Serum neurofilament light chain in distinct phenotypes of amyotrophic lateral sclerosis: A longitudinal, multicenter study.

OBJECTIVE: To assess the performance of serum neurofilament light chain (sNfL) in clinical phenotypes of amyotrophic lateral sclerosis (ALS). METHODS: In 2949 ALS patients at 16 ALS centers in Germany and Austria, clinical characteristics and sNfL were assessed. Phenotypes were differentiated for two anatomical determinants: (1) upper and/or lower motor involvement (typical, typMN; upper/lower motor neuron predominant, UMNp/LMNp; primary lateral sclerosis, PLS) and (2) region of onset and propagation of motor neuron dysfunction (bulbar, limb, flail-arm, flail-leg, thoracic onset). Phenotypes were correlated to sNfL, progression, and survival. RESULTS: Mean sNfL was - compared to typMN (75.7 pg/mL, n = 1791) - significantly lower in LMNp (45.1 pg/mL, n = 413), UMNp (58.7 pg/mL n = 206), and PLS (37.6 pg/mL, n = 84). Also, sNfL significantly differed in the bulbar (92.7 pg/mL, n = 669), limb (64.1 pg/mL, n = 1305), flail-arm (46.4 pg/mL, n = 283), flail-leg (53.6 pg/mL, n = 141), and thoracic (74.5 pg/mL, n = 96) phenotypes. Binary logistic regression analysis showed highest contribution to sNfL elevation for faster progression (odds ratio [OR] 3.24) and for the bulbar onset phenotype (OR 1.94). In contrast, PLS (OR 0.20), LMNp (OR 0.45), and thoracic onset (OR 0.43) showed reduced contributions to sNfL. Longitudinal sNfL (median 12 months, n = 2862) showed minor monthly changes (<0.2%) across all phenotypes. Correlation of sNfL with survival was confirmed (p < 0.001). CONCLUSIONS: This study underscored the correlation of ALS phenotypes - differentiated for motor neuron involvement and region of onset/propagation - with sNfL, progression, and survival. These phenotypes demonstrated a significant effect on sNfL and should be recognized as independent confounders of sNfL analyses in ALS trials and clinical practice.

Performance of serum neurofilament light chain in a wide spectrum of clinical courses of amyotrophic lateral sclerosis-a cross-sectional multicenter study.

BACKGROUND AND PURPOSE: The objective was to assess the performance of serum neurofilament light chain (sNfL) in amyotrophic lateral sclerosis (ALS) in a wide range of disease courses, in terms of progression, duration and tracheostomy invasive ventilation (TIV). METHODS: A prospective cross-sectional study at 12 ALS centers in Germany was performed. sNfL concentrations were age adjusted using sNfL Z scores expressing the number of standard deviations from the mean of a control reference database and correlated to ALS duration and ALS progression rate (ALS-PR), defined by the decline of the ALS Functional Rating Scale. RESULTS: In the total ALS cohort (n = 1378) the sNfL Z score was elevated (3.04; 2.46-3.43; 99.88th percentile). There was a strong correlation of sNfL Z score with ALS-PR (p < 0.001). In patients with long (5-10 years, n = 167) or very long ALS duration (>10 years, n = 94) the sNfL Z score was significantly lower compared to the typical ALS duration of <5 years (n = 1059) (p < 0.001). Furthermore, in patients with TIV, decreasing sNfL Z scores were found in correlation with TIV duration and ALS-PR (p = 0.002; p < 0.001). CONCLUSIONS: The finding of moderate sNfL elevation in patients with long ALS duration underlined the favorable prognosis of low sNfL. The strong correlation of sNfL Z score with ALS-PR strengthened its value as progression marker in clinical management and research. The lowering of sNfL in correlation with long TIV duration could reflect a reduction either in disease activity or in the neuroaxonal substrate of biomarker formation during the protracted course of ALS.

Small and long RNA transcriptome of whole human cerebrospinal fluid and serum as compared to their extracellular vesicle fractions reveal profound differences in expression patterns and impacts on biological processes.

