scholarly journals Constitutively active SARM1 variants that induce neuropathy are enriched in ALS patients

2022 ◽  
Vol 17 (1) ◽  
Author(s):  
A. Joseph Bloom ◽  
Xianrong Mao ◽  
Amy Strickland ◽  
Yo Sasaki ◽  
Jeffrey Milbrandt ◽  
...  
Keyword(s):  
Intrathecal Injection ◽  
Motor Dysfunction ◽  
Single Amino Acid ◽  
Reference Allele ◽  
Axon Loss ◽  
Als Patients ◽  
Multiple Patients ◽  
Lateral Sclerosis ◽  
Constitutively Active

Abstract Background In response to injury, neurons activate a program of organized axon self-destruction initiated by the NAD+ hydrolase, SARM1. In healthy neurons SARM1 is autoinhibited, but single amino acid changes can abolish autoinhibition leading to constitutively active SARM1 enzymes that promote degeneration when expressed in cultured neurons. Methods To investigate whether naturally occurring human variants might disrupt SARM1 autoinhibition and potentially contribute to risk for neurodegenerative disease, we assayed the enzymatic activity of all 42 rare SARM1 alleles identified among 8507 amyotrophic lateral sclerosis (ALS) patients and 9671 controls. We then intrathecally injected mice with virus expressing SARM1 constructs to test the capacity of an ALS-associated constitutively active SARM1 variant to promote neurodegeneration in vivo. Results Twelve out of 42 SARM1 missense variants or small in-frame deletions assayed exhibit constitutive NADase activity, including more than half of those that are unique to the ALS patients or that occur in multiple patients. There is a > 5-fold enrichment of constitutively active variants among patients compared to controls. Expression of constitutively active ALS-associated SARM1 alleles in cultured dorsal root ganglion (DRG) neurons is pro-degenerative and cytotoxic. Intrathecal injection of an AAV expressing the common SARM1 reference allele is innocuous to mice, but a construct harboring SARM1V184G, the constitutively active variant found most frequently among the ALS patients, causes axon loss, motor dysfunction, and sustained neuroinflammation. Conclusions These results implicate rare hypermorphic SARM1 alleles as candidate genetic risk factors for ALS and other neurodegenerative conditions.

scholarly journals Constitutively active SARM1 variants found in ALS patients induce neuropathy

2021 ◽  
Author(s):  
Joseph Bloom ◽  
Xianrong Mao ◽  
Amy Strickland ◽  
Yo Sasaki ◽  
Jeffrey Milbrandt ◽  
...  
Keyword(s):  
Intrathecal Injection ◽  
Motor Dysfunction ◽  
Single Amino Acid ◽  
Reference Allele ◽  
Naturally Occurring ◽  
Axon Loss ◽  
Als Patients ◽  
Multiple Patients ◽  
Lateral Sclerosis ◽  
Constitutively Active

In response to injury, neurons activate a program of organized axon self-destruction initiated by the NAD+ hydrolase SARM1. In healthy neurons SARM1 is autoinhibited, but single amino acid changes can abolish autoinhibition leading to constitutively-active SARM1 enzymes that promote degeneration when expressed in cultured neurons. To investigate whether naturally-occurring human variants might similarly disrupt SARM1 autoinhibition and potentially contribute to risk for neurodegenerative disease, we assayed the enzymatic activity of 29 rare SARM1 alleles identified among 8,507 amyotrophic lateral sclerosis (ALS) patients. Ten missense variants or small in-frame deletions exhibit constitutive NADase activity, including more than half of those that are unique to the ALS patients or that occur in multiple patients. Expression of these constitutively active ALS-associated SARM1 alleles in cultured dorsal root ganglion (DRG) neurons is pro-degenerative and cytotoxic. Intrathecal injection of an AAV expressing the common SARM1 reference allele is innocuous to mice, but a construct harboring SARM1V184G, the constitutively active variant found most frequently in the ALS patients, causes axon loss, motor dysfunction, and sustained neuroinflammation. These results implicate rare hypermorphic SARM1 alleles as candidate genetic risk factors for ALS and other neurodegenerative conditions.

