bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of Novel HLA-A2-Restricted Human Immunodeficiency Virus Type 1-Specific Cytotoxic T-Lymphocyte Epitopes Predicted by the HLA-A2 Supertype Peptide-Binding Motif
Authors: Altfeld Marcus A. , Livingston Brian , Reshamwala Neha , Nguyen Phuong T. , Addo Marylyn M. , Shea Amy , Newman Mark , Fikes John , Sidney John , Wentworth Peggy , Chesnut Robert , Eldridge Robert L. , Rosenberg Eric S. , Robbins Gregory K. , Brander Christian , Sax Paul E. , Boswell Steve , Flynn Theresa , Buchbinder Susan , Goulder Philip J. R. , Walker Bruce D. , Sette Alessandro , Kalams Spyros A.
Published: July 2002

Virus-specific cytotoxic T-lymphocyte (CTL) responses are critical in the control of human immunodeficiency virus type 1 (HIV-1) infection and will play an important part in therapeutic and prophylactic HIV-1 vaccines. The identification of virus-specific epitopes that are efficiently recognized by CTL is the first step in the development of future vaccines. Here we describe the immunological characterization of a number of novel HIV-1-specific, HLA-A2-restricted CTL epitopes that share a high degree of conservation within HIV-1 and a strong binding to different alleles of the HLA-A2 superfamily. These novel epitopes include the first reported CTL epitope in the Vpr protein. Two of the novel epitopes were immunodominant among the HLA-A2-restricted CTL responses of individuals with acute and chronic HIV-1 infection. The novel CTL epitopes identified here should be included in future vaccines designed to induce HIV-1-specific CTL responses restricted by the HLA-A2 superfamily and will be important to assess in immunogenicity studies in infected persons and in uninfected recipients of candidate HIV-1 vaccines.
Rapid Definition of Five Novel HLA-A∗3002-Restricted Human Immunodeficiency Virus-Specific Cytotoxic T-Lymphocyte Epitopes by Elispot and Intracellular Cytokine Staining Assays
Authors: Goulder Philip J. R. , Addo , Marylyn M. , Altfeld Marcus A. , Rosenberg Eric S. , Tang Yanhua , Govender Ugene , Mngqundaniso Nolwandle , Annamalai Ken , Vogel Thorsten U. , Hammond Mike , Bunce Michael , Coovadia Hoosen M. , Walker Bruce D.
Published: July 2002

Human immunodeficiency virus (HIV)-specific cytotoxic T lymphocytes (CTL) play a major role in control of viral replication. To understand the contribution of this antiviral response, an initial step is to fully define the specific epitopes targeted by CTL. These studies focused on CTL responses restricted by HLA-A∗3002, one of the HLA-A molecules most prominent in African populations. To avoid the time-consuming effort and expense involved in culturing CTL prior to defining epitopes and restricting alleles, we developed a method combining Elispot assays with intracellular gamma interferon staining of peripheral blood mononuclear cells to first map the optimal epitopes targeted and then define the HLA restriction of novel epitopes. In two A∗3002-positive subjects whose CTL responses were characterized in detail, the strongest response in both cases was to an epitope in p17 Gag, RSLYNTVATLY (residues 76 to 86). Using this method, CTL epitopes for which there were no motif predictions were optimized and the HLA restriction was established within 48 to 72 h of receipt of blood. This simple and convenient approach should prove useful especially in the characterization of CTL responses specific to HIV and other viruses, particularly in localities where performing cytotoxicity assays would be problematic.
Fluorescent Tagging of Herpes Simplex Virus Tegument Protein VP13/14 in Virus Infection
Authors: Donnelly Michelle , Elliott Gillian
Published: July 2002

The cellular site of herpesvirus tegument assembly has yet to be defined. We have previously used a recombinant herpes simplex virus type 1 expressing a green fluorescent protein (GFP)-tagged tegument protein, namely VP22, to show that VP22 is localized exclusively to the cytoplasm during infection. Here we have constructed a similar virus expressing another fluorescent tegument protein, YFP-VP13/14, and have visualized the intracellular localization of this second tegument protein in live infected cells. In contrast to VP22, VP13/14 is targeted predominantly to the nuclei of infected cells at both early and late times in infection. More specifically, YFP-13/14 localizes initially to the nuclear replication compartments and then progresses into intense punctate domains that appear at around 12 h postinfection. At even later times this intranuclear punctate fluorescence is gradually replaced by perinuclear micropunctate and membranous fluorescence. While the vast majority of YFP-13/14 seems to be targeted to the nucleus, a minor subpopulation also appears in a vesicular pattern in the cytoplasm that closely resembles the pattern previously observed for GFP-22. Moreover, at late times weak fluorescence appears at the cell periphery and in extracellular virus particles, confirming that YFP-13/14 is assembled into virions. This predominantly nuclear targeting of YFP-13/14 together with the cytoplasmic targeting of VP22 may imply that there are multiple sites of tegument protein incorporation along the virus maturation pathway. Thus, our YFP-13/14-expressing virus has revealed the complexity of the intracellular targeting of VP13/14 and provides a novel insight into the mechanism of tegument, and hence virus, assembly.
Expression of Human MxA Protein in Mosquito Cells Interferes with LaCrosse Virus Replication
Authors: Miura Tanya A. , Carlson Jonathan O. , Beaty Barry J. , Bowen Richard A. , Olson Ken E.
Published: July 2002