BACKGROUND: Next generation sequencing (NGS) of human specimen is expected to improve prognosis and diagnosis of human diseases, but its sensitivity urges for well-defined sampling and standardized protocols in order to avoid error-prone conclusions. METHODS: In this study, large volumes of pooled human cerebrospinal fluid (CSF) were used to prepare RNA from human CSF-derived extracellular vesicles (EV) and from whole CSF, as well as from whole human serum and serum-derived EV. In all four fractions small and long coding and non-coding RNA expression was analyzed with NGS and transcriptome analyses. RESULTS: We show, that the source of sampling has a large impact on the acquired NGS pattern, and differences between small RNA fractions are more distinct than differences between long RNA fractions. The highest percentual discrepancy between small RNA fractions and the second highest difference between long RNA fractions is seen in the comparison of CSF-derived EV and whole CSF. Differences between miR (microRNA) and mRNA fractions of EV and the respective whole body fluid have the potential to affect different cellular and biological processes. I.e. a comparison of miR in both CSF fractions reveals that miR from EV target four transcripts sets involved in neurobiological processes, whereas eight others, also involved in neurobiological processes are targeted by miR found in whole CSF only. Likewise, three mRNAs sets derived from CSF-derived EV are associated with neurobiological and six sets with mitochondrial metabolism, whereas no such mRNA transcript sets are found in the whole CSF fraction. We show that trace amounts of blood-derived contaminations of CSF can bias RNA-based CSF diagnostics. CONCLUSIONS: This study shows that the composition of small and long RNA differ significantly between whole body fluid and its respective EV fraction and thus can affect different cellular and molecular functions. Trace amounts of blood-derived contaminations of CSF can bias CSF analysis. This has to be considered for a meaningful RNA-based diagnostics. Our data imply a transport of EV from serum to CSF across the blood-brain barrier.

The Role of Axonal Transport in Glaucoma.

Glaucoma is a neurodegenerative disease that affects the retinal ganglion cells (RGCs) and leads to progressive vision loss. The first pathological signs can be seen at the optic nerve head (ONH), the structure where RGC axons leave the retina to compose the optic nerve. Besides damage of the axonal cytoskeleton, axonal transport deficits at the ONH have been described as an important feature of glaucoma. Axonal transport is essential for proper neuronal function, including transport of organelles, synaptic components, vesicles, and neurotrophic factors. Impairment of axonal transport has been related to several neurodegenerative conditions. Studies on axonal transport in glaucoma include analysis in different animal models and in humans, and indicate that its failure happens mainly in the ONH and early in disease progression, preceding axonal and somal degeneration. Thus, a better understanding of the role of axonal transport in glaucoma is not only pivotal to decipher disease mechanisms but could also enable early therapies that might prevent irreversible neuronal damage at an early time point. In this review we present the current evidence of axonal transport impairment in glaucomatous neurodegeneration and summarize the methods employed to evaluate transport in this disease.

Treatment expectations and perception of therapy in adult patients with spinal muscular atrophy receiving nusinersen.

BACKGROUND AND PURPOSE: This was an investigation of treatment expectations and of the perception of therapy in adult patients with 5q-associated spinal muscular atrophy (5q-SMA) receiving nusinersen. METHODS: A prospective, non-interventional observational study of nusinersen treatment in adult 5q-SMA patients was conducted at nine SMA centers in Germany. The functional status, treatment expectations and perceived outcomes were assessed using the Amyotrophic Lateral Sclerosis Functional Rating Scale-extended (ALS-FRS-ex), the Measure Yourself Medical Outcome Profile (MYMOP2), the Treatment Satisfaction Questionnaire for Medication (TSQM-9) and the Net Promoter Score (NPS). RESULTS: In all, 151 patients were included with a median age of 36 years (15-69 years). SMA type 3 (n = 90, 59.6%) prevailed, followed by type 2 (33.8%) and type 1 (6.6%). In SMA types 1-3, median ALS-FRS-ex scores were 25, 33 and 46 (of 60 scale points), respectively. MYMOP2 identified distinct treatment expectations: head verticalization (n = 13), bulbar function (n = 16), arm function (n = 65), respiration (n = 15), trunk function (n = 34), leg function (n = 76) and generalized symptoms (n = 77). Median symptom severity decreased during nusinersen treatment (median observational period 6.1 months, 0.5-16 months) from 3.7 to 3.3 MYMOP2 score points (p < 0.001). The convenience of drug administration was critical (49.7 of 100 TSQM-9 points, SD 22); however, the overall treatment satisfaction was high (74.3, SD 18) and the recommendation rating very positive (NPS +66). CONCLUSIONS: Nusinersen was administered across a broad range of ages, disease durations and motor function deficits. Treatment expectations were highly differentiated and related to SMA type and functional status. Patient-reported outcomes demonstrated a positive perception of nusinersen therapy in adult patients with 5q-SMA.

