Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019158 (hepatitis)
 30,205 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

As major antigen-presenting cells and effectors in the maintenance of tolerance, dendritic cells (DCs) are key cells of the immune system and can thus be envisioned to have roles in immunotherapy strategies. We, and others, previously showed that simian immunodeficiency virus (SIV)-derived lentiviral vectors were able to deliver a gene into human differentiated DCs. We describe here the upgrading of the SIV vector system and the improvements of the transduction protocol, which allowed us to transduce more than 90% of human monocyte-derived DCs. We developed new SIV lentiviral vectors carrying SIV splice regulatory elements and either the woodchuck hepatitis virus regulatory element (WPRE) or the murine phosphoglycerate-kinase 1 (PGK) promoter. We show that insertion of the WPRE in the SIV vector is detrimental to gene transfer in DCs, while this sequence increases transgene expression in 293T cells. Using an optimized SIV vector, high levels of transgene expression were obtained in more than 30% of human DCs at a multiplicity of infection (MOI) of 1, and close to 100% using a MOI of 20. VSV-G pseudotyped vectors generated with only gag, pol, tat, and rev helper functions failed to transduce DCs. This defect was completely rescued when the SIV accessory gene vpx was provided in trans in vector-producing cells. Genetically modified DCs were shown to behave as bona fide DCs in both allogenic and autologous mixed leukocyte reactions. These findings allow us to propose an optimal system for efficient and safe DC transduction with improved SIV vectors.
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PMID:High levels of transduction of human dendritic cells with optimized SIV vectors. 1186 18

Gene transfer systems based on lentiviruses have emerged as promising gene delivery vehicles for human gene therapy due to their ability to efficiently transduce nondividing target cells. Both primate and nonprimate lentiviruses have been used for construction of lentiviral vectors. An early generation of gene transfer system based on bovine immunodeficiency virus (BIV) has been developed (R. D. Berkowitz, H. Ilves, W. Y. Lin, K. Eckert, A. Coward, S. Tamaki, G. Veres, and I. Plavec, 2001, J. Virol. 75, 3371-3382). In this study, we mapped the BIV Rev response element (RRE) to 312 bp of the Env coding region. Furthermore, we compared transduction efficiencies of vectors containing different portions of the BIV Gag coding region and found that the first 104 bp of gag contains a functional part of the BIV packaging signal. These findings enabled the generation of a minimal BIV-based lentiviral vector. The minimal transfer vector construct consists of a self-inactivating long terminal repeats (LTR), minimal packaging sequence, putative central polypurine tract, minimal RRE, an internal promoter driving the gene of interest, and a woodchuck hepatitis posttranscriptional regulatory element. In addition, we constructed a BIV packaging construct containing gag/pol, minimal Rev/RRE, and the accessory gene vpy. The regulatory gene tat and the accessory genes vif and vpw have been inactivated or truncated. The current system has significantly reduced regions of homologies between the transfer vector and the packaging constructs. The vectors generated from this system achieved a titer of greater than 1 x 10(6) transducing units per milliliter and are fully functional as indicated by their ability to efficiently transduce both dividing and nondividing cells. These modifications should provide improved safety features for the BIV-based gene transfer system.
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PMID:Mapping of the bovine immunodeficiency virus packaging signal and RRE and incorporation into a minimal gene transfer vector. 1249 Mar 99

The genome of hepatitis delta virus (HDV) is a small single-stranded circular RNA that is replicated via RNA-directed RNA synthesis. This makes use of a host RNA polymerase, probably pol II, that normally transcribes DNA templates. In vivo, the host polymerase can initiate replication from transfected linear RNAs using intramolecular template-switching. The present studies report that the polymerase could also achieve intermolecular switching leading to "reconstitution" of full-length HDV RNAs following transfection with two linear RNAs that were less than full length and yet lacking different regions of the genome. These two RNAs were synthesized in vitro, gel purified, pre-annealed, and then transfected into delta293, a cell line conditionally expressing the small delta antigen that is essential for HDV replication. Northern analyses of total RNA harvested from transfected cells detected the accumulation of full-length HDV genomic and antigenomic RNAs. Such reconstitution of full-length replicating HDV RNA was also achieved using nine other pairs of antigenomic RNAs and three pairs of genomic RNAs. Annealing of the RNAs prior to transfection was required for detectable HDV reconstitution. A second cell line, Huh7, also supported reconstitution when a pair of RNAs was cotransfected together with mRNA for the small delta protein. Taken together, these results support a model that observed genome reconstitution is a special form of recombination involving intermolecular template switches and they provide insights into the mechanism of RNA-directed RNA transcription catalyzed by a host RNA polymerase.
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PMID:Reconstitution in cultured cells of replicating HDV RNA from pairs of less than full-length RNAs. 1557 17

HDV replicates its circular RNA genome using a double rolling-circle mechanism and transcribes a hepatitis delta antigen-encodeing mRNA from the same RNA template during its life cycle. Both processes are carried out by RNA-dependent RNA synthesis despite the fact that HDV does not encode an RNA-dependent RNA polymerase (RdRP). Cellular RNA polymerase II has long been implicated in these processes. Recent findings, however, have shown that the syntheses of genomic and antigenomic RNA strands have different metabolic requirements, including sensitives to alpha-amanitin and the site of synthesis. Evidence is summarized here for the involvement of other cellular polymerases, probably pol I, in the synthesis of antigenomic RNA strand. The ability of mammalian cells to replicate HDV RNA implies that RNA-dependent RNA synthesis was preserved throughout evolution.
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PMID:HDV RNA replication: ancient relic or primer? 1690 19