Human MxA protein inhibits LaCrosse virus (LAC virus; family
Bunyaviridae
) replication in vertebrate cells and
MxA
-transgenic mice. LAC virus is transmitted to humans by
Aedes triseriatus
mosquitoes. In this report, we have shown that transfected mosquito cells expressing the human
MxA
cDNA are resistant to LAC virus but permissive for Sindbis virus (family
Togaviridae
) infection.
Evaluation of Accumulation of Hepatitis C Virus Mutations in a Chronically Infected Chimpanzee: Comparison of the Core, E1, HVR1, and NS5b Regions
Authors: Lu Ling , Nakano Tatsunori , Orito Etsuro , Mizokami Masashi , Robertson Betty H.
Published: July 2002

Four hepatitis C virus genome regions (the core, E1, HVR1, and NS5b) were amplified and sequenced from yearly samples obtained from a chronically infected chimpanzee over a 12-year span. Nucleotide substitutions were found to accumulate in the core, E1, and HVR1 regions during the course of chronic infection; substitutions within the NS5b region were not detected for the first 8 years and were found to be minimal during the last 4 years. The rate of accumulation of mutations in the core and E1 regions, based on a direct comparison between the first 1979 sequence and the last 1990 sequence, was 1.120 × 10
−3
, while phylogenetic ancestral comparison using the 12 yearly sequences showed a rate of 0.816 × 10
−3
bases per site per year. Temporal evaluation of the sequences revealed that there appeared to be periods in which substitutions accumulated and became fixed, followed by periods with relative stasis or random substitutions that did not persist. Synonymous and nonsynonymous substitutions within the core, E1, and HVR1 regions were also analyzed. In the core and E1 regions, synonymous substitutions predominated and gradually increased over time. However, within the HVR1 region, nonsynonymous substitutions predominated but gradually decreased over time.
Modulation of Different Human Immunodeficiency Virus Type 1 Nef Functions during Progression to AIDS
Authors: Carl Silke , Greenough Thomas C. , Krumbiegel Mandy , Greenberg Michael , Skowronski Jacek , Sullivan John L. , Kirchhoff Frank
Published: July 2002

The human immunodeficiency virus type 1 (HIV-1) Nef protein has several independent functions that might contribute to efficient viral replication in vivo. Since HIV-1 adapts rapidly to its host environment, we investigated if different Nef properties are associated with disease progression. Functional analysis revealed that
nef
alleles obtained during late stages of infection did not efficiently downmodulate class I major histocompatibility complex but were highly active in the stimulation of viral replication. In comparison, functional activity in downregulation of CD4 and enhancement of HIV-1 infectivity were maintained or enhanced after AIDS progression. Our results demonstrate that various Nef activities are modulated during the course of HIV-1 infection to maintain high viral loads at different stages of disease progression. These findings suggest that all in vitro Nef functions investigated contribute to AIDS pathogenesis and indicate that
nef
variants with increased pathogenicity emerge in a significant number of HIV-1-infected individuals.
Ability of the V3 Loop of Simian Immunodeficiency Virus To Serve as a Target for Antibody-Mediated Neutralization: Correlation of Neutralization Sensitivity, Growth in Macrophages, and Decreased Dependence on CD4
Authors: Means Robert E. , Matthews Thomas , Hoxie James A. , Malim Michael H. , Kodama Toshiaki , Desrosiers Ronald C.
Published: July 2002

To better define the effects of sequence variation and tropism on the ability of the simian immunodeficiency virus SIVmac V3 loop to act as a target of antibody-mediated neutralization, a series of experiments were performed. Three SIV strains, SIVmac239, SIVmac316, and SIVmac155/T3, each with defined differences in
env
sequence and tropism, were used to construct a panel of viruses chimeric for a portion of envelope that includes the V2 and V3 regions. Peptides with sequences corresponding to the V3 loops of the parental viruses were used to immunize rabbits. The polyclonal rabbit antibodies and plasma from SIVmac239-infected animals were then used to assess the neutralization sensitivity of the parental and chimeric viruses. One of the parental viruses, SIVmac316, which is able to replicate to high titer in alveolar macrophages and can infect cells in a CD4-independent fashion, was highly sensitive to neutralization by plasma from SIVmac-infected rhesus macaques, with average 50% neutralization titers of 1:20,480; this same strain was also sensitive to neutralization by the anti-V3 loop peptide sera. Other parental and chimeric viruses were less sensitive to neutralization with this same panel of antibodies, but as seen with SIVmac316, those viruses that were able to productively replicate in alveolar macrophages were more sensitive to antibody-mediated neutralization. To further define the amino acids involved in increased sensitivity to neutralization, a panel of viruses was constructed by changing envelope residues in SIVmac316 to the corresponding SIVmac239 amino acids. The increased neutralization sensitivity observed for SIVmac316 was mapped principally to three amino acid changes spread throughout gp120. In addition, the increased sensitivity to neutralization by V3-directed antibodies correlated with the ability of the various viruses to replicate to high levels in alveolar macrophage cultures and a CD4-negative cell line, BC7/CCR5. These results demonstrate that the V3 loop of SIVmac Env can act as an efficient target of neutralizing antibodies in a fashion that is highly dependent on sequence context. In addition, these studies suggest a correlation between decreased dependence on CD4 and increased sensitivity to antibody-mediated neutralization.
Quintuple Deglycosylation Mutant of Simian Immunodeficiency Virus SIVmac239 in Rhesus Macaques: Robust Primary Replication, Tightly Contained Chronic Infection, and Elicitation of Potent Immunity against the Parental Wild-Type Strain
Authors: Mori Kazuyasu , Yasutomi Yasuhiro , Ohgimoto Shinji , Nakasone Tadashi , Takamura Shiki , Shioda Tatsuo , Nagai Yoshiyuki
Published: July 2002

