Superoxide production is inversely related to complex I activity in inherited complex I deficiency.

Contains fulltext : 53008.pdf (publisher's version ) (Closed access)Deficiency of NADH:ubiquinone oxidoreductase or complex I (CI) is the most common cause of disorders of the oxidative phosphorylation system in humans. Using life cell imaging and blue-native electrophoresis we quantitatively compared superoxide production and CI amount and activity in cultured skin fibroblasts of 7 healthy control subjects and 21 children with inherited isolated CI deficiency. Thirteen children had a disease causing mutation in one of the nuclear-encoded CI subunits, whereas in the remainder the genetic cause of the disease is not yet established. Superoxide production was significantly increased in all but two of the patient cell lines. An inverse relationship with the amount and residual activity of CI was observed. In agreement with this finding, rotenone, a potent inhibitor of CI activity, dose-dependently increased superoxide production in healthy control cells. Also in this case an inverse relationship with the residual activity of CI was observed. In sharp contrast, however, rotenone did not decrease the amount of CI. The data presented show that superoxide production is increased in inherited CI deficiency and that this increase is primarily a consequence of the reduction in cellular CI activity and not of a further leakage of electrons from mutationally malformed complexes

Calcium and ATP handling in human NADH:Ubiquinone oxidoreductase deficiency.

Contains fulltext : 80411.pdf (publisher's version ) (Closed access)Proper cell functioning requires precise coordination between mitochondrial ATP production and local energy demand. Ionic calcium (Ca(2+)) plays a central role in this coupling because it activates mitochondrial oxidative phosphorylation (OXPHOS) during hormonal and electrical cell stimulation. To determine how mitochondrial dysfunction affects cytosolic and mitochondrial Ca(2+)/ATP handling, we performed life-cell quantification of these parameters in fibroblast cell lines derived from healthy subjects and patients with isolated deficiency of the first OXPHOS complex (CI). In resting patient cells, CI deficiency was associated with a normal mitochondrial ([ATP](m)) and cytosolic ([ATP](c)) ATP concentration, a normal cytosolic Ca(2+) concentration ([Ca(2+)](c)), but a reduced Ca(2+) content of the endoplasmic reticulum (ER). Furthermore, cellular NAD(P)H levels were increased, mitochondrial membrane potential was slightly depolarized, reactive oxygen species (ROS) levels were elevated and mitochondrial shape was altered. Upon stimulation with bradykinin (Bk), the peak increases in [Ca(2+)](c), mitochondrial Ca(2+) concentration ([Ca(2+)](m)), [ATP](c) and [ATP](m) were reduced in patient cells. In agreement with these results, ATP-dependent Ca(2+) removal from the cytosol was slower. Here, we review the interconnection between cytosolic, endoplasmic reticular and mitochondrial Ca(2+) and ATP handling, and summarize our findings in patient fibroblasts in an integrative model