Search

A joint theoretical-experimental study on the optical properties of 5-N-succinimidyl-2.2 -bithiophene (NS-2T), a prototype system for a new class of biomarkers was conducted, and time-dependent density functional theory (TD-DFT) and approximate coupled-cluster single and doubles (CC2) calculations were performed in the ground and excited states. Results showed that CC2 and TD-DFT can very well describe the absorption and photoluminescence energies of the three systems, but the presence of several charge-transfer transitions in the TD-DFT spectrum of NS-2T required the use of a correlated method to validate the TD-DFT results.

Genetic expansions of the hexanucleotide repeats (GGGGCC) in the C9orf72 gene appear in approximately 40% of patients with familial ALS and 7% of patients with sporadic ALS in the European population, making this mutation one of the most prevalent genetic mutations in ALS. Here, we generated a human induced pluripotent stem cell (hiPSC) line from the dermal fibroblasts of a patient carrying a 56-repeat expansion in an ALS disease-causing allele of C9orf72. These iPSCs showed stable amplification in vitro with normal karyotype and high expression of pluripotent markers and differentiated spontaneously in vivo into three germ layers. [ABSTRACT FROM AUTHOR]

Background and purpose: The aim of our study was to describe patients with the p.D12Y variant (previously reported as D11Y) in SOD1 showing heterogeneous clinicopathological features. Methods: We performed clinical, electrophysiological, magnetic resonance imaging (MRI) and muscle pathology studies in four SOD1 p.D12Y variant‐positive patients. Results: The SOD1 p.D12Y clinical manifestations ranged from a benign phenotype characterized by distal distribution of muscular weakness and long survival to classic forms of amyotrophic lateral sclerosis with poor prognosis. Two patients with the distal clinical phenotype showed MRI and muscle pathology alterations indicating a concurrent muscle involvement. In one of these patients significant myopathic changes were associated with rimmed vacuolar pathology. Conclusions: We expand the clinical spectrum of SOD1 p.D12Y variant, including predominant lower motor neuron forms with long survival and classic forms with aggressive course. Some patients may have concomitant distal myopathy without other explanations. Given clinical, MRI and muscle pathology alterations, SOD1 should be considered in the differential diagnosis of molecularly undefined distal myopathies with rimmed vacuoles. [ABSTRACT FROM AUTHOR]

We have studied the optical properties of different thienyl-S,S-dioxide oligothiophenes under strong excitation. No stimulated emission is observed in neat films due to singlet-singlet bimolecular annihilation. The bimolecular recombination constant is quantified from a rate equation model in the limit of constant annihilation rate. We demonstrate that tunable stimulated emission in the range 490–660 nm, due to optical gain with an estimated cross section of the order of σg≈10-17 cm2, can be obtained by blending the active molecule with small quantities of inert polycarbonate poly(bisphenol-A-carbonate) (PC). The presence of amplified spontaneous emission (ASE) for a PC:active molecule relative concentration as small as 1:500 suggests that the polycarbonate role is to reduce the intermolecular diffusion rate rather than to isolate the active molecules. Moreover, for higher PC content, a continuous decrease of the bimolecular quenching role is observed. These results demonstrate that the absence of stimulated emission in neat films is not necessarily due to intrinsic molecular properties, as strong ASE can be obtained by slightly modifying the molecule interaction during the deposition process. This approach could allow a considerable extension of the number of molecules showing stimulated emission for organic laser applications. [ABSTRACT FROM AUTHOR]

We investigated the temperature dependence of the poly(9,9-dioctylfluorene) β phase photoluminescence (PL) spectra in spin coated thin films from tetrahydrofuran solutions. As the temperature increases from 18 to 300 K a continuous blueshift of the 0-0 PL peak of about 25 meV and an increase of the peak full width at half maximum (FWHM) of about 49 meV are observed. We show that the PL spectra temperature dependence is not due to a temperature dependent average conjugation length, as often assumed, but instead it can be quantitatively explained in the frame of a thermal quasiequilibrium model for excitons in an inhomogeneously broadened excited states distribution. We demonstrate that the emission blueshift and broadening are mainly due to the increase of the excitons’ temperature with the sample one. This effect is partially compensated by an increasing efficiency of the exciton energy migration. The interplay between these two processes quantitatively explains the observed temperature dependence of the PL peak energy and of its FWHM. On the contrary we show that the PL spectra are almost independent of the absorption blueshift with temperature. [ABSTRACT FROM AUTHOR]

The intramolecular radiative and nonradiative relaxation processes of three thiophene-S,S-dioxide derivatives with different molecular rigidity are investigated in different solutions and inert matrix. It is shown that the fluorescence quantum efficiency and the relaxation dynamics are strongly dependent on the environment viscosity, whereas they are almost independent of the environment polarity.

