bluuebunny/crossref_metadata_2025 huggingface dataset:
Interleukin-4 Gene Promoter Polymorphism [C590T] and Asthma in Kuwaiti Arabs
Authors: Hijazi Z. , Haider M.Z.
Published: April 2003

Background: Recent studies have demonstrated an association of the T allele of a promoter polymorphism [C590T] of interleukin-4 (IL-4) gene with asthma/atopy in the US and Japanese populations, but data from the UK and Australian families failed to find an association. We have investigated the association of this [C590T] promoter polymorphism in Kuwaiti Arabs with asthma. Methods: This study included 184 subjects (84 asthmatics and 100 non-asthmatics, 47 of whom were totally unrelated normal healthy controls with no history of asthma/atopy in three generations of near relatives). The [C590T] promoter polymorphism of the IL-4 gene was analyzed by a polymerase chain reaction-restriction fragment length polymorphism method using the restriction enzyme AvaII. Results: The frequency of the T allele was found to be 0.79 in asthmatics, 0.75 in non-asthmatic first-degree relatives and 0.76 in unrealted controls in Kuwaiti Arabs. No significant difference was detected in the distribution of the three genotypes (TT, CT and CC) between the asthmatic, non-asthmatic relatives and the unrelated controls. The incidence of co-existing phenotypes e.g. hay fever, eczema and a positive skin prick test was significantly higher in asthmatics than in the non-asthmatics irrespective of the [C590T] genotype. However, within the asthmatic group, a higher incidence of eczema was detected in asthmatics with a heterozygous CT genotype compared to the asthmatics with a TT genotype. Conclusions: Our results do not show an association between [C590T] promoter polymorphism of the IL-4 gene and asthma in Kuwaiti Arabs. Our results are in keeping with the reports from the UK and Australian families (no association) but in contrast to the findings from US and Japanese populations (an association reported).
IFN-γ Plays a Dominant Role in Upregulation of Candida-Specific IgE Synthesis in Patients with Atopic Dermatitis
Authors: Kimura Mitsuaki , Tsuruta Satoru , Yoshida Takami
Published: April 2003

Background: Although Candida albicans (CA) is known to induce Th1 clones that suppress IgE synthesis, serum IgE antibody against CA is often increased in atopic patients. This study aims to elucidate the mechanism of IgE synthesis against CA in atopic patients. Methods: We measured the production of IL-4 and IFN-γ by peripheral blood mononuclear cells (PBMCs) from atopic patients upon stimulation with CA and examined the correlation with the level of serum IgE antibody against CA. Results: The level of serum CA-specific IgE antibody (CA-IgE) was significantly higher in patients with atopic dermatitis (AD) than in patients with bronchial asthma (BA) (geometric mean = 3.6 vs. 0.27 UA/ml, p < 0.02) (UA = unit allergen), while there was no difference in the level of house dust mite-specific IgE antibody between them (67.6 vs. 87.1 UA/ml). Although IL-4 production by PBMCs upon stimulation with CA in patients with AD was not significantly different from that in patients with BA (mean = 359.1 vs. 515.3 fg/ml), IFN-γ production was significantly lower in the former than in the latter group (8.1 vs. 56.2 pg/ml, p < 0.001). Consequently, the ratio of IL-4/IFN-γ production was apparently higher in patients with AD than in those with BA, which corresponds to the difference between them in the level of serum CA-IgE. A significant negative correlation was seen in patients with AD between IFN-γ production by CA-stimulated PBMCs and the level of serum CA-IgE (p < 0.05). Conclusions: IgE synthesis against CA in atopic patients may be precipitated not by enhancing IL-4 production, but by reducing IFN-γ secretion.
Occurrence of IgE Antibody-Recognizing N-Linked Glycan Moiety of a Soybean Allergen, Gly m Bd 28K
Authors: Hiemori Miki , Bando Noriko , Ogawa Tadashi , Shimada Hisao , Tsuji Hideaki , Yamanishi Rintaro , Terao Junji
Published: April 2003

Background: It has been reported that N-linked glycan moieties of glycoproteins function as IgE-reactive determinants. Gly m Bd 28K, a soybean allergen, was a glycoprotein with glycan moieties, which are supposed to be the Man3GlcNAc2 backbone with the β1→2 xylose and α1→3 fucose branches. The purpose of the present study was to examine the IgE-binding ability of the glycan moiety of Gly m Bd 28K in the binding reaction with patients’ sera. Methods: A peptide containing the glycan moiety was prepared from Gly m Bd 28K by digestion with lysyl endopeptidase. The binding site of the glycan moiety was determined by amino acid sequence analyses. The glycan moiety of the allergen was characterized using anti-horseradish peroxidase antibody (anti-HRP) recognizing the N-linked glycan moieties of glycoproteins. The binding of patients’ IgE antibodies with their glycan moiety was examined by an immunostaining technique using the glycopeptide and its deglycosylated peptide derived from Gly m Bd 28K. Results: The binding site of the glycan moiety in Gly m Bd 28K was shown to be its Asn20 residue. Gly m Bd 28K did react with anti-HRP and the sera of soybean-sensitive patients, but the binding of IgE antibodies was inhibited by the preincubation with anti-HRP. Moreover, the glycopeptide also reacted with the sera of soybean-sensitive patients, but its deglycosylated peptide did not react with any IgE antibodies of patients’ sera. Conclusions: The specific IgE antibodies recognizing the N-linked glycan moieties of Gly m Bd 28K and other glycoproteins with homologous glycan moieties occur in the sera of soybean-sensitive patients. It was indicated that the N-linked glycan moieties such as that of Gly m Bd 28K may be one of the common IgE-reactive determinants distributed in various plant food proteins.
Dermatophagoides farinae (Der f 1) and Dermatophagoides pteronyssinus (Der p 1) Allergen Exposure among Subjects Living in Uberlândia, Brazil
Authors: Sopelete Mônica C. , Silva Deise A.O. , Arruda L. Karla , Chapman Martin D. , Taketomi Ernesto A.
Published: April 2003

