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Biomedical subjects

M Roggendorf

Publications and source records attributed to M Roggendorf.

At least 19 recordsLinked to original sources
PubMed2006

Immunology of HDV infection.

Hepatitis delta virus (HDV) infection may occur as coinfection with hepatitis B virus (HBV) or as superinfection of a chronically HBV-infected patient. A strong antibody response is mounted, which persists for many years; however, it is not able to modulate the course of infection. In most cases the superinfection takes a chronic course. In patients with inactive disease (HDV PCR negative) an oligospecific T-helper cell immune response and a cytotoxic T-cell response were found, which were absent in patients with persistent viremia. The role of the cellular immune response in liver injury during acute infection has not been investigated. Vaccination strategies tested in the woodchuck model induced specific B- and T-cell responses but failed to protect from HDV infection.

Animals↗
PubMed2005

Fluctuation of the cytokine expression in the liver during the chronic woodchuck hepatitis virus (WHV) infection is not related to viral load.

The woodchuck together with the woodchuck hepatitis virus (WHV) is an excellent model to study the pathogenesis of hepadnaviral infections. Chronic WHV infection causes severe liver disease and hepatocellular carcinoma in woodchucks. The mechanism of viral clearance is not fully understood, interferons seem to play a major role in down-regulating viral replication prior to elimination of infected hepatocytes. We investigated on the pattern of cytokine and T-cell-marker expression in livers of woodchucks chronically infected with WHV. RNase-protection-assay (RPA) was used to determine mRNA of woodchuck specific genes (TNF-alpha, IFN-gamma, IL-15, CD3, CD4, CD8). Serial liver biopsies were performed daily or weekly in eight chronic WHV-carrier woodchucks. Cytokine/T-cell-marker expression differed significantly between the time points up to +/-50% within each woodchuck. The different expression patterns of cytokines or T-cell-markers did not correlate to the (weak) fluctuations in the viremia but may explain the observed fluctuations in the WHV/HBV-load in chronically infected individuals. Furthermore, we observed associations between cytokine and T-cell-marker expression. The marginal fluctuations in viremia during the chronic infection may indicate, that, once the chronic hepadnaviral infection is established, cytokines/interferons expressed endogenously (i.e. not vector-borne or injected) play only a minor role.

Animals↗
PubMed2004

PCR for varicella zoster virus genome negative in corneal epithelial cells of patients with Thygeson's superficial punctate keratitis.

PURPOSE: To find out if varicella zoster virus is the causative agent of Thygeson's superficial punctuate keratitis. METHODS: Epithelial cells were harvested from the punctuate epithelial lesions 9 patients with Thygeson's superficial punctate keratitis. After DNA extraction polymerase chain reaction was carried out with varicella zoster virus primers. RESULTS: All samples were negative with regard to varicella zoster virus genome. CONCLUSIONS: This result suggests that varicella zoster virus is most probably not the causative agent of Thygeson's superficial punctate keratitis.

Adolescent↗
PubMed2004

Identification of a glycosylation site in the woodchuck hepatitis virus preS2 protein and its role in protein trafficking.

The middle surface antigen (M-sAg) of hepadnaviruses is one of three envelope proteins that share a common C-terminal S domain. M-sAg contains the preS2 domain in addition to the S region. The preS2 region of woodchuck hepatitis virus (WHV) contains a potential glycosylation site Asn-Gln-Thr at amino acid (aa) positions 3-5. In this study, we mutated this site by site-directed mutagenesis and confirmed that glycosylation occurs here. In in vitro translation assays, the mutation Thr to Asn at aa 5 significantly impaired glycosylation of M-sAg. The mutated M-sAg formed abnormal clustered structures in transfected cells as determined by immunofluorescent staining. Confocal microscopic analysis showed that an enrichment of this glycosylation-deficient protein in the Golgi apparatus occurred, which is not typical for the wild-type protein. These results are consistent with earlier findings that incorrect glycosylation of viral proteins may interfere with virus assembly.

