Objective The aim was to evaluate, in patients with atrial fibrillation (AF) and acute ischemic stroke, the association of prior anticoagulation with vitamin K antagonists (VKAs) or direct oral anticoagulants (DOACs) with stroke severity, utilization of intravenous thrombolysis (IVT), safety of IVT, and 3‐month outcomes. Methods This was a cohort study of consecutive patients (2014–2019) on anticoagulation versus those without (controls) with regard to stroke severity, rates of IVT/mechanical thrombectomy, symptomatic intracranial hemorrhage (sICH), and favorable outcome (modified Rankin Scale score 0–2) at 3 months. Results Of 8,179 patients (mean [SD] age, 79.8 [9.6] years; 49% women), 1,486 (18%) were on VKA treatment, 1,634 (20%) on DOAC treatment at stroke onset, and 5,059 controls. Stroke severity was lower in patients on DOACs (median National Institutes of Health Stroke Scale 4, [interquartile range 2–11]) compared with VKA (6, [2–14]) and controls (7, [3–15], p < 0.001; quantile regression: β −2.1, 95% confidence interval [CI] −2.6 to −1.7). The IVT rate in potentially eligible patients was significantly lower in patients on VKA (156 of 247 [63%]; adjusted odds ratio [aOR] 0.67; 95% CI 0.50–0.90) and particularly in patients on DOACs (69 of 464 [15%]; aOR 0.06; 95% CI 0.05–0.08) compared with controls (1,544 of 2,504 [74%]). sICH after IVT occurred in 3.6% (2.6–4.7%) of controls, 9 of 195 (4.6%; 1.9–9.2%; aOR 0.93; 95% CI 0.46–1.90) patients on VKA and 2 of 65 (3.1%; 0.4–10.8%, aOR 0.56; 95% CI 0.28–1.12) of those on DOACs. After adjustments for prognostic confounders, DOAC pretreatment was associated with a favorable 3‐month outcome (aOR 1.24; 1.01–1.51). Interpretation Prior DOAC therapy in patients with AF was associated with decreased admission stroke severity at onset and a remarkably low rate of IVT. Overall, patients on DOAC might have better functional outcome at 3 months. Further research is needed to overcome potential restrictions for IVT in patients taking DOACs. ANN NEUROL 2021;89:42–53

Axonopathies are a group of clinically diverse disorders characterized by the progressive degeneration of the axons of specific neurons. In hereditary spastic paraplegia (HSP), the axons of cortical motor neurons degenerate and cause a spastic movement disorder. HSP is linked to mutations in several loci known collectively as the spastic paraplegia genes (SPGs). We identified a heterozygous receptor accessory protein 1 (REEP1) exon 2 deletion in a patient suffering from the autosomal dominantly inherited HSP variant SPG31. We generated the corresponding mouse model to study the underlying cellular pathology. Mice with heterozygous deletion of exon 2 in Reep1 displayed a gait disorder closely resembling SPG31 in humans. Homozygous exon 2 deletion resulted in the complete loss of REEP1 and a more severe phenotype with earlier onset. At the molecular level, we demonstrated that REEP1 is a neuron-specific, membrane-binding, and membrane curvature-inducing protein that resides in the ER. We further show that Reep1 expression was prominent in cortical motor neurons. In REEP1-deficient mice, these neurons showed reduced complexity of the peripheral ER upon ultrastructural analysis. Our study connects proper neuronal ER architecture to long-term axon survival.