Background: The role of mite allergen exposure in sensitization and development of asthma has been widely recognized. Previous studies have shown that Dermatophagoides pteronyssinus and Blomia tropicalis were the most prevalent house dust mites in Brazil, while D. farinae was rarely found. The aim of this study was to measure Der f 1 and Der p 1 allergen levels in Brazilian asthmatics’ and controls’ homes. Methods: Sixty-four houses (32 asthmatic, 32 control) were visited for dust sampling from five sites. Der f 1 and Der p 1 levels were measured by two-site monoclonal-antibody-based ELISAs. Results: The highest levels of Der f 1 and Der p 1 allergens were found in bedding samples from both asthmatics’ and controls’ homes. However, the geometric mean of Der f 1 levels (15.8 μg/g of dust) was significantly higher than for Der p 1 (8.2 μg/g of dust) in these samples. In addition, allergen levels ≥10 μg/g of dust were found in 60–80% of the samples for Der f 1 and about 50% for Der p 1. Conclusions: These results indicate that high levels of Der f 1 allergen are present in both asthmatics’ and controls’ homes, in contrast to previously reported data. Therefore, studies on exposure to mites should be performed in different cities, seasons and times, since the mite fauna might be subject to variations. Knowledge of the mite fauna will certainly improve the means of investigating the association between allergen exposure and sensitization, allowing to establish the inclusion of new mite extracts in inhalant skin test sets, and even to detect monosensitized patients with respiratory allergy.
Synthesis of Factor D by Normal Human Hepatocytes
Authors: Kitano Etsuko , Kitamura Hajime
Published: April 2003

Background: Unlike most complement proteins, complement factor D is believed to be synthesized not by the liver but exclusively by adipose tissue. Method: Culture supernatants obtained from primary culture of normal human hepatocytes were assayed for factor D by ELISA and analyzed by Western blotting. Results: When normal hepatocytes were cultured in protein-free medium without addition of any stimulator for 5 days, factor D was detected in the supernatants at levels as high as 331.07 ± 41.38 μg/106 cells. Addition of TNF-α, IFN-γ, IL-1β or LPS to the medium did not result in any distinct effect on the amounts of secreted factor D. Reversible inhibition of factor D secretion by these cells was observed when cultured in the presence of cycloheximide. By immunoblot analysis, secreted factor D exhibited double bands, one with a molecular weight similar to factor D in normal human serum and the other with a slightly larger molecular weight. Conclusion: Normal human hepatocytes synthesize factor D constitutively. The liver may be a major source of plasma factor D.
The Idiotypic Network in Antinuclear-Antibody-Associated Diseases
Authors: Sherer Yaniv , Shoenfeld Yehuda
Published: April 2003

Antinuclear antibodies (ANA) entail a large group of autoantibodies (Abs) that bind certain nuclear antigens. The ANA test is a useful screening test for many autoimmune diseases and the presence of a specific binding pattern directs secondary testing for specific Abs associated with the suspected disease. Idiotypes (Ids) are the antigenic constitution of the variable region of an Ab, and they are recognized by anti-Ids Abs. The Id network is composed of interacting Abs in which the Id determinants of each Ab are complemented by those of another. It has a role in both physiologic and pathologic conditions. In this communication, we review the induction of autoimmune diseases via Id manipulation, the Ids of some ANA-associated Abs (DNA, SS-A, SS- B, Sm Abs), the pathogenic role of Abs carrying Ids, and the clinical implications of the Id network in autoimmunity.
Antinuclear Autoantibodies: Fluorescent Highlights on Structure and Function in the Nucleus
Authors: Hemmerich Peter , von Mikecz Anna
Published: April 2003

The eukaryotic nucleus is dynamically organized with respect to particular activities, such as RNA transcription, RNA processing or DNA replication. The spatial separation of metabolic activities is best reflected by the identification of functionally related proteins, in particular substructures of the nucleus. In a variety of human diseases, the integrity of such structures can be compromised, thus underlining the importance of a proper nuclear architecture for cell viability. Besides their clinical relevance, antinuclear autoantibodies (ANAs) have contributed to a large extent to the identification of subnuclear compartments, the isolation and cloning of their components (the autoantigens), as well a the characterization of their function. Although sophisticated techniques, such as confocal laser scanning microscopy (CLSM), fluorescence resonance energy transfer (FRET) and in vivo observation of cellular events have recently been established as valuable tools to study subnuclear architecture and function, cell biologists will continue to appreciate the specificity and power of ANAs for their research.
Sjögren’s Syndrome: Autoantibodies to Cellular Antigens
Authors: Mavragani Clio P. , Tzioufas Athanasios G. , Moutsopoulos Haralampos M.
Published: April 2003

Autoantibodies to cellular autoantigens are usually found in sera of patients with systemic autoimmune rheumatic diseases. Patients with Sjögren’s syndrome (SS) frequently present autoantibodies to both organ and non-organ-specific autoantigens. The most commonly detected autoantibodies are those directed against the ribonucleoproteins Ro/SSA and La/SSB. The presence of the antibodies in SS is associated with early disease onset, longer disease duration, parotid gland enlargement, higher frequency of extraglandular manifestations and more intense lymphocytic infiltration of the minor salivary glands. Over the past several years, the structure and function of these autoantigens have been extensively studied. Several centers, using different techniques, have investigated the B cell epitopes on the protein components Ro 60 kD, Ro 52kD, and La 48 kD. Finally, increased evidence of direct involvement of anti-Ro/SSA and anti-La/SSB autoantibodies in the pathogenesis of tissue injury has been contributed by several studies.
Diagnostic Importance of Anti-Proteasome Antibodies
Authors: Feist Eugen , Dörner Thomas , Kuckelkorn Ulrike , Scheffler Sonja , Burmester Gerd-R. , Kloetzel Peter-M.
Published: April 2003

