bluuebunny/crossref_metadata_2025_split huggingface dataset:
Epitope-Mapping Studies Define Two Major Neutralization Sites on the Vaccinia Virus Extracellular Enveloped Virus Glycoprotein B5R
Authors: Aldaz-Carroll Lydia , Whitbeck J. Charles , Ponce de Leon Manuel , Lou Huan , Hirao Lauren , Isaacs Stuart N. , Moss Bernard , Eisenberg Roselyn J. , Cohen Gary H. 
Published: April 2005

ABSTRACT
Vaccinia extracellular enveloped virus (EEV) is critical for cell-to-cell and long-range virus spread both in vitro and in vivo. The B5R gene encodes an EEV-specific type I membrane protein that is essential for efficient EEV formation. The majority of the B5R ectodomain consists of four domains with homology to short consensus repeat domains followed by a stalk. Previous studies have shown that polyclonal antibodies raised against the B5R ectodomain inhibit EEV infection. In this study, our goal was to elucidate the antigenic structure of B5R and relate this to its function. To do this, we produced multimilligram quantities of vaccinia virus B5R as a soluble protein [B5R(275t)] using a baculovirus expression system. We then selected and characterized a panel of 26 monoclonal antibodies (MAbs) that recognize B5R(275t). Five of these MAbs neutralized EEV and inhibited comet formation. Two other MAbs were able only to neutralize EEV, while five others were able only to inhibit comet formation. This suggests that the EEV neutralization and comet inhibition assays measure different viral functions and that at least two different antigenic sites on B5R are important for these activities. We further characterized the MAbs and the antigenic structure of B5R(275t) by peptide mapping and by reciprocal MAb blocking studies using biosensor analysis. The epitopes recognized by neutralizing MAbs were localized to SCR1-SCR2 and/or the stalk of B5R(275t). Furthermore, the peptide and blocking data support the concept that SCR1 and the stalk may be in juxtaposition and may be part of the same functional domain.
Discontinuous Subgenomic RNA Synthesis in Arteriviruses Is Guided by an RNA Hairpin Structure Located in the Genomic Leader Region
Authors: van den Born Erwin , Posthuma Clara C. , Gultyaev Alexander P. , Snijder Eric J. 
Published: April 2005

ABSTRACT

Nidoviruses produce an extensive 3′-coterminal nested set of subgenomic (sg) mRNAs, which are used to express structural proteins and sometimes accessory proteins. In arteriviruses and coronaviruses, these mRNAs contain a common 5′ leader sequence, derived from the genomic 5′ end. The joining of the leader sequence to different segments derived from the 3′-proximal part of the genome (mRNA bodies) presumably involves a unique mechanism of discontinuous minus-strand RNA synthesis in which base pairing between sense and antisense transcription-regulating sequences (TRSs) plays an essential role. The leader TRS is present in the loop of a hairpin structure that functions in sg mRNA synthesis. In this study, the minimal sequences in the 5′-proximal region of the
Equine arteritis virus
genome that are required for sg RNA synthesis were delimited through mutagenesis. A full-length cDNA clone was engineered in which this domain was duplicated, allowing us to make mutations and monitor their effects on sg RNA synthesis without seriously affecting genome replication and translation. The leader TRS present in the duplicated sequence was used and yielded novel sg mRNAs with significantly extended leaders. Our combined findings suggest that the leader TRS hairpin (LTH) and its immediate flanking sequences are essential for efficient sg RNA synthesis and form an independent functional entity that could be moved 300 nucleotides downstream of its original position in the genome. We hypothesize that a conformational switch in the LTH region regulates the role of the 5′-proximal region of the arterivirus genome in subgenomic RNA synthesis.
Sequence Variability, Gene Structure, and Expression of Full-Length Human Endogenous Retrovirus H
Authors: Jern Patric , Sperber Göran O. , Ahlsén Göran , Blomberg Jonas 
Published: April 2005