We previously generated a mutant of simian immunodeficiency virus (SIV) lacking 5 of a total of 22 N-glycans in its external envelope protein gp120 with no impairment in viral replication capability and infectivity in tissue culture cells. Here, we infected rhesus macaques with this mutant and found that it also replicated robustly in the acute phase but was tightly, though not completely, contained in the chronic phase. Thus, a critical requirement for the N-glycans for the full extent of chronic infection was demonstrated. No evidence indicating reversion to a wild type was obtained during the observation period of more than 40 weeks. Monkeys infected with the mutant were found to tolerate a challenge infection with wild-type SIV very well. Analyses of host responses following challenge revealed no neutralizing antibodies against the challenge virus but strong secondary responses of cytotoxic T lymphocytes against multiple antigens, including Gag-Pol, Nef, and Env. Thus, the quintuple deglycosylation mutant appeared to represent a novel class of SIV live attenuated vaccine.
Increased Efficacy of the Immunoglobulin G2a Subclass in Antibody-Mediated Protection against Lactate Dehydrogenase-Elevating Virus-Induced Polioencephalomyelitis Revealed with Switch Mutants
Authors: Markine-Goriaynoff Dominique , Coutelier Jean-Paul
Published: July 2002

Immunoglobulin G1 (IgG1), IgG2a, and IgG2b switch variants were derived from an IgG3 monoclonal antibody directed against the VP3 envelope glycoprotein of lactate dehydrogenase-elevating virus (LDV). Among the four antibodies, IgG2a delayed the onset and progression of LDV-induced polioencephalomyelitis more than did the other subclasses. This suggests that the IgG2a predominance observed in many IgG antibody responses elicited by live viruses could, at least under some circumstances, correspond to the selection of the best protection for the infected host.
Gammaherpesvirus Lytic Gene Expression as Characterized by DNA Array
Authors: Ahn Joo Wook , Powell Kenneth L. , Kellam Paul , Alber Dagmar G.
Published: July 2002

Gammaherpesviruses are associated with a number of diseases including lymphomas and other malignancies. Murine gammaherpesvirus 68 (MHV-68) constitutes the most amenable animal model for this family of pathogens. However experimental characterization of gammaherpesvirus gene expression, at either the protein or RNA level, lags behind that of other, better-studied alpha- and beta-herpesviruses. We have developed a cDNA array to globally characterize MHV-68 gene expression profiles, thus providing an experimental supplement to a genome that is chiefly annotated by homology. Viral genes started to be transcribed as early as 3 h postinfection (p.i.), and this was followed by a rapid escalation of gene expression that could be seen at 5 h p.i. Individual genes showed their own transcription profiles, and most genes were still being expressed at 18 h p.i. Open reading frames (ORFs) M3 (chemokine-binding protein), 52, and M9 (capsid protein) were particularly noticeable due to their very high levels of expression. Hierarchical cluster analysis of transcription profiles revealed four main groups of genes and allowed functional predictions to be made by comparing expression profiles of uncharacterized genes to those of genes of known function. Each gene was also categorized according to kinetic class by blocking de novo protein synthesis and viral DNA replication in vitro. One gene, ORF 73, was found to be expressed with α-kinetics, 30 genes were found to be expressed with β-kinetics, and 42 genes were found to be expressed with γ-kinetics. This fundamental characterization furthers the development of this model and provides an experimental basis for continued investigation of gammaherpesvirus pathology.
Deubiquitinating Function of Adenovirus Proteinase
Authors: Balakirev Maxim Y. , Jaquinod Michel , Haas Arthur L. , Chroboczek Jadwiga
Published: July 2002

The invasion strategy of many viruses involves the synthesis of viral gene products that mimic the functions of the cellular proteins and thus interfere with the key cellular processes. Here we show that adenovirus infection is accompanied by an increased ubiquitin-cleaving (deubiquitinating) activity in the host cells. Affinity chromatography on ubiquitin aldehyde (Ubal), which was designed to identify the deubiquitinating proteases, revealed the presence of adenovirus L3 23K proteinase (Avp) in the eluate from adenovirus-infected cells. This proteinase is known to be necessary for the processing of viral precursor proteins during virion maturation. We show here that in vivo Avp deubiquitinates a number of cellular proteins. Analysis of the substrate specificity of Avp in vitro demonstrated that the protein deubiquitination by this enzyme could be as efficient as proteolytic processing of viral proteins. The structural model of the Ubal-Avp interaction revealed some similarity between S1-S4 substrate binding sites of Avp and ubiquitin hydrolases. These results may reflect the acquisition of an advantageous property by adenovirus and may indicate the importance of ubiquitin pathways in viral infection.
The Envelope Glycoprotein of Human Endogenous Retrovirus Type W Uses a Divergent Family of Amino Acid Transporters/Cell Surface Receptors
Authors: Lavillette Dimitri , Marin Mariana , Ruggieri Alessia , Mallet François , Cosset François-Loïc , Kabat David
Published: July 2002

The human endogenous retrovirus type W (HERV-W) family includes proviruses with intact protein-coding regions that appear to be under selection pressure, suggesting that some HERV-W proviruses may remain active in higher primates. The envelope glycoprotein (Env) encoded by HERV-W is highly fusogenic, is naturally expressed in human placental syncytiatrophoblasts, and has been reported to function as a superantigen in lymphocyte cultures. Recent evidence suggested that HERV-W Env can mediate syncytium formation by interacting with the human sodium-dependent neutral amino acid transporter type 2 (hASCT2; gene name, SLC1A5) (J.-L. Blond, D. Lavillette, V. Cheynet, O. Bouton, G. Oriol, S. Chapel-Fernandez, B. Mandrand, F. Mallet, and F.-L. Cosset, J. Virol. 74:3321-3329, 2000) and that it can pseudotype human immunodeficiency virus cores (D. S. An, Y. Xie, and I. S. Y. Chen, J. Virol. 75:3488-3489, 2001). By using cell-cell fusion and pseudotype virion infection assays, we found that HERV-W Env efficiently uses both hASCT2 and the related transporter hASCT1 (gene name, SLC1A4) as receptors. In addition, although HERV-W Env mediates only slight syncytium formation or infection of mouse cells, it utilizes the mouse transporters mASCT1 and mASCT2 when their sites for N-linked glycosylation are eliminated by mutagenesis. Consistent with their role as a battlefield in host-virus coevolution, the viral recognition regions in ASCT1 and ASCT2 of humans and mice are highly divergent compared with other regions of these proteins, and their ratios of nonsynonymous to synonymous nucleotide sequence changes are extremely large. The recognition of ASCT1 and ASCT2 despite this divergence of their sequences strongly suggests that the use of both receptors has been highly advantageous for survival and evolution of the HERV-W family of retroviruses.
Identification of Two Cross-Neutralizing Linear Epitopes within the L1 Major Capsid Protein of Human Papillomaviruses
Authors: Combita Alba-Lucia , Touzé Antoine , Bousarghin Latifa , Christensen Neil D. , Coursaget Pierre
Published: July 2002