20S proteasome represents the proteolytic core complex for cytoplasmic protein degradation that is involved in the activation and regulation of the immune response. In this context, proteasome generates antigenic peptides for the MHC class I pathway and activates NF-κB. In a recent analysis, we could identify a frequent humoral autoimmune response directed against specific proteasomal subunits in patients with autoimmune myositis, systemic lupus erythematosus and primary Sjögren’s syndrome. The outer ring subunit HC9(α3) was identified as the predominant target of the anti-proteasome response in these entities. In addition to the reactivity against HC9(α3), patients with primary Sjögren’s syndrome expressed a more polyspecific recognition pattern of proteasomal subunits involving the active inner ring proteins. In follow-up analysis, anti-proteasome antibody titers revealed a correlation with disease activity in patients with autoimmune myositis and systemic lupus erythematosus. The current review summarizes recent data providing evidence that the 20S proteasome represents an important target of the humoral autoimmune response in systemic autoimmune diseases and extends insight into pathogenic aspects of these diseases.
Electrophoretic and Immunochemical Characterization of Allergenic Proteins in Buckwheat
Authors: Yoshimasu Mark Akira , Zhang Jie Wei , Hayakawa Shigeru , Mine Yoshinori
Published: April 2003

Background: Buckwheat allergies are not common, however, it is considered to be a very potent allergen. Ingestion of small amounts has been found to produce anaphylactic reactions, particularly in children. Identification and characterization of the major allergen(s) in buckwheat are currently underway, however, there are some discrepancies in the findings. Methods: Identification of the major allergen(s) was determined through Western blotting using buckwheat-allergic patients’ sera. Once the allergenic proteins were identified, they were purified, their IgE-binding activity assessed through an indirect ELISA and the N-terminal amino acid sequence completed. To assess the stability of the IgE-binding epitopes, protein fractions were exposed to various treatments and assayed using an indirect ELISA. Lastly, the presence of anti-buckwheat IgG in the patients’ sera was analyzed through Western blotting and ELISA. Results: IgE binding was detected to proteins with molecular masses of approximately 14 and 18 kDa. N-terminal sequencing was completed and found to share some homology with rice proteins associated with rice allergies and cross-allergenicity with buckwheat proteins. When the water-soluble protein fraction was heated, exposed to acidic and alkaline conditions and fully denatured, IgE-binding activity was reduced. When the fraction was partially denatured through urea, IgE-binding activity increased. Furthermore, IgG-binding activity was detected with proteins only above the 20 kDa region. Conclusions: Proteins with molecular masses around 14 and 18 kDa were identified as the major allergenic proteins in the buckwheat-allergic patients’ sera tested in this study. Results also indicate that these two proteins possess IgE-binding capability.
IgA Deficiency in Czech Healthy Individuals and Selected Patient Groups
Authors: Litzman J. , Ševčíková I. , Štikarovská D. , Pikulová Z. , Pazdírková A. , Lokaj J.
Published: April 2003

Background: Selective IgA deficiency (IgAD) is the most common immunoglobulin deficiency with a variety of clinical manifestations. The frequency of IgAD differs depending on the ethnic origin and clinical symptoms of investigated persons. Methods: The prevalence of IgAD (serum IgA level <0.05 g/l) was determined in 5,310 Czech blood donors, 10,326 patients who had undergone immunological investigation, and 246 first-degree relatives of IgAD and common variable immunodeficiency (CVID) patients. Results: IgAD was detected in 13 (1/408; 0.24%) of the blood donors. The prevalence of IgAD was increased both in children (48/3,113; 1.5%) and adults (33/3,824; 0.9%) referred for frequent respiratory tract infections (in both cases p < 0.001) compared to the healthy population. The frequency of IgAD was 12/189 (6%) in first-degree relatives of IgAD patients and 9/57 (16%) in relatives of CVID patients, with the highest frequency observed in children of CVID patients. Conclusions: The prevalence of IgAD in the Czech healthy population is comparable to that in other Caucasians. The frequency is increased in children with recurrent respiratory tract infections and especially in relatives of patients with immunoglobulin deficiencies.
Elaboration of Matrix Metalloproteinase Inhibitors by Human Skin in Organ Culture and by Skin Cells in Monolayer Culture: Relationship to Invasion
Authors: Chi Yiqing , Zeigler Mary E. , Walker James , Perone Patricia , Datta Subhash C. , Varani James
Published: April 2003

Functional and immunochemical approaches were used to assess matrix metalloproteinase (MMP) inhibitors, e.g., tissue inhibitor of metalloproteinases 1 and 2 (TIMP-1 and TIMP-2), in organ cultures of normal human skin maintained under growth factor free conditions or in medium supplemented with a combination of growth factors including epidermal growth factor, insulin, and pituitary extract. It has previously been shown that under growth factor free conditions, normal skin structure and function are maintained for several days, while in the presence of these exogenous growth factors, the epithelial cells invade the stroma [Invasion and Metastasis 1993;13:225–233]. TIMP-1 was detected in equivalent amounts in organ culture fluids under both conditions. TIMP-2 was not detected under either condition. Normal epidermal keratinocytes, normal dermal fibroblasts, and three different epithelial tumor cell lines were also examined for MMP inhibitor expression. Keratinocytes and fibroblasts produced high levels of both TIMP-1 and TIMP-2, but in neither cell type was there a significant difference between growth factor free and growth factor containing conditions. In contrast, the three epithelial tumor cell lines produced low to undetectable levels of both TIMP-1 and TIMP-2. These data suggest that acquisition of local invasive capacity is not dependent on a reduction in MMP inhibitor expression. A reduction in MMP inhibitors may accompany the transition from invasive to metastatic tumors.
Effects of All-trans-Retinoic Acid Incorporated into Low-Density Lipoprotein on Invasive Properties of Multidrug-Resistant MCF-7 Spheroids
Authors: Affoué M. , Akeli M.G. , Gueddari N. , Jardillier J.C. , Madoulet C.
Published: April 2003

