Journal: The Journal of Neuroscience

Article Title: MicroRNA Profiling Reveals Marker of Motor Neuron Disease in ALS Models

doi: 10.1523/JNEUROSCI.3582-16.2017

Figure Lengend Snippet: hSOD1 knockdown specifically and effectively preserves MN function and MN-enriched miRNAs are responsive to therapy. A, hSOD1 mRNA knockdown after treatment with hSOD1 ASO. There is an average 75% reduction in hSOD1 levels in the lumbar spinal cord of SOD1G93A rats treated with SOD1 ASO compared with aCSF-treated controls. n = 7/group. B, Hindlimb grip strength of aCSF, scrambled ASO, and hSOD1 ASO-treated SOD1G93A rats at 80, 115, and 150 d. C, miR-218 CSF levels are not significantly different in aCSF- and scrambled ASO-treated SOD1G93A littermates. n = 8–10. D, miR-218 in CSF is reduced in rats treated with SOD1-lowering ASO compared with aCSF-treated controls. n = 7/group. These aCSF vs scrambled ASO and aCSF vs hSOD1 ASO treatment cohorts were tested at separate time points. E, The neuronal miRNAs miR-132 and miR-124 are not responsive to ALS therapy. n = 7/group. F, miR-218 CSF levels have a strong and significant correlation with hSOD1 mRNA spinal cord levels. G, miR-218 CSF levels are not changed after treatment with aCSF, scrambled, or hSOD1 ASO in nontransgenic rats. n = 5/group. Values are expressed as mean ± SEM and are normalized to GAPDH (A) or a geometric mean of endogenous miRNA biological fluid controls, miR-103a, miR-24, and miR-191 (C–E, G). **p ≤ 0.01, ****p ≤ 0.0001, Student's unpaired, two-tailed t test (A, C, D, E), one-way ANOVA (G), two-way ANOVA (B), linear regression analysis (F).

Article Snippet: For CSF studies, Sprague Dawley nontransgenic, hSOD1 WT (provided by Pak Chan, Stanford University) and the SOD1 G93A (Taconic model 2148) rat lines were used.

Techniques: Two Tailed Test

Journal: Frontiers in Physiology

Article Title: Effect of Mutated Cu, Zn Superoxide Dismutase (SOD1 G93A ) on Modulation of Transductional Pathway Mediated by M1 Muscarinic Receptor in SK-N-BE and NSC-34 Cells

doi: 10.3389/fphys.2018.00611

Figure Lengend Snippet: (A) Presence of muscarinic acetylcholine M1 receptor in human neuroblastoma SK-NBE, oligodendrocytes MO3-13 and neuroblastoma-spinal motor neuron fusion cells NSC-34 M1 receptor protein bands normalized to α-tubulin. Effect of SOD1 on ERK1-2 (B) and AKT (C) activation in NSC-34 cells incubated with 400 ng/ml of SOD1 for 10 and 30 min. The data represent the means ± SEM of three independent experiments relative to control obtained by densitometric analysis of P-ERK1-2 and P-AKT protein bands normalized to ERK1-2 and AKT, respectively. ∗ p < 0.05 vs. Ctr; ∗∗ p < 0.001 vs. Ctr; # p < 0.001 vs. SOD1 10 min.

Article Snippet: Mutated form, SOD1 G93A , was provided by the “Recombinant Protein Service” of PRIMM srl (Milano, Italy).

Techniques: Activation Assay, Incubation, Control

Journal: Frontiers in Physiology

Article Title: Effect of Mutated Cu, Zn Superoxide Dismutase (SOD1 G93A ) on Modulation of Transductional Pathway Mediated by M1 Muscarinic Receptor in SK-N-BE and NSC-34 Cells

doi: 10.3389/fphys.2018.00611

Figure Lengend Snippet: Activation of ERK1-2 and AKT trasductional pathways in presence of SOD1 wt and SOD1 G93A in SK-N-BE cells. Western blotting analysis of P-ERK (A) and P-AKT (B) in cells incubated with 400 ng/ml of SOD1 wt and SOD1 G93A for 10 and 30 min. The histograms show the mean values (+SE) evaluated by densitometric analysis of three independent experiments. The results were normalized to ERK1-2 and AKT respectively. ∗ p < 0.05 vs. Ctr; ∗∗ p < 0.001 vs. Ctr; # p < 0.05 vs. 10 min SOD1 wt ; ## p < 0.001 vs. 10 min SOD1 wt ; ς p < 0.05 vs. 30 min SOD1 wt .