We demonstrate a static fabrication approach to make free-standing ordered arrays of fluorescent nanofibres through control of the transverse electrospinning field. The alignment and the density of the nanofibre arrays are optimised by careful design of both the source and collector electrode geometries which can control the transverse electric field over the full path of the jet. In doing so, we fabricate suspended fluorescent nanofibres with an aspect ratio of 104, and with a substantially increased density and order parameter (by a factor of ~10 compared to random deposition). Electrostatic modelling suggests that the field distribution of the component is the main contribution to the ordering between the plates. This method offers increased efficiency for the creation of ordered fibres collected over a small area and the characterisation of their photoluminescent properties. [ABSTRACT FROM AUTHOR]

Reversible oxygen induced emission quenching of both the Spontaneous Emission (SE) and the Amplified Spontaneous Emission (ASE) of poly(9,9-dioctylfluorene) waveguides is demonstrated. We showthat ASE shows a stronger quenching than SE, up to about 6.2 times, but also a stronger decrease when the excitation density increases. We conclude that the fast increase of the ASE decay rate is the main process in determining the ASE detection sensitivity, limiting the potentiality of sensitivity improvement of ASE with respect to SE. [ABSTRACT FROM AUTHOR]

Simultaneous photoluminescence (PL) and external quantum efficiency (EQE) confocal mapping is used to investigate the correlation between the local PL and the EQE in a regioregular poly(3-exylthiophene):poly(9,9-dioctylfluorene-co-benzothiadiazole) inverted bulk heterojunction solar cell. We show that the charge generation and charge collection are strongly non-uniform on a length scale up to 100 µm. Our results evidence that organic solar cells optimization requires not only the control of the submicrometric active materials arrangement but also the control of the large scale device uniformity. [ABSTRACT FROM AUTHOR]

The authors investigated the energy transfer between the chromophors of a white emitting polyspirobifluorene copolymer containing a blue emitting backbone, a green, and a red emitting dye. They demonstrate that the green dye is mainly excited by blue→green Förster resonant energy transfer (FRET), with a Förster radius R0bg=35 Å. The red dye is instead excited by direct blue→red FRET (R0br=21 Å) and by sequential blue→green→red FRET (R0gr=31 Å). Finally, the authors show that the FRET processes, fundamental to have white emission, are of intermolecular nature, while intramolecular energy migration and transfer are negligible. [ABSTRACT FROM AUTHOR]

We have studied the optical properties of a thienyl-S, S-dioxide oligothiophene under strong excitation. No stimulated emission is observed in neat films due to singlet-singlet bimolecular annihilation, with a bimolecular constant of A=(4.5±0.5)×10-11 cm3 s-1. On the contrary we show that strong stimulated emission is observed if the active molecule is blended to an inert polymer [poly(bisphenol A carbonate] (PC) even for a PC content as low as one PC molecule every 500 active molecules. We suggest that small PC quantities modify the film superamolecular packing, thus reducing the bimolecular recombination efficiency. These results are expected to be quite general and could allow a considerable extension of the number of molecules showing stimulated emission for organic laser applications. [ABSTRACT FROM AUTHOR]

We demonstrate that 20 nm thick indium tin oxide (ITO) layers deposited by pulsed laser deposition present sheet resistance as low as 130 Ω/□ and very uniform morphology, with an average roughness of about 0.4 nm, and peak-to-valley roughness as low as 8.2 nm. This good uniformity allowed us to realize a single layer polyfluorene active waveguide with both top and bottom ITO electrodes showing clear amplified spontaneous emission and electrode induced losses as low as 3.0 cm-1. We investigated the effects of hole injection in the ASE intensity concluding that complete gain suppression due to polaron absorption would take place for current density of about 360 mA cm-2. [ABSTRACT FROM AUTHOR]

We studied the microscopic dependence of poly(9,9-dioctylfluorene) photoluminescence (PL) on the deposition conditions. We show that in films spin coated from chloroform phase separation of β and glassy phases is present, with micrometric β phase clusters covering about 6% of the sample surface. The exposure to toluene vapors leads to the disappearance of the β phase clusters, but increases the β phase content in the films due to swelling induced polyfluorene chain planarization. The deposition from toluene solution leads to nonuniform PL intensity, dominated by the β phase emission, attributed to an interplay between aggregation during the solvent evaporation and solvent swelling induced chain planarization. [ABSTRACT FROM AUTHOR]

We report on random lasing in substituted quinquethienyl S,S-dioxide neat films. Despite the absence of highly efficient scatterers in the film, a fine structure with laser-like peaks as narrow as 5 Å is observed in the emission spectra. The far-field emission pattern is studied through angle-resolved emission measurements, demonstrating that random lasing emission is directional, with a 8° divergence but different individual emission patterns. The origin of the scattering centers providing the feedback for lasing has been analyzed through atomic force microscopy measurements of the film surface. We demonstrate that the random lasing is induced by sequential scattering from 50 nm diameter holes in the film with an average distance of 500 nm, while thickness fluctuations are not relevant. © 2003 American Institute of Physics. [ABSTRACT FROM AUTHOR]

A correction to the article "Interplay between stimulated emission and singlet singlet annihilation in oligothiophene dioxide thin films" which was published online on December 4, 2008 is presented.