ObjectiveAmyotrophic lateral sclerosis is an incurable disorder mainly characterized by motoneuron degeneration. Mutations in the superoxide dismutase 1 (SOD1) gene account for 20% of familial forms of the disease. Mutant SOD1 exerts multiple pathogenic effects through the gain of toxic properties in both neurons and glial cells. Here, we compare AAV-based gene therapy suppressing expression of mutant SOD1 in either motoneurons or astrocytes.MethodsAAV vectors encoding microRNA against human SOD1 were administered to G93ASOD1 mice either by intracerebroventricular injections in pups or by lumbar intrathecal injections in adults. Vector systems were designed to suppress SOD1 expression predominantly in either spinal motoneurons or astrocytes. Electrophysiological and behavioral tests were performed on treated animals to evaluate disease progression.ResultsFollowing vector injection in G93ASOD1 pups, efficient silencing of SOD1 expression was achieved in motoneurons and/or astrocytes. Most complete protection of motor units was obtained when targeting human SOD1 predominantly in motoneurons. Suppressing SOD1 mainly in astrocytes led to preserved muscle innervation despite only partial protection of spinal motoneurons. In both cases, injection in pups led to full recovery of neuromuscular function and significantly prolonged survival. Vector injections in adult mice also achieved significant protection of neuromuscular function, which was highest when motoneurons were targeted.InterpretationThese results suggest that AAV-mediated SOD1 silencing is an effective approach to prevent motoneuron degeneration caused by SOD1 mutation. AAV vectors suppressing SOD1 in motoneurons delay disease onset and show effective neuroprotection. On the other hand, AAV-based SOD1 silencing in astrocytes rescues neuromuscular function following initial denervation.

In the context of motoneuron diseases, gene delivery as an experimental or therapeutic approach is hindered by the challenge to specifically target cell populations that are widely distributed along the spinal cord. Further complicating the task, transgenes often need to be delivered to motoneurons and/or glial cells to address the non-cell-autonomous mechanisms involved in disease pathogenesis. Intracerebroventricular (ICV) injection of recombinant adeno-associated viruses (AAVs) in newborn mice allows distributing viral vectors throughout the central nervous system while limiting undesired transduction of peripheral organs. Here, we show that by combining the appropriate set of AAV serotype and promoter, specific transgene expression can be achieved in either motoneurons or astrocytes along the whole mouse spinal cord. ICV injection of recombinant AAV6 with the cytomegalovirus (cmv) promoter preferentially targets motoneurons, whereas AAV9 particles combined with the astrocyte-specific gfaABC₁D promoter lead to significant transgene expression selectively targeted to astrocytes. Importantly, ICV coinjection of both AAV6-cmv and AAV9-gfaABC₁D results in segregated expression of two different transgenes in motoneurons and astrocytes, respectively. Relevance of viral vector delivery via the cerebrospinal fluid was further investigated in young nonhuman primates. Intracisternal injection of recombinant AAV6-cmv led to robust cervical transduction of motoneurons, highlighting the potential of this approach for gene therapy and modeling of motoneuron diseases.

Amyotrophic lateral sclerosis (ALS) is the most common motor neuron disease. ALS is believed to be a non-cell autonomous condition, as other cell types, including astrocytes, have been implicated in disease pathogenesis. Hence, to facilitate the development of therapeutics against ALS, it is crucial to better understand the interactions between astrocytes and neural cells. Furthermore, cell culture assays are needed that mimic the complexity of cell to cell communication at the same time as they provide control over the different microenvironmental parameters. Here, we aim to validate a previously developed microfluidic system for an astrocyte-neuron cell culture platform, in which astrocytes have been genetically modified to overexpress either a human wild-type (WT) or a mutated form of the super oxide dismutase enzyme 1 (SOD1). Cortical neural cells were co-cultured with infected astrocytes and studied for up to two weeks. Using our microfluidic device that prevents direct cell to cell contact, we could evaluate neural cell response in the vicinity of astrocytes. We showed that neuronal cell density was reduced by about 45% when neurons were co-cultured with SOD-mutant astrocytes. Moreover, we demonstrated that SOD-WT overexpressing astrocytes reduced oxidative stress on cortical neurons that were in close metabolic contact. In contrast, cortical neurons in metabolic contact with SOD-mutant astrocytes lost their synapsin protein expression after severe glutamate treatment, an indication of the toxicity potentiating effect of the SOD-mutant enzyme.