Article Snippet: Mutated form, SOD1 G93A , was provided by the “Recombinant Protein Service” of PRIMM srl (Milano, Italy).

Techniques: Activation Assay, Western Blot, Incubation

Journal: Frontiers in Physiology

Article Title: Effect of Mutated Cu, Zn Superoxide Dismutase (SOD1 G93A ) on Modulation of Transductional Pathway Mediated by M1 Muscarinic Receptor in SK-N-BE and NSC-34 Cells

doi: 10.3389/fphys.2018.00611

Figure Lengend Snippet: Activation of PLC-PKC-ERK1-2-AKT-dependent transduction pathway in presence of SOD1 wt and SOD1 G93A in NSC-34 cells. Western Blotting analysis of the levels of P-ERK 1-2 (A) and P-AKT (B) in NSC-34 cells incubated with 400 ng/ml of SOD1 wt and with 400 ng/ml of SOD1 G93A for times of 10 and 30 min. The data represent the means ± SEM of three independent experiments relative to control obtained by densitometric analysis of P-ERK1-2 and P-AKT protein bands normalized to ERK1-2 and AKT, respectively. ∗ p < 0.05 vs. Ctr; ∗∗ p < 0.001 vs. Ctr; # p < 0.05 vs. 10 min SOD1 wt ; ## p < 0.001 vs. 10 min SOD1 wt ; ς p < 0.05 vs. 30 min SOD1 wt .

Article Snippet: Mutated form, SOD1 G93A , was provided by the “Recombinant Protein Service” of PRIMM srl (Milano, Italy).

Techniques: Activation Assay, Transduction, Western Blot, Incubation, Control

Journal: Frontiers in Physiology

Article Title: Effect of Mutated Cu, Zn Superoxide Dismutase (SOD1 G93A ) on Modulation of Transductional Pathway Mediated by M1 Muscarinic Receptor in SK-N-BE and NSC-34 Cells

doi: 10.3389/fphys.2018.00611

Figure Lengend Snippet: Activation of PLC-PKC-ERK 1-2-AKT-dependent transduction pathway in presence of SOD1 wt and SOD1 G93A . Western Blotting analysis of the levels of P-ERK 1-2 and P-AKT in SK-N-BE (A,B) and NSC-34 (C,D) cells incubated for 10 min with 400 ng/ml of SOD1 wt and with 400 ng/ml of SOD1 G93A in absence and in presence of Pirenzepine 10 μM for 5 min. The data represent the means ± SEM of three independent experiments relative to control obtained by densitometric analysis of P-ERK1/2 and P-AKT protein bands normalized to ERK1-2 and AKT respectively. ∗ p < 0.05 vs. Ctr; ∗∗ p < 0.01 vs. Ctr; ∗∗∗ p < 0.001 vs. Ctr; # p < 0.05 vs. SOD1 wt ; ## p < 0.001 vs. SOD1 wt ; ° p < 0.01 vs. SOD1 G93A .

Article Snippet: Mutated form, SOD1 G93A , was provided by the “Recombinant Protein Service” of PRIMM srl (Milano, Italy).

