Genome-wide RNA-seq of iPSC-derived motor neurons indicates selective cytoskeletal perturbation in Brown–Vialetto disease that is partially rescued by riboflavin

Abstract

Riboflavin is essential in numerous cellular oxidation/reduction reactions but is not synthesized by mammalian cells. Riboflavin absorption occurs through the human riboflavin transporters RFVT1 and RFVT3 in the intestine and RFVT2 in the brain. Mutations in these genes are causative for the Brown–Vialetto–Van Laere (BVVL), childhood-onset syndrome characterized by a variety of cranial nerve palsies as well as by spinal cord motor neuron (MN) degeneration. Why mutations in RFVTs result in a neural cell–selective disorder is unclear. As a novel tool to gain insights into the pathomechanisms underlying the disease, we generated MNs from induced pluripotent stem cells (iPSCs) derived from BVVL patients as an in vitro disease model. BVVL-MNs explained a reduction in axon elongation, partially improved by riboflavin supplementation. RNA sequencing profiles and protein studies of the cytoskeletal structures showed a perturbation in the neurofilament composition in BVVL-MNs. Furthermore, exploring the autophagy–lysosome pathway, we observed a reduced autophagic/mitophagic flux in patient MNs. These features represent emerging pathogenetic mechanisms in BVVL-associated neurodegeneration, partially rescued by riboflavin supplementation. Our data showed that this therapeutic strategy could have some limits in rescuing all of the disease features, suggesting the need to develop complementary novel therapeutic strategies.

Clinical, pathological and functional characterization of riboflavin-responsive neuropathy

“…Recently, reductions in axonal length and transcriptional signatures of altered cytoskeletal composition were identified in BVVL syndrome iPSC-derived motor neurons ( Rizzo et al , 2017 ). To test for altered axonal development and signatures of cytoskeletal dysfunction in vivo , we examined the neuromuscular junction of drift knockdown third instar larvae.…”

“…Indeed, although infants with early-onset riboflavin transporter deficiency rapidly become ventilator-dependent and usually die in the first year of life, treatment with high-dose riboflavin supplementation partially ameliorates the progression of this neurodegenerative condition, particularly if initiated soon after the onset of symptoms ( Foley et al , 2014 ). In a recent paper by Rizzo et al (2017) , motor neurons from induced pluripotent stem cells (iPSCs) were derived from two patients with BVVL syndrome carrying either SLC52A2 or SLC52A3 mutations. They found a reduction in axon elongation coupled with perturbations in neurofilament composition and reduced autophagic/mitophagic flux.…”

An update on the genetics, clinical presentation, and pathomechanisms of human riboflavin transporter deficiency

“…In a study by Rizzo et al, human‐induced pluripotent stem cell (hIPSC) lines were established from a RTD2 and a RTD3 patient and differentiated into motor neurons. RTD motor neurons displayed an increase in neurofilament heavy chain (NFH) expression and its aggregation in inclusions, something previously characterized as an early event leading to motor neuron degeneration in amyotrophic lateral sclerosis (ALS) .…”

Antioxidant Amelioration of Riboflavin Transporter Deficiency in Motoneurons Derived from Patient-Specific Induced Pluripotent Stem Cells

“…iPSC (induced pluripotent stem cell) technology has allowed the creation of disease models, drug screening and personalized treatments for many genetic disorders [ 42 , 43 , 44 , 45 , 46 ]. The special advantage of using iPSCs is creating patient-specific cellular models by differentiating iPSCs into the cell types affected in the disease, in this case, RTD-motoneurons (RTD-MNs) [ 47 ]. Since a murine model accurately recapitulating the human pathology is lacking, the RTD model of iPSC-derived motoneurons (MNs) is of particular interest.…”