One of the genetic mutations most associated with the onset of amyotrophic lateral sclerosis, both in sporadic and familial cases, is the expansion of the C9orf72 gene. The presence of more than 30 repeats (GGGGCC) correlates with uncertain ALS symptomatology. Here we collected a dermal biopsy from a subject carrying 36 hexanucleotide repeats and reprogrammed it into an induced pluripotent stem cell line. Despite the number of repeat elements, the subject had no symptoms at the age of the biopsy (76 years), thus resulting in a healthy carrier of the mutation., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Jessica Rosati reports financial support, article publishing charges, and equipment, drugs, or supplies were provided by Ministry of Health. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: In Amyotrophic Lateral Sclerosis (ALS) patients with SOD1 mutation the intrathecal administration of tofersen slowed down the progression of disease in a controlled clinical study, but results were not statistically significant., Methods: In this multicentre, observational study, we evaluated a cohort of 27 ALS-SOD1 patients who were treated with tofersen, focussing on 17 patients who were followed for at least 48 weeks (median period of 84 weeks, range 48-108). We compared the clinical slopes, as measured by ALSFRS-R, MRC scale and Forced Vital Capacity, during tofersen treatment with retrospective data at 1 year prior to therapy. Cerebrospinal fluid (CSF) and serum neurofilament light chains (NFL) were measured in all patients., Results: Cumulative evaluation of the ALSFRS-R and MRC progression rates showed a statistically significant change during treatment with respect to the period prior to therapy (p = 0.023 and p = 0.007, respectively). The analysis of individual patients showed that nine of the seventeen patients substantially stabilized or slightly improved. Four patients deteriorated during treatment, while in the remaining patients the very slow course did not allow to identify significant changes. CSF and serum NFL concentration markedly decreased in the near totality of patients. Increased levels of white blood cells and proteins in the CSF were found in 60% of patients. Such alterations were clinically asymptomatic in all but two patients who showed an acute pure motor radiculitis, which responded to steroid therapy., Conclusions: Clinical findings and NFL analysis strongly suggest that tofersen may have a disease-modifying effect in a subset of SOD1-ALS patients., (© 2024. Springer-Verlag GmbH Germany, part of Springer Nature.)

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease due to gradual motoneurons (MN) degeneration. Among the processes associated to ALS pathogenesis, there is the formation of cytoplasmic inclusions produced by aggregation of mutant proteins, among which the RNA binding protein FUS. Here we show that, in neuronal cells and in iPSC-derived MN expressing mutant FUS, such inclusions are significantly reduced in number and dissolve faster when the RNA m 6 A content is diminished. Interestingly, stress granules formed in ALS conditions showed a distinctive transcriptome with respect to control cells, which reverted to similar to control after m 6 A downregulation. Notably, cells expressing mutant FUS were characterized by higher m 6 A levels suggesting a possible link between m 6 A homeostasis and pathological aggregates. Finally, we show that FUS inclusions are reduced also in patient-derived fibroblasts treated with STM-2457, an inhibitor of METTL3 activity, paving the way for its possible use for counteracting aggregate formation in ALS., (© 2024. The Author(s).)

Introduction: High repeat expansion (HRE) alleles in C9orf72 have been linked to both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD); ranges for intermediate allelic expansions have not been defined yet, and clinical interpretation of molecular data lacks a defined genotype-phenotype association. In this study, we provide results from a large multicenter epidemiological study reporting the distribution of C9orf72 repeats in healthy elderly from the Italian population., Methods: A total of 967 samples were collected from neurologically evaluated healthy individuals over 70 years of age in the 13 institutes participating in the RIN (IRCCS Network of Neuroscience and Neurorehabilitation) based in Italy. All samples were genotyped using the AmplideXPCR/CE C9orf72 Kit (Asuragen, Inc.), using standardized protocols that have been validated through blind proficiency testing., Results: All samples carried hexanucleotide G 4 C 2 expansion alleles in the normal range. All samples were characterized by alleles with less than 25 repeats. In particular, 93.7% of samples showed a number of repeats ≤10, 99.9% ≤20 repeats, and 100% ≤25 repeats., Conclusion: This study describes the distribution of hexanucleotide G 4 C 2 expansion alleles in an Italian healthy population, providing a definition of alleles associated with the neurological healthy phenotype. Moreover, this study provides an effective model of federation between institutes, highlighting the importance of sharing genomic data and standardizing analysis techniques, promoting translational research. Data derived from the study may improve genetic counseling and future studies on ALS/FTD., Competing Interests: VS received compensation for consulting services and/or speaking activities from AveXis, Cytokinetics, Italfarmaco, Liquidweb S.r.l., Novartis Pharma AG, Zambon Biotech SA, and Asuragen. He receives or has received research supports form the Italian Ministry of Health, AriSLA, E-Rare Joint Transnational Call, and the ERN Euro-NMD. He is in the Editorial Board of Amyotrophic Lateral Sclerosis and Frontotemporal Degeneration, European Neurology, American Journal of Neurodegenerative Diseases, Frontiers in Neurology, and Exploration of Neuroprotective Therapy. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Giardina, Mandich, Ghidoni, Ticozzi, Rossi, Fenoglio, Tiziano, Esposito, Capellari, Nacmias, Mineri, Campopiano, Di Pilla, Sammarone, Zampatti, Peconi, De Angelis, Palmieri, Galandra, Nicodemo, Origone, Gotta, Ponti, Nicsanu, Benussi, Peverelli, Ratti, Ricci, Di Fede, Magri, Serpente, Lattante, Domi, Carrera, Saltimbanco, Bagnoli, Ingannato, Albanese, Tagliavini, Lodi, Caltagirone, Gambardella, Valente and Silani.)

