Fatty Acid Hydroxylase-associated Neurodegeneration Research Articles

Generation of the human iPSC lines AKOSi011-A carrying the mutation p.Pro65Ser/p.Asp35T and AKOSi012-A, carrying the mutation p.Tyr231His, derived from FAHN patient fibroblasts

Fatty acid hydroxylase-associated neurodegeneration (FAHN) is a hereditary neurodegenerative disease caused by mutations in the FA2H gene. Patients show a wide range of neurological symptoms and an abnormal myelination. Here we describe the generation of the human induced pluripotent stem cell (hiPSC) lines AKOSi011-A and AKOSi012-A, derived from FAHN-patient fibroblasts, carrying the compound heterozygous mutation p.Pro65Ser/p.Asp35Tyr and the homozygous mutation p.Tyr231His, respectively. The hiPSC lines were generated using a non-integrating Sendai virus. The obtained hiPSCs show an unobtrusive karyotype, carry the mutations of the original fibroblasts, express pluripotency markers and can differentiate into cells of the three germ layers.

The first reports of FA2H-associated neurodegeneration from two unrelated Iranian families.

NBIA (neurodegeneration with brain iron accumulation) is a diverse collection of neurodegenerative illnesses defined by iron accumulation in the basal ganglia. The fatty acid hydroxylase-associated neurodegeneration, or FAHN, is one of the uncommon subtypes of NBIAs, associated with inherited autosomal recessive mutations in gene coding the membrane-bound fatty acid 2 hydroxylase (FA2H) enzyme. Here, we report two cases with FAHN from two unrelated families from Iran confirmed by whole exome sequencing. FAHN is an uncommon variant of NBIA that may manifest as spastic paraparesis without signs of iron buildup on brain imaging. As a result, it should be taken into account while making a differential diagnosis of the hereditary spastic paraplegia (HSP) syndrome, especially in individuals who lack iron deposits.

Fatty Acid 2-Hydroxylase and 2-Hydroxylated Sphingolipids: Metabolism and Function in Health and Diseases.

Sphingolipids containing acyl residues that are hydroxylated at C-2 are found in most, if not all, eukaryotes and certain bacteria. 2-hydroxylated sphingolipids are present in many organs and cell types, though they are especially abundant in myelin and skin. The enzyme fatty acid 2-hydroxylase (FA2H) is involved in the synthesis of many but not all 2-hydroxylated sphingolipids. Deficiency in FA2H causes a neurodegenerative disease known as hereditary spastic paraplegia 35 (HSP35/SPG35) or fatty acid hydroxylase-associated neurodegeneration (FAHN). FA2H likely also plays a role in other diseases. A low expression level of FA2H correlates with a poor prognosis in many cancers. This review presents an updated overview of the metabolism and function of 2-hydroxylated sphingolipids and the FA2H enzyme under physiological conditions and in diseases.

A new model for fatty acid hydroxylase-associated neurodegeneration reveals mitochondrial and autophagy abnormalities.

Fatty acid hydroxylase-associated neurodegeneration (FAHN) is a rare disease that exhibits brain modifications and motor dysfunctions in early childhood. The condition is caused by a homozygous or compound heterozygous mutation in fatty acid 2 hydroxylase (FA2H), whose encoded protein synthesizes 2-hydroxysphingolipids and 2-hydroxyglycosphingolipids and is therefore involved in sphingolipid metabolism. A few FAHN model organisms have already been established and give the first insight into symptomatic effects. However, they fail to establish the underlying cellular mechanism of FAHN so far. Drosophila is an excellent model for many neurodegenerative disorders; hence, here, we have characterized and validated the first FAHN Drosophila model. The investigation of loss of dfa2h lines revealed behavioral abnormalities, including motor impairment and flying disability, in addition to a shortened lifespan. Furthermore, alterations in mitochondrial dynamics, and autophagy were identified. Analyses of patient-derived fibroblasts, and rescue experiments with human FA2H, indicated that these defects are evolutionarily conserved. We thus present a FAHN Drosophila model organism that provides new insights into the cellular mechanism of FAHN.