scholarly journals Involvement of cortico-efferent tracts in flail arm syndrome: a tract-of-interest-based DTI study

2021 ◽  
Author(s):  
Angela Rosenbohm ◽  
Kelly Del Tredici ◽  
Heiko Braak ◽  
Hans-Jürgen Huppertz ◽  
Albert C. Ludolph ◽  
...  
Keyword(s):  
White Matter ◽  
Diffusion Tensor ◽  
Statistical Comparison ◽  
Specific Analysis ◽  
Cerebral Pathology ◽  
Propagation Pattern ◽  
Als Patients ◽  
Lateral Sclerosis ◽  
Diffusion Tensor Imaging Dti

Abstract Background Flail arm syndrome is a restricted phenotype of motor neuron disease that is characterized by progressive, predominantly proximal weakness and atrophy of the upper limbs. Objective The study was designed to investigate specific white matter alterations in diffusion tensor imaging (DTI) data from flail arm syndrome patients using a hypothesis-guided tract-of-interest-based approach to identify in vivo microstructural changes according to a neuropathologically defined amyotrophic lateral sclerosis (ALS)-related pathology of the cortico-efferent tracts. Methods DTI-based white matter mapping was performed both by an unbiased voxel-wise statistical comparison and by a hypothesis-guided tract-wise analysis of fractional anisotropy (FA) maps according to the neuropathological ALS-propagation pattern for 43 flail arm syndrome patients vs 43 ‘classical’ ALS patients vs 40 matched controls. Results The analysis of white matter integrity demonstrated regional FA reductions for the flail arm syndrome group predominantly along the CST. In the tract-specific analysis according to the proposed sequential cerebral pathology pattern of ALS, the flail arm syndrome patients showed significant alterations of the specific tract systems that were identical to ‘classical’ ALS if compared to controls. Conclusions The DTI study including the tract-of-interest-based analysis showed a microstructural involvement pattern in the brains of flail arm syndrome patients, supporting the hypothesis that flail arm syndrome is a phenotypical variant of ALS.

scholarly journals In Vivo Characterization of High Basal Signaling from the Ghrelin Receptor

Endocrinology ◽  
2009 ◽  
Vol 150 (11) ◽  
pp. 4920-4930 ◽  
Author(s):  
Pia Steen Petersen ◽  
David P. D. Woldbye ◽  
Andreas Nygaard Madsen ◽  
Kristoffer L. Egerod ◽  
Chunyu Jin ◽  
...  
Keyword(s):  
Body Weight ◽  
Food Intake ◽  
Uncoupling Protein ◽  
Inverse Agonist ◽  
Single Amino Acid ◽  
Separate Experiment ◽  
Ghrelin Receptor ◽  
Control Peptide ◽  
Constitutively Active

The receptor for the orexigenic peptide, ghrelin, is one of the most constitutively active 7TM receptors known, as demonstrated under in vitro conditions. Change in expression of a constitutively active receptor is associated with change in signaling independent of the endogenous ligand. In the following study, we found that the expression of the ghrelin receptor in the hypothalamus was up-regulated approximately 2-fold in rats both during 48-h fasting and by streptozotocin-induced hyperphagia. In a separate experiment, to probe for the effect of the high basal signaling of the ghrelin receptor in vivo, we used intracerebroventricular administration by osmotic pumps of a peptide [D-Arg1, D-Phe5, D-Trp7,9, Leu11]-substance P. This peptide selectively displays inverse agonism at the ghrelin receptor as compared with an inactive control peptide with just a single amino acid substitution. Food intake and body weight were significantly decreased in the group of rats treated with the inverse agonist, as compared with the groups treated with the control peptide or the vehicle. In the hypothalamus, the expression of neuropeptide Y and uncoupling protein 2 was decreased by the inverse agonist. In a hypothalamic cell line that endogenously expresses the ghrelin receptor, we observed high basal activity of the cAMP response element binding protein, an important signaling transduction pathway for appetite regulation. The activation was further increased by ghrelin administration and decreased by administration of the inverse agonist. It is suggested that the high constitutive signaling activity is important for the in vivo function of the ghrelin receptor in the control of food intake and body weight.

