2.50
Hdl Handle:
http://hdl.handle.net/10755/157390
Type:
Presentation
Title:
GENE EXPRESSION DIFFERENCES IN AGING MOUSE MUSCLE TREATED WITH AZT
Abstract:
GENE EXPRESSION DIFFERENCES IN AGING MOUSE MUSCLE TREATED WITH AZT
Conference Sponsor:Western Institute of Nursing
Conference Year:2010
Author:Voss, Joachim G., PhD, RN
P.I. Institution Name:University of Washington School of Nursing
Title:Assistant Professor
Contact Address:Box 357266, Seattle, WA, 98195, USA
Co-Authors:Cassandra M. Steiner; Larry Adams; Jonathan Wanagat; David Marcinek; Peter S. Rabinovitch
PURPOSE: The purpose of this project was to determine the effects of azidothymidine (AZT) on gene expression of mitochondrially relevant genes in skeletal muscle of aging mice. A secondary aim was to develop a sensitive and reproducible protocol for reverse transcription polymerase chain reaction (RT-PCR).
BACKGROUND: Antiretroviral therapy (ART) significantly increased life expectancy in HIV disease. By 2020, 50% of HIV patients will be over 50 years of age. The interactions between ARTs and aging are poorly understood. The nucleoside reverse transcriptase inhibitor (NRTI) AZT is considered a backbone of ART. Unfortunately, the clinical usefulness of AZT is limited by its toxicity, notably being associated with dysfunction of mitochondria from inhibition of DNA replication. Mitochondria supply ATP for sustained muscle activity, impairment of mitochondrial ATP production RESULTS: in reduced muscle function. Similar pathology has been observed in aged muscle.
METHODS: Young (7 month) and old (27 month) wild type mice, were administered either 240 mg AZT/kg/day treatment in drinking water or water alone. Mice were sacrificed after 5 weeks and RNA was obtained from quadriceps skeletal muscle. Reverse transcription was compared between the Promega and ABI Reverse Transcription kits. RT- PCR was performed in triplicate and data was analyzed using the DeltaDelta cT method. Results were normalized to two housekeeping genes (18S rRNA and GAPDH). Genes investigated including: thymidine kinase 2 (TK2), DIABLO, cytochrome c oxidase subunit 2 (COX2), core protein II subunit of the mitochondrial cytochrome bc1 complex (UQCRC2), succinate dehydrogenase complex, subunit B (SDHB), and translocase of inner mitochondrial membrane 17 homolog B (Timm17b). These genes were selected based on published data.
RESULTS: After optimization of the protocol with a 1 hour RT amplification time and 18S as normalizing gene, RT-PCR data was established for 22 mice. Experiments were repeated twice under the same conditions to determine reproducibility of the protocol. Between the 2 studies, the expression levels of 4 of the 5 genes were consistent (TK2, TIMM17B, SDHB and Diablo). TK2 gene expression between young and old control animals was the same, while old animals treated with AZT showed a 50% increase in expression compared to young mice. SDHB expression was 40% lower in old vs. young controls. AZT further suppressed SDHB expression by another 30% in old AZT-treated compared to old control mice. Diabolo was 40% higher in old vs. young controls. When treated with AZT, DIABOLO expression was 100% higher in younger animals compared to old mice. The results from the TIMM17B gene expression indicate only a small decline in old animals with AZT compared to old controls; while no difference was observed in the young treated and control mice.
IMPLICATIONS: We confirmed previous results that mitochondrially relevant genes change with AZT administration. We demonstrated that age and AZT treatment have a significant impact on gene expression such as in TK2. Further studies will show whether the observed gene expression findings translate into protein changes and what the IMPLICATIONS: of these gene changes are for treatment and monitoring of older adults living with HIV.