Cultured cells grown as spheroids provide an in vitro model that is closer to an in vivo tumour than conventional monolayer techniques. Previous work from our laboratory has demonstrated that spheroids formed from multidrug-resistant MCF-7 cells exhibit invasive characteristics which were not present in their sensitive counterparts. The treatment of these spheroids by all-trans-retinoic acid (ATRA), a potent inducer of in vitro and in vivo differentiation, decreases their proteolytic activity and ability to invade Matrigel-coated filters. The efficiency of ATRA is enhanced by its incorporation into low-density lipoprotein (LDL) (LDL-ATRA). Indeed, invasion through a reconstituted basement membrane was reduced by 73% with 10–6 M ATRA and 3 × 10–8 M LDL-ATRA. Furthermore, inhibition of invasion was correlated with a decrease in several factors: (1) secreted matrix metalloproteinase-9 and enzymes degrading type IV collagen and Matrigel films, and (2) tissue plasminogen activator. The results observed were found with a concentration of LDL-ATRA 30 times lower than that of ATRA. This could be due to the protective effect of LDL and to a better targeting of cancer cells through their LDL receptors. LDL-ATRA may therefore represent a new and potent inhibitor of invasion that could be developed for clinical trials.
HHV-6 Infection in HIV-Infected Asymptomatic and AIDS Patients
Authors: Ablashi Dharam V. , Marsh Scott , Kaplan Mark , Whitman Jr. James E. , Pearson Gary R.
Published: April 2003

In order to investigate the levels of HHV-6 infection and elevated antibodies to HHV-6 in HIV-1-infected asymptomatic and symptomatic patients, peripheral blood mononuclear cells were (PBMC) cultured. As patients progressed from asymptomatic HIV infection to AIDS, there was a concurrent increase in replicating HHV-6. Plasma obtained from several of these patients showed the presence of IgM antibody and a significantly elevated level of HHV-6 IgG antibody. Serial samples of plasma from 10 AIDS patients collected over a period of 4 years were assayed for the detection of HHV-6 core protein (gp116/64/54) by antigen capture ELISA. The results demonstrated that either a persistent infection or reactivation can occur based on the degree of fluctuation in HHV-6 antigen detected. ELISA to HHV-6 purified viral proteins, i.e., early (p41/38) and late (gp110), demonstrated that IgG antibody to gp110 did not differentiate between HIV-1-infected and healthy donors. IgG and IgM antibody to p41/38, however, showed a significantly higher prevalence in HIV-1-infected individuals (56.7–85.3%) than in normal healthy donors (19.0%), suggesting virus activation. PBMC culture from the AIDS patients expressing significant peaks of HHV-6 core antigen (gp116/64/54) in their plasma showed that in most cases, HHV-6 early and late antigens were detectable; however, those patients with consistently low antigen peaks had no detectable antigens in their PBMC. Only 55% of PBMC cultures established from IgM antibody-positive HIV-1-infected asymptomatic and AIDS patients expressed HHV-6 antigens in the short-term cultures, but HHV-6 antigens could not be demonstrated in PBMC culture from 4 IgM-antibody-positive healthy donors. HHV-6 isolates obtained from the HIV-1-positive patients were predominantly HHV-6 variant A, compared to healthy donors. Based on the data presented here, it is evident that the levels of HHV-6 infection increased in HIV-1-infected asymptomatic individuals as they progressed to AIDS. Our immunovirological data on HHV-6-infected individuals with HIV infection support a role for HHV-6 in the pathogenesis of AIDS. We believe that simultaneous active infection with HIV-1 and HHV-6 may contribute to enhanced immune suppression perhaps leading to disease manifestations.
Evaluation of the New Automated Enzymun-Test® Anti-HBc Plus for the Detection of Hepatitis B Virus Core Antibody
Authors: Weber Bernard , Michl Ulrike , Mühlbacher Annelies , Paggi Giancarlo , Bossi Vincenzo
Published: April 2003

The new automated Enzymun-Test® Anti-HBc Plus for the detection of antibodies to hepatitis B virus (HBV) core antigen (anti-HBc) after pretreatment with reducing agents dithiothreitol (25°C; + DTT) or potassium bisulfite (37°C; + MBS) was evaluated by testing 571 serum and plasma samples. The panel included dilution series of reference standards and samples from chronic carriers, one seroconversion panel, samples from patients in acute or chronic stage of the disease or resolved hepatitis B, preselected sera from blood donors that were initially reactive in an anti-HBc assay without pretreatment, potentially cross-reactive serum samples obtained from patients suffering from other diseases or with passed infections other than HBV, pregnant women and individuals with HBV vaccination. The IMx CORETM (Abbott Diagnostics) served as reference assay. Discrepant samples were further investigated with two different commercial anti-HBc enzyme immunoassays, other HBV-specific serological markers and HBV DNA hybridization. The sensitivity of the Enzymun-Test Anti-HBc Plus (25°C; + DTT, and 37°C; + MBS) in comparison to the reference assay was 100%. The IMx CORE showed a twofold higher sensitivity than Enzymun-Test Anti-HBc Plus for anti-HBc detection in dilution series of serum samples from HBV carriers. The agreement in terms of specificity of the Enzymun-Test Anti-HBc Plus (25°C; + DTT, and 37°C; + MBS) and in comparison to IMx CORE was 97.4 and 96.4%, respectively. After resolution of discrepant results (3 samples were tested false negative with IMx CORE), the agreement in terms of specificity of the Enzymun-Test Anti-HBc Plus (25°C; + DTT, and 37°C; + MBS) in comparison to the combination of the comparative assays was 98.3 and 97.4%, respectively. In conclusion, the new automated Enzymun-Test Anti-HBc Plus with sample pretreatment with DTT (25°C; + DTT) or MBS (37°C; + MBS) permits a highly sensitive and specific detection of anti-HBc in diagnostic virology and blood donation testing.
Neutralization of Borna Disease Virus Depends upon Terminal Carbohydrate Residues (α-D-Man, β-D-GlcNAc) of Glycoproteins gp17 and gp94
Authors: Stoyloff Roman , Bode Liv , Borchers Kerstin , Ludwig Hanns
Published: April 2003