The neutralizing activities of polyclonal antibodies and monoclonal antibodies (MAbs) obtained by immunization of mice with L1 virus-like particles (VLPs) were investigated by using pseudovirion infectivity assays for human papillomavirus type 16 (HPV-16), HPV-31, HPV-33, HPV-45, HPV-58, and HPV-59 to obtain a better definition of cross-neutralization between high-risk HPVs. In this study, we confirmed and extended previous studies indicating that most genital HPV genotypes represent separate serotypes, and the results suggest that the classification of serotypes is similar to that of genotypes. In addition, three cross-neutralizing MAbs were identified (HPV-16.J4, HPV-16.I23, and HPV-33.E12). MAb HPV-16.J4 recognized a conserved linear epitope located within the FG loop of the L1 protein, and HPV-16.I23 recognized another located within the DE loop. The results suggested that reactivity of MAb HPV-16.I23 to L1 protein is lost when leucine 152 of the HPV-16 L1 protein is replaced by phenylalanine. This confirmed the existence of linear epitopes within the L1 protein that induce neutralizing antibodies, and this is the first evidence that such linear epitopes induce cross-neutralization. However, the cross-neutralization induced by L1 VLPs represents less than 1% of the neutralizing activity induced by the dominant conformational epitopes, and it is questionable whether this is sufficient to offer cross-protection in vivo.
Cross-Protection against Challenge with Puumala Virus after Immunization with Nucleocapsid Proteins from Different Hantaviruses
Authors: de Carvalho Nicacio Cristina , Gonzalez Della Valle Marcelo , Padula Paula , Björling Ewa , Plyusnin Alexander , Lundkvist Åke
Published: July 2002

Hantaviruses are rodent-borne agents that cause hemorrhagic fever with renal syndrome or hantavirus pulmonary syndrome in humans. The nucleocapsid protein (N) is relatively conserved among hantaviruses and highly immunogenic in both laboratory animals and humans, and it has been shown to induce efficient protective immunity in animal models. To investigate the ability of recombinant N (rN) from different hantaviruses to elicit cross-protection, we immunized bank voles with rN from Puumala (PUUV), Topografov (TOPV), Andes (ANDV), and Dobrava (DOBV) viruses and subsequently challenged them with PUUV. All animals immunized with PUUV and TOPV rN were completely protected. In the group immunized with DOBV rN, 7 of 10 animals were protected, while only 3 of 8 animals were protected in the group immunized with ANDV rN, which is more closely related to PUUV rN than DOBV rN. Humoral and cellular immune responses after rN immunization were also investigated. The highest cross-reactive humoral responses against PUUV antigen were detected in sera from ANDV rN-immunized animals, followed by those from TOPV rN-immunized animals, and only very low antibody cross-reactivity was observed in sera from DOBV rN-immunized animals. In proliferation assays, T lymphocytes from animals immunized with all heterologous rNs were as efficiently recalled in vitro by PUUV rN as were T lymphocytes from animals immunized with homologous protein. In summary, this study has shown that hantavirus N can elicit cross-protective immune responses against PUUV, and the results suggest a more important role for the cellular arm of the immune response than for the humoral arm in cross-protection elicited by rN.
Adenovirus Serotype 30 Fiber Does Not Mediate Transduction via the Coxsackie-Adenovirus Receptor
Authors: Law Lane K. , Davidson Beverly L.
Published: July 2002

Prior work by members of our laboratory and others demonstrated that adenovirus serotype 30 (Ad30), a group D adenovirus, exhibited novel transduction characteristics compared to those of serotype 5 (Ad5, belonging to group C). While some serotype D adenoviruses bind to the coxsackie-adenovirus receptor (CAR), the ability of Ad30 fiber to bind CAR is unknown. We amplified and purified Ad30 and cloned the Ad30 fiber by overlap PCR. Alignment of Ad30 fiber with Ad3, Ad35, Ad5, Ad9, and Ad17 revealed that Ad30, like Ad9 and Ad17, has a shortened fiber sequence relative to that of Ad5. The knob region of fiber was 45% identical to that of the Ad5 knob regions. We made a chimeric recombinant virus (Ad5GFPf30) in which the Ad5 fiber (amino acids [aa]47 to 582) was replaced with Ad30 fiber sequences (aa 46 to 372), and CAR-mediated viral entry was determined on CAR-expressing Chinese hamster ovary (CHO) cells. While CAR expression significantly increased Ad5GFP-mediated transduction in CHO cells (from 1 to 36%), it did not enhance Ad5GFPf30 gene transfer. Binding of radiolabeled Ad5GFPf30 or Ad30 wild-type virus was also not improved by the expression of CAR. These results suggest that Ad30 fiber is distinct from Ad5, Ad9, and Ad17 fibers in its inability to direct transduction via CAR.
Human Immunodeficiency Virus Type 1 (HIV-1)Quasispecies at the Sites ofMycobacterium tuberculosisInfectionContribute to Systemic HIV-1 Heterogeneity
Authors: Collins Kalonji R. , Quiñones-Mateu Miguel E. , Wu Mianda , Luzze Henry , Johnson John L. , Hirsch Christina , Toossi Zahra , Arts Eric J.
Published: July 2002

