[Abstract unavailable

A study is presented on the interaction of ceramide with the respiratory chain of rat heart mitochondria, and a comparison is made between the effects elicited by short- and long-chain ceramides. N-Acetylsphingosine (C(2)-ceramide) and N-palmitoylsphingosine (C(16)-ceramide) inhibited to the same extent the pyruvate+malate-dependent oxygen consumption. Succinate-supported respiration was also inhibited by ceramides, but this activity was substantially restored upon the addition of cytochrome c, which, on the contrary, was ineffective toward the ceramide-inhibited NADH-linked substrate oxidation. Direct measurements showed that short- and long-chain ceramides caused a large release of cytochrome c from mitochondria. The ceramide-dependent inhibition of pyruvate+malate and succinate oxidation caused reactive oxygen species to be produced at the level of either complex I or complex III. The activity of the cytochrome c oxidase, measured as ascorbate/TMPD oxidase activity, was significantly stimulated and inhibited by C(2)- and C(16)-ceramide, respectively. Similar effects were observed on the activity of the individual respiratory complexes isolated from bovine heart. Short- and long-chain ceramides had definitely different effects on the mitochondrial membrane potential. C(2)-ceramide caused an almost complete collapse of the respiration-dependent membrane potential, whereas C(16)-ceramide had a negligible effect. Similar results were obtained when the potential was generated in liposome-reconstituted complex III respiring at the steady-state. Furthermore, C(2)-ceramide caused a drop of the membrane potential generated by ATP hydrolysis instead of respiration, whereas C(16)-ceramide did not. Finally, only short-chain ceramides inhibited markedly the reactive oxygen species generation associated with membrane potential-dependent reverse electron flow from succinate to complex I. The emerging indication is that the short-chain ceramide-dependent collapse of membrane potential is a consequence of their ability to perturb the membrane structure, leading to an unspecific increase of its permeability.

Mitochondrial dysfunction and oxidative stress occur in Parkinson's disease (PD), but the molecular mechanisms controlling these events are not completely understood. Peroxisome proliferator-activated receptor-gamma coactivator-1α (PGC-1α) is a transcriptional coactivator known as master regulator of mitochondrial functions and oxidative metabolism. Recent studies, including one from our group, have highlighted altered PGC-1α activity and transcriptional deregulation of its target genes in PD pathogenesis suggesting it as a new potential therapeutic target. Resveratrol, a natural polyphenolic compound proved to improve mitochondrial activity through the activation of several metabolic sensors resulting in PGC-1α activation. Here we have tested in vitro the effect of resveratrol treatment on primary fibroblast cultures from two patients with early-onset PD linked to different Park2 mutations. We show that resveratrol regulates energy homeostasis through activation of AMP-activated protein kinase (AMPK) and sirtuin 1 (SIRT1) and raise of mRNA expression of a number of PGC-1α's target genes resulting in enhanced mitochondrial oxidative function, likely related to a decrease of oxidative stress and to an increase of mitochondrial biogenesis. The functional impact of resveratrol treatment encompassed an increase of complex I and citrate synthase activities, basal oxygen consumption, and mitochondrial ATP production and a decrease in lactate content, thus supporting a switch from glycolytic to oxidative metabolism. Moreover, resveratrol treatment caused an enhanced macro-autophagic flux through activation of an LC3-independent pathway. Our results, obtained in early-onset PD fibroblasts, suggest that resveratrol may have potential clinical application in selected cases of PD-affected patients.

No abstract

Mutations in the parkin gene are expected to play an essential role in autosomal recessive Parkinson's disease. Recent studies have established an impact of parkin mutations on mitochondrial function and autophagy. In primary skin fibroblasts from two patients affected by an early onset Parkinson's disease, we identified a hitherto unreported compound heterozygous mutation del exon2-3/del exon3 in the parkin gene, leading to the complete loss of the full-length protein. In both patients, but not in their heterozygous parental control, we observed severe ultrastructural abnormalities, mainly in mitochondria. This was associated with impaired energy metabolism, deregulated reactive oxygen species (ROS) production, resulting in lipid oxidation, and peroxisomal alteration. In view of the involvement of parkin in the mitochondrial quality control system, we have investigated upstream events in the organelles' biogenesis. The expression of the peroxisome proliferator-activated receptor gamma-coactivator 1-alpha (PGC-1α), a strong stimulator of mitochondrial biogenesis, was remarkably upregulated in both patients. However, the function of PGC-1α was blocked, as revealed by the lack of its downstream target gene induction. In conclusion, our data confirm the role of parkin in mitochondrial homeostasis and suggest a potential involvement of the PGC-1α pathway in the pathogenesis of Parkinson's disease. This article is part of a Special Issue entitled: Translating nuclear receptors from health to disease.