Borna disease virus (BDV) is an enveloped, nonsegmented, negative-stranded RNA virus that causes infections of the brain in a wide range of animal species and man. The third open reading frame codes for a protein of 17 kD (gp17) that is N-glycosylated and contains terminal α-D-mannose and N-acetyl-β-D-glucosamine residues. Rat sera raised against these carbohydrates (anti-sugar antisera) show high in vitro neutralization activity and were capable of precipitating BDV. The neutralizing capacity of sera derived from experimentally BDV-infected rabbits, in turn, decreased after adsorption with those carbohydrates. They partially inhibited infection of primary young rabbit brain cells in a dose-dependent manner. Furthermore, the anti-sugar antisera recognized a second virus-specific glycoprotein with an apparent molecular mass of 94 kD (gp94), providing indirect evidence that gp94 is involved in virus adsorption and/or entry into cells. Neutralization of BDV comprises a complex event and, as shown for the first time, involves the carbohydrate residues of both glycoproteins of BDV.
Should We Routinely Screen for Cytomegalovirus Antibody during Pregnancy?
Authors: Grangeot-Keros L. , Simon B. , Audibert F. , Vial M.
Published: April 2003

In order to evaluate the usefulness of cytomegalovirus (CMV) antibody screening during pregnancy, women attending for antenatal care at the Antoine Béclère Hospital (Clamart, France) were prospectively studied during 22 months (1995–1996). Forty-five percent of these women were CMV-seropositive. Twenty suspected or confirmed CMV primary infections were detected. Nine infected infants were born to these women. All infected infants are now between 10 months and 2.5 years old. They all are asymptomatic even those who initially presented abnormal biological parameters or slightly abnormal ultrasound scans during fetal life. Presently, screening for CMV antibody cannot be recommended because it induces economic, psychological and ethical problems. Furthermore, there is no efficient and safe treatment available so far. However, we do think that large studies must be performed to increase our knowledge about the natural history of intrauterine CMV infection. This is also important for an improved assessment of the value of ultrasound examination results as well as the biological parameters measured in fetal blood samples.
Phylogenetic Analysis of Hepatitis GB Virus Type C/Hepatitis G Virus in Spanish Patients with Chronic Hepatitis B or C Virus Infection
Authors: López-Alcorocho Juan Manuel , Amaro Maria José , Cabrerizo Maria , Fogeda Marta , Castillo Inmaculada , Bartolomé Javier , Carreño Vicente
Published: April 2003

The nucleotide sequence of hepatitis GB virus type C (HGBV-C)/hepatitis G virus (HGV) NS3/helicase and 5′-untranslated regions from 23 Spanish patients were analyzed to assign the HGV isolates one of the proposed HGBV-C/HGV genotypes. The analysis of the evolutionary distance frequency showed that the distances among all sequences in NS3/helicase region were distributed around a single peak of 0.20, suggesting that all included sequences belonged to the same HGBV-C/HGV genotype. By contrast, in the 5′-untranslated region, all the distances corresponding to our sequences and those of the HGBV-C/HGV types 2 and 3 were distributed around a major peak of 0.03. The remaining distances corresponding to the HGBV-C/HGV type 1 sequences were distributed around a minor peak of 0.11. The phylogenetic tree and pairwise comparison of evolutionary distances among the 5′-untranslated region of the infected patients and each HGBV-C/HGV genotype demonstrated that our HGBV-C/HGV isolates belonged to subtype 2a (17/23; 78%) and 2b (5/23; 22%). No relation was found between HGBV-C/HGV subtype and hepatitis B or C virus infection.
A Novel Protein Tag from Herpes Simplex Virus Type 1 DNA Polymerase
Authors: Schreiner Uwe , Hansen Jutta , Kühn Frank J.P. , Knopf Charles W.
Published: April 2003

An epitope (HPOL) derived from the so-called thumb region of the herpes simplex virus type 1 DNA polymerase in combination with a monoclonal antibody (MAb 1051c) was tested for protein tagging. Using a conventional expression vector, a DNA cassette encoding the HPOL epitope was fused to the C-terminus of the dihydrofolate reductase (DHFR) gene such that the recombinant DHFR contained both a N-terminal HIS-tag and a C-terminal HPOL tag. Expression of recombinant DHFR in Escherichia coli cells was compared by Western blot analysis using either mouse RGS·HIS antibody or MAb 1051c. Immunostaining revealed that both antibodies reacted specifically with DHFR, but the detection sensitivity achieved with MAb 1051c was about 15-fold greater using a standard staining protocol. An HPOL antibody column was successfully applied for affinity purification of DHFR, demonstrating the usefulness of the HPOL epitope/MAb 1051c system for protein tagging, expression monitoring and purification of HPOL-tagged recombinant proteins.
T Cell-Mediated and Non-Specific Inflammatory Mechanisms Contribute to the Skin Pathology of HPV 16 E6E7 Transgenic Mice
Authors: Hilditch-Maguire Paige A. , Lieppe Donna M. , West Donna , Lambert Paul F. , Frazer Ian H.
Published: April 2003