ABSTRACT

Recently, we identified and classified 926 human endogenous retrovirus H (HERV-H)-like proviruses in the human genome. In this paper, we used the information to, in silico, reconstruct a putative ancestral HERV-H. A calculated consensus sequence was nearly open in all genes. A few manual adjustments resulted in a putative 9-kb HERV-H provirus with open reading frames (ORFs) in
gag
,
pro
,
pol
, and
env
. Long terminal repeats (LTRs) differed by 1.1%, indicating proximity to an integration event. The
gag
ORF was extended upstream of the normal myristylation start site. There was a long leader (including a “pre-
gag
” ORF) region positioned like the N terminus of murine leukemia virus (MLV) “glyco-Gag,” potentially encoding a proline- and serine-rich domain remotely similar to MLV pp12. Another ORF, starting inside the 5′ LTR, had no obvious similarity to known protein domains. Unlike other hitherto described gammaretroviruses, the reconstructed Gag had two zinc finger motifs. Alternative splicing of sequences related to the HERV-H consensus was confirmed using dbEST data.
env
transcripts were most prevalent in colon tumors, but also in normal testis. We found no evidence for full length
env
transcripts in the dbEST. HERV-H had a markedly skewed nucleotide composition, disfavoring guanine and favoring cytidine. We conclude that the HERV-H consensus shared a gene arrangement common to gammaretroviruses with
gag
separated by stop codon from
pro-pol
in the same reading frame, while
env
resides in another reading frame. There was also alternative splicing. HERV-H consensus yielded new insights in gammaretroviral evolution and will be useful as a model in studies on expression and function.
Use of Confocal Immunofluorescence Microscopy To Localize Viral Nonstructural Proteins and Potential Sites of Replication in Pigs Experimentally Infected with Foot-and-Mouth Disease Virus
Authors: Monaghan P. , Simpson J. , Murphy C. , Durand S. , Quan M. , Alexandersen S. 
Published: April 2005

ABSTRACT
Replication of foot-and-mouth disease virus in infected pig epithelium has been studied by immunofluorescence labeling of the viral nonstructural protein 3ABC and confocal microscopy. The results were correlated with viral RNA copy numbers in tissue samples from adjacent sites determined by reverse transcription-PCR (RT-PCR). Lesion formation was seen in the tongues and coronary band epithelia of infected pigs 2 days after infection. Viral replication was observed in cells of the epithelium of the tongue and coronary band but not in the associated stromal cells. Infected epithelial cells were present in the stratum spinosum, away from the lesion, with small lesions formed above the basement membrane. Viral replication was markedly reduced in tongue epithelium by day 3 postinfection but remained apparent in the coronary band tissue up to 5 days postinfection. These results were confirmed by the RNA copy number determined by RT-PCR.
Interleukin-1 Is Responsible for Acute Lung Immunopathology but Increases Survival of Respiratory Influenza Virus Infection
Authors: Schmitz Nicole , Kurrer Michael , Bachmann Martin F. , Kopf Manfred 
Published: April 2005

ABSTRACTInterleukin-1α (IL-1α) and IL-1β are proinflammatory cytokines, which induce a plethora of genes and activities by binding to the type 1 IL-1 receptor (IL-1R1). We have investigated the role of IL-1 during pulmonary antiviral immune responses in IL-1R1−/−mice infected with influenza virus. IL-1R1−/−mice showed markedly reduced inflammatory pathology in the lung, primarily due to impaired neutrophil recruitment. Activation of CD4+T cells in secondary lymphoid organs and subsequent migration to the lung were impaired in the absence of IL-1R1. In contrast, activation of virus-specific cytotoxic T lymphocytes and killing of virus-infected cells in the lung were intact. Influenza virus-specific immunoglobulin G (IgG) and IgA antibody responses were intact, while the IgM response was markedly reduced in both serum and mucosal sites in IL-1R1−/−mice. We found significantly increased mortality in the absence of IL-1R1; however, lung viral titers were only moderately increased. Our results demonstrate that IL-1α/β mediate acute pulmonary inflammatory pathology while enhancing survival during influenza virus infection. IL-1α/β appear not to influence killing of virus-infected cells but to enhance IgM antibody responses and recruitment of CD4+T cells to the site of infection.
Acylation-Mediated Membrane Anchoring of Avian Influenza Virus Hemagglutinin Is Essential for Fusion Pore Formation and Virus Infectivity
Authors: Wagner Ralf , Herwig Astrid , Azzouz Nahid , Klenk Hans Dieter 
Published: April 2005

