Morphological and molecular course of mitochondrial pathology in cultured human cells exposed long-term to Zidovudine.
Long-term use of antiretroviral nucleoside reverse transcriptase inhibitors (NRTIs) as therapy for human immunodeficiency virus-1 (HIV-1) infection is limited by mitochondrial toxicity. Here we document mitochondrial pathology during the long-term culture of human HeLa cells in the presence or absence of the NRTI Zidovudine(R) (AZT, 800 muM) for up to 77-passages (p), with samples taken at early (p5-p11), middle (p36 and p37), and late (p70-p77) passages. Samples were analyzed for changes in mitochondrial morphology, mitochondrial (mt)DNA quantity, nuclear and mitochondrial gene expression, and mitochondrial membrane potential. Mitochondria showed abnormal proliferation at p5 and abnormal morphology >/=p36. mtDNA quantity was increased at p5 and p11, and 65% depleted at p71. Hierarchical clustering of nuclear gene expression, examined at p37 by the NCI cDNA microarray in AZT-exposed cells, showed down-regulation of 13 out of 16 lipid-metabolizing genes, and up-regulation of most oxidative phosphorylation (OXPHOS) genes. OXPHOS genes encoded by mtDNA, examined at p5, p36, and p75 using the Mitochondrial Gene Mini Array, revealed up-regulation of genes coding for polypeptides of NADH dehydrogenase, ATP synthase, and cytochrome c oxidase. Mitochondrial membrane potential, monitored by JC1 staining, was elevated at p10 and p32, and essentially completely absent at p71. The data show that during chronic exposure of HeLa cells to AZT, a compensatory response was induced at the earlier passages (p5-p37), and by p71 there was widespread mitochondrial morphological damage, severe mtDNA depletion, and a substantial loss of mitochondrial membrane potential.
Divi
Rao L
RL
Carcinogen-DNA Interactions Section, National Cancer Institute, NIH, Bethesda, Maryland 20892, USA. divir@exchange.nih.gov
Haverkos
Kathryn J
KJ
Humsi
Juliette A
JA
Shockley
Marie E
ME
Thamire
Chandrasekhar
C
Nagashima
Kunio
K
Olivero
Ofelia A
OA
Poirier
Miriam C
MC
D019380:Anti-HIV Agents / Q000494:pharmacology*; D053938:DNA Fragmentation; D004272:DNA, Mitochondrial / Q000032:analysis; D020869:Gene Expression Profiling; D006367:HeLa Cells; D006801:Humans; D050356:Lipid Metabolism / Q000187:drug effects; D046529:Microscopy, Electron, Transmission; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis; D010085:Oxidative Phosphorylation / Q000187:drug effects; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D015215:Zidovudine / Q000494:pharmacology*
D019380:Anti-HIV Agents; D004272:DNA, Mitochondrial; D018894:Reverse Transcriptase Inhibitors; D015215:Zidovudine