Elemental fingerprint as a cerebrospinal fluid biomarker for the diagnosis of Parkinson's disease.

The diagnosis of Parkinson's disease (PD) still lacks objective diagnostic markers independent of clinical criteria. Cerebrospinal fluid (CSF) samples from 36 PD and 42 age-matched control patients were subjected to inductively coupled plasma-sector field mass spectrometry and a total of 28 different elements were quantified. Different machine learning algorithms were applied to the dataset to identify a discriminating set of elements yielding a novel biomarker signature. Using 19 stably detected elements, the extreme gradient tree boosting model showed the best performance in the discrimination of PD and control patients with high specificity and sensitivity (78.6% and 83.3%, respectively), re-classifying the training data to 100%. The 10 times 10-fold cross-validation yielded a good area under the receiver operating characteristic curve of 0.83. Arsenic, magnesium, and selenium all showed significantly higher mean CSF levels in the PD group compared to the control group (p = 0.01, p = 0.04, and p = 0.03). Reducing the number of elements to a discriminating minimum, we identified an elemental cluster (Se, Fe, As, Ni, Mg, Sr), which most importantly contributed to the sample discrimination. Selenium was identified as the element with the highest impact within this cluster directly followed by iron. After prospective validation, this elemental fingerprint in the CSF could have the potential to be used as independent biomarker for the diagnosis of PD. Next to their value as a biomarker, these data also argue for a prominent role of these highly discriminating six elements in the pathogenesis of PD.

Alpha-synuclein and iron: two keys unlocking Parkinson's disease.

Current therapies for Parkinson's disease (PD) confer symptomatic relief and are particularly efficient in the treatment of motor symptoms in earlier disease stages. However, we are still unable to treat the causes of neurodegeneration by modification of the underlying mechanisms, which is partially due to their insufficient understanding. In this short review, we focus on two pivotal disease mechanisms: alpha-synuclein pathology and dysfunction of iron homeostasis as well as their intricate interaction. Both pathomechanisms have been extensively studied in the past and represent valid targets for disease-modifying pharmacological treatment approaches for PD. We summarize the current attempts to exploit iron chelation and modification of alpha-synuclein pathology as translational therapies in PD and discuss the chances and challenges of prospective disease-modifying approaches.

The role of autophagy in axonal degeneration of the optic nerve.

Different pathological conditions including glaucoma, optic neuritis, hereditary optic atrophy and traumatic injury lead to a degeneration of retinal ganglion cell axons in the optic nerve. Besides this clinical relevance, several experimental models employ the optic nerve as a model system to examine general mechanisms of axonal degeneration in the central nervous system. Several experimental studies have demonstrated that an activation of autophagy is a prominent feature of axonal degeneration in the optic nerve independent of the underlying pathological condition. However, the function of autophagy in axonal degeneration remains still unclear. Inhibition of autophagy was found to attenuate axonal degeneration within the first hours after optic nerve lesion. Other studies focusing on survival of retinal ganglion cells at later postlesional time points report contradicting results, where both inhibition and induction of autophagy were beneficial for survival, depending on the model system or examination time. Therefore, a more precise understanding of the role and the kinetics of autophagy in axonal degeneration is mandatory to develop new therapies for diseases of the optic nerve. Here, we review the literature on the pathophysiological role of autophagy in axonal degeneration in the optic nerve and discuss its implications for future therapeutic approaches in diseases of the eye and the central nervous system involving axonal degeneration.

AAV-mediated expression of BAG1 and ROCK2-shRNA promote neuronal survival and axonal sprouting in a rat model of rubrospinal tract injury.

A lesion to the rat rubrospinal tract is a model for traumatic spinal cord lesions and results in atrophy of the red nucleus neurons, axonal dieback, and locomotor deficits. In this study, we used adeno-associated virus (AAV)-mediated over-expression of BAG1 and ROCK2-shRNA in the red nucleus to trace [by co-expression of enhanced green fluorescent protein (EGFP)] and treat the rubrospinal tract after unilateral dorsal hemisection. We investigated the effects of targeted gene therapy on neuronal survival, axonal sprouting of the rubrospinal tract, and motor recovery 12 weeks after unilateral dorsal hemisection at Th8 in rats. In addition to the evaluation of BAG1 and ROCK2 as therapeutic targets in spinal cord injury, we aimed to demonstrate the feasibility and the limits of an AAV-mediated protein over-expression versus AAV.shRNA-mediated down-regulation in this traumatic CNS lesion model. Our results demonstrate that BAG1 and ROCK2-shRNA both promote neuronal survival of red nucleus neurons and enhance axonal sprouting proximal to the lesion.