ABSTRACT
Attachment of palmitic acid to cysteine residues is a common modification of viral glycoproteins. The influenza virus hemagglutinin (HA) has three conserved cysteine residues at its C terminus serving as acylation sites. To analyze the structural and functional roles of acylation, we have generated by reverse genetics a series of mutants (Ac1, Ac2, and Ac3) of fowl plague virus (FPV) containing HA in which the acylation sites at positions 551, 559, and 562, respectively, have been abolished. When virus growth in CV1 and MDCK cells was analyzed, similar amounts of virus particles were observed with the mutants and the wild type. Protein patterns and lipid compositions, characterized by high cholesterol and glycolipid contents, were also indistinguishable. However, compared to wild-type virus, Ac2 and Ac3 virions were 10 and almost 1,000 times less infectious, respectively. Fluorescence transfer experiments revealed that loss of acyl chains impeded formation of fusion pores, whereas hemifusion was not affected. When the affinity to detergent-insoluble glycolipid (DIG) domains was analyzed by Triton X-100 treatment of infected cells and virions, solubilization of Ac2 and Ac3 HAs was markedly facilitated. These observations show that acylation of the cytoplasmic tail, while not necessary for targeting to DIG domains, promotes the firm anchoring and retention of FPV HA in these domains. They also indicate that tight DIG association of FPV HA is essential for formation of fusion pores and thus probably for infectivity.
Differences in the Postfusion Conformations of Full-Length and Truncated Class II Fusion Protein E of Tick-Borne Encephalitis Virus
Authors: Stiasny Karin , Kössl Christian , Heinz Franz X. 
Published: April 2005

ABSTRACT

The trimeric postfusion structure of the C-terminally truncated fusion protein E of the flavivirus tick-borne encephalitis virus, a class II viral fusion protein, was previously determined (S. Bressanelli, K. Stiasny, S. L. Allison, E. A. Stura, S. Duquerroy, J. Lescar, F. X. Heinz, and F. A. Rey, EMBO J.
23:
728-738, 2004). In this study we compared the properties of this truncated form with the full-length trimer and found that the so-called stem-anchor region not only confers additional stability to the full-length molecule but also structurally modifies the protein domain carrying the fusion peptide loop. These data provide experimental evidence to support the model of a fusion process that leads to the interaction of the stem-anchor region with the fusion peptide loop in the postfusion trimer.
Characterization of Human Immunodeficiency Virus Type 1 (HIV-1) Envelope Variation and Neutralizing Antibody Responses during Transmission of HIV-1 Subtype B
Authors: Frost Simon D. W. , Liu Yang , Pond Sergei L. Kosakovsky , Chappey Colombe , Wrin Terri , Petropoulos Christos J. , Little Susan J. , Richman Douglas D. 
Published: April 2005

ABSTRACT
We analyzed neutralization sensitivity and genetic variation of transmitted subtype B human immunodeficiency virus type 1 (HIV-1) in eight recently infected men who have sex with men and the virus from the six subjects who infected them. In contrast to reports of heterosexual transmission of subtype C HIV-1, in which the transmitted virus appears to be more neutralization sensitive, we demonstrate that in our study population, relatively few phenotypic changes in neutralization sensitivity or genotypic changes in envelope occurred during transmission of subtype B HIV-1. We suggest that limited genetic variation within the infecting host reduces the likelihood of selective transmission of neutralization-sensitive HIV.
Expression of secreted Wnt antagonists in gastrointestinal tissues: potential role in stem cell homeostasis
Authors: Byun T , Karimi M , Marsh J L , Milovanovic T , Lin F , Holcombe R F 
Published: April 2005