Researchers studying Amyotrophic Lateral Sclerosis (ALS) have made significant efforts to find a unique mechanism to explain the etiopathology of the different forms of the disease. However, despite several mutations associated with ALS having been discovered in recent years, the link between the mutated genes and its onset has not yet been fully elucidated. Among the genes associated with ALS, superoxide dismutase 1 (SOD1) was the first to be identified, but its role in the etiopathogenesis of the disease is still unclear. In recent years, research has been focused on the non-coding part of the genome to fully understand the mechanisms underlying gene regulation. Non-coding RNAs are conserved molecules and are not usually translated in protein. A total of 98% of the human genome is composed of non-protein coding sequences with roles in the transcriptional and post-transcriptional regulation of gene expression. In this study, we characterized a divergent nuclear lncRNA (SOD1-DT) transcribed in the antisense direction from the 5' region of the SOD1 coding gene in both the SH-SY5Y cell line and fibroblasts derived from ALS patients. Interestingly, this lncRNA seems to regulate gene expression, since its inhibition leads to the upregulation of surrounding genes including SOD1. SOD1-DT represents a very complex molecule, displaying allelic and transcriptional variability concerning transposable elements (TEs) included in its sequence, widening the scenario of gene expression regulation in ALS disease.

Background and Objectives: Genetic variants in the gene TARDBP, encoding TDP-43 protein, are associated with amyotrophic lateral sclerosis (ALS) in familial (fALS) and sporadic (sALS) cases. Objectives of this study were to assess the contribution of TARDBP in a large cohort of Italian ALS patients, to determine the TARDBP-associated clinical features and to look for genotype-phenotype correlation and penetrance of the mutations., Methods: A total of 1992 Italian ALS patients (193 fALS and 1799 sALS) were enrolled in this study. Sanger sequencing of TARDBP gene was performed in patients and, when available, in patients' relatives., Results: In total, 13 different rare variants were identified in 43 index cases (10 fALS and 33 sALS) with a cumulative mutational frequency of 2.2% (5.2% of fALS, 1.8% of sALS). The most prevalent variant was the p.A382T followed by the p.G294V. Cognitive impairment was detected in almost 30% of patients. While some variants, including the p.G294V and the p.G376D, were associated with restricted phenotypes, the p.A382T showed a marked clinical heterogeneity regarding age of onset, survival and association with cognitive impairment. Investigations in parents, when possible, showed that the variants were inherited from healthy carriers and never occurred de novo., Conclusions: In our cohort, TARDBP variants have a relevant frequency in Italian ALS patients and they are significantly associated with cognitive impairment. Clinical presentation is heterogeneous. Consistent genotype-phenotype correlations are limited to some mutations. A marked phenotypic variability characterizes the p.A382T variant, suggesting a multifactorial/oligogenic pathogenic mechanism., (© 2023 The Authors. European Journal of Neurology published by John Wiley & Sons Ltd on behalf of European Academy of Neurology.)

Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disease caused by a complex interaction of genetic and environmental factors. Recently, a polymorphic intronic CA repeat in STMN2 gene has been proposed as risk factor for ALS. The presence of long/long CA genotype, especially if one allele had 24 CA, was reported to be significantly associated with the disease in a cohort of sporadic ALS patients. We tested an Italian cohort of 366 ALS patients and 353 healthy controls and we found no association between CA length and ALS risk.

Amyotrophic lateral sclerosis is a neurodegenerative disease characterized by the degeneration of motor neurons. There is no treatment for this disease that affects the ability to move, eat, speak and finally breathe, causing death. In an Italian family, a heterozygous pathogenic missense variant has been previously discovered in Exon 6 of the gene TARDBP encoding the TAR DNA-binding protein 43 protein. Here, we developed a potential therapeutic tool based on allele-specific small interfering RNAs for familial amyotrophic lateral sclerosis with the heterozygous missense mutation c.1127G>A. We designed a small interfering RNA that was able to diminish specifically the expression of the exogenous Green Fluorescent Protein (TAR DNA-binding protein 43 G376D mutant protein) in HEK-293T cells but not that of the Green Fluorescent Protein (TAR DNA-binding protein 43 wild-type). Similarly, this small interfering RNA silenced the mutated allele in fibroblasts derived from patients with amyotrophic lateral sclerosis but did not silence the wild-type gene in control fibroblasts. In addition, we established that silencing the mutated allele was able to strongly reduce the pathological cellular phenotypes induced by TAR DNA-binding protein 43 G376D expression, such as the presence of cytoplasmic aggregates. Thus, we have identified a small interfering RNA that could be used to silence specifically the mutated allele to try a targeted therapy for patients carrying the p.G376D TAR DNA-binding protein 43 mutation., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Guarantors of Brain.)

