1 cDNA. This suggests that there are several LSP1-related genes present in the human genome.     

Detection of amyloid beta protein precursor immunoreactivity in normal and Alzheimer's disease cerebrospinal fluid

The amyloid A4 (or beta protein), a 4.2 kD polypeptide, is a major component of amyloid deposits in the brains of patients with Alzheimer's Disease (AD). The self-aggregating amyloid A4 protein of AD is encoded as part of three larger proteins by the amyloid A4 precursor gene. The corresponding proteins have 695, 751 and 770 amino acid residues. To investigate the utility of amyloid beta protein precursor (A beta PP) as a diagnostic marker for AD an antiserum against a synthetic peptide (175-186), predicted from cDNA sequence for A beta PP, was used. The immunoreactivity of A beta PP in normal and AD cerebrospinal fluid (CSF) was measured by Western blot and detected with radiolabeled protein A. A total of fifty-seven CSF samples (AD = 27 and normal = 30) were analyzed for A beta PP immunoreactivity. A polyclonal antibody detected two major protein bands with apparent molecular weights of 105kD and 90kD both in normal and AD CSF. The difference between normal and AD CSF was not significant. These results indicate that immunoreactivity of A beta PP is present both in normal and AD CSF, and that the difference is too small to be used as a diagnostic marker.     

Molecular cloning and characterization of the human double-stranded RNA-activated protein kinase induced by interferon.

The double-stranded (ds) RNA-activated protein kinase from human cells is a 68 kd protein (p68 kinase) induced by interferon. On activation by dsRNA in the presence of ATP, the kinase becomes autophosphorylated and can catalyze the phosphorylation of the alpha subunit of eIF2, which leads to an inhibition of the initiation of protein synthesis. Here we report the molecular cloning and characterization of several related cDNAs from which can be deduced the full-length p68 kinase sequence. All of the cDNAs identify a 2.5 kb RNA that is strongly induced by interferon. The deduced amino acid sequence of the p68 kinase predicts a protein of 550 amino acids containing all of the conserved domains specific for members of the protein kinase family, including the catalytic domain characteristic of serine/threonine kinases. In vitro translation of a reconstructed full-length p68 kinase cDNA yields a protein of 68 kd that binds dsRNA, is recognized by a monoclonal antibody raised against the native p68 kinase, and is autophosphorylated.     

Supported phospholipid/alkanethiol biomimetic membranes: insulating properties.

A novel model lipid bilayer membrane is prepared by the addition of phospholipid vesicles to alkanethiol monolayers on gold. This supported hybrid bilayer membrane is rugged, easily and reproducibly prepared in the absence of organic solvent, and is stable for very long periods of time. We have characterized the insulating characteristics of this membrane by examining the rate of electron transfer and by impedance spectroscopy. Supported hybrid bilayers formed from phospholipids and alkanethiols are pinhole-free and demonstrate measured values of conductivity and resistivity which are within an order of magnitude of that reported for black lipid membranes. Capacitance values suggest a dielectric constant of 2.7 for phospholipid membranes in the absence of organic solvent. The protein toxin, melittin, destroys the insulating capability of the phospholipid layer without significantly altering the bilayer structure. This model membrane will allow the assessment of the effect of lipid membrane perturbants on the insulating properties of natural lipid membranes.     

Chloroplast DNA inheritance in theStellaria longipes complex (Caryophyllaceae)

Inheritance of chloroplast DNA (cpDNA) was examined in F₁ progenies derived from three crosses and three corresponding reciprocal crosses betweenStellaria porsildii andS. longifolia. Chloroplast DNA restriction fragments were analyzed using methods of nonradioactive digoxigenin-11-dUTP labeling and chemiluminescent detection with Lumi-Phos 530. Distinct interspecific restriction fragment polymorphisms were identified and used to demonstrate the mode of cpDNA inheritance. Mode of cpDNA inheritance differed among crosses. Two crosses in whichS. porsildii, SP2920-21, was the maternal parent exhibited three different types of plastids, maternal, paternal and biparental, among the F₁ hybrids, suggesting a biparental cpDNA inheritance and plastid sorting-out inStellaria.     

Spontaneous development of anti-hepatitis B virus envelope (anti-idiotypic) antibodies in animals immunized with human liver endonexin II or with the F(ab')2 fragment of anti-human liver endonexin II immunoglobulin G: evidence for a receptor-ligand-like relationship between small hepatitis B surface antigen and endonexin II.