Background:Wnt signalling dysregulation has been implicated in cancer, including colon and gastric cancer. Initiation of Wnt signalling is modulated by soluble Wnt antagonists (sWAs), including soluble frizzled related proteins, dickkopf (Dkk) proteins, and Wnt inhibitory factor-1 (Wif1).Aims:To evaluate the role of sWAs in upper (gastric) and lower (colon) gastrointestinal tract tumorigenesis.Methods:Dkk1–3, Wif1, and FrzB expression was evaluated by in situ RNA hybridisation on normal and malignant human gastric and colon tissues. Expression was graded semiquantitatively.Results:Wif1, Dkk1, and Dkk2 were not expressed in normal gastric tissue. Dkk3 was expressed in some samples, with stronger expression in deep gastric glands. FrzB was expressed in several normal gastric samples, but not in matched tumour specimens. In contrast, Dkk1 and FrzB were not expressed in normal colon. Wif1 was expressed in most colon samples, with stronger expression at crypt bases. Dkk3 and Dkk2 expression was also concentrated at crypt bases. There were no differences between sWA expression in malignant colon and matched normal tissue.Conclusions:sWA expression differed between upper and lower gastrointestinal tract. The loss of FrzB in gastric cancer suggests that it acts as a tumour suppressor. The graded expression of Dkk3 in gastric tissue, and Dkk2, Dkk3, and Wif1 in colon tissue, with increased expression in the deep gastric glands/colonic crypt bases, where gastrointestinal stem cells reside, suggests that sWAs may be crucial Wnt signalling regulators in these tissues, and may contribute to stem cell pool maintenance. sWAs are important components of the gastrointestinal proliferative regulatory network.
p16INK4A, CDC6, and MCM5: predictive biomarkers in cervical preinvasive neoplasia and cervical cancer
Authors: Murphy N , Ring M , Heffron C C B B , King B , Killalea A G , Hughes C , Martin C M , McGuinness E , Sheils O , O’Leary J J 
Published: April 2005

Aim:To analyse and compare expression patterns of three potential biomarkers—p16INK4A, CDC6, and MCM5—and evaluate their use as predictive biomarkers in squamous and glandular cervical preinvasive neoplasia.Methods:Immunocytochemical analysis of p16INK4A, MCM5, and CDC6 expression was performed on 20 normal, 38 cervical intraepithelial neoplasia 1 (CIN1), 33 CIN2, 46 CIN3, 10 squamous cell carcinoma, 19 cervical glandular intraepithelial neoplasia (cGIN), and 10 adenocarcinoma samples. Staining intensity was assessed using a 0–3 scoring system. p16INK4A, MCM5, and CDC6 expression was also examined in ThinPrep slides exhibiting mild, moderate, and severe dyskaryosis. Human papillomavirus (HPV) was detected using a modified SYBR green assay. Fluorogenic polymerase chain reaction (PCR) and solution phase PCR were used for specific HPV typing.Results:All three markers showed a linear correlation between expression and grade of dysplasia. p16INK4Aand MCM5 protein expression was upregulated in all grades of squamous and glandular dysplasia. CDC6 protein was preferentially expressed in high grade lesions and in invasive squamous cell carcinoma.Conclusion:p16INK4Aexpression was closely associated with high risk HPV infection—all grades of squamous and glandular cervical lesions were immunohistochemically positive. MCM5 staining intensity was independent of high risk HPV infection, highlighting its potential as a biomarker in both HPV dependent and independent cervical dysplasia. CDC6 may be a biomarker of high grade and invasive lesions of the cervix, with limited use in low grade dysplasia. p16INK4Awas the most reliable marker of cervical dysplasia. Combinations of dysplastic biomarkers may be useful in difficult diagnostic cases.
Denaturing high performance liquid chromatography: high throughput mutation screening in familial hypertrophic cardiomyopathy and SNP genotyping in motor neurone disease
Authors: Yu B , Sawyer N A , Caramins M , Yuan Z G , Saunderson R B , Pamphlett R , Richmond D R , Jeremy R W , Trent R J 
Published: April 2005

