Search

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by gradual degeneration and elimination of motor neurons (MNs) in the motor cortex, brainstem, and spinal cord. Some familial forms of ALS are caused by genetic mutations in superoxide dismutase 1 (SOD1) but the mechanisms driving MN disease are unclear. Identifying the naturally occurring pathology and understanding how this mutant SOD1 can affect MNs in translationally meaningful ways in a valid and reliable human cell model remains to be established. Here, using CRISPR/Cas9 genome editing system and human induced pluripotent stem cells (iPSCs), we generated highly pure, iPSC-derived MNs with a SOD1-G93A missense mutation. With the wild-type cell line serving as an isogenic control and MNs from a patient-derived iPSC line with an SOD1-A4V mutation as a comparator, we identified pathological phenotypes relevant to ALS. The mutant MNs accumulated misfolded and aggregated forms of SOD1 in cell bodies and processes, including axons. They also developed distinctive axonal pathologies. Mutants had axonal swellings with shorter axon length and less numbers of branch points. Moreover, structural and molecular abnormalities in presynaptic and postsynaptic size and density were found in the mutants. Finally, functional studies with microelectrode array demonstrated that the individual mutant MNs exhibited decreased number of spikes and diminished network bursting, but increased burst duration. Taken together, we identified spontaneous disease phenotypes relevant to ALS in mutant SOD1 MNs from genome-edited and patient-derived iPSCs. Our findings demonstrate that SOD1 mutations in human MNs cause cell-autonomous proteinopathy, axonopathy, synaptic pathology, and aberrant neurotransmission.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by gradual degeneration and elimination of motor neurons (MNs) in the motor cortex, brainstem, and spinal cord. Some familial forms of ALS are caused by genetic mutations in superoxide dismutase 1 (SOD1) but the mechanisms driving MN disease are unclear. Identifying the naturally occurring pathology and understanding how this mutant SOD1 can affect MNs in translationally meaningful ways in a valid and reliable human cell model remains to be established. Here, using CRISPR/Cas9 genome editing system and human induced pluripotent stem cells (iPSCs), we generated highly pure, iPSC-derived MNs with a SOD1-G93A missense mutation. With the wild-type cell line serving as an isogenic control and MNs from a patient-derived iPSC line with an SOD1-A4V mutation as a comparator, we identified pathological phenotypes relevant to ALS. The mutant MNs accumulated misfolded and aggregated forms of SOD1 in cell bodies and processes, including axons. They also developed distinctive axonal pathologies. Mutants had axonal swellings with shorter axon length and less numbers of branch points. Moreover, structural and molecular abnormalities in presynaptic and postsynaptic size and density were found in the mutants. Finally, functional studies with microelectrode array demonstrated that the individual mutant MNs exhibited decreased number of spikes and diminished network bursting, but increased burst duration. Taken together, we identified spontaneous disease phenotypes relevant to ALS in mutant SOD1 MNs from genome-edited and patient-derived iPSCs. Our findings demonstrate that SOD1 mutations in human MNs cause cell-autonomous proteinopathy, axonopathy, synaptic pathology, and aberrant neurotransmission. [ABSTRACT FROM AUTHOR]

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease characterized by degeneration and loss of motor neurons in the spinal cord, brainstem and motor cortex. Up to 10% of ALS cases are inherited (familial, fALS) and associated with mutations, frequently in the superoxide dismutase 1 (SOD1) gene. Rodent transgenic models of ALS are often used to elucidate a complex pathogenesis of this disease. Of importance, both ALS patients and animals carrying mutated human SOD1 gene show symptoms of oxidative stress and iron metabolism misregulation. The aim of our study was to characterize changes in iron metabolism in one of the most commonly used models of ALS - transgenic mice overexpressing human mutated SOD1G93A gene. We analyzed the expression of iron-related genes in asymptomatic, 2-month-old and symptomatic, 4-month-old SOD1G93A mice. In parallel, respective age-matched mice overexpressing human non-mutated SOD1 transgene and control mice were analyzed. We demonstrate that the overexpression of both SOD1 and SOD1G93A genes account for a substantial increase in SOD1 protein levels and activity in selected tissues and that not all the changes in iron metabolism genes expression are specific for the overexpression of the mutated form of SOD1. [ABSTRACT FROM AUTHOR]

Amyotrophic Lateral Sclerosis (ALS) is a fatal motoneuron disease that is characterized by the degradation of neurons throughout the central nervous system. Inflammation have been cited a key contributor to ALS neurodegeneration, but the timeline of cytokine upregulation remains unresolved. The goal of this study was to temporally examine the correlation between the varying levels of pro-inflammatory type I cytokines (IL-1ß, IL-1α, IL-12, TNF-α, and GFAP) and anti-inflammatory type II cytokines (IL-4, IL-6, IL-10) throughout the progression of ALS in the SOD1 G93A mouse model. Cytokine level data from high copy SOD1 G93A transgenic mice was collected from 66 peer-reviewed studies. For each corresponding experimental time point, the ratio of transgenic to wild type (TG/WT) cytokine was calculated. One-way ANOVA and t-tests with Bonferonni correction were used to analyze the data. Meta-analysis was performed for four discrete stages: early, pre-onset, post-onset, and end stage. A significant increase in TG cytokine levels was found when compared to WT cytokine levels across the entire SOD1 G93A lifespan for majority of the cytokines. The rates of change of the individual cytokines, and type I and type II were not significantly different; however, the mean fold change of type I was expressed at significantly higher levels than type II levels across all stages with the difference between the means becoming more pronounced at the end stage. An overexpression of cytokines occurred both before and after the onset of ALS symptoms. The trend between pro-inflammatory type I and type II cytokine mean levels indicate a progressive instability of the dynamic balance between pro- and anti-inflammatory cytokines as anti-inflammatory cytokines fail to mediate the pronounced increase in pro-inflammatory cytokines. Very early immunoregulatory treatment is necessary to successfully interrupt ALS-induced neuroinflammation. [ABSTRACT FROM AUTHOR]