Generation of the human iPSC line AKOSi010-A from fibroblasts of a female FAHN patient, carrying the compound heterozygous mutation p.Gly45Arg/p.His319Arg

Fatty acid hydroxylase-associated neurodegeneration (FAHN) is a rare childhood onset neurodegenerative disease caused by mutations in the FA2H gene. Patients display abnormal myelination, cerebellar atrophy and some have iron deposition in the central nervous system. Here we describe the generation of AKOSi010-A, a human induced pluripotent stem cell (hiPSC) line derived from fibroblasts of a female patient carrying the compound heterozygous p.Gly45Arg/p.His319Arg, using non-integrating Sendai virus. The generated iPSCs express pluripotency markers, can differentiate into cell types of the three germ layers and show a normal karyotype. This cell line displays a unique source to study the pathophysiology of FAHN.

Consensus clinical management guideline for beta-propeller protein-associated neurodegeneration.

This review provides recommendations for the evaluation and management of individuals with beta-propeller protein-associated neurodegeneration (BPAN). BPAN is one of several neurodegenerative disorders with brain iron accumulation along with pantothenate kinase-associated neurodegeneration, PLA2G6-associated neurodegeneration, mitochondrial membrane protein-associated neurodegeneration, fatty acid hydroxylase-associated neurodegeneration, and COASY protein-associated neurodegeneration. BPAN typically presents with global developmental delay and epilepsy in childhood, which is followed by the onset of dystonia and parkinsonism in mid-adolescence or adulthood. BPAN is an X-linked dominant disorder caused by pathogenic variants in WDR45, resulting in a broad clinical phenotype and imaging spectrum. This review, informed by an evaluation of the literature and expert opinion, discusses the clinical phenotype and progression of the disease, imaging findings, epilepsy features, and genetics, and proposes an approach to the initial evaluation and management of disease manifestations across the life span in individuals with BPAN. What this paper adds The complex epilepsy profile of beta-propeller protein-associated neurodegeneration (BPAN) often resolves in adolescence. The treatment for an individual with BPAN is supportive, with attention to sleep disorders, complex epilepsy, and behavioral problems. Individuals with BPAN have shifting needs throughout their life span requiring multidisciplinary care.

Decreased turnover of the CNS myelin protein Opalin in a mouse model of hereditary spastic paraplegia 35.

Spastic paraplegia 35 (SPG35) (OMIM: 612319) or fatty acid hydroxylase-associated neurodegeneration (FAHN) is caused by deficiency of fatty acid 2-hydroxylase (FA2H). This enzyme synthesizes sphingolipids containing 2-hydroxylated fatty acids, which are particularly abundant in myelin. Fa2h-deficient (Fa2h-/-) mice develop symptoms reminiscent of the human disease and therefore serve as animal model of SPG35. In order to understand further the pathogenesis of SPG35, we compared the proteome of purified CNS myelin isolated from wild type and Fa2h-/- mice at different time points of disease progression using tandem mass tag labeling. Data analysis with a focus on myelin membrane proteins revealed a significant increase of the oligodendrocytic myelin paranodal and inner loop protein (Opalin) in Fa2h-/- mice, whereas the concentration of other major myelin proteins was not significantly changed. Western blot analysis revealed an almost 6-fold increase of Opalin in myelin of Fa2h-/- mice aged 21-23months. A concurrent unaltered Opalin gene expression suggested a decreased turnover of the Opalin protein in Fa2h-/- mice. Supporting this hypothesis, Opalin protein half-life was reduced significantly when expressed in CHO cells synthesizing 2-hydroxylated sulfatide, compared to cells synthesizing only non-hydroxylated sulfatide. Degradation of Opalin was inhibited by inhibitors of lysosomal degradation but unaffected by proteasome inhibitors. Taken together, these results reveal a new function of 2-hydroxylated sphingolipids namely affecting the turnover of a myelin membrane protein. This may play a role in the pathogenesis of SPG35.