We have recently reported an increased heterogeneity in the human immunodeficiency virus type 1 (HIV-1) envelope gene (env) in HIV-1-infected patients with pulmonary tuberculosis (TB) compared to patients with HIV-1 alone. This increase may be a result of dissemination of lung-derived HIV-1 isolates from sites ofMycobacterium tuberculosisinfection and/or the systemic activation of the immune system in response to TB. To distinguish between these two mechanisms, blood and pleural fluid samples were obtained from HIV-1-infected patients with active pleural TB in Kampala, Uganda (CD4 cell counts of 34 to 705 cells/μl, HIV-1 plasma loads of 2,400 to 280,000 RNA copies/ml, and HIV-1 pleural loads of 7,600 to 4,500,000 RNA copies/ml). The C2-C3 coding region of HIV-1envwas PCR amplified from lysed peripheral blood mononuclear cells and pleural fluid mononuclear cells and reverse transcriptase-PCR amplified from plasma and pleural fluid HIV-1 virions of eight HIV-1 patients with pleural TB. Phylogenetic and phenetic analyses revealed a compartmentalization of HIV-1 quasispecies between blood and pleural space in four of eight patients, with migration events between the compartments. There was a trend for a greater genetic heterogeneity in the pleural space, which may be the result of anM. tuberculosis-mediated increase in HIV-1 replication and/or selection pressure at the site of infection. Collectively, these findings suggest that HIV-1 quasispecies in theM. tuberculosis-infected pleural space may leak into the systemic circulation and lead to increased systemic HIV-1 heterogeneity during TB.
Baculovirus Replication Factor LEF-1 Is a DNA Primase
Authors: Mikhailov Victor S. , Rohrmann George F.
Published: July 2002

The baculovirus replication factors LEF-1 and LEF-2 of the
Autographa californica
multinucleocapsid nucleopolyhedrovirus were overexpressed as fusions containing a hemagglutinin (HA) epitope and a HIS
6
tag using recombinant baculoviruses. LEF-1 was purified to near homogeneity and found to have primase activity in an indirect assay employing
Escherichia coli
DNA polymerase I (Klenow enzyme) and poly(dT) template. The LEF-1 primase products were also directly characterized by electrophoresis in 20% polyacrylamide-8 M urea gels and agarose gels. Primer synthesis was time dependent, and products of several hundred nucleotides or more were observed from the M13 single-stranded DNA (ssDNA) template. The LEF-1 primase was absolutely dependent on divalent cations (Mg
2+
), and optimal activity was supported by 10 mM MgCl
2
. An alkaline pH (8.8 to 9.4) was optimal, whereas monovalent salt (KCl) was inhibitory. Mutation of an invariant aspartic acid in a putative primase domain caused LEF-1 activity to be abolished. Upon ultracentrifugation in glycerol gradients, LEF-1 was found to have a sedimentation coefficient of 3S that is consistent with its being present as a monomer. Elution profiles of LEF-1 and LEF-2 from ssDNA-cellulose and DEAE resin suggested that LEF-2 may bind to both DNA and LEF-1.
Magnitude of Functional CD8+T-Cell Responses to the Gag Protein of Human Immunodeficiency Virus Type 1 Correlates Inversely with Viral Load in Plasma
Authors: Edwards Bradley H. , Bansal Anju , Sabbaj Steffanie , Bakari Janna , Mulligan Mark J. , Goepfert Paul A.
Published: July 2002

The importance of CD8+T-cell responses in the control of human immunodeficiency virus type 1 (HIV-1) infection has been demonstrated, yet few studies have been able to correlate these responses with markers of HIV-1 disease progression. This study measured cell-mediated immune responses using peripheral blood mononuclear cells (PBMC) obtained from 27 patients with chronic HIV-1 infection, the majority of whom were off antiretroviral therapy. The ELISPOT assay was used to detect gamma interferon-secreting PBMC after stimulation with overlapping HIV-1 peptides spanning the Gag, Pol, Env, and Nef proteins in addition to the baculovirus-derived p24 and gp160 proteins. All volunteers had responses to at least one HIV-1-specific peptide. All but one of the subjects (96%) responded to the Gag peptide pool, and 86% responded to the Pol and/or Nef peptide pools. The magnitude and the breadth of T-cell responses directed to either the Gag or p24 peptide pools correlated inversely with viral load in plasma (r= −0.60,P< 0.001 andr= −0.52,P< 0.005, respectively) and directly with absolute CD4+T-cell counts (r= 0.54,P< 0.01 andr= 0.39,P< 0.05, respectively) using the Spearman rank correlation test. Responses to the Pol and integrase peptide pools also correlated with absolute CD4+T-cell counts (r= 0.45,P< 0.05 andr= 0.49,P< 0.01, respectively). No correlation with markers of disease progression was seen with specific T-cell responses directed toward the Env or Nef peptides. These data serve as strong evidence that major histocompatibility complex class I presentation of Gag peptides is an essential feature for any HIV-1 vaccine designed to elicit optimal CD8+T-cell responses.
Nucleoprotein Structure of Immediate-Early Promoters Zp and Rp and of oriLyt of Latent Epstein-Barr Virus Genomes
Authors: Niller Hans Helmut , Salamon Daniel , Uhlig Jörg , Ranf Stefanie , Granz Marcus , Schwarzmann Fritz , Wolf Hans , Minarovits Janos
Published: July 2002

