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

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.

Human L-ferritin deficiency is characterized by idiopathic generalized seizures and atypical restless leg syndrome.

The ubiquitously expressed iron storage protein ferritin plays a central role in maintaining cellular iron homeostasis. Cytosolic ferritins are composed of heavy (H) and light (L) subunits that co-assemble into a hollow spherical shell with an internal cavity where iron is stored. The ferroxidase activity of the ferritin H chain is critical to store iron in its Fe3+ oxidation state, while the L chain shows iron nucleation properties. We describe a unique case of a 23-yr-old female patient affected by a homozygous loss of function mutation in the L-ferritin gene, idiopathic generalized seizures, and atypical restless leg syndrome (RLS). We show that L chain ferritin is undetectable in primary fibroblasts from the patient, and thus ferritin consists only of H chains. Increased iron incorporation into the FtH homopolymer leads to reduced cellular iron availability, diminished levels of cytosolic catalase, SOD1 protein levels, enhanced ROS production and higher levels of oxidized proteins. Importantly, key phenotypic features observed in fibroblasts are also mirrored in reprogrammed neurons from the patient's fibroblasts. Our results demonstrate for the first time the pathophysiological consequences of L-ferritin deficiency in a human and help to define the concept for a new disease entity hallmarked by idiopathic generalized seizure and atypical RLS.