Aims:To evaluate the usefulness of denaturing high performance liquid chromatography (DHPLC) as a high throughput tool in: (1) DNA mutation detection in familial hypertrophic cardiomyopathy (FHC), and (2) single nucleotide polymorphism (SNP) discovery and validation in sporadic motor neurone disease (MND).Methods:The coding sequence and intron–exon boundaries of the cardiac β myosin heavy chain gene (MYH7) were screened by DHPLC for mutation identification in 150 unrelated patients diagnosed with FHC. One hundred and forty patients with sporadic MND were genotyped for the A67T SNP in the poliovirus receptor gene. All DHPLC positive signals were confirmed by conventional methods.Results:Mutation screening of MYH7 covered 10 kb with a total of 5700 amplicons, and more than 6750 DHPLC injections were completed within 35 days. The causative mutation was identified in 14% of FHC cases, including seven novel missense mutations (L227V, E328G, K351E, V411I, M435T, E894G, and E927K). Genotyping of the A67T SNP was performed at two different temperatures both in MND cases and 280 controls. This coding SNP was found more frequently in MND cases (13.6%) than in controls (6.8%). Furthermore, 19 and two SNPs were identified in MYH7 and the poliovirus receptor gene, respectively, during DHPLC screening.Conclusions:DHPLC is a high throughput, sensitive, specific, and robust platform for the detection of DNA variants, such as disease causing mutations or SNPs. It enables rapid and accurate screening of large genomic regions.
Angiocentric immunoproliferative lesion and angiocentric lymphoma of lymph node in children. A report of two cases
Authors: Drut R , Drut R M 
Published: April 2005

Aim:To report two examples of an angiocentric immunoproliferative lesion (AIL) and angiocentric angiodestructive lymphoma (AL) presenting in lymph nodes in children. Most commonly involving extranodal sites, AIL/AL rarely presents in the spleen and lymph nodes.Methods/Results:Case 1 presented as a cervical lymphadenopathy in a 3 year old girl being treated for pre-B cell acute lymphoblastic leukaemia. Histological and immunohistochemistry studies revealed an Epstein-Barr virus positive (EBV+), large B cell (CD20 and CD30+) AIL with large areas of necrosis, the whole resembling lymphomatoid granulomatosis. Case 2 presented as a large supraclavicular lymphadenopathy in a 13 year old boy. Histology and immunohistochemistry revealed an EBV–, large T cell (CD45RO, CD56, and CD30+) AL, presenting the features of so called angiocentric T cell/natural killer cell lymphoma, nasal type.Conclusions:The term AIL/AL refers to a heterogeneous group of conditions not unique to a particular type of lymphoid cell. These lesions are easily recognised by the histopathologist because of their extremely unusual angiocentric pattern. Although rare, AIL/AL may present as nodal lesions in children ab initio.
Analysis of Bcl-2 protein expression in choroidal melanomas
Authors: Hussein M R 
Published: April 2005