Rho kinase inhibition modulates microglia activation and improves survival in a model of amyotrophic lateral sclerosis.

Disease progression in amyotrophic lateral sclerosis (ALS) is characterized by degeneration of motoneurons (MN) and their axons, but is also influenced by neighboring cells such as astrocytes and microglial cells. The role of microglia in ALS is complex as it switches from an anti-inflammatory and neuroprotective phenotype in early disease to a proinflammatory and neurotoxic phenotype in later stages. Our previous studies in models of neurodegeneration identified rho kinase (ROCK) as a target, which can be manipulated to beneficially influence disease progression. Here, we examined the neuroprotective potential of the ROCK inhibitor Fasudil to target the central pathogenic features of ALS. Application of Fasudil to kainic acid-lesioned primary MN in vitro resulted in a strong prosurvival effect. In vivo, SOD1(G93A) mice benefited from oral treatment with Fasudil showing prolonged survival and improved motor function. These findings were correlated to an improved survival of motor neurons and a pronounced alteration of astroglial and microglial cell infiltration of the spinal cord under Fasudil treatment. Modeling a proinflammatory microglial phenotype by stimulation with LPS in vitro, Fasudil decreased the release of proinflammatory cytokines and chemokines TNFα, Il6, CCL2, CCL3, and CCL5 while CXCL1 release was only transiently suppressed. In sciatic nerve motor axons, neuromuscular junction remodeling processes were increased. In conclusion, we provide preclinical and neurobiological evidence that inhibition of ROCK by the clinically approved small molecule inhibitor Fasudil may be a novel therapeutic approach in ALS combining both neuroprotection and immunomodulation for the cure of this devastating disease.

Axonal autophagic vesicle transport in the rat optic nerve in vivo under normal conditions and during acute axonal degeneration.

Neurons pose a particular challenge to degradative processes like autophagy due to their long and thin processes. Autophagic vesicles (AVs) are formed at the tip of the axon and transported back to the soma. This transport is essential since the final degradation of the vesicular content occurs only close to or in the soma. Here, we established an in vivo live-imaging model in the rat optic nerve using viral vector mediated LC3-labeling and two-photon-microscopy to analyze axonal transport of AVs. Under basal conditions in vivo, 50% of the AVs are moving with a majority of 85% being transported in the retrograde direction. Transport velocity is higher in the retrograde than in the anterograde direction. A crush lesion of the optic nerve results in a rapid breakdown of retrograde axonal transport while the anterograde transport stays intact over several hours. Close to the lesion site, the formation of AVs is upregulated within the first 6 h after crush, but the clearance of AVs and the levels of lysosomal markers in the adjacent axon are reduced. Expression of p150Glued, an adaptor protein of dynein, is significantly reduced after crush lesion. In vitro, fusion and colocalization of the lysosomal marker cathepsin D with AVs are reduced after axotomy. Taken together, we present here the first in vivo analysis of axonal AV transport in the mammalian CNS using live-imaging. We find that axotomy leads to severe defects of retrograde motility and a decreased clearance of AVs via the lysosomal system.

Synchrotron-Based Fourier-Transform Infrared Micro-Spectroscopy of Cerebrospinal Fluid from Amyotrophic Lateral Sclerosis Patients Reveals a Unique Biomolecular Profile.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease, with the most common adult-onset neurodegenerative disorder affecting motoneurons. Although disruptions in macromolecular conformation and homeostasis have been described in association with ALS, the underlying pathological mechanisms are still not completely understood, and unambiguous biomarkers are lacking. Fourier Transform Infrared Spectroscopy (FTIR) of cerebrospinal fluid (CSF) is appealing to extensive interest due to its potential to resolve biomolecular conformation and content, as this approach offers a non-invasive, label-free identification of specific biologically relevant molecules in a few microliters of CSF sample. Here, we analyzed the CSF of 33 ALS patients compared to 32 matched controls using FTIR spectroscopy and multivariate analysis and demonstrated major differences in the molecular contents. A significant change in the conformation and concentration of RNA is demonstrated. Moreover, significantly increased glutamate and carbohydrates are found in ALS. Moreover, key markers of lipid metabolism are strongly altered; specifically, we find a decrease in unsaturated lipids and an increase in peroxidation of lipids in ALS, whereas the total amount of lipids compared to proteins is reduced. Our study demonstrates that FTIR characterization of CSF could represent a powerful tool for ALS diagnosis and reveals central features of ALS pathophysiology.

