•ALS disease heterogeneity presents challenges for therapeutic development.•Animal modeling has provided initial insights into cell subtype contributions to ALS.•iPSC show potential for dissecting ...contributions of cell subtypes to ALS progression.•3D modeling and precision medicine approaches with IPSC are powerful new platforms.•iPSC have helped identify drugs currently in clinical trial for ALS.
Amongst the most important discoveries in ALS pathobiology are the works demonstrating that multiple cell types contribute to disease onset and progression. However, a significant limitation in ALS research is the inability to obtain tissues from ALS patient brain and spinal cord during the course of the disease. In vivo modeling has provided insights into the role of these cell subtypes in disease onset and progression. However, in vivo models also have shortcomings, including the reliance on a limited number of models based upon hereditary forms of the disease. Therefore, using human induced pluripotent stem cells (iPSC) reprogrammed from somatic cells of ALS patients, with both hereditary and sporadic forms of the disease, and differentiated into cell subtypes of both the central nervous system (CNS) and peripheral nervous system (PNS), have become powerful complementary tools for investigating basic mechanisms of disease as well as a platform for drug discovery. Motor neuron and other neuron subtypes, as well as non-neuronal cells have been differentiated from human iPSC and studied for their potential contributions to ALS pathobiology. As iPSC technologies have advanced, 3D modeling with multicellular systems organised in microfluidic chambers or organoids are the next step in validating the pathways and therapeutic targets already identified. Precision medicine approaches with iPSC using either traditional strategies of screening drugs that target a known pathogenic mechanism as well as “blind-to-target” drug screenings that allow for patient stratification based on drug response rather than clinical characteristics are now being employed.
Amyotrophic Lateral Sclerosis (ALS) is a complex multifactorial disorder, characterized by motor neuron loss with involvement of several other cell types, including astrocytes, oligodendrocytes and ...microglia. Studies in vivo and in in vitro models have highlighted that the contribution of non‐neuronal cells to the disease is a primary event and ALS pathogenesis is driven by both cell‐autonomous and non‐cell autonomous mechanisms. The advancements in genetics and in vitro modeling of the past 10 years have dramatically changed the way we investigate the pathogenic mechanisms involved in ALS. The identification of mutations in transactive response DNA‐binding protein gene (TARDBP), fused in sarcoma (FUS) and, more recently, a GGGGCC‐hexanucleotide repeat expansion in chromosome 9 open reading frame 72 (C9ORF72) and their link with familial ALS have provided new avenues of investigation and hypotheses on the pathophysiology of this devastating disease. In the same years, from 2007 to present, in vitro technologies to model neurological disorders have also undergone impressive developments. The advent of induced pluripotent stem cells (iPSCs) gave the field of ALS the opportunity to finally model in vitro not only familial, but also the larger part of ALS cases affected by sporadic disease. Since 2008, when the first human iPS‐derived motor neurons from patients were cultured in a petri dish, several different techniques have been developed to produce iPSC lines through genetic reprogramming and multiple direct conversion methods have been optimised. In this review, we will give an overview of how human in vitro models have been used so far, what discoveries they have led to since 2007, and how the recent advances in technology combined with the genetic discoveries, have tremendously widened the horizon of ALS research.
The blood-brain barrier (BBB) is a highly specialized and dynamic compartment which regulates the uptake of molecules and solutes from the blood. The relevance of the maintenance of a healthy BBB ...underpinning disease prevention as well as the main pathomechanisms affecting BBB function will be detailed in this review. Barrier disruption is a common aspect in both neurodegenerative diseases, such as amyotrophic lateral sclerosis, and neurodevelopmental diseases, including autism spectrum disorders. Throughout this review, conditions altering the BBB during the earliest and latest stages of life will be discussed, revealing common factors involved. Due to the barrier's role in protecting the brain from exogenous components and xenobiotics, drug delivery across the BBB is challenging. Potential therapies based on the BBB properties as molecular Trojan horses, among others, will be reviewed, as well as innovative treatments such as stem cell therapies. Additionally, due to the microbiome influence on the normal function of the brain, microflora modulation strategies will be discussed. Finally, future research directions are highlighted to address the current gaps in the literature, emphasizing the idea that common therapies for both neurodevelopmental and neurodegenerative pathologies exist.