Background:Bcl-2 protooncogene alterations are involved in tumorigenesis and may have prognostic ramifications.Aims:To investigate normal ocular structures and choroidal melanoma for: (1) Bcl-2 protein expression (semiquantitative staining values: SI, staining intensity; PP, percentage of positive cells; and IRS, immunoreactivity score) and (2) any associations between the staining values and clinicopathological features in these lesions.Materials/Methods:Bcl-2 protein expression was analysed in 24 choroidal melanomas using immunoperoxidase staining methods.Results:Bcl-2 protein expression was seen in corneal epithelium, lens epithelium, the ciliary body, and retinal cells. In these structures, the mean (SEM) values were: 1.1 (0.1), 1.6 (0.3), 1.1 (0.1), and 2.3 (0.3), respectively, for SI; 1.6 (0.2), 1.7 (0.1), 1.7 (0.2), and 1.7 (0.2) for PP, respectively; and 1.9 (0.4), 2.7 (0.5), 1.9 (0.1), and 4.0 (0.8), respectively, for IRS. Based on Bcl-2 immunoreactivity, the lesions were divided into two groups. The first group comprised 12 tumours with Bcl-2 expression. Bcl-2 expression was significantly higher in this group compared with normal ocular structures (1.5 (0.1)v2.8 (0.2), 1.7 (0.1)v3.5 (0.1), and 2.6 (0.3)v9.3 (0.9) for mean (SEM) SI, PP, and IRS scores, respectively; p  =  0.00). The second group comprised 12 tumours lacking Bcl-2 protein expression. There was no significant correlation between Bcl-2 protein expression and most of the clinicopathological features of these lesions.Conclusions:Bcl-2 protein expression is altered in choroidal melanomas.
Overexpression/amplification of HER-2/neu is uncommon in hepatocellular carcinoma
Authors: Xian Z-H , Zhang S-H , Cong W-M , Wu W-Q , Wu M-C 
Published: April 2005

Background:Hepatocellular carcinoma (HCC) is one of the most prevalent fatal cancers in the world. Despite advances in early diagnosis and improvements in surgical techniques, the survival of patients with HCC even after resection is poor because of the high incidence of recurrences. Therefore, the identification of prognostic factors may be helpful in the development of new treatment protocols.Aims:To investigate HER-2/neu status in HCC by immunohistochemistry (IHC) and fluorescence in situ hybridisation (FISH), and to explore the possibility of using trastuzumab in the treatment of HCC.Methods:Eight hundred and sixty eight surgical samples from patients with primary HCC were examined for their HER-2/neu status. IHC for HER-2/neu was performed with the HercepTest kit; FISH analysis was performed with the PathVysion HER-2 DNA probe kit. The correlations between HER-2/neu overexpression and clinicopathological characteristics were analysed statistically.Results:HER-2/neu overexpression was detected in 21 (2.42%) of the 868 primary HCCs. Only one specimen showed HER-2/neu gene amplification by FISH. No significant associations were found between HER-2/neu overexpression and the clinicopathological parameters.Conclusions:There is a low frequency of HER-2/neu overexpression/amplification in HCC. There appears to be no role for HER-2/neu as a prognostic marker and no benefit of anti-HER-2/neu trastuzumab treatment in patients with HCC.
Voltage-Dependent and Frequency-Independent Inhibition of Recombinant Ca<sub>v</sub>3.2 T-Type Ca<sup>2+</sup> Channel by Bepridil
Authors: Uchino Tomoko , Lee Tae-Seong , Kaku Toshihiko , Yamashita Noboru , Noguchi Takayuki , Ono Katsushige 
Published: April 2005

Effects of bepridil on the low voltage-activated T-type Ca<sup>2+</sup> channel (Ca<sub>V</sub>3.2) current stably expressed in human embryonic kidney (HEK)-293 cells were examined using patch-clamp techniques. Bepridil potently inhibited I<sub>Ca,T</sub> with a markedly voltage-dependent manner; the IC<sub>50</sub> of bepridil was 0.4 µmol/l at the holding potential of –70 mV, which was 26 times as potent as that at –100 mV (10.6 µmol/l). Steady-state inactivation curve (8.4 ± 1.7 mV) and conductance curve (5.9 ± 1.9 mV) were shifted to the hyperpolarized potential by 10 µmol/l bepridil. Bepridil exerted the tonic blocking action but not the use-dependent block. Bepridil had no effect on the recovery from inactivation of T-type Ca<sup>2+</sup> channels. Thus, high efficacy of bepridil for terminating atrial fibrillation and atrial flutter may be considered to be attributed, at least in a part, to the T-type Ca<sup>2+</sup> channel-blocking actions.