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease that selectively affects motor neurons. In 20% of cases, ALS appears in comorbidity with frontotemporal dementia (FTD). We generated patient-derived-induced Pluripotent Stem Cells (iPSCs), from an ALS/FTD patient. The patient had a familial form of the disease and a missense variant in TARDBP gene. We used an established protocol based on Sendai virus to reprogram fibroblasts. We confirmed the stemness and the pluripotency of the iPSC clones, thus generating embryoid bodies. We believe that the iPSC line carrying a TARDBP mutation could be a valuable tool to investigate TDP-43 proteinopathy linked to ALS., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the loss of the upper and lower motor neurons (MNs). About 10% of patients have a family history (familial, fALS); however, most patients seem to develop the sporadic form of the disease (sALS). SOD1 (Cu/Zn superoxide dismutase-1) is the first studied gene among the ones related to ALS. Mutant SOD1 can adopt multiple misfolded conformation, lose the correct coordination of metal binding, decrease structural stability, and form aggregates. For all these reasons, it is complicated to characterize the conformational alterations of the ALS-associated mutant SOD1, and how they relate to toxicity. In this work, we performed a multilayered study on fibroblasts derived from two ALS patients, namely SOD1 L145F and SOD1 S135N , carrying the p.L145F and the p.S135N missense variants, respectively. The patients showed diverse symptoms and disease progression in accordance with our bioinformatic analysis, which predicted the different effects of the two mutations in terms of protein structure. Interestingly, both mutations had an effect on the fibroblast energy metabolisms. However, while the SOD1 L145F fibroblasts still relied more on oxidative phosphorylation, the SOD1 S135N fibroblasts showed a metabolic shift toward glycolysis. Our study suggests that SOD1 mutations might lead to alterations in the energy metabolism.

Background: Koolen-de Vries syndrome (KdVS) is a multisystem neurodevelopmental disorder caused by 17q21.31 deletions or mutations in KANSL1 . It was mainly described in children., Methods: A retrospective study on 9 subjects aged 19-45 years and revision of 18 literature patients, with the purpose to get insights into the phenotypic evolution with time, and into the clinical manifestations in adulthood., Results: Seven patients had a 17q21.31 deletion and two a point mutation in KANSL1 . All had intellectual disability, which was mild in five (56%) and moderate in four (44%). Epilepsy was diagnosed in four subjects (44%), with onset from 1 to 7 years and full remission before 9 years in 3/4 patients. Scoliosis affected seven individuals (77.7%) and it was substantially stable with age in 5/7 patients, allowing for simple daily activities. Two subjects had severely progressive scoliosis, which was surgically corrected. Overweight or true obesity did occur after puberty in six patients (67%). Behaviour abnormalities were recorded in six patients (67%). The facial phenotype slightly evolved with time to include thick eyebrows, elongated nose and pronounced pointed chin. Despite behaviour abnormalities, happy disposition and sociable attitudes were common. Half of patients had fluent language and were good at writing and reading. Rich language, although limited to single words or short sentences, and very limited or absent skills in writing and reading were observed in the remaining patients. Autonomy in daily activities and personal care was usually limited., Conclusions: Distinctive features in adult KdVS subjects include intellectual disability, overweight/obesity, behaviour abnormalities with preserved social interest, ability in language, slight worsening of the facial phenotype and no seizures., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. No commercial re-use. See rights and permissions. Published by BMJ.)

Mutations in FUS and TBK1 often cause aggressive early-onset amyotrophic lateral sclerosis (ALS) or a late-onset ALS and/or frontotemporal dementia (FTD) phenotype, respectively. Co-occurrence of mutations in two or more Mendelian ALS/FTD genes has been repeatedly reported. However, little is known how two pathogenic ALS/FTD mutations in the same patient interact to shape the final phenotype. We screened 28 ALS patients with a known FUS mutation by whole-exome sequencing and targeted evaluation for mutations in other known ALS genes followed by genotype-phenotype correlation analysis of FUS/TBK1 double-mutant patients. We report on new and summarize previously published FUS and TBK1 double-mutant ALS/FTD patients and their families. We found that, within a family, mutations in FUS cause ALS while TBK1 single mutations are observed in FTD patients. FUS/TBK1 double mutations manifested as ALS and without a manifest difference regarding age at onset and disease duration when compared to FUS single-mutant individuals. In conclusion, TBK1 and FUS variants do not seem to interact in a simple additive way. Rather, the phenotype of FUS/TBK1 double-mutant patients appears to be dominated by the FUS mutation., (© 2021. The Author(s).)