Techniques: Activation Assay, Transduction, Western Blot, Incubation, Control

Journal: Frontiers in Physiology

Article Title: Effect of Mutated Cu, Zn Superoxide Dismutase (SOD1 G93A ) on Modulation of Transductional Pathway Mediated by M1 Muscarinic Receptor in SK-N-BE and NSC-34 Cells

doi: 10.3389/fphys.2018.00611

Figure Lengend Snippet: Fluorimetric Determination of Reactive oxygen species (ROS) levels. SK-N-BE (A) and NSC-34 (B) semiconfluent cells were incubated for 18 h in medium containing 0.2% FBS and then for 30 min with 400 ng/ml SOD1 wt and mutated SOD1 G93A . To determine Reactive Oxygen Species (ROS) levels, SK-N-BE (A) and NSC-34 (B) cells were incubated with 10 μM of the ROS sensitive probe DCHF-DA and ROS levels were measured by fluorometric analysis. The graphs on the left show the mean ± SEM values ( n = 6) relative to control of the indicated time points. The histograms on the right show the values of 30 min point and the relative ANOVA analysis. ∗ p < 0.05 vs. Ctr; ∗∗ p < 0.001 vs. Ctr; # p < 0.001 vs. SOD wt .

Article Snippet: Mutated form, SOD1 G93A , was provided by the “Recombinant Protein Service” of PRIMM srl (Milano, Italy).

Techniques: Incubation, Control

Journal: Frontiers in Physiology

Article Title: Effect of Mutated Cu, Zn Superoxide Dismutase (SOD1 G93A ) on Modulation of Transductional Pathway Mediated by M1 Muscarinic Receptor in SK-N-BE and NSC-34 Cells

doi: 10.3389/fphys.2018.00611

Figure Lengend Snippet: Effect of SOD1 wt and mutated SOD1 G93A on intracellular calcium levels in neuroblastoma SK-N-BE cells (A) and in NSC-34 cells (B) . The data are compared with control and with carbachol-induced intracellular calcium increase (positive control). The graphs on the left show the mean ± SEM values ( n = 6) relative to control of the indicated time points. The histograms on the right show the values of 10 s point and the relative ANOVA analysis. ∗ p < 0.001 vs. Ctr; # p < 0.01 vs. SOD wt .

Article Snippet: Mutated form, SOD1 G93A , was provided by the “Recombinant Protein Service” of PRIMM srl (Milano, Italy).

Techniques: Control, Positive Control

Journal: Frontiers in Physiology

Article Title: Effect of Mutated Cu, Zn Superoxide Dismutase (SOD1 G93A ) on Modulation of Transductional Pathway Mediated by M1 Muscarinic Receptor in SK-N-BE and NSC-34 Cells

doi: 10.3389/fphys.2018.00611

Figure Lengend Snippet: SK-N-BE and NSC-34 semiconfluent cells were preincubated with 15 μM of pirenzepine or with 10 μM of BAPTA-AM for 30 and 5 min respectively. Then, the cells were treated for 4 h with 400 ng/ ml of SOD1 wt or mutated SOD1 G93A . Induction of apoptosis by SOD1 G93A on SK-N-BE (A) and NSC-34 (B) cells is showed by Western Blotting analysis of cleaved form of PARP-1 protein bands. Evaluation of cell viability by trypan blue assay in SK-N-BE and NSC-34 cells are showed in (C,D) , respectively; the data are reported as per cent variation compared to control. The data represent the means ± SEM relative to control obtained by densitometric analysis of cleaved PARP protein bands normalized to α-tubulin of three independent experiments. ∗ p < 0.005 vs. Ctr; # p < 0.005 vs. SOD G93A .

Article Snippet: Mutated form, SOD1 G93A , was provided by the “Recombinant Protein Service” of PRIMM srl (Milano, Italy).

Techniques: Western Blot, Control

Journal: PLoS ONE

Article Title: EGFR Inhibitor Erlotinib Delays Disease Progression but Does Not Extend Survival in the SOD1 Mouse Model of ALS

doi: 10.1371/journal.pone.0062342

Figure Lengend Snippet: Kaplan-Meier survival plot showing the lifespan of the SOD1 Tg mice treated with vehicle (blue) or erlotinib (red). The lifespan of each mouse is represented by a black dot. All mice were included in the statistical analysis. Mice that were censored in the analysis because of a non-ALS death are represented by dots that are not accompanied by a lowering of the curve.