Remote digital assessment of amyotrophic lateral sclerosis functional rating scale - a multicenter observational study.

OBJECTIVE: Remote self-assessment of the revised amyotrophic lateral sclerosis functional rating scale (ALSFRS-R) using digital data capture was investigated for its feasibility as an add-on to ALSFRS-R assessments during multidisciplinary clinic visits. METHODS: From August 2017 to December 2021, at 12 ALS centers in Germany, an observational study on remote assessment of the ALSFRS-R was performed. In addition to the assessment of ALSFRS-R during clinic visits, patients were offered a digital self-assessment of the ALSFRS-R - either on a computer or on a mobile application ("ALS-App"). RESULTS: An estimated multicenter cohort of 4,670 ALS patients received care at participating ALS centers. Of these patients, 971 remotely submitted the ALSFRS-R, representing 21% of the multicenter cohort. Of those who opted for remote assessment, 53.7% (n = 521) completed a minimum of 4 ALSFRS-R per year with a mean number of 10.9 assessments per year. Different assessment frequencies were found for patients using a computer (7.9 per year, n = 857) and mobile app (14.6 per year, n = 234). Patients doing remote assessments were more likely to be male and less functionally impaired but many patients with severe disability managed to complete it themselves or with a caregiver (35% of remote ALSFRS-R cohort in King's Stage 4). CONCLUSIONS: In a dedicated ALS center setting remote digital self-assessment of ALSFRS-R can provide substantial data which is complementary and potentially an alternative to clinic assessments and could be used for research purposes and person-level patient management. Addressing barriers relating to patient uptake and adherence are key to its success.

Economic evaluation of Motor Neuron Diseases: a nationwide cross-sectional analysis in Germany.

BACKGROUND AND OBJECTIVES: Motor Neuron Diseases (MND) are rare diseases but have a high impact on affected individuals and society. This study aims to perform an economic evaluation of MND in Germany. METHODS: Primary patient-reported data were collected including individual impairment, the use of medical and non-medical resources, and self-rated Health-Related Quality of Life (HRQoL). Annual socio-economic costs per year as well as Quality-Adjusted Life Years (QALYs) were calculated. RESULTS: 404 patients with a diagnosis of Amyotrophic Lateral Sclerosis (ALS), Spinal Muscular Atrophy (SMA) or Hereditary Spastic Paraplegia (HSP) were enrolled. Total annual costs per patient were estimated at 83,060€ in ALS, 206,856€ in SMA and 27,074€ in HSP. The main cost drivers were informal care (all MND) and disease-modifying treatments (SMA). Self-reported HRQoL was best in patients with HSP (mean EuroQoL Five Dimension Five Level (EQ-5D-5L) index value 0.67) and lowest in SMA patients (mean EQ-5D-5L index value 0.39). QALYs for patients with ALS were estimated to be 1.89 QALYs, 23.08 for patients with HSP and 14.97 for patients with SMA, respectively. Cost-utilities were estimated as follows: 138,960€/QALY for ALS, 525,033€/QALY for SMA, and 49,573€/QALY for HSP. The main predictors of the high cost of illness and low HRQoL were disease progression and loss of individual autonomy. CONCLUSION: As loss of individual autonomy was the main cost predictor, therapeutic and supportive measures to maintain this autonomy may contribute to reducing high personal burden and also long-term costs, e.g., care dependency and absenteeism from work.

Improvements in Walking Distance during Nusinersen Treatment - A Prospective 3-year SMArtCARE Registry Study.