One of three lines of mice transgenic for the E6 and E7 genes of human papillomavirus type 16 (HPV16) expressed from an αA-crystallin promoter also expresses the transgene ectopically in the skin. This line, designated αACE6E7#19, develops skin disease from 3 months of age, characterised by epidermal hyperplasia and eventual skin loss. Administration of complete Freund’s adjuvant (CFA) to αACE6E7#19 mice, but not to non-transgenic littermate controls, induced local epidermal hyperplasia which was histologically similar to the spontaneously arising skin pathology. Local application of 2,4-dinitrochlorobenzene (DNCB) to DNCB-sensitised αACE6E7#19 mice, but not DNCB-sensitised controls, also induced hyperplasia. Treatment with cyclosporin A (CsA) or systemic depletion of CD4+ cells significantly reduced the incidence of skin disease. These data suggest that local inflammation, and cytokines produced by T helper cells, contribute to the induction of hyperplastic skin disease in αACE6E7#19 mice. Spontaneous skin disease with similar histological appearance, frequency, age of onset and severity in αACE6E7#19 mice was observed in scid–/– αACE6E7#19 mice, despite immune paresis. Antigen-specific immune responses and T-cell cytokines are therefore not necessary for the induction of skin disease. We propose that epidermal hyperplasia associated with HPV16 E6 and E7 expression in skin is accelerated by local secretion of pro-inflammatory cytokines, whose production can be enhanced by activated CD4+ T cells.
A Novel Unenveloped DNA Virus (TT Virus) Associated with Acute and Chronic Non-A to G Hepatitis
Authors: Okamoto Hiroaki , Nishizawa Tsutomu , Ukita Masato
Published: April 2003

In 1997, a novel DNA virus was isolated from the serum of a patient with posttransfusion hepatitis of unknown etiology in Japan, and it was named TT virus (TTV) after the initials of the index patient. TTV is a nonenveloped, single-stranded and circular DNA virus, and its entire sequence of ∼3.9 kb has been determined. For being a DNA virus, TTV has a wide range of sequence divergence, allowing the classification into at least 16 genotypes separated by a sequence difference of >30% from one another. The nucleotide sequence of the noncoding region of the TTV genome is conserved, whereas that of the coding region is highly variable. TTV strains with extremely high sequence divergence are common in the same individuals, thereby indicating a mixed infection of TTV strains of different genotypes. An association is found between hepatitis of unknown etiology and the TTV genotypes which are detectable by PCR with primers deduced from the N22 region (genotype 1) in the open reading frame 1 encoding the capsid protein. It would be important to select the primers for specific detection of the TTV genotypes associated with clinical diseases, to further evaluate the capacity of TTV to induce acute and chronic liver disease as well as extrahepatic manifestations.
Hepatitis E Virus: An Update on Its Molecular, Clinical and Epidemiological Characteristics
Authors: Irshad Mohammad
Published: April 2003

The present overview gives a summary of HEV infection comprising its discovery, a characterisation of the virus, its diagnosis, epidemiology, course of the disease, and finally its prevention.
High Frequency of HCV Infection in Individuals with Isolated Antibody to Hepatitis B Core Antigen
Authors: Berger Annemarie , Doerr Hans Wilhelm , Rabenau Holger F. , Weber Bernard
Published: April 2003

Although isolated antibody to hepatitis B core antigen (anti-HBc) is frequently nonspecific or may be the only serological marker of past self-limiting hepatitis B, where antibodies against the surface antigen have disappeared, isolated anti-HBc seropositivity is frequently associated with chronic hepatitis B in HIV- and HCV-infected individuals. Of 5,520 samples that tested positive for anti-HBc (IMx and AxSYM CORE, Abbott, Delkenheim, Germany) at the Institute of Virology, University Clinic Frankfurt during the time interval from January 1994 to February 1996, 643 (11.6%) were isolated anti-HBc-reactive in the IMx and AxSYM CORE assays (inhibition values >90%). There was a statistically significant association between isolated anti-HBc seropositivity and HCV and HIV/HCV coinfection (p < 0.05). A total of 190 samples were available for further testing. Six (3.2%) of 190 isolated anti-HBc-positive samples were considered false-positive since they were only positive in the AxSYM or IMx CORE assay and a linear decrease of the measured signal could not be observed in dilution series. Of 184 serum samples tested with nested PCR using primers of the S genome region, only 6 (3.3%) were HBV DNA-positive. Anti-HBc-IgM antibody could be detected in 3 (1.6 %) of the tested samples using the IMx CORE-M. With the more sensitive VIDAS HBc IgM specific IgM antibody was detected in 15 (8.5%) of 177 samples at concentrations ranging from 10 to >200 Paul Ehrlich Institute U/ml. HIV or HCV coinfection was present in 28.1% and 37.5% of isolated anti-HBc-positive individuals, respectively. We conclude from our observations that only a limited proportion of anti-HBc-isolated individuals are potentially infectious, however anti-HBc-IgM which is detectable in any form of liver disease associated with HBV infection was present in more than 8% of the individuals. Of isolated anti-HBc-positive sera 37% were positive for anti-HCV, suggesting that anti-HCV antibody testing should be performed in isolated anti-HBc-positive individuals.
Identification of a Single Nucleotide Polymorphism in the MxA Gene Promoter (G/T at nt –88) Correlated with the Response of Hepatitis C Patients to Interferon
Authors: Hijikata Minako , Ohta Yasuhiko , Mishiro Shunji
Published: April 2003