Article Snippet: SOD1 G93A high copy number transgenic mice (SOD1 Tg) and their non-transgenic littermates (SOD1 WT) were bred at Genentech.

Techniques:

Journal: PLoS ONE

Article Title: EGFR Inhibitor Erlotinib Delays Disease Progression but Does Not Extend Survival in the SOD1 Mouse Model of ALS

doi: 10.1371/journal.pone.0062342

Figure Lengend Snippet: Censoring information for survival study: animal n by cause of death/euthanasia for SOD1 Tg mice.

Article Snippet: SOD1 G93A high copy number transgenic mice (SOD1 Tg) and their non-transgenic littermates (SOD1 WT) were bred at Genentech.

Techniques: Injection

Journal: The Neuroscientist : a review journal bringing neurobiology, neurology and psychiatry

Article Title: Expanding axonal transcriptome brings new functions for axonally-synthesized proteins in health and disease

doi: 10.1177/1073858417712668

Figure Lengend Snippet: Axonal Transcriptome Profiles

Article Snippet: The reader should bear in mind that this is a rapidly evolving area and additional diseases and/or disease mechanisms linked to axonal protein synthesis will undoubtedly be uncovered in the near future. table ft1 table-wrap mode="anchored" t5 caption a7 Disease/Disorder mRNAs Altered Mechanism References Alzheimer disease ATF4 Aβ1-42 induced transcription and anterograde transport ( Baleriola and others 2014 ) Amyotrophic lateral sclerosis (ALS) Nefl Decreased transport with TDP-43 mutation ( Alami and others 2014 ) 271 decreased + 176 increased Altered axonal localization with TDP-43 A315T expression ( Rotem and others 2017 ) 80 decreased + 95 increased Altered axonal localization with SOD1 G93A expression APC mRNA ligands Altered axonal localization with FUS/TLS expression ( Yasuda and others 2017 ) Spinal Muscular Atrophy (SMA) β-actin, GAP43, Ca v 2.2 Decreased axonal levels with SMN depletion ( Fallini and others 2016 ; Jablonka and others 2007 ) Increased and decreased mRNAs SMN depletion from motor neurons substantially alters the axonal transcriptome ( Saal and others 2014 ) Neuropathic pain CREB1 Capsaicin and IL-6 injection increase intra-axonal translation through mTOR pathways ( Melemedjian and others 2010 ; Melemedjian and others 2013 ; Melemedjian and others 2014 ) Neurotropic viral infection ?

Techniques: Microarray, Cell Differentiation, Transduction, Knockdown, RNA Binding Assay, RNA Sequencing, Expressing, Control, Binding Assay

Journal: The Neuroscientist : a review journal bringing neurobiology, neurology and psychiatry

Article Title: Expanding axonal transcriptome brings new functions for axonally-synthesized proteins in health and disease

doi: 10.1177/1073858417712668

Figure Lengend Snippet: A, Where are mRNAs stored in axons? There is clear evidence for storage of mRNAs in axons, where they are somehow sequestered away from the translational machinery and kept dormant until needed. The mechanisms underlying this are not known. How does an RNP disengage from the motor proteins used for transport into axons and avoid translation of its mRNA? Binding of additional proteins to the RNP (1) or replacement of RNP proteins with other RBPs (2) are possible mechanisms. Additionally, there must be stimulus-dependent mechanisms to recruit axonal mRNAs from their storage site (3).