BACKGROUND AND OBJECTIVES: Disease progression in patients with spinal muscular atrophy (SMA) has changed dramatically within the past years due to the approval of three different disease-modifying treatments. Nusinersen was the first drug to be approved for the treatment of SMA patients. Clinical trials provided data from infants with SMA type 1 and children with SMA type 2, but there is still insufficient evidence and only scarcely reported long-term experience for nusinersen treatment in ambulant patients. Here, we report data from the SMArtCARE registry of ambulant patients under nusinersen treatment with a follow-up period of up to 38 months. METHODS: SMArtCARE is a disease-specific registry in Germany, Austria and Switzerland. Data are collected as real-world data during routine patient visits. Our analysis included all patients under treatment with nusinersen able to walk independently before start of treatment with focus on changes in motor function. RESULTS: Data from 231 ambulant patients were included in the analysis. During the observation period, 31 pediatric walkers (27.2%) and 31 adult walkers (26.5%) experienced a clinically meaningful improvement of≥30 m in the 6-Minute-Walk-Test. In contrast, only five adult walkers (7.7%) showed a decline in walking distance≥30 m, and two pediatric walkers (1.8%) lost the ability to walk unassisted under treatment with nusinersen. HFMSE and RULM scores improved in pediatric and remained stable in adult patients. CONCLUSION: Our data demonstrate a positive effect of nusinersen treatment in most ambulant pediatric and adult SMA patients. We not only observed a stabilization of disease progression or lack of deterioration, but clinically meaningful improvements in walking distance.

Cytoskeletal assembly in axonal outgrowth and regeneration analyzed on the nanoscale.

The axonal cytoskeleton is organized in a highly periodic structure, the membrane-associated periodic skeleton (MPS), which is essential to maintain the structure and function of the axon. Here, we use stimulated emission depletion microscopy of primary rat cortical neurons in microfluidic chambers to analyze the temporal and spatial sequence of MPS formation at the distal end of growing axons and during regeneration after axotomy. We demonstrate that the MPS does not extend continuously into the growing axon but develops from patches of periodic ßII-spectrin arrangements that grow and coalesce into a continuous scaffold. We estimate that the underlying sequence of assembly, elongation, and subsequent coalescence of periodic ßII-spectrin patches takes around 15 h. Strikingly, we find that development of the MPS occurs faster in regenerating axons after axotomy and note marked differences in the morphology of the growth cone and adjacent axonal regions between regenerating and unlesioned axons. Moreover, we find that inhibition of the spectrin-cleaving enzyme calpain accelerates MPS formation in regenerating axons and increases the number of regenerating axons after axotomy. Taken together, we provide here a detailed nanoscale analysis of MPS development in growing axons.

Use and subjective experience of the impact of motor-assisted movement exercisers in people with amyotrophic lateral sclerosis: a multicenter observational study.

Motor-assisted movement exercisers (MME) are devices that assist with physical therapy in domestic settings for people living with ALS. This observational cross-sectional study assesses the subjective experience of the therapy and analyzes users' likelihood of recommending treatment with MME. The study was implemented in ten ALS centers between February 2019 and October 2020, and was coordinated by the research platform Ambulanzpartner. Participants assessed symptom severity, documented frequency of MME use and rated the subjective benefits of therapy on a numerical scale (NRS, 0 to 10 points, with 10 being the highest). The Net Promotor Score (NPS) determined the likelihood of a participant recommending MME. Data for 144 participants were analyzed. Weekly MME use ranged from 1 to 4 times for 41% of participants, 5 to 7 times for 42%, and over 7 times for 17%. Particularly positive results were recorded in the following domains: amplification of a sense of achievement (67%), diminution of the feeling of having rigid limbs (63%), diminution of the feeling of being immobile (61%), improvement of general wellbeing (55%) and reduction of muscle stiffness (52%). Participants with more pronounced self-reported muscle weakness were more likely to note a beneficial effect on the preservation and improvement of muscle strength during MME treatment (p < 0.05). Overall, the NPS for MME was high (+ 61). High-frequency MME-assisted treatment (defined as a minimum of five sessions a week) was administered in the majority of participants (59%) in addition to physical therapy. Most patients reported having achieved their individual therapeutic objectives, as evidenced by a high level of satisfaction with MME therapy. The results bolster the justification for extended MME treatment as part of a holistic approach to ALS care.

Hepatocyte growth factor protects retinal ganglion cells by increasing neuronal survival and axonal regeneration in vitro and in vivo.

