Biology subjects

De Vos, W. H.

Publications and source records attributed to De Vos, W. H..

9 recordsLinked to original sources

bioRxiv2025

KCNQ2 Loss-of-Function variants disrupt neuronal maturation via early hyperexcitability followed by maladaptive network remodeling

Loss-of-function (LOF) variants in the potassium channel subunit KCNQ2 cause a spectrum of neonatal epilepsies from self-limiting familial neonatal epilepsy (SeLFNE) to severe developmental and epileptic encephalopathy (DEE). To dissect the developmental consequences of KCNQ2 LOF, we conducted a longitudinal and multimodal comparative analysis in a human neuronal model generated from patients with KCNQ2-DEE and KCNQ2-SeLFNE. KCNQ2-LOF induced a biphasic network dysfunction, with early Kv7-driven hyperexcitability rescued by acute Retigabine (RTG) treatment, followed by maladaptive remodeling in the opposite direction. Transcriptomic analysis mirrored this biphasic dynamic trajectory, revealing an initial upregulation followed by a subsequent downregulation of synaptic genes. Structural analysis showed a steeper decline in presynaptic density alongside a distal shift in the axon initial segment (AIS) throughout maturation, and impaired AIS plasticity at later stages. Overall, KCNQ2-LOF disrupts human neuronal maturation through dynamic, biphasic changes in function, gene expression and structure, offering insights into disease mechanisms and therapeutic options.

neuroscience↗

bioRxiv2024

A human-specific, concerted repression of microcephaly genes contributes to radiation-induced growth defects in forebrain organoids

AbstractPrenatal radiation-induced DNA damage poses a significant threat to normal brain development, resulting in microcephaly which primarily affects the cerebral cortex. It is unclear which molecular mechanisms are at the basis of this defect in humans as the few mechanistic studies performed so far were done in animals. Here, we leveraged human embryonic stem cell- derived forebrain organoids as a model for human corticogenesis. Organoids were X-irradiated with a moderate and a high dose at different time points, representing very early and mid corticogenesis. Irradiation caused a dose- and developmental-timing-dependent reduction in organoid size, which was more prominent in developmentally younger organoids. This coincided with a dose-dependent canonical p53-DREAM-dependent DNA damage response (DDR), consisting of cell cycle arrest, DNA repair and apoptosis. The DDR was delayed and less pronounced in the older organoids. Besides the DDR, we observed radiation-induced premature differentiation of neural progenitors and changes in metabolism. Importantly, our transcriptomic analysis furthermore demonstrated a concerted p53-E2F4-dependent repression of primary microcephaly genes. We found that this was a human-specific feature, as it was not observed in mouse embryonic brains or primary mouse neural progenitor cells. Thus, human forebrain organoids are an excellent model to investigate prenatal DNA damage-induced microcephaly and to uncover potentially targetable human-specific pathways.

developmental biology↗

bioRxiv2023

Evolution of aberrant brain-wide spatiotemporal dynamics of resting-state networks in a Huntington's disease mouse model

Distinct resting-state networks (RSNs) are differentially altered in the course of Huntingtons disease (HD). However, these RSN changes are depicted using traditional functional connectivity analyses which ignore the dynamic brain states that constitute these RSNs and their time-dependent relationship. Dynamic states are represented by recurring spatiotemporal patterns of propagating cortical and subcortical brain activity observed in low-frequency BOLD fluctuations, called quasi-periodic patterns (QPPs). In this study, we used resting-state fMRI to investigate QPPs in the zQ175DN mouse model of HD at 3, 6 and 10 months of age. We identified age- and genotype-specific short (3s) QPPs, representative of the lateral cortical network (LCN) and the default mode-like network (DMLN), and a long (10s) QPP, the homolog of the human primary QPP, exhibiting the propagation of activity between the LCN and the DMLN. Hyperactivity was present in the caudate putamen and the somatosensory cortex in zQ175DN mice at 3 and 6 months of age. Moreover, DMLN-wide reduction in activation was observed at all ages, where at 6 and 10 months of age the reduced activity gradually advanced into a breakdown of the LCN-to-DMLN propagation. We then investigated the relationship in the timing of peak activity of six brain regions involved in the long QPPs and found that the retrosplenial cortex, a transmodal region which orchestrates multisensory integration, has a premature peak of BOLD activation in zQ175DN mice at 6 months of age, as compared to age-matched controls. Irrespective of either LCN or DMLN activation, this resulted in an asynchrony of the retrosplenial cortex in the peak timing relationships relative to other regions during the long (10s) QPPs. Finally, the normative, age-dependent, wild-type QPPs were significantly decreased in occurrence in the zQ175DN group at each age, indicating the presence of phenotypically-driven LCN and DMLN states as captured with QPPs. As BOLD-dependent variations result from neurovascular coupling, we assessed mutant huntingtin (mHTT) deposition in astrocytes and pericytes, known components of the neurogliovascular unit. These analyses showed increased cell-type dependent deposition starting at 6 months in the caudate putamen, somatosensory and motor cortex, regions that are prominently involved in HD pathology as seen in humans. Our findings provide meaningful insights into the development and progression of altered functional brain dynamics in this HD model, opening potential new avenues for its application in clinical HD research. SUMMARYHuntingtons disease (HD) is marked by irreversible loss of neuronal function for which currently no availability for disease-modifying treatment exists. Advances in the understanding of disease progression can aid biomarker development, which in turn can accelerate therapeutic discovery. We characterized the progression of altered dynamics of whole-brain network states in the zQ175DN mouse model of HD using a dynamic functional connectivity (FC) approach to resting-state fMRI and identified quasi-periodic patterns (QPPs) of brain activity constituting the most prominent resting-state networks. The occurrence of the normative QPPs, as observed in healthy controls, was reduced in the HD model as the phenotype progressed. This uncovered progressive cessation of synchronous brain activity with phenotypic progression, which is not observed with the conventional static FC approaches. This work opens new avenues in assessing the dynamics of whole brain states, through QPPs, in clinical HD research.