scholarly journals Targeting S100A4 with niclosamide attenuates inflammatory and profibrotic pathways in models of amyotrophic lateral sclerosis

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Martina Milani ◽  
Eleonora Mammarella ◽  
Simona Rossi ◽  
Chiara Miele ◽  
Serena Lattante ◽  
...  
Keyword(s):  
Amyotrophic Lateral Sclerosis ◽  
Cell Types ◽  
Specific Protein ◽  
Beneficial Effects ◽  
Hexanucleotide Repeat ◽  
Hexanucleotide Repeat Expansion ◽  
Als Patients ◽  
Different Cell Types ◽  
Lateral Sclerosis

Abstract Background An increasing number of studies evidences that amyotrophic lateral sclerosis (ALS) is characterized by extensive alterations in different cell types and in different regions besides the CNS. We previously reported the upregulation in ALS models of a gene called fibroblast-specific protein-1 or S100A4, recognized as a pro-inflammatory and profibrotic factor. Since inflammation and fibrosis are often mutual-sustaining events that contribute to establish a hostile environment for organ functions, the comprehension of the elements responsible for these interconnected pathways is crucial to disclose novel aspects involved in ALS pathology. Methods Here, we employed fibroblasts derived from ALS patients harboring the C9orf72 hexanucleotide repeat expansion and ALS patients with no mutations in known ALS-associated genes and we downregulated S100A4 using siRNA or the S100A4 transcriptional inhibitor niclosamide. Mice overexpressing human FUS were adopted to assess the effects of niclosamide in vivo on ALS pathology. Results We demonstrated that S100A4 underlies impaired autophagy and a profibrotic phenotype, which characterize ALS fibroblasts. Indeed, its inhibition reduces inflammatory, autophagic, and profibrotic pathways in ALS fibroblasts, and interferes with different markers known as pathogenic in the disease, such as mTOR, SQSTM1/p62, STAT3, α-SMA, and NF-κB. Importantly, niclosamide in vivo treatment of ALS-FUS mice reduces the expression of S100A4, α-SMA, and PDGFRβ in the spinal cord, as well as gliosis in central and peripheral nervous tissues, together with axonal impairment and displays beneficial effects on muscle atrophy, by promoting muscle regeneration and reducing fibrosis. Conclusion Our findings show that S100A4 has a role in ALS-related mechanisms, and that drugs such as niclosamide which are able to target inflammatory and fibrotic pathways could represent promising pharmacological tools for ALS.

scholarly journals Niclosamide Targets Inflammatory and Profibrotic Pathways in Amyotrophic Lateral Sclerosis

2021 ◽  
Author(s):  
Martina Milani ◽  
Eleonora Mammarella ◽  
Simona Rossi ◽  
Serena Lattante ◽  
Mario Sabatelli ◽  
...  
Keyword(s):  
Amyotrophic Lateral Sclerosis ◽  
Cell Types ◽  
Specific Protein ◽  
Beneficial Effects ◽  
Hexanucleotide Repeat ◽  
Sporadic Als ◽  
Als Patients ◽  
Different Cell Types ◽  
Lateral Sclerosis

