Biology subjects

Svendsen, C. N.

Publications and source records attributed to Svendsen, C. N..

12 recordsLinked to original sources

bioRxiv2026

Differentiation protocol and maturation shape the axial identity and synaptic state in human iPSC-derived spinal motor neurons

Human induced pluripotent stem cell (iPSC)-derived motor neurons are widely used for disease modeling. Stem cells can be differentiated into motor neurons by exposure to specific patterning morphogens, which can ultimately determine terminal cellular composition and developmental state. We compared two small molecule workflows for human iPSC maturation into motor neurons: a rapid direct protocol (diMN protocol) and an extended protocol (purMN protocol), with both protocols matched for 20 days in maturation medium. While the diMN protocol generated mixed neural cultures, the purMN protocol provided cultures with greater motor neuron enrichment, stronger caudal spinal identity, broader synaptic and cholinergic programs, and reduced progenitor-associated signatures. By extending the purMN protocol to 32 days in maturation media, cultures developed reinforced spinal, postsynaptic, presynaptic, calcium-signaling, and cholinergic features. Across four independent reference frameworks, deconvolution aligned diMN cultures with anterior and progenitor-associated states and purMN cultures with spinal and post-mitotic states. In a microfluidic co-culture system, purMNs formed denser, more highly branched distal neurite networks and produced more innervated acetylcholine receptor clusters compared to diMNs. Revealing how purity and maturation jointly define the transcriptomic and structural state of human iPSC-derived spinal motor neuron cultures will help the field choose optimal models for studies of neurodevelopment and neurodegenerative diseases.

neuroscience↗

bioRxiv2026

Multiple motor proteins regulate ALS-linked TDP-43 anterograde axonal transport

TDP-43 is an RNA-binding protein essential for RNA metabolism. Under physiological conditions, it predominantly resides in the nucleus but is also expressed in the cytoplasm, where it regulates mRNA trafficking and local translation. In nearly 97% of Amyotrophic Lateral Sclerosis (ALS) cases, TDP-43 undergoes nuclear depletion and cytoplasmic aggregation. While its nuclear functions are well characterized, its axonal roles remain poorly understood, despite axonal degeneration being a hallmark of ALS pathology. To address this gap, we investigated TDP-43 localization and transport dynamics in axons of H9-derived human neurons. We compared fluorescently labeled TDP-43 with three well-characterized axonal cargoes, Rab5, synaptophysin, and APP, and examined protein-protein interactions between TDP-43 and the axonal transport machinery. Our analyses revealed that TDP-43 exhibits active anterograde axonal transport and interacts with multiple kinesin motor proteins, including all three KIF5 isoforms, through the adaptor KLC1, and the synaptic vesicles motor KIF1A. This multi-motor engagement suggests a flexible transport system that ensures mRNA delivery to distal axons. In ALS, where TDP-43 accumulates abnormally in the cytoplasm, this flexibility may become compromised, with multiple transport mechanisms simultaneously affected. This could contribute to progressive accumulation of non-functional TDP-43 granules, disrupting mRNA trafficking and local translation. Our findings provide a foundation for understanding how physiological TDP-43 transport mechanisms may be impaired during disease, highlighting axonal TDP-43 transport pathways as potential therapeutic targets.

neuroscience↗

bioRxiv2026

Single-cell proteomic maps of human induced pluripotent stem cells and their differentiation into motor neurons

The majority of single cell-studies use RNA to identify the cell state. However, many RNAs are transient and decrease in response to elevated protein to control homeostasis. Thus, final cell states are largely defined by their unique and dynamic protein composition not RNA. Here a single-cell proteomics approach was used to identify the proteomic profile of human induced pluripotent stem cells (iPSCs) before and during their in vitro differentiation to motor neurons. By measuring up to 1000 proteins in each cell, novel clusters of growing iPSCs could be characterized along with a new proteomic pathway that defines motor neuron development. Interestingly, there was a dynamic and cell type-specific discordance between the protein levels and their corresponding messenger RNAs. This lays the foundation for drawing new single-cell proteomic maps of developing human tissues. IN BRIEFIn this article, we report the first single-cell proteomic maps of induced pluripotent stem cells (iPSCs) and their differentiation into motor neurons (MNs). By identifying proteomes of individual cells, we resolve iPSC and MN states, their proteomic, metabolomic and organellar heterogeneity. We show considerable, stage-specific discordance between the transcriptome and the proteome in differentiated neurons.

neuroscience↗

bioRxiv2025

Human iPSC-derived prostate organoids with germline BRCA2 mutation undergo tumorigenic transformations

The lack of physiologically relevant in vitro prostate models has impeded studies of organ development and prostate tumorigenesis. We reprogrammed peripheral blood mononuclear cells (PBMCs) from individuals with and without pathogenic-germline BRCA2 mutation (MUT_BRCA2, CON_BRCA2) into induced pluripotent stem cells (iPSCs), which showed no differences in morphology, proliferation, or pluripotency markers. Differentiation of MUT_BRCA2 iPSCs into prostate organoids (iPROS) using defined growth factors and signaling molecules resulted in disrupted morphology, impaired polarity, increased proliferation, and elevated prostate-specific antigen (PSA) secretion compared to CON_BRCA2 iPROS. Transcriptomic profiling revealed early prostate cancer (PCa) signatures. Upon exposure to dietary carcinogens, MUT_BRCA2 iPROS showed further PSA elevation, enhanced proliferation, AMACR upregulation, p63 reducetion are markers of aggressive PCa. In vivo, MUT_BRCA2 iPROS formed tumors in immunodeficient mice. This patient-derived iPROS-platform recapitulates human-prostate mopphology and function, models early tumorigenesis events, and provides a valuable tool for studying PCa biology and enabling personalized drug discovery. IN BRIEFIn this study, we developed patients iPSC-derived prostate organoids (iPROS) with or without a pathogenic BRCA2 germline mutation that display human-prostate like morphology and function. MUT_BRCA2 iPROS displayed disrupted morphology, early tumorigenic changes, and formed tumors in mice. Upon carcinogen exposure, they showed markers of aggressive prostate cancer. This platform models early prostate tumorigenesis and enables personalized studies of cancer initiation and therapeutic response.