Hepatocyte growth factor (HGF) is known to promote the survival and foster neuritic outgrowth of different subpopulations of CNS neurons during development. Together with its corresponding receptor c-mesenchymal-epithelial transition factor (Met), it is expressed in the developing and the adult murine, rat and human CNS. We have studied the role of HGF in paradigms of retinal ganglion cell (RGC) regeneration and cell death in vitro and in vivo. After application of recombinant HGF in vitro, survival of serum-deprived RGC-5 cells and of growth factor-deprived primary RGC was significantly increased. This was shown to be correlated to the phosphorylation of c-Met and subsequent activation of serine/threonine protein kinase Akt and MAPK downstream signalling pathways involved in neuronal survival. Furthermore, neurite outgrowth of primary RGC was stimulated by HGF. In vivo, c-Met expression in RGC was up-regulated after optic nerve axotomy lesion. Here, treatment with HGF significantly improved survival of axotomized RGC and enhanced axonal regeneration after optic nerve crush. Our data demonstrates that exogenously applied HGF has a neuroprotective and regeneration-promoting function for lesioned CNS neurons. We provide strong evidence that HGF may represent a trophic factor for adult CNS neurons, which may play a role as therapeutic target in the treatment of neurotraumatic and neurodegenerative CNS disorders.

AAV-mediated inhibition of ULK1 promotes axonal regeneration in the central nervous system in vitro and in vivo.

Axonal damage is an early step in traumatic and neurodegenerative disorders of the central nervous system (CNS). Damaged axons are not able to regenerate sufficiently in the adult mammalian CNS, leading to permanent neurological deficits. Recently, we showed that inhibition of the autophagic protein ULK1 promotes neuroprotection in different models of neurodegeneration. Moreover, we demonstrated previously that axonal protection improves regeneration of lesioned axons. However, whether axonal protection mediated by ULK1 inhibition could also improve axonal regeneration is unknown. Here, we used an adeno-associated viral (AAV) vector to express a dominant-negative form of ULK1 (AAV.ULK1.DN) and investigated its effects on axonal regeneration in the CNS. We show that AAV.ULK1.DN fosters axonal regeneration and enhances neurite outgrowth in vitro. In addition, AAV.ULK1.DN increases neuronal survival and enhances axonal regeneration after optic nerve lesion, and promotes long-term axonal protection after spinal cord injury (SCI) in vivo. Interestingly, AAV.ULK1.DN also increases serotonergic and dopaminergic axon sprouting after SCI. Mechanistically, AAV.ULK1.DN leads to increased ERK1 activation and reduced expression of RhoA and ROCK2. Our findings outline ULK1 as a key regulator of axonal degeneration and regeneration, and define ULK1 as a promising target to promote neuroprotection and regeneration in the CNS.

Increased chitotriosidase 1 concentration following nusinersen treatment in spinal muscular atrophy.

BACKGROUND: Studies regarding the impact of (neuro)inflammation and inflammatory response following repetitive, intrathecally administered antisense oligonucleotides (ASO) in 5q-associated spinal muscular atrophy (SMA) are sparse. Increased risk of hydrocephalus in untreated SMA patients and a marginal but significant increase of the serum/CSF albumin ratio (Qalb) with rare cases of communicating hydrocephalus during nusinersen treatment were reported, which confirms the unmet need of an inflammatory biomarker in SMA. The aim of this study was to investigate the (neuro)inflammatory marker chitotriosidase 1 (CHIT1) in SMA patients before and following the treatment with the ASO nusinersen. METHODS: In this prospective, multicenter observational study, we studied CSF CHIT1 concentrations in 58 adult and 21 pediatric patients with SMA type 1, 2 or 3 before treatment initiation in comparison to age- and sex-matched controls and investigated its dynamics during nusinersen treatment. Concurrently, motor performance and disease severity were assessed. RESULTS: CHIT1 concentrations were elevated in treatment-naïve SMA patients as compared to controls, but less pronounced than described for other neurodegenerative diseases such as amyotrophic lateral sclerosis. CHIT1 concentration did not correlate with disease severity and did not distinguish between clinical subtypes. CHIT1 concentration did show a significant increase during nusinersen treatment that was unrelated to the clinical response to nusinersen therapy. CONCLUSIONS: CHIT1 elevation in treatment-naïve SMA patients indicates the involvement of (neuro)inflammation in SMA. The lacking correlation of CHIT1 concentration with disease severity argues against its use as a marker of disease progression. The observed CHIT1 increase during nusinersen treatment may indicate an immune response-like, off-target reaction. Since antisense oligonucleotides are an establishing approach in the treatment of neurodegenerative diseases, this observation needs to be further evaluated.