Abstract BackgroundAn increasing number of studies evidence that amyotrophic lateral sclerosis (ALS) is characterized by extensive alterations in different cell types and in different regions besides the CNS. We previously reported the up-regulation in ALS models of a gene called fibroblast-specific protein (FSP)-1 or S100A4, generally recognized as a pro-inflammatory and profibrotic factor. Since inflammation and fibrosis are often mutual-sustaining events that contribute to establish a hostile environment for organ functioning, the comprehension of the elements responsible for these interconnected pathways is crucial to disclose novel aspects involved in ALS pathology.MethodsHere we employed fibroblasts derived from ALS patients harboring the C9orf72 hexanucleotide repeat expansion and sporadic ALS patients with no mutations in known ALS-associated genes and we downregulated S100A4 using siRNA or the S100A4 transcriptional inhibitor niclosamide. Mice overexpressing human FUS were adopted to assess the effects of niclosamide in vivo on ALS pathology.ResultsWe demonstrated that S100A4 underlies impaired autophagy and a profibrotic phenotype, which characterize ALS fibroblasts. Indeed, its inhibition reduces inflammatory, autophagic and profibrotic pathways in ALS fibroblasts, and to interfere with different markers known as pathogenic in the disease, such as mTOR, SQSTM1/p62, STAT3, α-SMA and NF-κB. Importantly, niclosamide in vivo treatment of ALS-FUS mice reduces the expression of S100A4, α-SMA and PDGFRβ in the spinal cord, as well as gliosis in central and peripheral nervous tissues, together with axonal impairment and displays beneficial effects on muscle atrophy, by promoting muscle regeneration and reducing fibrosis.ConclusionOur findings show that S100A4 has a role in ALS-related mechanisms, and that drugs such as niclosamide that are able to target inflammatory and fibrotic pathways could represent promising pharmacological tools for ALS.

scholarly journals The Cyanotoxin 2,4-DAB Reduces Viability and Causes Behavioral and Molecular Dysfunctions Associated with Neurodegeneration in Larval Zebrafish

2022 ◽  
Author(s):  
Rubia M. Martin ◽  
Michael S. Bereman ◽  
Kurt C. Marsden
Keyword(s):  
Metabolic Pathways ◽  
Protein Profile ◽  
Acoustic Startle ◽  
Motor Dysfunction ◽  
Zebrafish Model ◽  
Larval Zebrafish ◽  
Neurotoxic Effects ◽  
And Behavior ◽  
Lateral Sclerosis

AbstractExposure to cyanotoxins has been linked to neurodegenerative diseases, including amyotrophic lateral sclerosis, Alzheimer’s, and Parkinson’s disease. While the cyanotoxin β-methylamino-L-alanine (BMAA) has received much attention, cyanobacteria produce many cyanotoxic compounds, several of which have been detected in nature alongside BMAA, including 2,4-diaminobutyric acid (2,4-DAB) and N-(2-aminoethyl)glycine (AEG). Thus, the question of whether 2,4-DAB and AEG also cause neurotoxic effects in vivo is of great interest, as is the question of whether they interact to enhance toxicity. Here, we evaluate the toxic and neurotoxic effects of these cyanotoxins alone or in combination by measuring zebrafish larval viability and behavior after exposure. 2,4-DAB was the most potent cyanotoxin as it decreased larval viability by approximately 50% at 6 days post fertilization, while BMAA and AEG decreased viability by just 16% and 8%, respectively. Although we only observed minor neurotoxic effects on spontaneous locomotion, BMAA and AEG enhanced acoustic startle sensitivity, and they interacted in an additive manner to exert their effects. 2,4-DAB; however, only modulated startle kinematics, an indication of motor dysfunction. To investigate the mechanisms of 2,4-DAB’s effects, we analyzed the protein profile of larval zebrafish exposed to 500 µM 2,4-DAB at two time points and identified molecular signatures consistent with neurodegeneration, including disruption of metabolic pathways and downregulation of the ALS-associated genes SOD1 and UBQLN4. Together, our data demonstrate that BMAA and its isomers AEG and 2,4-DAB cause neurotoxic effects in vivo, with 2,4-DAB as the most potent of the three in the zebrafish model.

scholarly journals Looking backward to move forward: a meta-analysis of stem cell therapy in amyotrophic lateral sclerosis

2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Cynthia Morata-Tarifa ◽  
Garikoitz Azkona ◽  
Jonathan Glass ◽  
Letizia Mazzini ◽  
Rosario Sanchez-Pernaute
Keyword(s):  
Amyotrophic Lateral Sclerosis ◽  
Mononuclear Cells ◽  
Meta Analysis ◽  
Intrathecal Injection ◽  
Current Evidence ◽  
Eligibility Criteria ◽  
Before And After ◽  
Als Patients ◽  
Positive Effect ◽  
Lateral Sclerosis

