Recruitment of mitochondrial cyclophilin to the mitochondrial inner membrane under conditions of oxidative stress that enhance the opening of a calcium-sensitive non-specific channel.

Binding of mitochondrial matrix cyclophilin (CyP) to the rat liver mitochondrial membranes was detected by SDS/PAGE and Western blotting with suitable antipeptide antibodies. Binding was not affected by prior exposure of mitochondria to Ca2+, adenine nucleotides or inhibitors of the adenine nucleotide translocase, but was greatly increased by t-butyl hydroperoxide (tBH), phenylarsine oxide or diamide. These all sensitized the opening of the non-specific mitochondrial pore to [Ca2+], and the effect of tBH was shown to be maintained after washing away the tBH, consistent with it being caused by the enhanced CyP binding. The bound CyP did not demonstrate peptidyl-prolyl cis-trans isomerase activity. CyP-binding was prevented by 5 microM cyclosporin A, but not reversed by cyclosporin treatment of the membranes. The effect of tBH on binding was concentration-dependent and maximal within 30 s

Spinal cord mitochondria display lower calcium retention capacity compared with brain mitochondria without inherent differences in sensitivity to cyclophilin D inhibition

The mitochondrial permeability transition (mPT) is a potential pathogenic mechanism in neurodegeneration. Varying sensitivity to calcium-induced mPT has been demonstrated for regions within the CNS possibly correlating with vulnerability following insults. The spinal cord is selectively vulnerable in e.g. amyotrophic lateral sclerosis and increased mPT sensitivity of mitochondria derived from the spinal cord has previously been demonstrated. In this study, we introduce whole-body hypothermia prior to removal of CNS tissue to minimize the effects of differential tissue extraction prior to isolation of spinal cord and cortical brain mitochondria. Spinal cord mitochondria were able to retain considerably less calcium when administered as continuous infusion, which was not related to a general increased sensitivity of the mPT to calcium, its desensitization to calcium by the cyclophilin D inhibitor cyclosporin-A, or to differences in respiratory parameters. Spinal cord mitochondria maintained a higher concentration of extramitochondrial calcium during infusion than brain mitochondria possibly related to an increased set-point concentration for calcium uptake. A hampered transport and retention capacity of calcium may translate into an increased susceptibility of the spinal cord to neurodegenerative processes involving calcium-mediated damage

Doxorubicin-induced thiol-dependent alteration of cardiac mitochondrial permeability transition and respiration

Abstract Doxorubicin (DOX) is a highly effective treatment for several forms of cancer. However, clinical experience shows that DOX induces a cumulative and dose-dependent cardiomyopathy that has been ascribed to redox-cycling of the drug on the mitochondrial respiratory chain generating free radicals and oxidative stress in the process. Mitochondrial dysfunction including induction of the mitochondrial permeability transition (MPT) and inhibition of mitochondrial respiration have been implicated as major determinants in the pathogenesis of DOX cardiotoxicity. The present work was aimed at investigating whether the inhibition of mitochondrial respiration occurs secondarily to MPT induction in heart mitochondria isolated from DOX-treated rats and whether one or both consequences of DOX treatment are related with oxidation of protein thiol residues. DOX-induced oxidative stress was associated with the accumulation of products of lipid peroxidation and the depletion of a-tocopherol in cardiac mitochondrial membranes. No changes in mitochondrial coenzyme Q9 and Q10 concentrations were detected in hearts of DOX-treated rats. Cardiac mitochondria from DOX-treated rats were more susceptible to diamide-dependent induction of the MPT. Although DOX treatment did not affect state 4 respiration, state 3 respiration was decreased in heart mitochondria isolated from DOX-treated rats, which was reversed in part by adding either cyclosporin A or dithiothreitol, but not Trolox. The results suggest that in DOX-treated rats, (i) induction of the MPT is at least in part responsible for decreased mitochondrial respiration, (ii) heart mitochondria are more susceptible to diamide induced-MPT, (iii) thiol-dependent alteration of mitochondrial respiration is partially reversible ex vivo with dithiothreitol. Collectively, these data are consistent with the thesis that thiol-dependent alteration of MPT and respiration is an important factor in DOX-induced mitochondrial dysfunction