Article Snippet: The reader should bear in mind that this is a rapidly evolving area and additional diseases and/or disease mechanisms linked to axonal protein synthesis will undoubtedly be uncovered in the near future. table ft1 table-wrap mode="anchored" t5 caption a7 Disease/Disorder mRNAs Altered Mechanism References Alzheimer disease ATF4 Aβ1-42 induced transcription and anterograde transport ( Baleriola and others 2014 ) Amyotrophic lateral sclerosis (ALS) Nefl Decreased transport with TDP-43 mutation ( Alami and others 2014 ) 271 decreased + 176 increased Altered axonal localization with TDP-43 A315T expression ( Rotem and others 2017 ) 80 decreased + 95 increased Altered axonal localization with SOD1 G93A expression APC mRNA ligands Altered axonal localization with FUS/TLS expression ( Yasuda and others 2017 ) Spinal Muscular Atrophy (SMA) β-actin, GAP43, Ca v 2.2 Decreased axonal levels with SMN depletion ( Fallini and others 2016 ; Jablonka and others 2007 ) Increased and decreased mRNAs SMN depletion from motor neurons substantially alters the axonal transcriptome ( Saal and others 2014 ) Neuropathic pain CREB1 Capsaicin and IL-6 injection increase intra-axonal translation through mTOR pathways ( Melemedjian and others 2010 ; Melemedjian and others 2013 ; Melemedjian and others 2014 ) Neurotropic viral infection ?

Techniques: Binding Assay

Journal: Stem Cells International

Article Title: Human Motor Neuron Progenitor Transplantation Leads to Endogenous Neuronal Sparing in 3 Models of Motor Neuron Loss

doi: 10.4061/2011/207230

Figure Lengend Snippet: hMNP transplantation spares endogenous neurons. (a) Comparison of NeuN positive cells in cranial ( n = 13 hMNP; n = 15 vehicle) and caudal ( n = 14 hMNP; n = 15 vehicle) transverse sections in SCI animals. (b) Comparison of ChAT positive cells in cranial ( n = 10 hMNP; n = 12 vehicle) and caudal ( n = 9 hMNP; n = 12 vehicle) transverse sections of ventral horns in SCI animals. (c) Comparison of NeuN positive cells in cranial transverse sections of SOD1 G93A animals ( n = 5 hMNP; n = 5 vehicle). (d) Comparison of NeuN positive cells in cranial ( n = 3 hMNP; n = 3 vehicle) and caudal ( n = 3 hMNP; n = 2 vehicle) transverse sections of Δ7SMN animals. NeuN stained histological sections of lumbar spinal cord from hMNP (e) and vehicle (f) injected into SOD1 G93A mice clearly show a difference in neuronal numbers. Data is expressed as mean ± standard error. * P < .05. Bar = 50 μ m.

Article Snippet: The SOD1 G93A mutant mice were supplied and bred for ALS-TDI by GTC Biotherapeutics.

Techniques: Transplantation Assay, Comparison, Staining, Injection

Journal: Frontiers in Neurology

Article Title: Elevation of inositol pyrophosphate IP 7 in the mammalian spinal cord of amyotrophic lateral sclerosis

doi: 10.3389/fneur.2023.1334004

Figure Lengend Snippet: Elevated IP 7 level in the spinal cord of SOD1(G93A) TG mice in the ALS late stage. (A) Schematic illustration of the experimental workflow. SOD1(G93A) TG and their littermate WT mice at 12-week (before ALS onset), 15-week (ALS early-middle stage) and 18-week (ALS late stage) were sacrificed to harvest central nervous system (CNS) for LC-MS analysis. (B) The concentrations of IP 6 , IP 7 , and IP 7 /IP 6 (product-to-precursor) ratio in the cerebrum, cerebellum and spinal cord of SOD1(G93A) TG and their littermate WT mice. The values shown represent the mean ± SD of four independent experiments and are expressed relative to the WT mice. P -values calculated by Student's t -test are given in parenthesis. Asterisks indicate statistical significance ( p < 0.05) compared with WT mice. (C) Representative SRM chromatograms of IP 6 , IP 7 , and internal standard ITPP-d 6 in the spinal cord of SOD1(G93A) TG and their littermate WT mice. The arrowheads indicate the SRM peaks of the corresponding analytes. IS, internal standard.

Article Snippet: SOD1 (G93A) transgenic (TG) mice and littermate wild-type (WT) mice were obtained from Clea Japan (Tokyo, Japan) and maintained at an ambient temperature of 23 ± 2°C and humidity of 55 ± 15% with a 12 h light-dark cycle.

Techniques: Liquid Chromatography with Mass Spectroscopy