Repository Posting Date:
26-Oct-2011
Date of Publication:
17-Oct-2011
Sponsors:
Western Institute of Nursing

Full metadata record

DC FieldValue Language
dc.typePresentationen_GB
dc.titleGENE EXPRESSION DIFFERENCES IN AGING MOUSE MUSCLE TREATED WITH AZTen_GB
dc.identifier.urihttp://hdl.handle.net/10755/157390-
dc.description.abstract<table><tr><td colspan="2" class="item-title">GENE EXPRESSION DIFFERENCES IN AGING MOUSE MUSCLE TREATED WITH AZT</td></tr><tr class="item-sponsor"><td class="label">Conference Sponsor:</td><td class="value">Western Institute of Nursing</td></tr><tr class="item-year"><td class="label">Conference Year:</td><td class="value">2010</td></tr><tr class="item-author"><td class="label">Author:</td><td class="value">Voss, Joachim G., PhD, RN</td></tr><tr class="item-institute"><td class="label">P.I. Institution Name:</td><td class="value">University of Washington School of Nursing</td></tr><tr class="item-author-title"><td class="label">Title:</td><td class="value">Assistant Professor</td></tr><tr class="item-address"><td class="label">Contact Address:</td><td class="value">Box 357266, Seattle, WA, 98195, USA</td></tr><tr class="item-email"><td class="label">Email:</td><td class="value">vossj@u.washington.edu</td></tr><tr class="item-co-authors"><td class="label">Co-Authors:</td><td class="value">Cassandra M. Steiner; Larry Adams; Jonathan Wanagat; David Marcinek; Peter S. Rabinovitch</td></tr><tr><td colspan="2" class="item-abstract">PURPOSE: The purpose of this project was to determine the effects of azidothymidine (AZT) on gene expression of mitochondrially relevant genes in skeletal muscle of aging mice. A secondary aim was to develop a sensitive and reproducible protocol for reverse transcription polymerase chain reaction (RT-PCR).<br/>BACKGROUND: Antiretroviral therapy (ART) significantly increased life expectancy in HIV disease. By 2020, 50% of HIV patients will be over 50 years of age. The interactions between ARTs and aging are poorly understood. The nucleoside reverse transcriptase inhibitor (NRTI) AZT is considered a backbone of ART. Unfortunately, the clinical usefulness of AZT is limited by its toxicity, notably being associated with dysfunction of mitochondria from inhibition of DNA replication. Mitochondria supply ATP for sustained muscle activity, impairment of mitochondrial ATP production RESULTS: in reduced muscle function. Similar pathology has been observed in aged muscle.<br/>METHODS: Young (7 month) and old (27 month) wild type mice, were administered either 240 mg AZT/kg/day treatment in drinking water or water alone. Mice were sacrificed after 5 weeks and RNA was obtained from quadriceps skeletal muscle. Reverse transcription was compared between the Promega and ABI Reverse Transcription kits. RT- PCR was performed in triplicate and data was analyzed using the DeltaDelta cT method. Results were normalized to two housekeeping genes (18S rRNA and GAPDH). Genes investigated including: thymidine kinase 2 (TK2), DIABLO, cytochrome c oxidase subunit 2 (COX2), core protein II subunit of the mitochondrial cytochrome bc1 complex (UQCRC2), succinate dehydrogenase complex, subunit B (SDHB), and translocase of inner mitochondrial membrane 17 homolog B (Timm17b). These genes were selected based on published data. <br/>RESULTS: After optimization of the protocol with a 1 hour RT amplification time and 18S as normalizing gene, RT-PCR data was established for 22 mice. Experiments were repeated twice under the same conditions to determine reproducibility of the protocol. Between the 2 studies, the expression levels of 4 of the 5 genes were consistent (TK2, TIMM17B, SDHB and Diablo). TK2 gene expression between young and old control animals was the same, while old animals treated with AZT showed a 50% increase in expression compared to young mice. SDHB expression was 40% lower in old vs. young controls. AZT further suppressed SDHB expression by another 30% in old AZT-treated compared to old control mice. Diabolo was 40% higher in old vs. young controls. When treated with AZT, DIABOLO expression was 100% higher in younger animals compared to old mice. The results from the TIMM17B gene expression indicate only a small decline in old animals with AZT compared to old controls; while no difference was observed in the young treated and control mice.<br/> IMPLICATIONS: We confirmed previous results that mitochondrially relevant genes change with AZT administration. We demonstrated that age and AZT treatment have a significant impact on gene expression such as in TK2. Further studies will show whether the observed gene expression findings translate into protein changes and what the IMPLICATIONS: of these gene changes are for treatment and monitoring of older adults living with HIV.<br/></td></tr></table>en_GB
dc.date.available2011-10-26T19:49:46Z-
dc.date.issued2011-10-17en_GB
dc.date.accessioned2011-10-26T19:49:46Z-
dc.description.sponsorshipWestern Institute of Nursingen_GB
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