Amino Acid Sequence↗
PubMed2003

Hepatitis C virus genotypes in Hungarian and Austrian patients with chronic hepatitis C.

Hepatitis C virus (HCV) genotypes are relevant to epidemiological questions, vaccine development, and clinical management of chronic HCV infection. The aim of this study was to determine HCV genotypes of South Hungarian and Southeast Austrian patients with chronic hepatitis C. Results were obtained by the largely automated TruGene HCV 5'NC Genotyping Kit (Visible Genetics, Toronto, Ontario) and by phylogenetic analysis. All of the 20 Hungarian patients and 15 out of 20 Austrian patients were infected with genotype 1. The remaining Austrian patients were infected with genotypes 3 or 2. With the commercially available assay, it was not possible to determine the HCV subtype in a total of three patients. The TruGene HCV 5'NC Genotyping assay for the determination of HCV genotypes proved to be useful for a high-throughput routine diagnostic laboratory.

Alanine Transaminase↗
PubMed2003

Hepatitis B virus (HBV) and hepatitis C virus (HCV) infections in health care workers (HCWs): guidelines for prevention of transmission of HBV and HCV from HCW to patients.

The transmission of viral hepatitis from health care workers (HCW) to patients is of worldwide concern. Since the introduction of serologic testing in the 1970s there have been over 45 reports of hepatitis B virus (HBV) transmission from HCW to patients, which have resulted in more than 400 infected patients. In addition there are six published reports of transmissions of hepatitis C virus (HCV) from HCW to patients resulting in the infection of 14 patients. Additional HCV cases are known of in the US and UK, but unpublished. At present the guidelines for preventing HCW to patient transmission of viral hepatitis vary greatly between countries. It was our aim to reach a Europe-wide consensus on this issue. In order to do this, experts in blood-borne infection, from 16 countries, were questioned on their national protocols. The replies given by participating countries formed the basis of a discussion document. This paper was then discussed at a meeting with each of the participating countries in order to reach a Europe-wide consensus on the identification of infected HCWs, protection of susceptible HCWs, management and treatment options for the infected HCW. The results of that process are discussed and recommendations formed. The guidelines produced aim to reduce the risk of transmission from infected HCWs to patients. The document is designed to complement existing guidelines or form the basis for the development of new guidelines. This guidance is applicable to all HCWs who perform EPP, whether newly appointed or already in post.

DNA, Viral↗
PubMed2003

Molecular genetic and biochemical analysis of woodchuck (Marmota monax) MHC class I polymorphism.

The woodchuck (Marmota monax) is an animal model that is used in the study of human hepatitis B virus ( HBV ) infection. A knowledge of woodchuck MHC class I (Mamo-I) genes and gene products is therefore essential for understanding the antigen-specific T-cell responses in this animal model. A number of Mamo-I genes have been identified by molecular cloning and sequencing. However, the allelic nature of these genes has not been proven by classical genetics like the segregation analysis in families. In this study, we analyzed the allelic diversity of Mamo-I in two three-generation woodchuck families including 15 members by sequencing of Mamo-I genes and immunoblotting of Mamo-I proteins after one-dimensional isoelectric focusing (1D-IEF). In addition to four published Mamo-I alleles, six new alleles that belonged to the same locus as the known Mamo-I alleles (Mamo-A) were found within the two woodchuck families. A typical Mendelian segregation of Mamo-I gene and antigens was observed in the families studied. For simple and rapid detection of allelic variability of Mamo-I gene, a typing method based on the detection of PCR products amplified by sequence specific primers (SSP) has been developed and tested in 41 unrelated animals. The most prevalent allele was Mamo-A*01 with a frequency of 21.9% followed by Mamo-A*07 (12.2%). Our study established Mamo-A as a classical MHC class I locus by the polymorphic and allelic nature of Mamo-I gene in the woodchuck.