Genomic footprints across Rp, Zp, and oriLyt of Epstein-Barr virus (EBV) have been conducted in a panel of latently infected B-cell lines. Close protein-base contacts were found about 360 nucleotides upstream of the Zp initiation site. Gel shifts and transient transfection assays indicated that an Sp1-NF1 locus may serve as a repressive transcriptional element against Zp induction from latent EBV genomes.
Synthesis of Peptides Influencing Growth Hormone Release
Authors: Hlaváček Jan , Smékal Otto , Pospíšek Jan , Barth Tomislav
Published: July 2002

A solid phase peptide synthesis of 17 growth hormone (GH) releasing peptide analogues Ia - IIIc for their use in a pharmacological assay on GH release is described. While the linear peptide amides Ia - Ij were synthesized on p-methylbenzhydrylamine resin using Boc strategy, and cleaved by HF in the presence of scavengers the linear peptide amides Ik and Il were prepared on Merrifield benzyl ester type resin using Fmoc strategy and cleaved by ammonolysis. The deleted peptide amides IIa and IIb were obtained as by-products during HPLC purification of analogues Ic and Id. The linear precursors of cyclic peptides IIIa - IIIc were also prepared on Merrifield resin and cleaved under mild alkaline conditions. Their cyclization was performed in solution by diphenylphosphoryl azide.
Interfacial Tension Along the Binodal Curve in the Benzene-Ethanol-Water and n-Hexane-Acetone-Water Ternary Systems at 25 °C
Authors: Čechová Markéta , Bartovská Lidmila
Published: July 2002

The surface tensions, interfacial tensions and densities of conjugate solutions of compositions lying in the miscibility gap were measured for two ternary systems, viz. benzene-ethanol-water and n-hexane-acetone-water, at 25 °C.
Improved Methods for Obtaining Immonium Perchlorates and Enamines of Solanidine Type Steroidal Alkaloids
Authors: Penov Gaši Katarina M. , Čolic Dušica Rackov , Arcson Otto N. , Sakač Zvonimir O. , Djurendić Evgenija A. , Sakač Marija N. , Medic-Mijačevic Ljubica , Miljković Dušan A.
Published: July 2002

Oxidation of solanidine and some of its derivatives with mercury(II) acetate in acetone afforded corresponding N-22 immonium perchlorates or 22(23)-enamines. It has been found that the effect of solvents on regioselectivity e.g. formation N-22 and N-16 immonium salts, was remarkable. The isomerisation of 22(23)-enamines into corresponding 20(22)-enamines, potentialy important intermediates in planned chemical degradation of solanidine to steroidal hormones, was performed in 0.1% acetic acid in ether, or in dichloromethane (chloroform).
Use of the Ion-Exchange Properties of Layered Double Hydroxides for Water Purification
Authors: Houri Boumadiane , Legrouri Ahmed , Barroug Allal , Forano Claude , Besse Jean-Pierre
Published: July 2002

The sorption of chromate and dichromate ions by a [Zn-Al-Cl] layered double hydroxide was investigated. The anionic clay, prepared by coprecipitation at pH 8, was found to be pure upon characterization by X-ray diffraction, Fourier-transform infrared spectroscopy, and thermogravimetry and to have low BET surface area and small pore volume. The interaction of the clay with both chromate and dichromate ions has been studied by UV-VIS spectroscopy. The treatment of water containing 15-300 ppm of CrO42- or Cr2O72- was successful. The sorption capacities of the clay for CrO42- and Cr2O72- are close to 1 and 0.6 mmol/g, respectively.
Syntheses, Spectroelectrochemistry and Photoinduced Electron-Transfer Processes of Novel Ru and Os Dyad and Triad Complexes with Functionalized Diimide Ligands
Authors: Hossain M. Delower , Haga Masa-aki , Gholamkhass Bobak , Nozaki Koichi , Tsushima Minoru , Ikeda Noriaki , Ohno Takeshi
Published: July 2002

A series of mono- and dinuclear Ru complexes containing a bridging ligand, L-diimide-L (L = 2-(2-pyridyl)benzimidazole; diimide = benzene-1,2:4,5-bis(dicarboximide) (bdi; pyromellitimide) or naphthalene-1,8:4,5-bis(dicarboximide) (ndi), with either propane or xylene group as linkers, have been prepared. The mono- and dinuclear Ru complexes containing the bdi or ndi ligand, exhibit characteristic metal-to-ligand charge transfer (MLCT) transition at 458 nm. The mono- and dinuclear Ru/Os complexes exhibit a rich redox chemistry arising from both M(II) to M(III) oxidation and ligand-based consecutive reduction of diimide and 2,2'-bipyridine (bpy) ligands. The emission decays fit well with double- or triple-exponential decay models. The non-exponential decay curve reveals the existence of several conformers in solution due to the flexible propane or p-xylene linker. The much shorter emission lifetimes of the [M(bpy)2(L-diimide-L)] complexes compared with the parent [M(bpy)2L] indicates the intramolecular electron-transfer from the excited M(bpy)2 moiety to the diimide. The fastest rates of electron transfer (3 · 1010 s-1) are attributable to the folded conformers suitable for the electron donor/acceptor through-space interaction. A time-resolved absorption spectroscopic study of the dinuclear bdi and ndi complexes revealed appearance of the electron-transfer products, M(III) and the diimide radical anion, and their rapid disappearance. The effect of the linkers of the ligand L-diimide-L on the rates of electron transfer and the back electron transfer is also discussed.
Avulsion fracture of the extensor carpi radialis longus in a rugby player: a case report
Authors: Jena D , Giannikas K A , Din R
Published: July 2002

Avulsion fracture of the base of the second metacarpal is an unusual injury, and the cause in the few cases reported in the literature was a fall on a volarly flexed wrist. A case of this rare injury suffered in a sport related accident by a semiprofessional rugby player is reported. It was treated with open reduction and internal fixation after failure of conservative management.
Incidence of injury in amateur rugby league sevens
Authors: Gabbett T J
Published: July 2002

Objectives: To investigate the incidence, site, and nature of injuries sustained in amateur rugby league sevens tournaments.

