Liver mitochondrial membrane crosslinking and destruction in a rat model of Wilson disease
Hans Zischka, Josef Lichtmannegger, Sabine Schmitt, Nora Jägemann, Sabine Schulz, Daniela Wartini, Luise Jennen, Christian Rust, Nathanael Larochette, Lorenzo Galluzzi, Veronique Chajes, Nathan Bandow, Valérie S. Gilles, Alan A. DiSpirito, Irene Esposito, Martin Goettlicher, Karl H. Summer, Guido Kroemer
Hans Zischka, Josef Lichtmannegger, Sabine Schmitt, Nora Jägemann, Sabine Schulz, Daniela Wartini, Luise Jennen, Christian Rust, Nathanael Larochette, Lorenzo Galluzzi, Veronique Chajes, Nathan Bandow, Valérie S. Gilles, Alan A. DiSpirito, Irene Esposito, Martin Goettlicher, Karl H. Summer, Guido Kroemer
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Research Article Hepatology

Liver mitochondrial membrane crosslinking and destruction in a rat model of Wilson disease

Abstract

Wilson disease (WD) is a rare hereditary condition that is caused by a genetic defect in the copper-transporting ATPase ATP7B that results in hepatic copper accumulation and lethal liver failure. The present study focuses on the structural mitochondrial alterations that precede clinical symptoms in the livers of rats lacking Atp7b, an animal model for WD. Liver mitochondria from these Atp7b–/– rats contained enlarged cristae and widened intermembrane spaces, which coincided with a massive mitochondrial accumulation of copper. These changes, however, preceded detectable deficits in oxidative phosphorylation and biochemical signs of oxidative damage, suggesting that the ultrastructural modifications were not the result of oxidative stress imposed by copper-dependent Fenton chemistry. In a cell-free system containing a reducing dithiol agent, isolated mitochondria exposed to copper underwent modifications that were closely related to those observed in vivo. In this cell-free system, copper induced thiol modifications of three abundant mitochondrial membrane proteins, and this correlated with reversible intramitochondrial membrane crosslinking, which was also observed in liver mitochondria from Atp7b–/– rats. In vivo, copper-chelating agents reversed mitochondrial accumulation of copper, as well as signs of intra-mitochondrial membrane crosslinking, thereby preserving the functional and structural integrity of mitochondria. Together, these findings suggest that the mitochondrion constitutes a pivotal target of copper in WD.

Authors

Hans Zischka, Josef Lichtmannegger, Sabine Schmitt, Nora Jägemann, Sabine Schulz, Daniela Wartini, Luise Jennen, Christian Rust, Nathanael Larochette, Lorenzo Galluzzi, Veronique Chajes, Nathan Bandow, Valérie S. Gilles, Alan A. DiSpirito, Irene Esposito, Martin Goettlicher, Karl H. Summer, Guido Kroemer

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Figure 5

Simulation of pathological mitochondrial alterations in a cell-free system.

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Simulation of pathological mitochondrial alterations in a cell-free syst...
(A) Copper and calcium induce mitochondrial swelling as assessed by optical density measurements of mitochondrial suspensions at 540 nm. In contrast to calcium, copper-triggered swelling could be inhibited by reduced GSH and dithiols such as DTT and DTE. Each data curve represents the average of 4 individual measurements for copper and duplicates for calcium. (B) In the presence of DTT, repetitive copper dosing induced a staircase-like increase in light refraction, indicative of mitochondrial contraction. This increase could be reversed by repetitive equimolar doses of the copper chelator BCDS. Measurements of mitochondrial suspensions were performed at 540 nm in a fluorescence spectrometer. For clarity, fluorescence peaks due to additions made during the measurement were removed from the displayed graph. (C–G) Electron micrographs of control mitochondria treated with copper in the presence of DTT. Note that copper was added repeatedly to the same mitochondrial suspension and samples withdrawn after the additions: (C) untreated starting control, (D) 100 μM copper/1 mM DTT, (E) 200 μM copper/1 mM DTT, (F) 300 μM copper/1 mM DTT, (G) 400 μM copper/1 mM DTT. Mitochondria with progressively enlarged cristae and intermembrane spaces were found. At high copper doses matrix remnants gathered at the mitochondrial membranes (arrows). Scale bars: 0.5 μm.