P.86Depends on how you look: hereditary spastic paraplegia, leukodystrophy or brain iron accumulation

Fatty acid hydroxylase-associated neurodegeneration (FAHN), a progressive disorder once classified as hereditary spastic paraplegia (SPG35; 612319), has recently linked to neurodegeneration with brain iron accumulation and leukodystrophies. Two boys (6.5 and 7 years) from unrelated consanguineous families were detected to have homozygous mutations in FA2Hgene through whole exome sequencing. Case 1, born at term, showed normal early development, but presented seizures and acute vision loss at 3.5 years. Vision partially recovered in 3 days. Initially asymmetric, then bilateral progressive weakness in lower extremities, foot deformity, gait disturbance, then speech and perception impairment manifested. On examination he had strabismus, bilateral optic atrophy, spastic-ataxic gait, foot inversion, dysarthria, hyperactive deep tendon reflexes and Babinski sign. Case 2, born at 33th gestational week, had been floppy and delayed in motor and language domains since birth. He walked at 2 years, but gait deteriorated after 2.5 years old with frequent falls, ultimately loss of ambulation and communication. When 7 years old, he had bilateral optic atrophy, central hypotonia, increased tone and scissoring of lower extremities, foot inversion, genu recurvatum, and upper motor neuron findings. Both cases' MRI, around 3 years of age, revealed white matter lesions/thin corpus callosum. Phenotype was most consistent with neuroaxonal dystrophies, leukodystrophies or mitochondrial disorders. WES showed homozygous missense (c.443C>T (p.Pro148Leu)) and deletion (c.685_687delATC (p.Ile229del) mutations in FA2H gene, respectively. FA2H encodes fatty acid 2-hydroxylase, enzyme participating in the synthesis fatty acids of galactolipids. These being an important part of the myelin sheath explains the broad clinical spectrum: spasticity, ataxia, dystonia, optic atrophy, progressive intellectual impairment and seizures, illustrated by our two cases.

FAHN/SPG35: a narrow phenotypic spectrum across disease classifications.

The endoplasmic reticulum enzyme fatty acid 2-hydroxylase (FA2H) plays a major role in the formation of 2-hydroxy glycosphingolipids, main components of myelin. FA2H deficiency in mice leads to severe central demyelination and axon loss. In humans it has been associated with phenotypes from the neurodegeneration with brain iron accumulation (fatty acid hydroxylase-associated neurodegeneration, FAHN), hereditary spastic paraplegia (HSP type SPG35) and leukodystrophy (leukodystrophy with spasticity and dystonia) spectrum. We performed an in-depth clinical and retrospective neurophysiological and imaging study in a cohort of 19 cases with biallelic FA2H mutations. FAHN/SPG35 manifests with early childhood onset predominantly lower limb spastic tetraparesis and truncal instability, dysarthria, dysphagia, cerebellar ataxia, and cognitive deficits, often accompanied by exotropia and movement disorders. The disease is rapidly progressive with loss of ambulation after a median of 7 years after disease onset and demonstrates little interindividual variability. The hair of FAHN/SPG35 patients shows a bristle-like appearance; scanning electron microscopy of patient hair shafts reveals deformities (longitudinal grooves) as well as plaque-like adhesions to the hair, likely caused by an abnormal sebum composition also described in a mouse model of FA2H deficiency. Characteristic imaging features of FAHN/SPG35 can be summarized by the 'WHAT' acronym: white matter changes, hypointensity of the globus pallidus, ponto-cerebellar atrophy, and thin corpus callosum. At least three of four imaging features are present in 85% of FA2H mutation carriers. Here, we report the first systematic, large cohort study in FAHN/SPG35 and determine the phenotypic spectrum, define the disease course and identify clinical and imaging biomarkers.

Identification of progesterone receptor membrane component-1 as an interaction partner and possible regulator of fatty acid 2-hydroxylase.

The fatty acid 2-hydroxylase (FA2H) is essential for synthesis of 2-hydroxylated fatty acids in myelinating and other cells, and deficiency of this enzyme causes a complicated form of hereditary spastic paraplegia also known as fatty acid hydroxylase-associated neurodegeneration. Despite its important role in sphingolipid metabolism, regulation of FA2H and its interaction with other proteins involved in the same or other metabolic pathways is poorly understood. To identify potential interaction partners of the enzyme, quantitative mass spectrometry using stable isotope labeling of cells was combined with formaldehyde cross-linking and proximity biotinylation, respectively. Besides other enzymes involved in sphingolipid synthesis and intermembrane transfer of ceramide, and putative redox partners of FA2H, progesterone receptor membrane component-1 (PGRMC1) and PGRMC2 were identified as putative interaction partners. These two related heme-binding proteins are known to regulate several cytochrome P450 enzymes. Bimolecular fluorescence complementation experiments confirmed the interaction of FA2H with PGRMC1. Moreover, the PGRMC1 inhibitor AG-205 significantly reduced synthesis of hydroxylated ceramide and glucosylceramide in FA2H-expressing cells. This suggests that PGRMC1 may regulate FA2H activity, possibly through its heme chaperone activity.

