Molecular and phenotypic characterization of two monogenetic neurodevelopmental disorders: MCT8 deficiency & FOXG1 syndrome

Einleitung: Die menschliche Gehirnentwicklung ist komplex. Einzelne Mutationen können zu schweren Entwicklungsstörungen führen. In diesem Dissertationsprojekt habe ich zwei Erkrankungen untersucht. (I) Lokal ist die Gehirnentwicklung durch Transkriptionsfaktoren (TF) reguliert. Inaktivierende Mutationen des TF FOXG1 führen zu psychomotorischer Retardierung. Thesen zur Pathophysiologie stützten sich bisher auf Mausmodelle und klinische Daten. Neuropathologische Analysen existierten keine. (II) Neben lokalen Einflüssen regulieren endokrine Faktoren, z.B. Schilddrüsenhormone (SD), die zerebrale Genexpression. SD müssen mittels Transportern Zellmembranen der neurovaskulären Einheit überwinden, um intrazelluläre Rezeptoren zu binden. Mutationen des Transporters MCT8 führen zu einem Syndrom mit Entwicklungs-/Bewegungsstörungen. Zur gezielten Therapieentwicklung ist es notwendig, die genaue MCT8 Lokalisation zu identifizieren; für die topographische Diagnostik und für Erkrankungsspezifische Perzentilen, den neurologischen Phänotyp exakt zu beschreiben. Methoden: (I) Wir führten immunhistochemische Analysen an Gehirnen zweier Feten mit FOXG1 Syndrom und eine systematische Literatursuche zu den Grundlagen der Pathophysiologie durch. (II) Wir untersuchten die MCT8 Expression mittels Immunfluoreszenz in adulten humanen Gehirnen. Zur qualitativen und semi-quantitativen Analyse dynamischer Veränderungen, schlossen wir eine Altersserie murinen Gewebes und humane Organoide ein. Zur Bewegungsstörungsanalyse zeichneten wir Patientenvideos auf und werteten diese mittels einer Dystonie Skala aus. Ergebnisse: (I) Es zeigten sich ein reduziertes Gehirnvolumen, kortikale Schichtungsdefekte, veränderte neuronale Projektionen und weniger Interneurone. Ich konnte 35 Studien zur Pathophysiologie in Mausmodellen identifizieren. (II) MCT8 war konstant in „Barrieren des ZNS“ (Bluthirnschranke, Plexus choroideus, Tanyzyten) nachweisbar. In Neuronen konnte MCT8 früh postnatal im Gewebe der Maus, in neuronalen Progenitorzellen der Organoide, nicht aber in adulten Gehirnen detektiert werden. Analysen der Bewegungsstörungen stellten die Dystonie als vorwiegende, jedoch Hypotonie als schwerwiegendste Störung dar. Diskussion: (I) Beim FOXG1 Syndrom scheinen kortikale Schichtungsdefekte neuronale Projektionen zu verändern und eine verfrühte neuronale Differenzierung zu einem reduzierten Gehirnvolumen zu führen, assoziiert mit Entwicklungsstörungen. Frühe Strukturierungsdefekte des Gehirns scheinen eine erhöhte Ratio von exzitatorischer/inhibitorischer neuronaler Aktivität zu bedingen und somit zu Hyperkinesen zu führen. (II) MCT8 scheint wichtig zu sein, für den SD-Transport über die Bluthirnschranke und für die prä- und früh postnatale neuronale Entwicklung. Bewegungsstörungen der Patienten deuten auf eine wichtige pathophysiologische Rolle des MCT8 in den Basalganglien hin. Das humane „Zeitfenster der MCT8 Expression“ und die Rolle des motorischen Systems sollten Themen zukünftiger Studien sein.Introduction: The achievement of developmental milestones in children depends on healthy brain development. Single gene mutations may have devastating consequences, leading to neurodevelopmental diseases (NDDs). In this dissertation project, I investigated two NDDs. (I) Locally, brain development is regulated by transcription factors (TF). Loss-of-function mutations of the TF FOXG1 cause a syndrome with psychomotor retardation. Knowledge about the pathophysiology so far relied on mouse models and clinical descriptions. Neuropathological data did not exist. (II) Besides local signals, endocrine factors such as thyroid hormones (THs) regulate cerebral gene expression. To bind their intracellular receptors, THs must be moved across membranes of the neurovascular unit via transporters. Mutations of the TH transporter MCT8 lead to a severe NDD, called MCT8 deficiency. For targeted therapies, it is necessary to identify the exact localization of MCT8 and, for topographic diagnostics and disease-specific percentiles to describe the neurological phenotype precisely. Methods: (I) We performed immunohistochemical analyses of two fetal brains with FOXG1 syndrome and systematically searched for murine studies on its pathophysiology. (II) We analyzed the MCT8 expression in adult human brains by immunostaining. To test dynamic changes (qualitatively and semi-quantitively), we included a series of murine brains of different age and human organoids. Videos of patients with movement disorders (MDs) were captured and analyzed with a Dystonia Rating Scale. Results: (I) FOXG1+/- brains presented with reduced brain volumes, layering defects, deficient neuronal projections, and fewer interneurons. 35 articles were identified studying the pathophysiology. (II) MCT8 immunosignals were constantly detectable in the “barriers of the CNS” (blood-brain barrier, choroid plexus, tanycytes), but were only visible in murine neuronal populations early postnatally. Correspondingly, in human organoids, MCT8 immunosignals were present in solitary neuronal progenitor cells, but not in neurons of adult individuals. The analysis of MDs revealed dystonia to be a consistent feature, hypotonia, however, seemed to be the clinically most relevant disability. Discussion: (I) In FOXG1 syndrome, premature neuronal differentiation may cause a microcephaly that is associated with developmental delay. Impaired corticogenesis may result in deficient neuronal projections and early patterning defects may lead to an increased exhibition-to-inhibition ratio triggering hyperkinetic symptoms. (II) MCT8 seems to facilitate the transport over the blood-brain barrier independent of age and over specific neuronal cell membranes in the pre- and early postnatal phase. MDs of patients indicate a substantial pathophysiological role of the basal ganglia and the white matter. The human “time window of MCT8 expression” and the role of basal ganglia/ white matter still needs to be further defined

Ten simple rules for implementing open and reproducible research practices after attending a training course

Open, reproducible, and replicable research practices are a fundamental part of science. Training is often organized on a grassroots level, offered by early career researchers, for early career researchers. Buffet style courses that cover many topics can inspire participants to try new things; however, they can also be overwhelming. Participants who want to implement new practices may not know where to start once they return to their research team. We describe ten simple rules to guide participants of relevant training courses in implementing robust research practices in their own projects, once they return to their research group. This includes (1) prioritizing and planning which practices to implement, which involves obtaining support and convincing others involved in the research project of the added value of implementing new practices; (2) managing problems that arise during implementation; and (3) making reproducible research and open science practices an integral part of a future research career. We also outline strategies that course organizers can use to prepare participants for implementation and support them during this process

Ten simple rules for implementing open and reproducible research practices after attending a training course

Training in robust research practices is becoming increasingly common. However, many course participants may encounter challenges in implementation of what they learned after returning to their research groups. In this piece, we summarize insights and "lessons learned" from a group of former course participants. We offer practical tips on implementation and cultural change that may be useful for researchers at any career stage. In addition, we provide a list of considerations for course instructors to help them support course attendees after training is over