title: Mitochondrial gene expression is impaired by ethanol exposure in cultured chick cardiac myocytes.
abstract: OBJECTIVE
  A depression in cytochrome c oxidase (COX) activity occurs following chronic embryonic ethanol exposure in vivo. The aim of this study was to examine the effect of chronic ethanol exposure on COX activity in isolated cardiac cells maintained in vitro. Additionally, the mechanism by which ethanol produces an impairment in COX activity was evaluated by examining mitochondrial gene expression.
  
  
  METHODS
  Spontaneously beating cardiac myocyte cultures were established from 10-day embryonic chick hearts. Various concentrations of ethanol (0-250 mM) were introduced at the time of plating and cells were harvested over 7 days. COX activity was determined in myocyte homogenates. The levels of nuclear-encoded (COXIV) and mitochondrial-encoded (COXII) subunit proteins were measured by Western blotting. Relative levels of mitochondrial DNA and the mitochondrially-encoded COXIII mRNA were determined by Southern and Northern blotting.
  
  
  RESULTS
  A consistent decrease in COX activity in ethanol-exposed cardiac myocytes of approximately 30% was observed with an ethanol concentration of 25 mM. Increasing the ethanol concentration to 250 mM produced only a minor enhancement of this effect, while severely decreasing cellular viability. The content of the mitochondrially-encoded COXII subunit was reduced by ethanol exposure, while that of the nuclear-encoded COXIV subunit was unchanged. The content of the mitochondrially-encoded COXIII mRNA was unchanged by ethanol exposure. However, prolonged ethanol exposure produced an increase in mitochondrial DNA levels in cardiac myocytes.
  
  
  CONCLUSIONS
  Ethanol exposure of cardiac myocytes produces deficits in COX activity in the absence of systemic variables, indicating that ethanol has a direct effect on cardiac mitochondria. The ethanol-induced decrease in COX activity is correlated with a specific decrease in at least one mitochondrially-encoded gene product, COXII. No changes were observed in the level of the nuclear-encoded COXIV subunit, indicating that expression of this nuclear-encoded gene is not impaired by ethanol exposure.
authors.affiliation: University of Illinois at Chicago, Department of Physiology and Biophysics 60612-7342, USA. jkennedy@uic.edu
authors.forename: J M
authors.initials: JM
authors.lastname: Kennedy
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D015139:Blotting, Southern; D015153:Blotting, Western; D002467:Cell Nucleus / Q000201:enzymology; D002478:Cells, Cultured; D002642:Chick Embryo; D004272:DNA, Mitochondrial / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D003576:Electron Transport Complex IV / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D000431:Ethanol / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects; D006321:Heart / Q000196:embryology; D007150:Immunohistochemistry; D008929:Mitochondria, Heart / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D009206:Myocardium / Q000201:enzymology / Q000648:ultrastructure; D012333:RNA, Messenger / Q000187:drug effects*; D013997:Time Factors
chemical_list: D004272:DNA, Mitochondrial; D012333:RNA, Messenger; D000431:Ethanol; D003576:Electron Transport Complex IV