Considering Big tau as a novel and specific biomarker for spinal motor neuron pathology.

Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease caused by the selective loss of both spinal and upper motor neurons. One strategy in treating ALS is to use stem cells to replace lost spinal motor neurons. However, transplanted stem cell-derived motor neurons may not survive when exposed to the harsh microenvironment in the spinal cord of ALS. In particular, dysfunctional astrocytes and overactivated microglia in ALS may limit the survival of motor neurons generated from cell replacement therapy. On the other hand, stem cells may provide large quantities of motor neurons that can be used for studying glia-mediated toxic mechanisms and potential therapies in ALS. Here we will review methods and molecular factors for directed differentiation of stem cells into spinal motor neurons, the potential uses of these models for dissecting the mechanisms underlying glia-induced motor neuron degeneration and screening for new therapeutics aimed at protecting motor neurons in ALS, as well as discuss challenges facing the development of motor neuron replacement-based cell therapies for recovery in ALS.

The etiological underpinnings of amyotrophic lateral sclerosis (ALS) are complex and incompletely understood, although contributions to pathogenesis by regulators of proteolytic pathways have become increasingly apparent. Here, we present a novel variant in UBQLN4 that is associated with ALS and show that its expression compromises motor axon morphogenesis in mouse motor neurons and in zebrafish. We further demonstrate that the ALS-associated UBQLN4 variant impairs proteasomal function, and identify the Wnt signaling pathway effector beta-catenin as a UBQLN4 substrate. Inhibition of beta-catenin function rescues the UBQLN4 variant-induced motor axon phenotypes. These findings provide a strong link between the regulation of axonal morphogenesis and a new ALS-associated gene variant mediated by protein degradation pathways.DOI: http://dx.doi.org/10.7554/eLife.25453.001

A Stem Cell-Based Screening Platform Identifies Compounds that Desensitize Motor Neurons to Endoplasmic Reticulum Stress.

Mutations of the lipid phosphatase FIG4 that regulates PI(3,5)P(2) are responsible for the recessive peripheral-nerve disorder CMT4J. We now describe nonsynonymous variants of FIG4 in 2% (9/473) of patients with amyotrophic lateral sclerosis (ALS) and primary lateral sclerosis (PLS). Heterozygosity for a deleterious allele of FIG4 appears to be a risk factor for ALS and PLS, extending the list of known ALS genes and increasing the clinical spectrum of FIG4-related diseases.

Enrichment of C-Terminal Fragments in TAR DNA-Binding Protein-43 Cytoplasmic Inclusions in Brain but not in Spinal Cord of Frontotemporal Lobar Degeneration and Amyotrophic Lateral Sclerosis

DNA damage is implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS). However, relationships between DNA damage accumulation, DNA damage response (DDR), and upper and lower motor neuron vulnerability in human ALS are unclear; furthermore, it is unknown whether epigenetic silencing of DNA repair pathways contributes to ALS pathogenesis. We tested the hypotheses that DNA damage accumulates in ALS motor neurons along with diminished DDR, and that DNA repair genes undergo hypermethylation. Human postmortem CNS tissue was obtained from ALS cases (N = 34) and age-matched controls without neurologic disease (N = 15). Compared to age-matched controls, abasic sites accumulated in genomic DNA of ALS motor cortex and laser capture microdissection-acquired spinal motor neurons but not in motor neuron mitochondrial DNA. By immunohistochemistry, DNA damage accumulated significantly in upper and lower motor neurons in ALS cases as single-stranded DNA and 8-hydroxy-deoxyguanosine (OHdG) compared to age-matched controls. Significant DDR was engaged in ALS motor neurons as evidenced by accumulation of c-Abl, nuclear BRCA1, and ATM activation. DNA damage and DDR were present in motor neurons at pre-attritional stages and throughout the somatodendritic attritional stages of neurodegeneration. Motor neurons with DNA damage were also positive for activated p53 and cleaved caspase-3. Gene-specific promoter DNA methylation pyrosequencing identified the DNA repair genes Ogg1, Apex1, Pnkp and Aptx as hypomethylated in ALS. In human induced-pluripotent stem cell (iPSC)-derived motor neurons with familial ALS SOD1 mutations, DNA repair capacity was similar to isogenic control motor neurons. Our results show that vulnerable neurons in human ALS accumulate DNA damage, and contrary to our hypothesis, strongly activate and mobilize response effectors and DNA repair genes. This DDR in ALS motor neurons involves recruitment of c-Abl and BRCA1 to the nucleus in vivo, and repair of DNA double-strand breaks in human ALS motor neurons with SOD1 mutations in cell culture.