The G4C2-repeat expansion in C9orf72 is the most common cause of frontotemporal dementia and of amyotrophic lateral sclerosis. The variability of age at onset and phenotypic presentations is a hallmark of C9orf72 disease. In this study, we aimed to identify modifying factors of disease onset in C9orf72 carriers using a family-based approach, in pairs of C9orf72 carrier relatives with concordant or discordant age at onset. Linkage and association analyses provided converging evidence for a locus on chromosome Xq27.3. The minor allele A of rs1009776 was associated with an earlier onset (P = 1 × 10-5). The association with onset of dementia was replicated in an independent cohort of unrelated C9orf72 patients (P = 0.009). The protective major allele delayed the onset of dementia from 5 to 13 years on average depending on the cohort considered. The same trend was observed in an independent cohort of C9orf72 patients with extreme deviation of the age at onset (P = 0.055). No association of rs1009776 was detected in GRN patients, suggesting that the effect of rs1009776 was restricted to the onset of dementia due to C9orf72. The minor allele A is associated with a higher SLITRK2 expression based on both expression quantitative trait loci (eQTL) databases and in-house expression studies performed on C9orf72 brain tissues. SLITRK2 encodes for a post-synaptic adhesion protein. We further show that synaptic vesicle glycoprotein 2 and synaptophysin, two synaptic vesicle proteins, were decreased in frontal cortex of C9orf72 patients carrying the minor allele. Upregulation of SLITRK2 might be associated with synaptic dysfunctions and drives adverse effects in C9orf72 patients that could be modulated in those carrying the protective allele. How the modulation of SLITRK2 expression affects synaptic functions and influences the disease onset of dementia in C9orf72 carriers will require further investigations. In summary, this study describes an original approach to detect modifier genes in rare diseases and reinforces rising links between C9orf72 and synaptic dysfunctions that might directly influence the occurrence of first symptoms., (© The Author(s) (2021). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Importance: Juvenile amyotrophic lateral sclerosis (ALS) is a rare form of ALS characterized by age of symptom onset less than 25 years and a variable presentation., Objective: To identify the genetic variants associated with juvenile ALS., Design, Setting, and Participants: In this multicenter family-based genetic study, trio whole-exome sequencing was performed to identify the disease-associated gene in a case series of unrelated patients diagnosed with juvenile ALS and severe growth retardation. The patients and their family members were enrolled at academic hospitals and a government research facility between March 1, 2016, and March 13, 2020, and were observed until October 1, 2020. Whole-exome sequencing was also performed in a series of patients with juvenile ALS. A total of 66 patients with juvenile ALS and 6258 adult patients with ALS participated in the study. Patients were selected for the study based on their diagnosis, and all eligible participants were enrolled in the study. None of the participants had a family history of neurological disorders, suggesting de novo variants as the underlying genetic mechanism., Main Outcomes and Measures: De novo variants present only in the index case and not in unaffected family members., Results: Trio whole-exome sequencing was performed in 3 patients diagnosed with juvenile ALS and their parents. An additional 63 patients with juvenile ALS and 6258 adult patients with ALS were subsequently screened for variants in the SPTLC1 gene. De novo variants in SPTLC1 (p.Ala20Ser in 2 patients and p.Ser331Tyr in 1 patient) were identified in 3 unrelated patients diagnosed with juvenile ALS and failure to thrive. A fourth variant (p.Leu39del) was identified in a patient with juvenile ALS where parental DNA was unavailable. Variants in this gene have been previously shown to be associated with autosomal-dominant hereditary sensory autonomic neuropathy, type 1A, by disrupting an essential enzyme complex in the sphingolipid synthesis pathway., Conclusions and Relevance: These data broaden the phenotype associated with SPTLC1 and suggest that patients presenting with juvenile ALS should be screened for variants in this gene.

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease affecting upper and lower motor neurons. We generated patient-derived-induced Pluripotent Stem Cells (iPSCs), from an ALS patient affected by an early-onset and aggressive form of the disease, carrying a missense pathogenic variant in FUS gene. We reprogrammed somatic cells using an established Sendai virus protocol and we obtained clones of iPSC. We confirmed their stemness and further generated embryoid bodies, showing their potential of differentiating in all three germ layers. This iPSC line, carrying a pathogenic FUS variant, is a valuable tool to deeply investigate pathogenic mechanisms leading to ALS., (Copyright © 2021. Published by Elsevier B.V.)

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.

Amyotrophic lateral sclerosis (ALS) is an incurable neurodegenerative condition with phenotypic and genetic heterogeneity. It is characterized by the selective vulnerability and the progressive loss of the neural population. Here, an induced pluripotent stem cell (iPSC) line was generated from dermal fibroblasts of an individual carrying the p.G376D mutation in the TDP-43 protein. Fibroblasts were reprogrammed using non-integrating episomal plasmids. There were no karyotype abnormalities, and iPSCs successfully differentiated into all three germ layers. This cell line may prove useful in the study of the pathogenic mechanisms that underpin ALS syndrome., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