AbstractTransplantation of several types of stem cells (SC) for the treatment of amyotrophic lateral sclerosis (ALS) has been evaluated in numerous Phase I/II clinical trials with inconclusive results. Here, we conducted a meta-analysis to systematically assess the outcome of SC therapy trials which report the evolution of each patient before and after cell administration. In this way, we aimed to determine the effect of the SC intervention despite individual heterogeneity in disease progression. We identified 670 references by electronic search and 90 full-text studies were evaluated according to the eligibility criteria. Eleven studies were included comprising 220 cell-treated patients who received mesenchymal (M) SC (n = 152), neural (N) SC (n = 57), or mononuclear cells (MNC: CD34, CD117, and CD133 positive cells) (n = 11). Our analyses indicate that whereas intrathecal injection of mesenchymal stromal cells appears to have a transient positive effect on clinical progression, as measured by the ALS functional rating score, there was a worsening of respiratory function measured by forced vital capacity after all interventions. Based on current evidence, we conclude that optimal cell product and route of administration need to be determined in properly controlled preclinical models before further advancing into ALS patients. In addition, in-depth understanding of disease mechanisms in subsets of patients will help tailoring SC therapy to specific targets and increase the likelihood of improving outcomes.

scholarly journals Mutational Analysis of the Residue at Position 48 in the Salmonella enterica Serovar Typhimurium PhoQ Sensor Kinase

2003 ◽  
Vol 185 (6) ◽  
pp. 1935-1941 ◽  
Author(s):  
Sarah Sanowar ◽  
Alexandre Martel ◽  
Hervé Le Moual
Keyword(s):  
Amino Acid ◽  
Phosphatase Activity ◽  
Salmonella Enterica ◽  
Mutational Analysis ◽  
Regulatory System ◽  
Single Amino Acid ◽  
Serovar Typhimurium ◽  
Constitutively Active

ABSTRACT The PhoP/PhoQ two-component regulatory system of Salmonella enterica serovar Typhimurium plays an essential role in controlling virulence by mediating the adaptation to Mg2+ depletion. The pho-24 allele of phoQ harbors a single amino acid substitution (T48I) in the periplasmic domain of the PhoQ histidine kinase sensor. This mutation has been shown to increase net phosphorylation of the PhoP response regulator. We analyzed the effect on signaling by PhoP/PhoQ of various amino acid substitutions at this position (PhoQ-T48X [X = A, S, V, I, or L]). Mutations T48V, T48I, and T48L were found to affect signaling by PhoP/PhoQ both in vivo and in vitro. Mutations PhoQ-T48V and PhoQ-T48I increased both the expression of the mgtA::lacZ transcriptional fusion and the net phosphorylation of PhoP, conferring to cells a PhoP constitutively active phenotype. In contrast, mutation PhoQ-T48L barely responded to changes in the concentration of external Mg2+, in vivo and in vitro, conferring to cells a PhoP constitutively inactive phenotype. By analyzing in vitro the individual catalytic activities of the PhoQ-T48X sensors, we found that the PhoP constitutively active phenotype observed for the PhoQ-T48V and PhoQ-T48I proteins is solely due to decreased phosphatase activity. In contrast, the PhoP constitutively inactive phenotype observed for the PhoQ-T48L mutant resulted from both decreased autokinase activity and increased phosphatase activity. Our data are consistent with a model in which the residue at position 48 of PhoQ contributes to a conformational switch between kinase- and phosphatase-dominant states.

scholarly journals The cyanotoxin 2,4-DAB enhances mortality and causes behavioral and molecular dysfunctions associated with neurodegeneration in larval zebrafish

2021 ◽  
Author(s):  
Rubia M. Martin ◽  
Michael S. Bereman ◽  
Kurt C. Marsden
Keyword(s):  
Metabolic Pathways ◽  
Protein Profile ◽  
Acoustic Startle ◽  
Motor Dysfunction ◽  
Zebrafish Model ◽  
Larval Zebrafish ◽  
Neurotoxic Effects ◽  
And Behavior ◽  
Lateral Sclerosis