The interferon (IFN)-inducible MxA protein is known to play an important role in the host defense against certain viruses. We aimed to see if any genetic polymorphism in the promoter region of the MxA gene is associated with the IFN responsiveness of hepatitis C virus (HCV)-infected patients. Initially we sequenced the promoter region of the MxA gene in 12 subjects and found a polymorphic site. We then constructed a specific PCR-RFLP system for this site and subjected 63 samples from chronic hepatitis C patients who were nonresponders (NR) to IFN therapy to it, 52 with sustained response (SR), and 42 healthy controls. Subjects were all Japanese, and unrelated. A single nucleotide polymorphism (SNP) was identified in the MxA promoter region: G/T alleles at nt position –88. Interestingly, this SNP was involved in a genetic element highly homologous to the IFN-stimulated response element consensus sequence, and the G-to-T change there makes this homology a little greater. The rate of G·G homozygosity was 31% in the SR patients, significantly lower than in the NR patients (62%, p = 0.0009), while that of healthy controls was between the two groups (48%). Differences in HCV genotypes did not influence this result. Based on these findings, we propose that the SNP of the MxA promoter at nt –88 identified in this study affects the expression of MxA protein, and may thus be associated with the response of HCV patients to IFN.
Enhancement of Herpes Simplex Virus-Induced Polykaryocyte Formation by 12-O-Tetradecanoyl Phorbol 13-Acetate: Association with the Reorganization of Actin Filaments and Cell Motility
Authors: Yura Yoshiaki , Kusaka Jun , Bando Takashi , Yamamoto Sin , Yoshida Hideo , Sato Mitsunobu
Published: April 2003

A morphological change induced by syn– herpes simplex virus type 1 (HSV-1), polykaryocyte formation, was enhanced by treatment with 12-O-tetradecanoyl phorbol 13-acetate (TPA) in A431 cells. TPA treatment decreased the number of stress fibers, but led to the development of spike-like filopodia and actin-containing long projections. Similar reorganization of actin filaments was observed in HSV-1-induced polykaryocytes. The actin filament-disrupting drug cytochalasin D, but not the microtubule-disrupting drug nocodazole, inhibited the effect of TPA on polykaryocyte formation, indicating that the actin microfilament system plays a key role in this event. HSV-1 glycoprotein D (gD) was present in the cytoplasm of HSV-1-infected cells and gD gene-transfected cells; its expression became prominent at long cell projections in the presence of TPA. These findings suggest that the reorganization of actin filaments and cell motility are associated with the enhancing effect of TPA on HSV-1-induced polykaryocyte formation.
Modulation of Arterial Growth of the Rabbit Carotid Artery Associated with Experimental Elevation of Blood Flow
Authors: Di Stefano Ida , Koopmans Donna R. , Langille B. Lowell
Published: April 2003

We examined the growth of the right common carotid artery of young rabbits after ligating the left common carotid artery at 3 weeks of age, a procedure that approximately doubled right carotid blood flows. Flow increased from 0.065 ± 0.003 to 0.096 ± 0.009 ml/s within 1 h and, at 15 weeks of age, carotid blood flows in experimental animals (0.747 ± 0.102 ml/s) were more than double of those of sham-operated control animals (0.334 ± 0.053 ml/s). Contralateral carotid ligation resulted in more rapid increases in diameter of the artery with growth in the experimental animals. At 15 weeks of age, the vessel was 15% larger than that of sham-operated controls (2.70 ± 0.09 vs. 2.34 ± 0.05 mm). This more rapid growth of diameter resulted in shear stresses that were not different from controls despite the higher blood flow rates. Interestingly, however, shear stresses in control arteries fell from 17.4 ± 3.4 to 9.19 ± 1.16 dyn/cm2 over the experimental period (p < 0.05). Elastin accumulation in the experimental artery was much more rapid than in controls and elastin contents were 49% more than in controls at 15 weeks of age. DNA and collagen contents were not significantly affected by contralateral carotid ligation. Previously, we found that experimental manipulations that decreased flow in the same artery of weanling rabbits substantially affected elastin and DNA accumulation, but had no effect on collagen contents. We conclude that increased blood flow is associated with arterial growth and specifically with accumulation of elastin, a wall constituent that bears much of the wall tension at resting blood pressure, and therefore is a primary determinant of resting vessel dimensions.
Induction of Nitric Oxide and Tetrahydrobiopterin Synthesis by Lipoteichoic Acid from Staphylococcus aureus in VascularSmooth Muscle Cells
Authors: Hattori Yoshiyuki , Kasai Kikuo , Nakanishi Nobuo , Gross Steven S. , Thiemermann Christoph
Published: April 2003

Lipoteichoic acid (LTA), a component of the membrane of gram-positive bacteria, induces an isoform of nitric oxide (NO) synthase (iNOS) in vascular smooth muscle; this process may be associated with the vascular failure observed in gram-positive septic shock. The aim of the present work was to study the cellular mechanisms involved in the induction of NO synthesis by LTA from Staphylococcus aureus in cultured rat vascular smooth muscle cells. LTA induces the gene expression of iNOS and GTP cyclohydrolase I (GTPCH) as well as the formation of NO and tetrahydrobiopterin (BH4), effects which are synergistic with interferon-γ. 2,4-Diamino-6-hydroxypyrimidine (DAHP), a selective inhibitor of GTPCH, inhibits both the increase in cellular levels of BH4 as well as the concomitant formation of NO caused by LTA in combination with interferon-γ. This inhibition by DAHP is reversed by co-addition of sepiapterin which is a substrate for BH4 synthesis. Thus, BH4 synthesis is an absolute requirement for the induction of NO synthesis by LTA in vascular smooth muscle. Our findings also suggest that interrupting pterin synthesis may be an effective target for pharmacologic interventions aimed at limiting NO overproduction in gram-positive shock.
Regulation of Insulin-Like Growth Factor-1 Expression in Vascular Endothelial Cells by the Inflammatory Cytokine Interleukin-1
Authors: Glazebrook Heather , Hatch Timothy , Brindle Nicholas P.J.
Published: April 2003