In the last few years, NEK1 has been identified as a new gene related to amyotrophic lateral sclerosis (ALS). Loss-of-function variants have been mostly described, although several missense variants exist, which pathogenic relevance remains to be established. We attempted to determine the contribution of NEK1 gene in an Italian cohort of 531 sporadic and familial amyotrophic lateral sclerosis (ALS) patients applying massive parallel sequencing technologies. We filtered results of NEK1 gene and identified 20 NEK1 rare variants (MAF < 0.01) in 22 patients. In particular, we found two novel frameshift variants (p.Glu929Asnfs*12 and p.Val1030Ilefs*23), 18 missense variants, including the p.Arg261His in three patients, and a novel variant in the start codon, the p.Met1?, which most likely impairs translation initiation. To clarify the role of NEK1 missense variants we investigated NEK1 expression in primary fibroblast cultures. We obtained skin biopsies from four patients with NEK1 variants and we assessed NEK1 expression by western blot and immunofluorescence. We detected a decrease in NEK1 expression in fibroblasts from patients with NEK1 variants, suggesting that missense variants in NEK1 gene may have a pathogenic role. Moreover, we observed additional variants in ALS related genes in seven patients with NEK1 variants (32%), further supporting an oligogenic ALS model., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The development of high-throughput sequencing technologies and screening of big patient cohorts with familial and sporadic amyotrophic lateral sclerosis (ALS) led to the identification of a significant number of genetic variants, which are sometimes difficult to interpret. The American College of Medical Genetics and Genomics (ACMG) provided guidelines to help molecular geneticists and pathologists to interpret variants found in laboratory testing. We assessed the application of the ACMG criteria to ALS-related variants, combining data from literature with our experience. We analyzed a cohort of 498 ALS patients using massive parallel sequencing of ALS-associated genes and identified 280 variants with a minor allele frequency < 1%. Examining all variants using the ACMG criteria, thus considering the type of variant, inheritance, familial segregation, and possible functional studies, we classified 20 variants as "pathogenic". In conclusion, ALS's genetic complexity, such as oligogenic inheritance, presence of genes acting as risk factors, and reduced penetrance, needs to be considered when interpreting variants. The goal of this work is to provide helpful suggestions to geneticists and clinicians dealing with ALS.

The main hallmark of many forms of familiar and sporadic amyotrophic lateral sclerosis (ALS) is a reduction in nuclear TDP-43 protein and its inclusion in cytoplasmic aggregates in motor neurons. In order to understand which cellular and molecular mechanisms underlie the mislocalization of TDP-43, we examined human skin fibroblasts from two individuals with familial ALS, both with mutations in TDP-43, and two individuals with sporadic ALS, both without TDP-43 mutations or mutations in other ALS related genes. We found that all ALS fibroblasts had a partially cytoplasmic localization of TDP-43 and had reduced cell metabolism as compared to fibroblasts from apparently healthy individuals. ALS fibroblasts showed an increase in global protein synthesis and an increase in 4E-BP1 and rpS6 phosphorylation, which is indicative of mTORC1 activity. We also observed a decrease in glutathione (GSH), which suggests that oxidative stress is elevated in ALS. ERK1/2 activity regulated the extent of oxidative stress and the localization of TDP-43 in the cytoplasm in all ALS fibroblasts. Lastly, ALS fibroblasts showed reduced stress granule formation in response to H 2 O 2 stress. In conclusion, these findings identify specific cellular and molecular defects in ALS fibroblasts, thus providing insight into potential mechanisms that may also occur in degenerating motor neurons., Competing Interests: Declaration of Competing Interest The authors declare they have no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Variants in tank-binding kinase 1 (TBK1) are responsible for a significant proportion of amyotrophic lateral sclerosis (ALS) cases. In the present study, we analyzed variants in TBK1 extracted by targeted sequencing of 32 genes in a group of 406 Italian patients with ALS. We identified 7 different TBK1 variants in 7 sporadic cases, resulting in a frequency of 1.7%. Three patients had missense variants (p.R357Q, p.R358H, and p.R724C), one patient had a small deletion (p.E618del), and 3 had truncating variants (p.Y482*, p.R229*, and p.N681*). Notably, we found that 4 patients had an additional variant in ALS-related genes: 2 in OPTN and 2 in the 3'UTR region of FUS. By studying an independent group of 7 TBK1-mutated patients previously reported, we found another variant in the 3'UTR region of FUS in one patient. The presence of a second variant in TBK1 variant carriers is an interesting finding that needs to be investigated in larger cohorts of patients. These findings suggest that TBK1 belongs to the category of genes conferring a significantly increased risk but not sufficient to cause disease., (Copyright © 2019 Elsevier Inc. All rights reserved.)

S100A4, belonging to a large multifunctional S100 protein family, is a Ca 2+ -binding protein with a significant role in stimulating the motility of cancer and immune cells, as well as in promoting pro-inflammatory properties in different cell types. In the CNS, there is limited information concerning S100A4 presence and function. In this study, we analyzed the expression of S100A4 and the effect of the S100A4 transcriptional inhibitor niclosamide in murine activated primary microglia. We found that S100A4 was strongly up-regulated in reactive microglia and that niclosamide prevented NADPH oxidase 2, mTOR (mammalian target of rapamycin), and NF-κB (nuclear factor-kappa B) increase, cytoskeletal rearrangements, migration, and phagocytosis. Furthermore, we found that S100A4 was significantly up-regulated in astrocytes and microglia in the spinal cord of a transgenic rat SOD1-G93A model of amyotrophic lateral sclerosis. Finally, we demonstrated the increased expression of S100A4 also in fibroblasts derived from amyotrophic lateral sclerosis (ALS) patients carrying SOD1 pathogenic variants. These results ascribe S100A4 as a marker of microglial reactivity, suggesting the contribution of S100A4-regulated pathways to neuroinflammation, and identify niclosamide as a possible drug in the control and attenuation of reactive phenotypes of microglia, thus opening the way to further investigation for a new application in neurodegenerative conditions.