neuroscience↗

bioRxiv2021

MYCN-induced nucleolar stress drives an early senescence-like transcriptional program in hTERT-immortalized RPE cells

MYCN is an oncogenic driver in neural crest-derived neuroblastoma and medulloblastoma. To better understand the early effects of MYCN activation in a neural-crest lineage context, we profiled the transcriptome of immortalized human retina pigment epithelial cells with inducible MYCN activation. Gene signatures associated with elevated MYC/MYCN activity were induced after 24 h of MYCN activation, which attenuated but sustained at later time points. Unexpectedly, MYCN activation was accompanied by reduced cell growth. Gene set enrichment analysis revealed a senescence-like signature with strong induction of p53 and p21 but in the absence of canonical hallmarks of senescence such as beta-galactosidase positivity, suggesting incomplete cell fate commitment. When scrutinizing the putative drivers of this growth attenuation, differential gene expression analysis identified several regulators of nucleolar stress. This process was also reflected by phenotypic correlates such as cytoplasmic granule accrual and nucleolar coalescence. Hence, we propose that the induction of MYCN congests the translational machinery, causing nucleolar stress and driving cells into a transient pre-senescent state. Our findings shed new light on the early events induced by MYCN activation and may help unravelling which factors are required for cells to tolerate unscheduled MYCN overexpression during early malignant transformation. HighlightsO_LIActivation of MYCN attenuates proliferation in RPE1 cells C_LIO_LIGrowth arrest is associated with an early senescence-like transcriptional signature C_LIO_LITranscriptional and phenotypic evidence of nucleolar stress C_LIO_LICDKN1A upregulation in G2 phase primes cells for faltering in subsequent G1 C_LI

cancer biology↗

bioRxiv2020

Large-scale curvature sensing by epithelial monolayers depends on active cell mechanics and nuclear mechanoadaptation

While many tissues fold in vivo in a highly reproducible and robust way, epithelial folds remain difficult to reproduce in vitro, so that the effects and underlying mechanisms of local curvature on the epithelial tissue remains unclear. Here, we photoreticulated polyacrylamide hydrogels though an optical photomask to create corrugated hydrogels with isotropic wavy patterns, allowed us to show that concave and convex curvatures affect cellular and nuclear shape. By culturing MDCK epithelial cells at confluency on corrugated hydrogels, we showed that the substrate curvature leads to thicker epithelial zones in the valleys and thinner ones on the crest, as well as corresponding density, which can be generically explained by a simple 2D vertex model, leading us to hypothesize that curvature sensing could arise from resulting density/thickness changes. Additionally, positive and negative local curvatures lead to significant modulations of the nuclear morphology and positioning, which can also be well-explained by an extension of vertex models taking into account membrane-nucleus interactions, where thickness/density modulation generically translate into the corresponding changes in nuclear aspect ratio and position, as seen in the data. Consequently, we find that the spatial distribution of Yes associated proteins (YAP), the main transcriptional effector of the Hippo signaling pathway, is modulated in folded epithelial tissues according to the resulting thickness modulation, an effect that disappears at high cell density. Finally, we showed that these deformations are also associated with changes of A-type and B-type lamin expression, significant chromatin condensation and to lower cell proliferation rate. These findings show that active cell mechanics and nuclear mechanoadaptation are key players of the mechanistic regulation of epithelial monolayers to substrate curvature, with potential application for a number of in vivo situations.Competing Interest StatementThe authors have declared no competing interest.View Full Text

