title: Polarographic evaluation of mitochondrial enzymes activity in isolated mitochondria and in permeabilized human muscle cells with inherited mitochondrial defects.
abstract: Inherited disturbances of the mitochondrial energy generating system represent a heterogeneous group of disorders associated with a broad spectrum of metabolic abnormalities and clinical symptoms. We used the polarographic and spectrophotometric method for detection of mitochondrial disorders, because these two techniques provide a different insight into mitochondrial function. In six patients suspected of mitochondrial disease we found defects of complex I (two patients), complex III (one patient), complex IV (two patients) and a combination of defect of complex III and IV (one patient). Citrate synthase activity, used as the reference enzyme, was not changed. A comparison of the two methods showed several differences in evaluation of mitochondrial enzymes activity due to the fact that both methods used different conditions for enzyme activity measurements. In contrast to oxygen consumption measurements, where the function of the whole-integrated respiratory chain is characterized, spectrophotometric measurements characterize activities of isolated complexes in disintegrated membranes. However, it may be concluded from our experiments that both methods provide useful and complementary data about mitochondrial energetic functions. Whereas spectrophotometric data are suitable for evaluation of maximal enzyme activities of mitochondrial enzyme complexes, polarographic data provide better information about enzyme activities in cells with mitochondrial defects under in situ conditions.
authors.affiliation: Center for Integrated Genomics, Department of Pediatrics, Charles University, Prague, Czech Republic.
authors.forename: L
authors.initials: L
authors.lastname: Wenchich
authors.forename: Z
authors.initials: Z
authors.lastname: Drahota
authors.forename: T
authors.initials: T
authors.lastname: Honzík
authors.forename: H
authors.initials: H
authors.lastname: Hansíková
authors.forename: M
authors.initials: M
authors.lastname: Tesarová
authors.forename: J
authors.initials: J
authors.lastname: Zeman
authors.forename: J
authors.initials: J
authors.lastname: Houstek
mesh_terms: D000293:Adolescent; D000328:Adult; D002463:Cell Membrane Permeability / Q000187:drug effects; D002648:Child; D002675:Child, Preschool; D045222:Electron Transport Chain Complex Proteins / Q000378:metabolism; D042967:Electron Transport Complex I / Q000378:metabolism; D042963:Electron Transport Complex II / Q000378:metabolism; D014450:Electron Transport Complex III / Q000378:metabolism; D003576:Electron Transport Complex IV / Q000378:metabolism; D030342:Genetic Diseases, Inborn / Q000378:metabolism* / Q000503:physiopathology; D006801:Humans; D007223:Infant; D007625:Kearns-Sayre Syndrome / Q000378:metabolism / Q000503:physiopathology; D017241:MELAS Syndrome / Q000235:genetics / Q000378:metabolism / Q000503:physiopathology; D008931:Mitochondria, Muscle / Q000201:enzymology* / Q000378:metabolism; D028361:Mitochondrial Diseases / Q000378:metabolism* / Q000503:physiopathology; D018482:Muscle, Skeletal / Q000201:enzymology* / Q000378:metabolism / Q000503:physiopathology; D010101:Oxygen Consumption / Q000187:drug effects / Q000502:physiology; D011048:Polarography / Q000379:methods*; D013053:Spectrophotometry
chemical_list: D045222:Electron Transport Chain Complex Proteins; D042963:Electron Transport Complex II; D003576:Electron Transport Complex IV; D042967:Electron Transport Complex I; D014450:Electron Transport Complex III