Apoptosis has been implicated in many neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS). We previously demonstrated a role for G1 to S phase cell cycle regulators in ALS with increased levels of hyperphosphorylated retinoblastoma (ppRb) and E2F-1 in ALS spinal cord motor neurons. In this study we examined the levels of the cell cycle checkpoint tumor suppressor protein p53 with concurrent changes in cell death markers during ALS. Expression and subcellular distribution of p53, retinoblastoma, Bax, Fas, and caspases were explored by immunoblot, immunohistochemistry and double-label confocal microscopy in the spinal cord and motor cortex of ALS and control subjects. We identified elevated levels of p53 in ALS spinal cord motor neurons but not neurons in the motor cortex. In addition, there was an increase in Bax, Fas, caspases-8 and -3 proteins in ALS spinal motor neurons. While caspase-3 and TUNEL labeled neurons were positive for ppRb, E2F-1 and p53 in spinal motor neurons, and Fas co-localized with caspase-8 in spinal motor neurons, we failed to observe these results in large neurons in the motor cortex of ALS subjects. We have linked p53 and activation of G1 to S phase cell cycle regulators to an apoptotic mode of cell death ALS spinal cord motor neurons.

Upper motor neuron (UMN) and lower motor neuron (LMN) involvement represent the core clinical features of amyotrophic lateral sclerosis (ALS). Several studies divided patients into prevalent UMN and LMN impairment phenotypes to investigate the association between motor systems impairments and ALS clinical course. However, this distinction was somehow heterogeneous and significantly affected the comparability across studies. This study aimed to investigate whether patients spontaneously segregate based on the extent of UMN and LMN involvement without a-priori categorization and to identify potential clinical and prognostic features of different clusters. Eighty-eight consecutive spinal-onset ALS patients were referred to an ALS tertiary center between 2015 and 2022. UMN and LMN burden was assessed with the Penn Upper Motor Neuron scale (PUMNS) and the Devine score, respectively. PUMNS and LMN scores were normalized into 0-1 and analyzed using a two-step cluster analysis and the Euclidean distance measure. The Bayesian Information Criterion was used to determine the cluster number. Demographic and clinical variables were tested for differences among the clusters. Three distinct clusters emerged at cluster analysis. Patients in "cluster-1" showed moderate UMN and severe LMN involvement, corresponding to the typical ALS phenotype. Patients in "cluster-2" showed mild LMN and severe UMN damage, corresponding to a predominant UMN phenotype, while "cluster-3" patients showed mild UMN and moderate LMN damage, corresponding to a predominant LMN phenotype. Patients in "cluster-1" and "cluster-2" showed a higher prevalence of definite ALS than those in "cluster-3" (61% and 46 vs 9%, p < 0.001). "Cluster-1" patients had a lower median ALSFRS-r score compared to both "cluster-2" and 3 patients (27 vs 40 and 35, < 0.001). "Cluster-1" (HR: 8.5; 95% CI 2.1-35.1 and p = 0.003) and 3 (HR: 3.2; 95% CI 1.1-9.1; p = 0.03) were associated with shorter survival than those in "cluster-2". Spinal-onset ALS can be categorized into three groups according to LMN and UMN burden. The UMN burden is related to higher diagnostic certainty and broader disease spread, while LMN involvement is associated with higher disease severity and shorter survival.

