Aromatic L-amino acid decarboxylase deficiency: a patient-derived neuronal model for precision therapies

Aromatic L-amino acid decarboxylase (AADC) deficiency is a complex inherited neurological disorder of monoamine synthesis which results in dopamine and serotonin deficiency. The majority of affected individuals have variable, though often severe cognitive and motor delay, with a complex movement disorder and high risk of premature mortality. For most, standard pharmacological treatment provides only limited clinical benefit. Promising gene therapy approaches are emerging, though may not be either suitable or easily accessible for all patients. In order to better characterize the underlying disease pathophysiology and guide precision therapies, we generated a patient-derived midbrain dopaminergic (mDA) neuronal model of AADC deficiency from induced pluripotent stem cells (iPSCs). The neuronal model recapitulates key disease features, including absent AADC enzyme activity and dysregulated dopamine metabolism. We observed developmental defects affecting synaptic maturation and neuronal electrical properties, which were improved by lentiviral gene therapy. Bioinformatic and biochemical analyses on recombinant AADC predicted that the activity of one variant could be improved by L-3,4-dihydroxyphenylalanine (L-DOPA) administration; this hypothesis was corroborated in the patient-derived neuronal model, where L-DOPA treatment leads to amelioration of dopamine metabolites. Our study has shown that patient-derived disease modelling provides further insight into the neurodevelopmental sequelae of AADC deficiency, as well as a robust platform to investigate and develop personalised therapeutic approaches

Aromatic l-amino acid decarboxylase deficiency: a patient-derived neuronal model for precision therapies

Aromatic l-amino acid decarboxylase (AADC) deficiency is a complex inherited neurological disorder of monoamine synthesis which results in dopamine and serotonin deficiency. The majority of affected individuals have variable, though often severe cognitive and motor delay, with a complex movement disorder and high risk of premature mortality. For most, standard pharmacological treatment provides only limited clinical benefit. Promising gene therapy approaches are emerging, though may not be either suitable or easily accessible for all patients. To characterize the underlying disease pathophysiology and guide precision therapies, we generated a patient-derived midbrain dopaminergic neuronal model of AADC deficiency from induced pluripotent stem cells. The neuronal model recapitulates key disease features, including absent AADC enzyme activity and dysregulated dopamine metabolism. We observed developmental defects affecting synaptic maturation and neuronal electrical properties, which were improved by lentiviral gene therapy. Bioinformatic and biochemical analyses on recombinant AADC predicted that the activity of one variant could be improved by l-3,4-dihydroxyphenylalanine (l-DOPA) administration; this hypothesis was corroborated in the patient-derived neuronal model, where l-DOPA treatment leads to amelioration of dopamine metabolites. Our study has shown that patient-derived disease modelling provides further insight into the neurodevelopmental sequelae of AADC deficiency, as well as a robust platform to investigate and develop personalized therapeutic approaches

Foamy Virus Vectors Transduce Visceral Organs and Hippocampal Structures following In Vivo Delivery to Neonatal Mice

Viral vectors are rapidly being developed for a range of applications in research and gene therapy. Prototype foamy virus (PFV) vectors have been described for gene therapy, although their use has mainly been restricted to ex vivo stem cell modification. Here we report direct in vivo transgene delivery with PFV vectors carrying reporter gene constructs. In our investigations, systemic PFV vector delivery to neonatal mice gave transgene expression in the heart, xiphisternum, liver, pancreas, and gut, whereas intracranial administration produced brain expression until animals were euthanized 49 days post-transduction. Immunostaining and confocal microscopy analysis of injected brains showed that transgene expression was highly localized to hippocampal architecture despite vector delivery being administered to the lateral ventricle. This was compared with intracranial biodistribution of lentiviral vectors and adeno-associated virus vectors, which gave a broad, non-specific spread through the neonatal mouse brain without regional localization, even when administered at lower copy numbers. Our work demonstrates that PFV can be used for neonatal gene delivery with an intracranial expression profile that localizes to hippocampal neurons, potentially because of the mitotic status of the targeted cells, which could be of use for research applications and gene therapy of neurological disorders

Application of near-infrared tissue oxymetry to the diagnosis of peripheral vascular disease.

Near-infrared spectroscopy (NIRS) is a noninvasive technique to measure the tissue oxygenation in real time. This optical method has many advantages over the invasive analysis currently used for clinical tests. Among the possible applications of near-infrared oxymetry, we report three protocols (exercise, venous occlusion and tilting table) in conjunction with NIRS, and discuss their applicability in the diagnosis of peripheral vascular disease (PVD)

LGI1 downregulation increases neuronal circuit excitability

OBJECTIVE: Leucine‐rich glioma‐inactivated 1 (LGI1) is a secreted transsynaptic protein that interacts presynaptically with Kv1.1 potassium channels and a disintegrin and metalloprotease (ADAM) protein 23, and postsynaptically influences α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionate receptors through a direct link with the ADAM22 cell adhesion protein. Haploinsufficiency of LGI1 or autoantibodies directed against LGI1 are associated with human epilepsy, generating the hypothesis that a subacute reduction of LGI1 is sufficient to increase network excitability. METHODS: We tested this hypothesis in ex vivo hippocampal slices and in neuronal cultures, by subacutely reducing LGI1 expression with shRNA. RESULTS: Injection of shRNA‐LGI1 in the hippocampus increased dentate granule cell excitability and low‐frequency facilitation of mossy fibers to CA3 pyramidal cell neurotransmission. Application of the Kv1 family blocker, α‐dendrotoxin, occluded this effect, implicating the involvement of Kv1.1. This subacute reduction of LGI1 was also sufficient to increase neuronal network activity in neuronal primary culture. SIGNIFICANCE: These results indicate that a subacute reduction in LGI1 potentiates neuronal excitability and short‐term synaptic plasticity, and increases neuronal network excitability, opening new avenues for the treatment of limbic encephalitis and temporal lobe epilepsies