Chapter 19 - Neurodegeneration with brain iron accumulation

Neurodegeneration with brain iron accumulation (NBIA) comprises a clinically and genetically heterogeneous group of disorders affecting children and adults. These rare disorders are often first suspected when increased basal ganglia iron is observed on brain magnetic resonance imaging. For the majority of NBIA disorders the genetic basis has been delineated, and clinical testing is available. The four most common NBIA disorders include pantothenate kinase-associated neurodegeneration (PKAN) due to mutations in PANK2, phospholipase A2-associated neurodegeneration caused by mutation in PLA2G6, mitochondrial membrane protein-associated neurodegeneration from mutations in C19orf12, and beta-propeller protein-associated neurodegeneration due to mutations in WDR45. The ultrarare NBIA disorders are caused by mutations in CoASY, ATP13A2, and FA2H (causing CoA synthase protein-associated neurodegeneration, Kufor-Rakeb disease, and fatty acid hydroxylase-associated neurodegeneration, respectively). Together, these genes account for disease in approximately 85% of patients diagnosed with an NBIA disorder. New NBIA genes are being recognized with increasing frequency as a result of whole-exome sequencing, which is also facilitating early ascertainment of patients whose phenotype is often nonspecific.

Clinical phenotype and genetic mutation of fatty acid hydroxylase - associated neurodegeneration: analysis of four cases

Objective To report 4 cases of fatty acid hydroxylase - associated neurodegeneration (FAHN) and to summarize the clinical and genetic characteristics of FAHN by literatures review. Methods Four cases of FAHN patients' clinical and family data were collected in detail. The gDNA of patients and their parents were extracted from peripheral blood. FA2H gene was conducted and followed by Sanger sequencing. Results Among the 4 cases, 3 cases (Case 2, Case 3, Case 4) presented typical manifestations of FAHN while the other (Case 1) was atypical. Genetic sequencing showed FA2H gene mutation in all affected patients. Compound heterozygous mutation c.461G > A (p.Arg154His) and c.794T > G (p.Phe265Cys) were seen in Case 1. In Case 2, only one documented heterozygous mutation c.703C > T (p.Arg235Cys) was found, and dificit mutation was not found in single nucleotide polymorphism (SNP) chip test of the patient and her mother. Compound heterozygous mutation c.688G > A (p.Glu230Lys) and insertion mutation c.172_173insGGGCCAGGAC (p.Ile58ArgfsX47) were presented in Case 3. In Case 4, compound heterozygous mutation c.688G > A (p.Glu230Lys), c.968C > A (p.Pro323Gln) and c.976G > A (p. Gly326Asp) were seen, while his father was the carrier of c.688G > A (p.Glu230Lys) mutation and his mother was the carrier of c.968C > A (p.Pro323Gln) and c.976G > A (p.Gly326Asp) mutation. According to the standard of American College of Medical Genetics and Genomics (ACMG), c.461G > A (p.Arg154His) and c.794T > G (p.Phe265Cys) in Case 1, and c.703C > T (p.Arg235Cys) in Case 2 were considered as likely while FA2H gene compound heterozygous mutation c.688G > A (p.Glu230Lys), insertion mutation c.172_173insGGGCCAGGAC (p.Ile58ArgfsX47) in Case 3 was as pathogenic, and in Case 4, the FA2H gene mutation c.688G > A (p.Glu230Lys) and c.968C > A (p.Pro323Gln) were pathogenic and c.976G > A (p.Gly326Asp) was likely pathogenic. Conclusions FAHN has highly clinical and genetic heterogenieity in which spastic paraplegia is the main clinical presentation. In typing diagnosis for patietns with autosomal recessive herditary spastic paraplegia (HSP), especially combined with dyslalia, dysnoesia, and clinical features of white matter lesion and cerebellar atrophy, FA2H gene mutation-induced FAHN should be considered. DOI: 10.3969/j.issn.1672-6731.2017.07.010

A diagnostic approach for neurodegeneration with brain iron accumulation: clinical features, genetics and brain imaging.