Inhibition of superoxide dismutase selectively suppresses growth of rat spinal motor neurons: Comparison with phosphorylated neurofilament-containing spinal neurons

Rapid and Efficient Generation of Functional Motor Neurons From Human Pluripotent Stem Cells Using Gene Delivered Transcription Factor Codes

Evaluation Of Electrophysiological Changes In Primary Spinal Cord Neuronal Cultures Exposed To ALS-CSF Using Microelectrode Array.

Objective To quantitatively estimate upper motor neuron (UMN) lesion with the triple stimulation technique (TST) in amyotrophic lateral sclerosis (ALS). Methods Fifty ALS patients and 22 normal controls were enrolled in the study. Patients were examined clinically with conventional transcranial magnetic stimulation (TMS) and TST at abductor digiti minimi. Central motor conduction time (CMCT),motor evoked potential ( MEP), resting motor threshold ( RMT), compound muscle action potential ( CMAP), modified ashworth scale ( MAS), Medical Research Council scale and modified ALS scale were assessed and their correlation to disease progress was analyzed. Results The TST amplitude ratio was significantly decreased in ALS patients with UMN signs(28 cases 62. 0% (40. 7%, 75.9% ) ), compared with controls ( 96. 9% ± 2. 6% ) and ALS patients without UMN signs ( 22 cases 95.6% ( 85.4%,100. 0% ) ;Z = -4. 827, -5.435, both P =0. 000). The abnormal rates of the TST amplitude in ALS with UMN signs, ALS without UMN signs and controls were 89. 3%, 27.3% ,9. 1% respectively. The abnormal rates of the TST amplitude, the latency of MEP, CMCT, RMT in detecting UMN lesions were 89. 3%,64. 3%, 53.6%, 64. 3% , 78. 6%, respectively. The TST amplitude was significantly correlated to tendon reflex in right arm ( r = 0. 690, P = 0. 000), with modified ashworth scale ( MAS, r = - 0. 772,P = 0. 000),with diagnostic degree ( r = 0. 483, P = 0. 000), with RMT ( r = - 0. 774,P = 0. 000), the latency of MEP (r = - 0. 444, P = 0. 005 ), motor evoked potential/compound muscle action potential of erb' s ( MEP/ CMAPerb, r = 0. 685, P = 0. 000 ), MEP/CMAPerb in facilitation ( r = 0. 770, P = 0. 000). Conclusions TST appears to be a more accurate and sensitive measure of detecting and quantifying UMN abnormality in ALS patients than the other parameters. TST may reveal the subclinical UMN impairment in ALS and provide an accurate diagnosis assessment for UMN loss in ALS and an objective scale for monitoring the progression of disease. Key words: Amyotrophic lateral sclerosis; Transcranial magnetic stimulation; Motor neurons

There are no effective cures for upper motor neuron (UMN) diseases, such as amyotrophic lateral sclerosis (ALS), primary lateral sclerosis, and hereditary spastic paraplegia. Here, we show UMN loss occurs independent of spinal motor neuron degeneration and that UMNs are indeed effective cellular targets for gene therapy, which offers a potential solution especially for UMN disease patients.UCHL1 (ubiquitin C-terminal hydrolase-L1) is a deubiquitinating enzyme crucial for maintaining free ubiquitin levels. Corticospinal motor neurons (CSMN, a.k.a UMNs in mice) show early, selective, and profound degeneration in Uchl1nm3419 (UCHL1−/−) mice, which lack all UCHL1 function. When UCHL1 activity is ablated only from spinal motor neurons, CSMN remained intact. However, restoring UCHL1 specifically in CSMN of UCHL1−/− mice via directed gene delivery, was sufficient to improve CSMN integrity to the healthy control levels. In addition, when UCHL1 gene was delivered selectively to CSMN that are diseased due to misfolded SOD1 toxicity and TDP-43 pathology via AAV-mediated retrograde transduction, the disease causing misfolded SOD1 and mutant human TDP-43 were reduced in hSOD1G93A and prpTDP-43A315T models, respectively. Diseased CSMN retained their neuronal integrity and cytoarchitectural stability in two different mouse models that represent two distinct causes of neurodegeneration in ALS.

Excitation BolsTORs Motor Neurons in ALS Mice