Increasing evidence supports the involvement of DNA damage in several neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS). Elevated levels of DNA damage are consistently ...observed in both sporadic and familial forms of ALS and may also play a role in Western Pacific ALS, which is thought to have an environmental cause. The cause of DNA damage in ALS remains unclear but likely differs between genetic subgroups. Repeat expansion in the
C9ORF72
gene is the most common genetic cause of familial ALS and responsible for about 10% of sporadic cases. These genetic mutations are known to cause R-loops, thus increasing genomic instability and DNA damage, and generate dipeptide repeat proteins, which have been shown to lead to DNA damage and impairment of the DNA damage response. Similarly, several genes associated with ALS including
TARDBP
,
FUS
,
NEK1
,
SQSTM1
and
SETX
are known to play a role in DNA repair and the DNA damage response, and thus may contribute to neuronal death via these pathways. Another consistent feature present in both sporadic and familial ALS is the ability of astrocytes to induce motor neuron death, although the factors causing this toxicity remain largely unknown. In this review, we summarise the evidence for DNA damage playing a causative or secondary role in the pathogenesis of ALS as well as discuss the possible mechanisms involved in different genetic subtypes with particular focus on the role of astrocytes initiating or perpetuating DNA damage in neurons.
Amyotrophic lateral sclerosis (ALS) is a genetically diverse disease. At least 15 ALS-associated gene loci have so far been identified, and the causative gene is known in approximately 30% of ...familial ALS cases. Less is known about the factors underlying the sporadic form of the disease. The molecular mechanisms of motor neuron degeneration are best understood in the subtype of disease caused by mutations in superoxide dismutase 1, with a current consensus that motor neuron injury is caused by a complex interplay between multiple pathogenic processes. A key recent finding is that mutated TAR DNA-binding protein 43 is a major constituent of the ubiquitinated protein inclusions in ALS, providing a possible link between the genetic mutation and the cellular pathology. New insights have also indicated the importance of dysregulated glial cell-motor neuron crosstalk, and have highlighted the vulnerability of the distal axonal compartment early in the disease course. In addition, recent studies have suggested that disordered RNA processing is likely to represent a major contributing factor to motor neuron disease. Ongoing research on the cellular pathways highlighted in this Review is predicted to open the door to new therapeutic interventions to slow disease progression in ALS.
Spinal muscular atrophy (SMA) is the most frequent lethal genetic neurodegenerative disorder in infants. The disease is caused by low abundance of the survival of motor neuron (SMN) protein leading ...to motor neuron degeneration and progressive paralysis. We previously demonstrated that a single intravenous injection (IV) of self-complementary adeno-associated virus-9 carrying the human SMN cDNA (scAAV9-SMN) resulted in widespread transgene expression in spinal cord motor neurons in SMA mice as well as nonhuman primates and complete rescue of the disease phenotype in mice. Here, we evaluated the dosing and efficacy of scAAV9-SMN delivered directly to the cerebral spinal fluid (CSF) via single injection. We found widespread transgene expression throughout the spinal cord in mice and nonhuman primates when using a 10 times lower dose compared to the IV application. Interestingly, in nonhuman primates, lower doses than in mice can be used for similar motor neuron targeting efficiency. Moreover, the transduction efficacy is further improved when subjects are kept in the Trendelenburg position to facilitate spreading of the vector. We present a detailed analysis of transduction levels throughout the brain, brainstem, and spinal cord of nonhuman primates, providing new guidance for translation toward therapy for a wide range of neurodegenerative disorders.
Ubiquitous expression of amyotrophic lateral sclerosis (ALS)-causing mutations in superoxide dismutase 1 (SOD1) provokes noncell autonomous paralytic disease. By combining ribosome affinity ...purification and high-throughput sequencing, a cascade of mutant SOD1-dependent, cell type-specific changes are now identified. Initial mutant-dependent damage is restricted to motor neurons and includes synapse and metabolic abnormalities, endoplasmic reticulum (ER) stress, and selective activation of the PRKR-like ER kinase (PERK) arm of the unfolded protein response. PERK activation correlates with what we identify as a naturally low level of ER chaperones in motor neurons. Early changes in astrocytes occur in genes that are involved in inflammation and metabolism and are targets of the peroxisome proliferator-activated receptor and liver X receptor transcription factors. Dysregulation of myelination and lipid signaling pathways and activation of ETS transcription factors occur in oligodendrocytes only after disease initiation. Thus, pathogenesis involves a temporal cascade of cell type-selective damage initiating in motor neurons, with subsequent damage within glia driving disease propagation.
