Document Detail


Use of new T-cell-based cell lines expressing two luciferase reporters for accurately evaluating susceptibility to anti-human immunodeficiency virus type 1 drugs.
MedLine Citation:
PMID:  17182760     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Two new T-cell-based reporter cell lines were established to measure human immunodeficiency virus type 1 (HIV-1) infectivity. One cell line naturally expresses CD4 and CXCR4, making it susceptible to X4-tropic viruses, and the other cell line, in which a CCR5 expression vector was introduced, is susceptible to both X4- and R5-tropic viruses. Reporter cells were constructed by transfecting the human T-cell line HPB-Ma, which demonstrates high susceptibility to HIV-1, with genomes expressing two different luciferase reporters, HIV-1 long terminal repeat-driven firefly luciferase and cytomegalovirus promoter-driven renilla luciferase. Upon HIV infection, the cells expressed firefly luciferase at levels that were highly correlated (r2=0.91 to 0.98) with the production of the capsid antigen p24. The cells also constitutively expressed renilla luciferase, which was used to monitor cell numbers and viability. The reliability of the cell lines for two in vitro applications, drug resistance phenotyping and drug screening, was confirmed. As HIV-1 efficiently replicated in these cells, they could be used for multiple-round replication assays as an alternative method to a single-cycle replication protocol. Coefficients of variation for drug susceptibility evaluated with the cell lines ranged from 17 to 41%. The new cell lines were beneficial for evaluating antiretroviral drug resistance. Firefly luciferase gave a wider dynamic range for evaluating virus infectivity, and the introduction of renilla luciferase improved assay reproducibility. The cell lines were also beneficial for screening new antiretroviral agents, as false inhibition caused by the cytotoxicity of test compounds was easily detected by monitoring renilla luciferase activity.
Authors:
Tomoko Chiba-Mizutani; Hideka Miura; Masakazu Matsuda; Zene Matsuda; Yoshiyuki Yokomaku; Kosuke Miyauchi; Masako Nishizawa; Naoki Yamamoto; Wataru Sugiura
Publication Detail:
Type:  Evaluation Studies; Journal Article; Research Support, Non-U.S. Gov't     Date:  2006-12-20
Journal Detail:
Title:  Journal of clinical microbiology     Volume:  45     ISSN:  0095-1137     ISO Abbreviation:  J. Clin. Microbiol.     Publication Date:  2007 Feb 
Date Detail:
Created Date:  2007-02-05     Completed Date:  2007-03-29     Revised Date:  2013-06-06    
Medline Journal Info:
Nlm Unique ID:  7505564     Medline TA:  J Clin Microbiol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  477-87     Citation Subset:  IM    
Affiliation:
AIDS Research Center, National Institute of Infectious Diseases, 4-7-1 Gakuenn, Musashimurayama, Tokyo 2080011, Japan.
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MeSH Terms
Descriptor/Qualifier:
Anti-HIV Agents / pharmacology*
Cell Line
Drug Resistance, Viral
Genes, Reporter*
HIV-1 / drug effects*,  pathogenicity
Humans
Luciferases / genetics,  metabolism*
Microbial Sensitivity Tests
Receptors, CCR5 / metabolism
Receptors, CXCR4 / metabolism
T-Lymphocytes / virology*
Transfection
Virus Replication
Chemical
Reg. No./Substance:
0/Anti-HIV Agents; 0/Receptors, CCR5; 0/Receptors, CXCR4; EC 1.13.12.-/Luciferases
Comments/Corrections

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