Exposure to cyanotoxins has been linked to neurodegenerative diseases, including amyotrophic lateral sclerosis, Alzheimer's, and Parkinson's disease. While the cyanotoxin beta-methylamino-L-alanine (BMAA) has received much attention, cyanobacteria produce many cyanotoxic compounds, several of which have been detected in nature alongside BMAA including 2,4-diaminobutyric acid (2,4-DAB), and N-(2-aminoethyl)glycine (AEG). Thus, the question of whether DAB and AEG also cause neurotoxic effects in vivo is of great interest, as is the question of whether they interact to enhance toxicity. Here, we evaluate the toxic and neurotoxic effects of these cyanotoxins alone or in combination by measuring zebrafish larval viability and behavior after exposure. 2,4-DAB was the most potent cyanotoxin as it decreased larval viability by approximately 50% at 6 days post fertilization, while BMAA and AEG decreased viability by just 16% and 8%, respectively. Although we only observed minor neurotoxic effects on spontaneous locomotion, BMAA and AEG enhanced acoustic startle sensitivity, and they interacted in an additive manner to exert their effects. 2,4-DAB, however, only modulated the startle kinematics, an indication of motor dysfunction. To investigate the mechanisms of 2,4-DAB's effects, we analyzed the protein profile of larval zebrafish exposed to 500 uM 2,4-DAB at two time points and identified molecular signatures consistent with neurodegeneration, including disruption of metabolic pathways and downregulation of the ALS-associated genes SOD1 and UBQLN4. Together, our data demonstrate that BMAA and its isomers AEG and 2,4-DAB cause neurotoxic effects in vivo, with 2,4-DAB as the most potent of the three in the zebrafish model.

Riluzole Exhibits No Therapeutic Efficacy on a Transgenic Rat model of Amyotrophic Lateral Sclerosis

2020 ◽  
Vol 17 (3) ◽  
pp. 275-285 ◽  
Author(s):  
Si Chen ◽  
Qiao Liao ◽  
Ke Lu ◽  
Jinxia Zhou ◽  
Cao Huang ◽  
...  
Keyword(s):  
Amyotrophic Lateral Sclerosis ◽  
Motor Neurons ◽  
Rat Model ◽  
Microglial Activation ◽  
Transgenic Rat ◽  
Intragastric Administration ◽  
Behavioral Deficits ◽  
Als Patients ◽  
Lateral Sclerosis ◽  
Transgenic Rat Model

Background: Amyotrophic lateral sclerosis (ALS) is a neurological disorder clinically characterized by motor system dysfunction, with intraneuronal accumulation of the TAR DNAbinding protein 43 (TDP-43) being a pathological hallmark. Riluzole is a primarily prescribed medicine for ALS patients, while its therapeutical efficacy appears limited. TDP-43 transgenic mice are existing animal models for mechanistic/translational research into ALS. Methods: We developed a transgenic rat model of ALS expressing a mutant human TDP-43 transgene (TDP-43M337V) and evaluated the therapeutic effect of Riluzole on this model. Relative to control, rats with TDP-43M337V expression promoted by the neurofilament heavy subunit (NEF) gene or specifically in motor neurons promoted by the choline acetyltransferase (ChAT) gene showed progressive worsening of mobility and grip strength, along with loss of motor neurons, microglial activation, and intraneuronal accumulation of TDP-43 and ubiquitin aggregations in the spinal cord. Results: Compared to vehicle control, intragastric administration of Riluzole (30 mg/kg/d) did not mitigate the behavioral deficits nor alter the neuropathologies in the transgenics. Conclusion: These findings indicate that transgenic rats recapitulate the basic neurological and neuropathological characteristics of human ALS, while Riluzole treatment can not halt the development of the behavioral and histopathological phenotypes in this new transgenic rodent model of ALS.

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