Methods: A total of 168 players competing in three amateur rugby league sevens tournaments were studied. All injuries sustained during matches were recorded. Information recorded included the name of the injured player and the time, cause, anatomical site, and nature of the injury.

Results: The incidence of injury was 283.5 per 1000 playing hours. Some 40% (113.4 per 1000 playing hours) of all injuries sustained were to the lower limb (χ2= 5.3, df = 1, p<0.05). Contusions were the most common type of injury (113.4 per 1000 hours, 40%, χ2= 9.5, df = 4, p<0.05), with most (198.4 per 1000 hours, 70%, χ2= 31.5, df =4, p<0.001) occurring in physical collisions and tackles. An increasing injury incidence was observed over the first (99.2 per 1000 hours), second (198.4 per 1000 hours), third (347.2 per 1000 hours), and fourth (694.4 per 1000 hours) matches played during the tournaments (χ2= 9.2, df = 3, p<0.05).

Conclusions: The results of this study suggest that amateur rugby league sevens tournaments, which require players to compete repeatedly on the same day, may hasten the onset of fatigue and predispose to injury.
Incidence of lymphohaematopoietic malignancies in a petrochemical industry cohort: 1983–94 follow up
Authors: Huebner Wendy W , Chen Vivien W , Friedlander Barry R , Wu Xiao C , Jorgensen Gail , Bhojani Faiyaz A , Friedmann Craig H , Schmidt Beth A , Sales Eugene A , Joy Javed A , Correa Catherine N
Published: July 2002

OBJECTIVES

In response to a previous finding of increased mortality from lymphohaematopoietic (LH) malignancies, this study examines incidence of LH malignancy in a petrochemical industry cohort. Emphasis is on chronic lymphocytic leukaemia (CLL) and on comparisons by period of first employment.

METHOD

The study cohort consists of 8942 employees who were active in the period 1970–92 and alive on 31 December 1982. Record linkage with the Louisiana tumour registry (LTR) provided information on cancer for cases occurring between 1983 and 1994. Standardised incidence ratios (SIR), with the south Louisiana population as a comparison, were computed for all cancers, all LH malignancies and specific LH subtypes. Analyses were conducted for sex and race categories, and by period of first employment, job type, duration of employment, and latency.

RESULTS

672 Cases of cancer were identified, including 59 LH malignancies. Women (n=1169) had an overall cancer SIR below unity and four LH malignancies versus 2.28 expected. Among the 7773 men, those first employed before 1950 had no overall cancer excess, a significant 1.4-fold increase in overall LH malignancies (43 observed versus 30.78 expected), and four CLL cases versus 3.27 expected. Findings for men first employed after 1950 are based on fewer cases, but there was no indication of excesses of overall cancer or LH malignancy. Numbers were too small in the group first employed after 1950 for meaningful analysis of LH malignancy subtypes such as CLL (one case).

CONCLUSION

These findings do not suggest a continuing excess of CLL but do suggest a small increase in incidence of overall LH malignancy for workers first employed before 1950. This may reflect associations with earlier workplace conditions, although work related patterns are mixed. Interpretation is limited by the diverse group of diseases within LH malignancies, and the lack of control for non-work factors other than sex, age, race, and period of diagnosis. This study has a major advantage of more complete and reliable cancer ascertainment compared with the mortality investigation, and shows the feasibility and benefits of using cancer registry incidence data in an occupational cohort study.
Gallstones and cholecystectomy in modern Britain
Authors: Bateson M C
Published: July 2002

Background

It has been suggested that gallstone disease is now commoner, and that this might explain an increase in cholecystectomy rates, though conclusive evidence has been lacking.

Methods

All the non-forensic necropsy results for Dundee 1953–98 were examined to assess the prevalence of gallstone disease. The NHS Scotland annual cholecystectomy figures were extracted from their earliest availability in 1961 up to the present. The subgroup of patients from Dundee was analysed separately, as were laparoscopic procedures, which were recorded from 1991.

Results

Gallstone disease was much commoner in 1974–98 than in 1953–73. Increasing age was the main determinant of gallstone disease. Though gallstone disease was commoner in women than men aged 40–89, there was no sex difference under 40 or over 90 years. Cholecystectomy became much commoner in the 1960s when frequency of gallstone disease did not change. It increased further in the 1970s, peaking in 1977–8. There was a gradual fall in rates in the 1980s when gallstone prevalence remained high. There was a further moderate rise in the 1990s after the wide introduction of laparoscopic cholecystectomy. Cholecystectomy is now much commoner in young women and this change started in the 1960s. By contrast, cholecystectomy in men has become more prevalent in the older age group.