Amino Acid Sequence↗
PubMed2003

Molecular characterization of CD28 and cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) of woodchuck (Marmota monax).

Eastern woodchuck (Marmota monax) became an important animal model to study the immunological processes in hepatitis B virus infection. To facilitate further study of T-cell responses in this model, we cloned and sequenced the cDNAs of Woodchuck CD28 and cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4), which play important roles for the regulation of T-cell activation by delivering the costimulation signals. According to the deduced amino-acid sequences, Woodchuck CD28 showed a similarity of 70% to 80% to its mammalian homologues. Woodchuck CTLA-4 has a higher similarity of 74% to 85% to corresponding mammalian CTLA-4 molecules. The strict conservation of critical amino-acid residues like cystein and asparagine residues in Woodchuck CD28 and CTLA-4 suggests that both molecules may structurally resemble their human or mouse homologues. A hexapeptide motif, MYPPPY, which has been supposed to be essential for the interaction with CD80, is present in both Woodchuck CD28 and CTLA-4. The cloned cDNAs of Woodchuck CD28 and CTLA-4 were placed under the control of the cytomegalovirus (CMV) promoter of the mammalian expression vector pcDNA3. Both proteins were expressed and detected by respective crossreactive antibodies in transiently transfected mammalian cells. By immunohistochemical staining with these antibodies, CD28 and CTLA-4 were also detected on cultured woodchuck peripheral blood lymphocytes. The molecular characterization of Woodchuck CD28 and CTLA-4 will facilitate studies on the T-cell response to hepadnavirus in the woodchuck model.

Animals↗
PubMed2003

High prevalence of human metapneumovirus infection in young children and genetic heterogeneity of the viral isolates.

RNA of the newly identified human metapneumovirus (HMPV) was detected in nasopharyngeal aspirates of 11 of 63 (17.5%) young children with respiratory tract disease. Markers of infection caused by another member of the Pneumovirinae subfamily of the family Paramyxoviridae, respiratory syncytial virus (RSV), were identified in 15 of these patients (23.8%). Three patients were simultaneously infected with HMPV and RSV. Studies of the clinical characteristics of HMPV-infected children did not reveal any difference between HMPV-infected patients and a control population of RSV-infected patients with regard to disease severity, but the duration of symptoms was significantly shorter for HMPV-infected patients. Phylogenetic analysis of the amplified viral genome fragments confirmed the existence and simultaneous circulation within one epidemic season of HMPV isolates belonging to two genetic lineages.

Genetic Variation↗
PubMed2002

Quantitation of hepatitis C virus RNA by third generation branched DNA-based signal amplification assay.

Quantitation of hepatitis C virus (HCV) RNA has become an important tool in different clinical settings and is used extensively for pretreatment evaluation of patients infected chronically with HCV. In this study, the performance characteristics of the third generation branched DNA-based signal amplification assay (bDNA 3.0) for HCV RNA quantitation were established. The new assay version showed an analytical specificity of 98%. Mean intra- and between-run imprecisions were 6.8 and 11.2%, respectively. The assay was linear over its entire dynamic range. Quantitation appeared to be unaffected by the genotypic variability of HCV. A comparison of bDNA 3.0 with the second generation bDNA assay calibrated against the international WHO HCV RNA standard, and the PCR-based Cobas Amplicor HCV Monitor 2.0 revealed a fairly good correlation among the assays. Twenty-nine and 11% of the paired quantitative results differed by more than log(10)0.5 (i.e. three-fold). All three assays after calibration against the WHO standard also yielded clinically comparable results with regard to the tailoring of interferon alpha/ribavirin treatment duration in patients infected by HCV genotypes 1, 4, and 5.

Adolescent↗
PubMed2002

RT-PCR-based detection of circulating calcitonin-producing cells in patients with advanced medullary thyroid cancer.