Insulin-like growth factor-1 (IGF-1) has important roles in vascular physiology and pathologies such as atherosclerosis. However, the factors that control expression of this growth factor in human vascular cells are largely unknown. In this study the effects of the inflammatory cytokine interleukin-1 (IL-1) on IGF-1 mRNA and peptide synthesis was examined in human endothelial cells. Cultured human umbilical vein endothelial (HUVE) cells were challenged with interleukin-1 and IGF-1 Ea and Eb mRNA levels were determined by nuclease protection assays and reverse transcription/polymerase chain reaction. IL-1 caused a dramatic increase in IGF-1 Ea mRNA in HUVE cells. Transcript levels peaked between 2 and 4 h of IL-1 treatment and declined over the subsequent 4 h. Consistent with its effect on mRNA, the inflammatory cytokine causes a 3-fold stimulation in production of IGF-1 peptide from endothelial cells. These results demonstrate increased IGF-1 mRNA and peptide in vascular endothelial cells stimulated with IL-1. This suggests that under conditions of local inflammation at the vessel wall the direct action of IL-1 on endothelium can lead to induction of IGF-1 which could promote the intimal hyperplasia often seen in these circumstances.
Expression of mRNA for Phospholipase A2, Cyclooxygenases, and Lipoxygenases in Cultured Human Umbilical Vascular Endothelial and Smooth Muscle Cells and in Biopsies from Umbilical Arteries and Veins
Authors: Öst Martin , Uhl Eberhard , Carlsson Maria , Gidlöf Anders , Söderkvist Peter , Sirsjö Allan
Published: April 2003

Arachidonic acid (AA) is released by phospholipase A2 (PLA2) and then converted into vasoactive and inflammatory eicosanoids by cyclooxygenases (COX) and lipoxygenases (LOX). These eicosanoids are important paracrine regulators of vascular permeability, blood flow, local pro- and anticoagulant activity and they play a major role in the local inflammatory response. We have investigated the presence of mRNAs for PLA2 and for isoforms of COX and LOX in both human endothelial cells (EC) and in human smooth muscle cells (SMC) in culture and in vascular biopsies of human umbilical veins (HUVB) and arteries (HUAB) by using the reversed transcription-polymerase chain reaction (RT-PCR) technique. Results show detectable levels of PLA2 type IV (cPLA2) in cultured EC and SMC and in vascular wall biopsies from HUAB and HUVB. The cultured EC and SMC demonstrate higher levels of both COX-1 and COX-2 with PCR analyses than do vascular wall biopsies from HUAB and HUVB. This indicates a difference in the native expression of COX-1 and COX-2 in cultures of EC and SMC compared to that in biopsies from intact vessel walls. The EC and SMC in culture do not express mRNA for 5-LOX, that was, however, expressed in the vascular wall biopsies. This speaks in favour of a constitutive, i.e. in vivo expression of 5-LOX in SMC in the vascular wall of both umbilical vein and arteries. Thus results from in vitro studies of constitutive COX and LOX expression in EC and vascular SMC in culture cannot simply be extrapolated to represent in vivo conditions.
A Platelet-Activating Factor Antagonist (WEB 2170) Preserves Endothelium-Dependent Vasodilatation and Prevents Development of a Neo-Intima Induced by a Periarterial Collar in Rabbit Carotid Arteries
Authors: Yin Zhan-Li , Hickey Haruyo , Dusting Gregory J.
Published: April 2003

Platelet-activating factor (PAF) may be involved in adhesion of leucocytes and migration of cells during vascular remodelling for it is expressed in leucocytes after cytokine priming and is required for cell adhesion. We studied the effects of WEB 2170, a potent PAF antagonist, on the development of an atheroma-like neo-intima induced by a peri-arterial collar in rabbits. Either WEB 2170 (3 mg/kg/day) or vehicle was given by subcutaneous injection once a day for 4 or 9 days, and on day 3 peri-arterial collars were applied to both carotid arteries in all animals. Two or 7 days after implanting the collars vasodilator responses to the endothelium-dependent vasodilator, acetylcholine and the endothelium-independent vasodilator, sodium nitroprusside were studied in isolated artery rings from both groups of rabbits. Neo-intima formation after 7 days (day 10 of treatment) was measured by light microscopy as the ratio of cross-sectional areas of intima and media, and expression of inducible nitric oxide synthase (iNOS) was studied by immunohistochemistry. PAF-induced platelet aggregation ex vivo was inhibited specifically in WEB 2170-treated rabbits. At day 5, acetylcholine-induced vasorelaxation in collared artery rings was markedly impaired as compared to control sections from both vehicle- and WEB 2170-treated rabbits. At day 10, acetylcholine-induced vasorelaxation in collared artery rings from vehicle rabbits was markedly less than in controls, but in WEB 2170-treated rabbits, the acetylcholine response in collared arteries was similar to control sections. Intimal thickening was much reduced in WEB 2170-treated rabbits, ratios of intima/media areas being vehicle: 0.21 ± 0.02 (n = 5) and WEB 2170: 0.07 ± 0.01 (n = 7; p < 0.01). Immunofluorescence showed expression of iNOS only in the neo-intima of vehicle-treated, collared arteries, but not in the residual neo-intima of WEB 2170-treated, collared arteries. These results suggest that WEB 2170 is effective in preserving endothelial function, prevents the development of neo-intima and blocks iNOS expression in the neo-intima in this model.