title: Changes in mitochondrial RNA production in cells treated with nucleoside analogues.
abstract: BACKGROUND
  To investigate mitochondrial (mt) toxicity of antiretroviral drugs further, we developed a novel real-time PCR-based assay for the quantification of mtRNA. We analysed the effects of stavudine (d4T), didanosine (ddl) and zidovudine (AZT) on the production of mtRNAs in different human cell lines and compared the production with the amount of mtDNA present in the same cells.
  
  
  METHODS
  HUT78, CEM and U937 cells were exposed to different nucleoside reverse transcriptase inhibitors (NRTIs) for 7 days. Thereafter, nucleic acids were isolated and Taqman-based real-time PCR was used to quantify mtDNA and three different mtRNAs (ND1, CYTB and ND6 gene products).
  
  
  RESULTS
  Different amounts of mtRNAs exist in different cell lines. When mtRNA was measured in cells exposed to an NRTI, a marked decrease was observed in cells treated with d4T, but not with ddl or AZT. Changes in mtRNA production did not always correspond to modifications in mtDNA content: 1 microM d4T significantly changed mtRNA but not mtDNA content.
  
  
  CONCLUSIONS
  d4T, but not ddl or AZT, significantly alters mtRNA quantity and quality. The method we have developed can reveal changes that are not observed by measuring mtDNA content only, and can be used for ex vivo studies on drug toxicity.
authors.affiliation: Section of General Pathology, Department of Biomedical Sciences, University of Modena and Reggio Emilia, Modena, Italy.
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Galluzzi
authors.forename: Marcello
authors.initials: M
authors.lastname: Pinti
authors.forename: Leonarda
authors.initials: L
authors.lastname: Troiano
authors.forename: Nicole
authors.initials: N
authors.lastname: Prada
authors.forename: Milena
authors.initials: M
authors.lastname: Nasi
authors.forename: Roberta
authors.initials: R
authors.lastname: Ferraresi
authors.forename: Paolo
authors.initials: P
authors.lastname: Salomoni
authors.forename: Cristina
authors.initials: C
authors.lastname: Mussini
authors.forename: Andrea
authors.initials: A
authors.lastname: Cossarizza
mesh_terms: D000998:Antiviral Agents / Q000633:toxicity*; D002460:Cell Line; D004272:DNA, Mitochondrial / Q000032:analysis / Q000235:genetics; D016049:Didanosine / Q000633:toxicity; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods; D012313:RNA / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics; D000077278:RNA, Mitochondrial; D018894:Reverse Transcriptase Inhibitors / Q000633:toxicity; D018119:Stavudine / Q000633:toxicity; D020298:U937 Cells; D015215:Zidovudine / Q000633:toxicity
chemical_list: D000998:Antiviral Agents; D004272:DNA, Mitochondrial; D000077278:RNA, Mitochondrial; D018894:Reverse Transcriptase Inhibitors; D015215:Zidovudine; D012313:RNA; D018119:Stavudine; D016049:Didanosine