Amyotrophic lateral sclerosis (ALS) causes motor neuron degeneration, paralysis, and death. Accurate disease modeling, identifying disease mechanisms, and developing therapeutics is urgently needed. ...We previously reported motor neuron toxicity through postmortem ALS spinal cord-derived astrocytes. However, these cells can only be harvested after death, and their expansion is limited. We now report a rapid, highly reproducible method to convert adult human fibroblasts from living ALS patients to induced neuronal progenitor cells and subsequent differentiation into astrocytes (i-astrocytes). Non-cell autonomous toxicity to motor neurons is found following coculture of i-astrocytes from familial ALS patients with mutation in superoxide dismutase or hexanucleotide expansion in C9orf72 (ORF 72 on chromosome 9) the two most frequent causes of ALS. Remarkably, i-astrocytes from sporadic ALS patients are as toxic as those with causative mutations, suggesting a common mechanism. Easy production and expansion of i-astrocytes now enables rapid disease modeling and high-throughput drug screening to alleviate astrocyte-derived toxicity.
Oligodendrocytes have recently been implicated in the pathophysiology of amyotrophic lateral sclerosis (ALS). Here we show that, in vitro, mutant superoxide dismutase 1 (SOD1) mouse oligodendrocytes ...induce WT motor neuron (MN) hyperexcitability and death. Moreover, we efficiently derived human oligodendrocytes from a large number of controls and patients with sporadic and familial ALS, using two different reprogramming methods. All ALS oligodendrocyte lines induced MN death through conditioned medium (CM) and in coculture. CM-mediated MN death was associated with decreased lactate production and release, whereas toxicity in coculture was lactate-independent, demonstrating that MN survival is mediated not only by soluble factors. Remarkably, human SOD1 shRNA treatment resulted in MN rescue in both mouse and human cultures when knockdown was achieved in progenitor cells, whereas it was ineffective in differentiated oligodendrocytes. In fact, early SOD1 knockdown rescued lactate impairment and cell toxicity in all lines tested, with the exclusion of samples carrying chromosome 9 ORF 72 (C9orf72) repeat expansions. These did not respond to SOD1 knockdown nor did they show lactate release impairment. Our data indicate that SOD1 is directly or indirectly involved in ALS oligodendrocyte pathology and suggest that in this cell type, some damage might be irreversible. In addition, we demonstrate that patients with C9ORF72 represent an independent patient group that might not respond to the same treatment.
It is important to understand how the disease process affects the metabolic pathways in amyotrophic lateral sclerosis and whether these pathways can be manipulated to ameliorate disease progression. ...To analyse the basis of the metabolic defect in amyotrophic lateral sclerosis we used a phenotypic metabolic profiling approach. Using fibroblasts and reprogrammed induced astrocytes from C9orf72 and sporadic amyotrophic lateral sclerosis cases we measured the production rate of reduced nicotinamide adenine dinucleotides (NADH) from 91 potential energy substrates simultaneously. Our screening approach identified that C9orf72 and sporadic amyotrophic lateral sclerosis induced astrocytes have distinct metabolic profiles compared to controls and displayed a loss of metabolic flexibility that was not observed in fibroblast models. This loss of metabolic flexibility, involving defects in adenosine, fructose and glycogen metabolism, as well as disruptions in the membrane transport of mitochondrial specific energy substrates, contributed to increased starvation induced toxicity in C9orf72 induced astrocytes. A reduction in glycogen metabolism was attributed to loss of glycogen phosphorylase and phosphoglucomutase at the protein level in both C9orf72 induced astrocytes and induced neurons. In addition, we found alterations in the levels of fructose metabolism enzymes and a reduction in the methylglyoxal removal enzyme GLO1 in both C9orf72 and sporadic models of disease. Our data show that metabolic flexibility is important in the CNS in times of bioenergetic stress.
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