biophysics↗

bioRxiv2020

p53 drives premature neuronal differentiation in response to radiation-induced DNA damage during early neurogenesis

p53 regulates the cellular DNA damage response (DDR). Hyperactivation of p53 during embryonic development, however, can lead to a range of developmental defects including microcephaly. Here, we induce microcephaly by acute irradiation of mouse fetuses at the onset of neurogenesis. Besides a classical DDR culminating in massive apoptosis, we observe ectopic neurons in the subventricular zone in the brains of irradiated mice, indicative of premature neuronal differentiation. A transcriptomic study indicates that p53 activates both DDR genes and differentiation-associated genes. In line with this, mice with a targeted inactivation of Trp53 in the dorsal forebrain, do not show this ectopic phenotype and partially restore brain size after irradiation. Irradiation furthermore induces an epithelial-to-mesenchymal transition-like process resembling the radiation-induced proneural-mesenchymal transition in glioma and glioma stem-like cells. Our results demonstrate a critical role for p53 beyond the DDR as a regulator of neural progenitor cell fate in response to DNA damage.

neuroscience↗

bioRxiv2020

Systematic quantification of synapses in primary neuronal culture

A vast set of neurological disorders is associated with impaired synaptic connectivity. Therefore, modulation of synapse formation could have therapeutic relevance. However, the high density and small size of synapses make their quantification a challenging task. To improve the reliability of synapse-oriented drug screens, we evaluated a panel of synapse-targeting antibodies for their labeling specificity on hippocampal and cortical cell cultures using quantitative immunofluorescence microscopy. For those antibodies that passed multiparametric validation, we assessed pairwise colocalization, an often-used readout for established synapses. We found that even when two pan-synaptic markers were used, the overlap was incomplete, and the presence of spurious signals limited the dynamic range. To circumvent this problem, we implemented a proximity ligation-based approach, that only leads to a signal when two pre- and postsynaptic markers are sufficiently close. We demonstrate that this approach can be applied to different synaptic marker combinations and can be successfully used for quantification of synapse density in cultures of different maturity stage in healthy or pathological conditions. Thus, the unbiased analysis of synapse labeling and exploitation of resident protein proximity, allows increasing the sensitivity of synapse quantifications in neuronal culture and therefore represents a valuable extension of the analytical toolset for in vitro synapse screens.

neuroscience↗

bioRxiv2019

Chemogenetic silencing of neurons in the mouse anterior cingulate area modulates neuronal activity and functional connectivity

The anterior cingulate area (ACA) is an integral part of the prefrontal cortex in mice and has been implicated in several cognitive functions. Previous anatomical and functional imaging studies demonstrated that the ACA is highly interconnected with numerous brain regions acting as a hub region in functional networks. However, the importance of the ACA in regulating functional network activity and connectivity remains to be elucidated. Recently developed neuromodulatory techniques, such as Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) allow for precise control of neuronal activity. In this study, we used an inhibitory kappa-opioid receptor DREADDs (KORD) to temporally inhibit neuronal firing in the right ACA of mice and assessed functional network activity and connectivity using non-invasive functional MRI. We demonstrated that KORD-induced inhibition of the right ACA induced blood oxygenation-level dependent (BOLD) signal decreases and increases in connected brain regions throughout of hemispheres. Furthermore, these modulations in neuronal activity were associated with decreased intra- and interhemispheric functional connectivity. These results demonstrate that the combination of the DREADD technology and non-invasive functional imaging methods is a valuable tool for unraveling the underlying mechanisms of network function and dysfunction.

neuroscience↗

bioRxiv2019

Deep coverage microscopy exposes a pharmacological window for modifiers of neuronal network connectivity

BackgroundTherapeutic developments for neurodegenerative disorders are redirecting their focus to the mechanisms that contribute to synaptic plasticity and the loss thereof. Identification of novel regulators requires a method to quantify neuronal network connectivity with high accuracy and throughput. To meet this demand, we have established a microscopy-based pipeline that integrates morphological and functional correlates of connectivity in primary neuronal culture. ResultsWe unveiled a connectivity signature that was specific to the cell type and culture age. We defined a score that accurately reports on the degree of neuronal connectivity and we validated this score by targeted perturbation of microtubule stability and selective depletion of anti-oxidants. With a focused compound screen, we discovered that inhibition of dual leucine zipper kinase activity increased neuronal connectivity in otherwise unperturbed cultures and exerted neuroprotective effects in cultures grown under sub-optimal or challenged conditions. ConclusionsOur results illustrate that profiling microscopy images with deep coverage enables sensitive interrogation of neuronal connectivity and allows exposing a dose and time window for pharmacological interventions. Therefore, the current approach holds promise for identifying pathways and compounds that preserve or rescue neuronal connectivity in neurodegenerative disorders.

neuroscience↗