Recent guidelines emphasise the risk of hepatitis B virus (HBV) reactivation among patients with hematologic malignancies of B lineage, in which HBV has been recently hypothesised to play a pathogenetic role. We aimed to determine the prevalence of occult HBV infection (OBI) of peripheral blood mononuclear cells, defined as detection of sequences from >or=2 HBV genes in subjects lacking hepatitis B surface antigen, among patients with treatment-naive chronic lymphocytic leukemia (CLL). HBV DNA sequences from four HBV genes (S, X, core and pol) were searched for in archival material obtained at diagnosis (N = 173), and from age and sex-matched controls. OBI was observed in 17/173 (10%) patients and 5/173 (3%) controls (OR = 3.6, 95% CI 1.37-9.79, p = 0.014). OBI was not associated with differences on 5-year survival and biological predictors, but patients with CLL with OBI had significantly lower peripheral blood lymphocyte count. After 8 years of observation without treatment, one OBI positive patient with CLL converted into positive HBsAg serology and developed active hepatitis. In conclusion, OBI is significantly more prevalent among patients with CLL than in age and sex-matched controls, and may contribute to the susceptibility of patients with CLL to HBV reactivation, whether exposed or not to biological agents.
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PMID:Occult hepatitis B virus infection of peripheral blood mononuclear cells among treatment-naive patients with chronic lymphocytic leukemia. 1937 59

To understand how DNA-dependent RNA polymerase II (pol II) recognizes hepatitis delta virus (HDV) RNA as a template, it is first necessary to identify the HDV sequence that acts as a promoter of pol II-initiated RNA synthesis. Therefore, we isolated the pol II-response element from HDV cDNA and examined the regulation by hepatitis delta antigens (HDAgs). Two HDV cDNA fragments containing bidirectional promoter activity were identified. One was located at nt 1582-1683 (transcription-promoter region 1, TR-P1) and the other at nt 1223-1363 (transcription-internal region 5, TR-I5). The promoter activities of these two regions were enhanced by HDAgs to differing degrees. Next, the role of these sequences in an HDV cDNA-free RNA replication system was characterized by site-directed mutagenesis. Our data showed that: (i) the AUG codon at the HDAg ORF of HDV RNA (nt 1599-1601) that mutates to UAG (amber stop codon) results in loss of dimeric but not monomeric HDV RNA synthesis. (ii) A 5 nt mutation of TR-P1 (P1-m5, nt 1670-1674) abolishes RNA replication completely. Two-nucleotide-mutated RNA (P1-m2, nt 1662-1663) is able to synthesize short RNAs but not monomeric HDV RNA. (iii) A mutation in 5 nt at the TR-I5 region (I5-m5, nt 1351-1355) also abolishes HDV replication. Mutants with 2 nt mutations (I5-m2, nt 1351-1352) or 3 nt mutations (I5-m3, nt 1353-1355) inhibit HDV dimeric but not monomeric RNA synthesis. Furthermore, large HDAg is expressed in cells transfected with I5-m3 and I5-m2 RNAs and that demonstrate the RNA-editing event in the monomeric HDV RNA. These results provide further understanding of the double rolling-circle mechanism in HDV RNA replication.
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PMID:Multiple genomic sequences of hepatitis delta virus are associated with cDNA promoter activity and RNA double rolling-circle replication. 2215 80

Fecal specimens, including swabs and litter extracts, collected from chickens, domestic ducks, turkeys, and Canadian geese were tested using degenerate primers targeting regions encoding for conserved amino acid motifs (YGDD and DY(T/S)(R/K/G)WDST) in calicivirus RNA-dependent RNA polymerases. Similar motifs are also present in other RNA viruses. Two fecal specimens and 18 litter extracts collected from chickens and turkeys yielded RT-PCR products. BLAST search and phylogenetic analysis revealed that all amplicons represented picornaviruses that clustered into two major groups. Four chicken and one turkey samples yielded 250 bp amplicons with 84-91% nucleotide identity to the recently described turkey hepatitis viruses, while 280 and 283 bp amplicons obtained from 11 chicken and 4 turkey samples represented novel picornaviruses with the closest nucleotide identity to kobuviruses (54-61%) and turdiviruses (47-54%). Analysis of 2.2-3.2 kb extended genome sequences including the partial P2 (2C) and complete P3 (3A, 3B (VPg), 3C(pro), and 3D(pol)) regions of selected strains indicated that viruses yielding the 280/283 bp amplicons represent a putative new genus of Picornaviridae. The 3'-non-translated region (NTR) of the turkey hepatitis-like viruses described in this study was significantly longer (641-654 nt) than that of any of the other piconaviruses and included a putative short open reading frame (ORF). In summary, we report the molecular detection of novel picornaviruses that appear to be endemic in both chickens and turkeys.
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PMID:Molecular detection of novel picornaviruses in chickens and turkeys. 2216 Aug 27


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