Conclusions

Gallstones were definitely more common in both sexes at all ages over 40 in the last 25 years. Changes in the cholecystectomy rates are only partly explained by changes in gallstone prevalence, and are more determined by surgical practice.
Mitochondrial cytopathy presenting with focal segmental glomerulosclerosis, hypoparathyroidism, sensorineural deafness, and progressive neurological disease
Authors: Hameed R , Raafat F , Ramani P , Gray G , Roper H P , Milford D V
Published: July 2002

A 6 year old boy who presented with steroid unresponsive nephrotic syndrome is reported. He was found to have focal segmental glomerulosclerosis and associated hypoparathyroidism and sensorineural deafness. The child progressed to end stage renal failure and was successfully managed by dialysis and cadaveric renal transplantation. He later developed progressive neurological deterioration and mitochondrial myopathy and neuropathy was diagnosed.
Hydroelectric development and translocation ofGalaxias brevipinnis: a cloud at the end of the tunnel?
Authors: Waters Jonathan M , Shirley Michael , Closs Gerard P
Published: July 2002

Two major drainages of southeastern Australia, the Snowy River and the Murray River, were artificially linked by a major hydroelectric project during the early 20th century. This development diverts Snowy River flow into tributaries of the Murray River via a series of extensive tunnels. In 1990, fish surveys of the upper Murray River system recorded Galaxias brevipinnis, an aggressive migratory species previously unrecorded from the drainage. We used genetic analysis to discriminate between alternative hypotheses for Murray River G. brevipinnis: (i) anthropogenic translocation via the Snowy River diversion or (ii) a previously undiscovered natural population. Landlocked G. brevipinnis from the Murray River (43 fish, eight control region haplotypes) and Snowy River (39 fish, 11 haplotypes) exhibit similar levels of mtDNA diversity, share six haplotypes, and are not significantly differentiated for microsatellite loci (p = 0.0884). Coastal samples exhibit higher haplotypic diversity (40 fish, 20 haplotypes) but share only three haplotypes with Murray River and are significantly differentiated from Murray River samples for microsatellite loci (p = 0.0008). Our data are consistent with the translocation hypothesis but are generally inconsistent with a natural origin for Murray River G. brevipinnis. The suggested human-mediated translocation represents a risk to native fauna.
Effects of captopril and omapatrilat on early post-myocardial infarction survival and cardiac hemodynamics in rats: interaction with cardiac cytokine expression
Authors: Blais Jr. Charles , Lapointe Nathalie , Rouleau Jean-Lucien , Clément Robert , Bachvarov Dimcho R , Adam Albert
Published: July 2002

The purpose of this study was to evaluate and compare the effects of simultaneous angiotensin-converting enzyme (ACE) and neutral endopeptidase 24.11 (NEP) inhibition by the vasopeptidase inhibitor omapatrilat (10 and 40 mg·kg1·day1) with those of the selective ACE inhibitor captopril (160 mg·kg1·day1) on survival, cardiac hemodynamics, and cytokine mRNA expression in left ventricular (LV) tissues 4 days after myocardial infarction (MI) in rats. The effects of the co-administration of both B1 and B2 kinin receptor antagonists (2.5 mg·kg1·day1 each) with and without omapatrilat were also evaluated to assess the role of bradykinin (BK) during this post-MI period. Both omapatrilat and captopril treatments improve early (4 days) post-MI survival when started 4 h post-MI. The use of kinin receptor antagonists had no significant effect on survival in untreated MI rats and omapatrilat-treated MI rats. This improvement in survival with omapatrilat and captopril is accompanied by a reduced LV end-diastolic pressure (LVEDP) and pulmonary congestion. The use of kinin receptor antagonists had little effect on cardiac hemodynamics or morphologic measurements. Acute MI significantly increased the expression of cardiac cytokines (TNF-α, TGF-β1, and IL-10). Captopril significantly attenuated this activation, while omapatrilat had variable effects: sometimes increasing but generally not changing activation depending on the cytokine measured and the dose of omapatrilat used. The co-administration of both kinin receptor antagonists attenuates the increase in expression of cardiac TNF-α and TGF-β1 after omapatrilat treatment. Taken together, these results would suggest that despite very marked differences in the way these drugs modified the expression of cardiac cytokines, both omapatrilat and captopril improved early (4 days) post-MI survival and cardiac function to a similar extent.Key words: ACE inhibitor, cytokines, kinins, myocardial infarction, vasopeptidase inhibitor.
Is there a relationship between fluctuating asymmetry and reproductive investment in perch (Perca fluviatilis)?
Authors: Øxnevad S A , Heibo E , Vøllestad L A
Published: July 2002

Fluctuating asymmetry (FA), or random deviation from perfect bilateral symmetry, is often used as an indicator of perturbed development. Several studies attempt to correlate FA with components of individual fitness or population viability. In this study we test for a correlation between FA and four fitness traits in female Eurasian perch (Perca fluviatilis) inhabiting acidified or non-acidified lakes. Three bilateral meristic characters were counted on each side of the fish: number of gill rakers on the lower first branchial arch, number of gill rakers on the upper first branchial arch, and number of pectoral-fin rays. An asymmetry index summarizing the numbers of asymmetric characters per fish was also calculated. Four traits related to fitness were measured: gonad dry mass, egg mass, gonadosomatic index, and fecundity. There were significant differences in FA among the five perch populations for the characters number of pectoral-fin rays and number of upper gill rakers, and also for the FA index. Asymmetry was generally greater in perch living in acidified lakes than in those in non-acidified lakes. However, there was no significant correlation between FA and any of the four fitness-related traits within populations. Therefore, asymmetry in the traits measured here may not be a good indicator of individual fitness in perch.
CODING WITH MINIMAL PROGRAMS
Authors: CALUDE CRISTIAN S. , ISHIHARA HAJIME , YAMAGUCHI TAKESHI
Published: July 2002

According to the Algorithmic Coding Theorem, minimal programs of any universal machine are prefix-codes asymptotically optimal (i.e. optimal up to at most an additive, unknown constant) with respect to the machine algorithmic probabilities. A stronger version of this result will be proven for a class of machines, not necessarily universal, and any semi-distribution. Furthermore, minimal programs with respect to universal machines will be shown to be almost optimal (i.e. optimal up to an additive constant less than or equal to 2) for any semi-computable semi-distribution. Finally, a complete characterization of all machines satisfying the Algorithmic Coding Theorem is given.