In a previous study, we have identified endonexin II (E-II) on human liver plasma membranes as a specific, Ca(2+)-dependent, small hepatitis B surface antigen (HBsAg)-binding protein. In this article, we describe the spontaneous development of anti-HBs antibodies in rabbits immunized with native or recombinant human liver E-II and in chickens immunized with the F(ab')2 fragment of rabbit anti-human liver E-II immunoglobulin G. Anti-HBs activity was not observed in rabbits immunized with rat liver E-II. Cross-reactivity of anti-E-II antibodies to HBsAg epitopes was excluded, since anti-HBs and anti-E-II activities can be separated by E-II affinity chromatography. The existence of an anti-idiotypic antibody is further demonstrated by competitive binding of human liver E-II and this antibody (Ab2) to small HBsAg, suggesting that Ab2 mimics a specific E-II epitope that interacts with small HBsAg. In addition, it was demonstrated that anti-HBs antibodies developed in rabbits after immunization with intact human liver E-II or in chickens after immunization with F(ab')2 fragments of rabbit anti-human liver E-II immunoglobulin G recognize the same epitopes on small HBsAg. These findings strongly indicate that human liver E-II is a very specific small HBsAg-binding protein and support the assumption that human liver E-II is the hepatitis B virus receptor protein.     

质体醌重组的D1／D2／Cytb559复合物的荧光衰减动力学和光破坏作用

光系统Ⅱ反应中心Ｄ１／Ｄ２／Ｃｙｔｂ５５９ 在分离纯化过程中失去了电子受体ＱＡ 和ＱＢ,人工合成的质体醌可以与Ｄ１／Ｄ２／Ｃｙｔｂ５５９ 复合物发生重组。癸基质体醌（ＤＰＱ）与Ｄ１／Ｄ２／Ｃｙｔｂ５９９ 复合物的重组导致该复合物的荧光强度下降及发射光谱蓝移,同时两个与光化学活性相关的长寿命（２４ ｎｓ和７３ ｎｓ）荧光衰减组分占整个荧光的百分数下降,这些结果表明ＤＰＱ作为Ｐｈｅｏ－ 的电子受体,限制了Ｐ６８０＋ ·Ｐｈｅｏ－ 的电荷重组。ＤＰＱ 的加入对Ｄ１／Ｄ２／Ｃｙｔｂ５５９复合物中Ｃｈｌａ 分子的光破坏敏感性影响不大,但β－胡萝卜素在加入ＤＰＱ 之后可以被光照破坏,这个过程可能与β－胡萝卜素的生理功能相关。     

A shortened dengue IgM capture ELISA using simultaneous incubation of antigen and peroxidase-labeled monoclonal antibody

A shortened IgM capture ELISA for the detection of dengue IgM antibodies using simultaneous incubation of antigen and peroxidase-labeled monoclonal antibody was described. The shortened two-step assay was compared with the four-step IgM capture ELISA on sera from dengue patients confirmed by the hemagglutination inhibition (HI) test. When paired acute and convalescent sera were tested, the shortened ELISA showed 100% agreement with HI results. It detected dengue IgM antibodies in the acute sera of 66% of patients with a primary dengue infection, 60% of patients with a secondary infection, and 98% of patients with a presumptive secondary infection. When the results of 151 dengue patients were combined, 75% of the acute sera were positive by the shortened IgM capture ELISA.     

大鼠脑神经元特异性烯醇化酶基因的cDNA克隆及序列分析

用ＲＴ－ＰＣＲ方法快速克隆了Ｗｉｓｔａｒ大鼠脑神经元特异性烯醇化酶（ＮＳＥ）的ｃＤＮＡ,将此包括编码全长ＮＳＥ４３３个氨基醚的ＤＮＡ片段重组入ｐＵＣ质粒,并用ＰＣＲ方法测定了全部顺序,经重复实验,发现Ｗｉｓｔａｒ大鼠与Ｆｏｒｓｓ－Ｐｅｔｔｅｒ报导的ＳＤ大鼠ＮＳＥ基因顺序,有两外单碱基的差别,其中一个涉及氨基酸的改变。同时还对ＲＮＡ的提取及长片段ＤＮＡ的ＲＴ－ＰＣＲ扩增进行了方法学的探讨。