In patients with medullary thyroid carcinoma (MTC) the clinical course of disease ranges from rapid tumor progression to long-lasting stable disease. The purpose of the present study was to investigate whether circulating tumor cells can be detected in the peripheral blood of patients with MTC by RT-PCR targeted to calcitonin (CT) mRNA and whether the results of this method are correlated with disease manifestation and metastatic potential. Blood samples from 19 consecutive patients with MTC and elevated CT levels were analyzed. Four had newly diagnosed MTC, and 15 patients had undergone total thyroidectomy. Six of 19 patients had detectable CT mRNA by RT-PCR. CT levels in the CT mRNA-positive patients were significantly higher than those in CT mRNA-negative patients [2,239-265,313 pg/ml; median 80,921 pg/ml (n = 6) vs. 70-46,787 pg/ml; median, 932 pg/ml (n = 13); P = 0.006]. CT mRNA was detectable in 5 of 8 patients with distant metastases, in 1 of 6 patients with local/regional lymph node metastases, but in none of the patients with newly diagnosed, organ-confined MTC (n = 2) or surgically treated MTC without tumor manifestation by various imaging studies (n = 3). In peripheral blood from 10 healthy volunteers and 21 patients with benign thyroid nodules, no CT RNA could be detected. In conclusion, an RT-PCR-based procedure was established to detect circulating CT-producing cells in the peripheral blood of patients with MTC. RT-PCR results seem to reflect tumor spread and aggressiveness and thus may help with early identification of patients with disseminated and rapidly progressive disease.

Adult↗
PubMed2001

Induction of cytotoxic T lymphocyte responses against hepatitis delta virus antigens which protect against tumor formation in mice.

The cellular immune response is a crucial defense mechanism against hepatotropic viruses and in chronic viral hepatitis prevention. Moreover, hepatitis delta virus (HDV) immunogenicity may be an important component in the development of prophylactic and therapeutic vaccines. Therefore, we evaluated the immunogenicity of the small (HDAg) or large delta antigen (LHDAg) to be used as a DNA-based vaccine. We immunized different mouse haplotypes, determined cellular immune responses, and tested protection of animals against tumor formation using syngeneic tumor cells stably expressing the delta antigens. Both LHDAg and HDAg primed CD4+ and CD8+ T cell immunity against both forms of delta antigens. CD8+ T cell frequencies were about 1% and antigen-specific CD8+ T cells remained detectable directly ex vivo for at least 35 days post-injection. No anti-delta antibody responses could be detected despite multiple detection systems and varied immunization approaches. We observed protection against syngeneic tumor formation and growth in mice immunized with DNA plasmids encoding secreted or intracellular forms of HDAg and LHDAg but not with recombinant HDAg establishing the generation of significant cellular immunity in vivo. Both CD4+ and CD8+ T cells were required for antitumoral activity as determined by in vivo T cell depletion experiments. The results indicate that DNA-based immunization with genes encoding LHDAg and HDAg induces strong T cell responses and, therefore, is an attractive approach for the construction of therapeutic and prophylactic T cell vaccines against HDV.

Animals↗
PubMed2001

Immunization of woodchucks (Marmota monax) with hepatitis delta virus DNA vaccine.

We investigated the DNA immunization approach in order to induce a protective immune response against hepatitis delta virus (HDV) superinfection of chronically woodchuck hepatitis virus (WHV) infected woodchucks. The animals were immunized with an expression vector encoding HDAg by gene gun. T cell and humoral immune responses induced by this protocol were determined and compared with those induced by HDAg immunization using a CpG oligonucleotide as an adjuvant. After immunization the woodchucks were challenged with 10(6) genome equivalents of HDV. The protein immunization with HDAg induced good humoral and T helper cell responses in the woodchucks, but did not protect them from HDV superinfection. The DNA immunized woodchucks were also not protected from HDV superinfection, however, the course of infection was modified: HDV viremia occurred later, the typical fluctuation of the HDV RNA titer with several peaks was absent, and antibodies to HDV were not detectable.

Animals↗