Neurodegeneration with brain iron accumulation (NBIA) represents a heterogeneous and complex group of inherited neurodegenerative diseases, characterized by excessive iron accumulation, particularly in the basal ganglia. Common clinical features of NBIA include movement disorders, particularly parkinsonism and dystonia, cognitive dysfunction, pyramidal signs, and retinal abnormalities. The forms of NBIA described to date include pantothenase kinase-associated neurodegeneration (PKAN), phospholipase A2 associated neurodegeneration (PLAN), neuroferritinopathy, aceruloplasminemia, beta-propeller protein-associated neurodegeneration (BPAN), Kufor-Rakeb syndrome, mitochondrial membrane protein-associated neurodegeneration (MPAN), fatty acid hydroxylase-associated neurodegeneration (FAHN), coenzyme A synthase protein-associated neurodegeneration (CoPAN) and Woodhouse-Sakati syndrome. This review is a diagnostic approach for NBIA cases, from clinical features and brain imaging findings to the genetic etiology.

Brain iron takes off: a new propeller protein links neurodegeneration with autophagy

‘Neurodegeneration with brain iron accumulation’ (NBIA) is a clinically and genetically heterogeneous group of disorders presenting with progressive extrapyramidal dysfunction, and as a common feature, with brain iron deposition in the basal ganglia, particularly in the globus pallidus and substantia nigra (Gregory and Hayflick, 2013 a ). Over recent years an increasing number of mutations in novel disease genes have been identified in NBIA, facilitated by new genetic technologies. Mutations in nine different genes have been shown to cause NBIA to date, with a spectrum of overlapping clinical phenotypes (Fig. 1, Gregory and Hayflick, 2013 a ). The identification of novel disease genes has improved our understanding of major disease mechanisms leading to iron deposition as a potential common pathway, although the direct link between iron accumulation and clinical presentation requires further work. Figure 1 Summary of the distinguishing features in different types of NBIA. Different coloured arrows represent the primary pathomechanisms. aCP = aceruloplasminemia; BPAN = beta-propeller protein-associated neurodegeneration; FAHN = fatty acid hydroxylase-associated neurodegeneration; INAD = infantile neuroaxonal dystrophy; KRD = Kufor-Rakeb disease; MPAN = mitochondrial membrane protein-associated neurodegeneration; PKAN = pantothenate kinase associated neurodegeneration; WSS = Woodhouse-Sakati syndrome. The clinical ‘hallmarks’ of NBIA are progressive dystonia, dysarthria, spasticity and parkinsonism. Optic atrophy, retinal degeneration and peripheral neuropathy may be associated features in a number of NBIA syndromes. Characteristic MRI findings may be helpful in the diagnosis, but they may appear only later in the disease course. However some specific signs on MRI may facilitate the diagnosis (Fig. 1). The age of manifestation (childhood–adulthood) and the inheritance pattern (autosomal recessive in seven forms; autosomal dominant in neuroferritinopathy; and X-linked in WDR45 deficiency) may be helpful in the differential diagnosis of NBIA (Fig. 1). In a large proportion of cases (∼40%), the underlying genetic basis of NBIA has yet to be defined, …

Exome sequencing and SNP analysis detect novel compound heterozygosity in fatty acid hydroxylase-associated neurodegeneration

Fatty acid hydroxylase-associated neurodegeneration due to fatty acid 2-hydroxylase deficiency presents with a wide range of phenotypes including spastic paraplegia, leukodystrophy, and/or brain iron deposition. All previously described families with this disorder were consanguineous, with homozygous mutations in the probands. We describe a 10-year-old male, from a non-consanguineous family, with progressive spastic paraplegia, dystonia, ataxia, and cognitive decline associated with a sural axonal neuropathy. The use of high-throughput sequencing techniques combined with SNP array analyses revealed a novel paternally derived missense mutation and an overlapping novel maternally derived ~28-kb genomic deletion in FA2H. This patient provides further insight into the consistent features of this disorder and expands our understanding of its phenotypic presentation. The presence of a sural nerve axonal neuropathy had not been previously associated with this disorder and so may extend the phenotype.