Mitochondrial iron and energetic dysfunction distinguish fibroblasts and induced neurons from pantothenate kinase-associated neurodegeneration patients

Abstract

Pantothenate kinase-associated neurodegeneration is an early onset autosomal recessive movement disorder caused by mutation of the pantothenate kinase-2 gene, which encodes a mitochondrial enzyme involved in coenzyme A synthesis. The disorder is characterised by high iron levels in the brain, although the pathological mechanism leading to this accumulation is unknown. To address this question, we tested primary skin fibroblasts from three patients and three healthy subjects, as well as neurons induced by direct fibroblast reprogramming, for oxidative status, mitochondrial functionality and iron parameters. The patients' fibroblasts showed altered oxidative status, reduced antioxidant defence, and impaired cytosolic and mitochondrial aconitase activities compared to control cells. Mitochondrial iron homeostasis and functionality analysis of patient fibroblasts indicated increased labile iron pool content and reactive oxygen species development, altered mitochondrial shape, decreased membrane potential and reduced ATP levels. Furthermore, analysis of induced neurons, performed at a single cell level, confirmed some of the results obtained in fibroblasts, indicating an altered oxidative status and signs of mitochondrial dysfunction, possibly due to iron mishandling. Thus, for the first time, altered biological processes have been identified in vitro in live diseased neurons. Moreover, the obtained induced neurons can be considered a suitable human neuronal model for the identification of candidate therapeutic compounds for this disease.

Iron overload induced death of osteoblasts in vitro: involvement of the mitochondrial apoptotic pathway

“…In iron-overload conditions, excess labile iron enters the mitochondria via the calcium uniporter, and then interacts with reactive oxygen intermediates leaked from mitochondrial respiratory chain through Fenton reactions, catalyzing powerful ROS to damage mitochondria (Chen et al, 2014; Sripetchwandee et al, 2014; Pelizzoni et al, 2011; Uchiyama et al, 2008). ROS catalyzed by labile iron elicits a range of detrimental effects in mitochondria, as following (1) impairment of mitochondrial respiratory enzyme activity; (2) decrease in ATP production; (3) loss of MMP; and (4) damage to mitochondrial DNA (mtDNA) (Ward et al, 2014; Mallikarjun et al, 2014; Al-Qenaei et al, 2014; Santambrogio et al, 2015; Rouault, 2016; Rines & Ardehali, 2013). In this experiment, the inhibition of mitochondrial dehydrogenase activity was observed in osteoblasts exposed to FAC.…”

Augmentation of glycolytic metabolism by meclizine is indispensable for protection of dorsal root ganglion neurons from hypoxia-induced mitochondrial compromise

“…Assessment of reduced intracellular glutathione (GSH) levels was with Thioltracker Violet (Life Technologies) cell-permeant thiol-reactive fluorescent probe as described [40, 41]. DRG neurons were seeded in black wall 96-well plates (10 4 /well).…”

Down-regulation of coasy, the gene associated with NBIA-VI, reduces Bmp signaling, perturbs dorso-ventral patterning and alters neuronal development in zebrafish

“…It may well be that distinctive features of PANK2, and particularly its localization in the mitochondrial intermembrane space, play a relevant role in determining the starting pathogenic mechanism and the specificity of PKAN neuropathology. Interestingly, morphological and functional perturbations of mitochondria have been found in fibroblasts from patients as well as in Drosophila and mouse knock-out models101516. These features are often associated with signs of lipid dyshomeostasis, implying possible perturbation in mitochondrial membrane remodeling, and disruption of iron balance1517.…”

“…Interestingly, morphological and functional perturbations of mitochondria have been found in fibroblasts from patients as well as in Drosophila and mouse knock-out models101516. These features are often associated with signs of lipid dyshomeostasis, implying possible perturbation in mitochondrial membrane remodeling, and disruption of iron balance1517. To gain new information about the functional connection between enzyme defects and pathology, we recently performed a thorough analysis of pank2 role during zebrafish embryonic development13.…”