Background: Activating pathogenic variants in PIK3CA gene usually occur at a mosaic status and underlie a variety of segmental overgrowth phenotypes. Germline variants in PIK3CA have been rarely reported, described in a total of 12 patients with macrocephaly to date. Clinical and prognostic features of these germline variants have not been described in detail yet., Methods: Targeted deep sequencing by custom panel of the 21 genes involved in the PI3K/AKT/mTOR pathway was performed in a 13-year-old boy with macrocephaly and physical overgrowth. PI3K/AKT/mTOR pathway analysis was performed in fibroblasts by Western blot. The effects of miransertib (AKT inhibitor) and rapamycin (mTOR inhibitor) were assessed., Results: A de novo pathogenic variant (c.1090G>C; p.Gly364Arg) in PIK3CA gene was detected in a non-mosaic status in peripheral blood cells, buccal smears, and skin fibroblasts. Increased levels of phosphorylated AKT residues were observed in fibroblasts, rescued by miransertib., Conclusion: Germline variants in PIK3CA are associated to a mild phenotype characterized by overgrowth, severe macrocephaly, mild intellectual disability, and few dysmorphic features. Investigations of PI3K/AKT/mTOR pathway should be performed in patients with severe macrocephaly and unspecific physical overgrowth. Longitudinal studies to assess prognosis and cancer predisposition are recommended., (© 2019 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.)

A (GGGGCC) n repeat expansion in C9orf72 gene is the major cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). The relations between the repeats size and the age at disease onset (AO) or the clinical phenotype (FTD vs. ALS) were investigated in 125 FTD, ALS, and presymptomatic carriers. Positive correlations were found between repeats number and the AO (p < 10 e-4 ) but our results suggested that the association was mainly driven by age at collection (p < 10 e-4 ). A weaker association was observed with clinical presentation (p = 0.02), which became nonsignificant after adjustment for the age at collection in each group. Importantly, repeats number variably expanded or contracted over time in carriers with multiple blood samples, as well as through generations in parent-offspring pairs, conversely to what occurs in several expansion diseases with anticipation at the molecular level. Finally, this study establishes that measure of repeats number in lymphocytes is not a reliable biomarker predictive of the AO or disease outcome in C9orf72 long expansion carriers., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

Fibroblasts isolated from an Amyotrophic Lateral Sclerosis (ALS)-patient carrying a mutation in Matrin-3 (p.Q66K -MATR3) gene were reprogrammed to the pluripotency stage by using non-integrating episomal plasmids. We generated the Q66K#44DRM induced pluripotent stem cell (iPSC) line that showed regular karyotype, expressed pluripotency-associated markers and were able to properly differentiate into the three germ layers. The heterozygous missense mutation in the MATR3 gene (p.Q66K), which is associated to ALS disease, was present in the generated iPSC line. Resource table., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

The diverse clinical phenotypes of Wolf-Hirschhorn syndrome (WHS) are the result of haploinsufficiency of several genes, one of which, LETM1 , encodes a protein of the mitochondrial inner membrane of uncertain function. Here, we show that LETM1 is associated with mitochondrial ribosomes, is required for mitochondrial DNA distribution and expression, and regulates the activity of an ancillary metabolic enzyme, pyruvate dehydrogenase. LETM1 deficiency in WHS alters mitochondrial morphology and DNA organization, as does substituting ketone bodies for glucose in control cells. While this change in nutrient availability leads to the death of fibroblasts with normal amounts of LETM1, WHS-derived fibroblasts survive on ketone bodies, which can be attributed to their reduced dependence on glucose oxidation. Thus, remodeling of mitochondrial nucleoprotein complexes results from the inability of mitochondria to use specific substrates for energy production and is indicative of mitochondrial dysfunction. However, the dysfunction could be mitigated by a modified diet-for WHS, one high in lipids and low in carbohydrates., (© 2018 The Authors. Published under the terms of the CC BY 4.0 license.)

To clarify the possible involvement of intermediate ATXN1 alleles as risk factors for amyotrophic lateral sclerosis (ALS), we tested ATXN1 in a cohort of 1146 Italian ALS patients, previously screened for variants in other ALS genes, and in 529 controls. We detected ATXN1 alleles with ≥33 polyglutamine repeats in 105 of 1146 patients (9.16%) and 29 of 529 controls (5.48%) (p = 0.003). The frequency of ATXN1 alleles with ≥33 polyglutamine repeats was particularly high in the group of ALS patients carrying the C9orf72 expansion (12/59, 20.3%). We confirmed this result in an independent cohort of C9orf72 Italian patients (10/80 cases, 12.5%), thus finding a cumulative frequency of ATXN1 expansion of 15.82% in C9orf72 carriers (p = 2.40E-05). Our results strongly support the hypothesis that ATXN1 could act as a disease risk gene in ALS, mostly in C9orf72 expansion carriers. Further studies are needed to confirm our results and to define the mechanism by which ATXN1 might contribute to neuronal degeneration leading to ALS., (Copyright © 2017 Elsevier Inc. All rights reserved.)