cancer biology↗

bioRxiv2025

Dynamic Transcriptomic Remodeling in Human Neural Progenitor Cells Reveals Mechanisms for Vision Preservation in Retinitis Pigmentosa Model

Human neural progenitor cells (hNPCs) have shown promise in slowing down retinal degeneration in animal models and are currently being tested in clinical trials for treating retinitis pigmentosa (RP). However, the status of grafted hNPCs and their interaction with host retinal cells over time is largely unknown. Here, we investigated single-cell transcriptomic changes in grafted hNPCs and host retinal cells following injection into a rodent model for RP. Grafted hNPCs and host retinal cells undergo dynamic transcriptomic changes in the degenerative retinal environment. Grafted hNPCs protect vision through multiple mechanisms, including trophic factor support, modulation of metabolic activity, reduction of apoptosis, oxidative stress, and inflammation, alongside extracellular matrix remodeling. CellChat analysis revealed a progressive decline in intercellular signaling and communication strength between hNPCs and host retinal cells over time. This study indicates that enhancing trophic factor supports and improving host retinal environment are key targets to enable long-term vision preservation.

neuroscience↗

bioRxiv2023

Disease related changes in ATAC-Seq of more than 450 iPSC-derived motor neuron lines from ALS patients and controls

Amyotrophic Lateral Sclerosis (ALS), like many other neurodegenerative diseases, is highly heritable, but with only a small fraction of cases explained by monogenic disease alleles. To better understand sporadic ALS, we report epigenomic profiles, as measured by ATAC-seq, of motor neuron cultures derived from a diverse group of 380 ALS patients and 80 healthy controls. We find that chromatin accessibility is heavily influenced by sex, the iPSC cell type of origin, ancestry, and the inherent variance arising from sequencing. Once these covariates are corrected for, we are able to identify robust ALS-specific signals in the data. Additionally, we find that the ATAC-seq data is able to predict ALS disease progression rates with similar accuracy to methods based on biomarkers and clinical status. These results suggest that iPSC-derived motor neurons recapitulate important disease-relevant epigenomic changes.

neuroscience↗

bioRxiv2021

AAV9-MCT8 delivery at juvenile stage ameliorates neurological and behavioral deficits in an AHDS mouse model

Allan-Herndon-Dudley syndrome (AHDS) is a severe X-linked intellectual and psychomotor disability disorder accompanied by abnormal thyroid hormone (TH) levels. AHDS is caused by inactivating mutations in the monocarboxylate transporter 8 (MCT8), a specific TH transporter widely expressed in the central nervous system. MCT8 gene mutations cause impaired transport of TH across brain barriers, leading to insufficient neural TH supply. There is currently no successful therapy for the neurological symptoms. AAV9-based gene therapy is a promising approach to treat monogenic neurological disorders. Here, the potential of this approach was tested in the well-established double knockout (dKO) Mct8-/y;Oatp1c1-/- mouse model of AHDS, which displays disease-relevant neurological and TH phenotypes. Systemic intravenous delivery of AAV9-MCT8 at a juvenile stage led to improved locomotor and cognitive function, as well as rescue of T3-brain content and T3-related gene expression. This preclinical study indicates that this gene therapy may improve the neurological symptoms of AHDS patients.

neuroscience↗

bioRxiv2020

Antiviral Drug Screen of Kinase inhibitors Identifies Cellular Signaling Pathways Critical for SARS-CoV-2 Replication

Emergence of a highly contagious novel coronavirus, SARS-CoV-2 that causes COVID-19, has precipitated the current global health crisis with over 479,000 deaths and more than 9.3 million confirmed cases. Currently, our knowledge of the mechanisms of COVID-19 disease pathogenesis is very limited which has hampered attempts to develop targeted antiviral strategies. Therefore, we urgently need an effective therapy for this unmet medical need. Viruses hijack and dysregulate cellular machineries in order for them to replicate and infect more cells. Thus, identifying and targeting dysregulated signaling pathways that have been taken over by viruses is one strategy for developing an effective antiviral therapy. We have developed a high-throughput drug screening system to identify potential antiviral drugs targeting SARS-CoV-2. We utilized a small molecule library of 430 protein kinase inhibitors, which are in various stages of clinical trials. Most of the tested kinase antagonists are ATP competitive inhibitors, a class of nucleoside analogs, which have been shown to have potent antiviral activity. From the primary screen, we have identified 34 compounds capable of inhibiting viral cytopathic effect in epithelial cells. Network of drug and protein relations showed that these compounds specifically targeted a limited number of cellular kinases. More importantly, we have identified mTOR-PI3K-AKT, ABL-BCR/MAPK, and DNA-Damage Response (DDR) pathways as key cellular signaling pathways critical for SARS-CoV-2 infection. Subsequently, a secondary screen confirmed compounds such as Berzosertib (VE-822), Vistusertib (AZD2014), and Nilotinib with anti SARS-CoV-2 activity. Finally, we found that Berzosertib, an ATR kinase inhibitor in the DDR pathway, demonstrated potent antiviral activity in a human epithelial cell line and human induced pluripotent stem cell (hIPSC)-derived cardiomyocytes. These inhibitors are already in clinical trials of phase 2 or 3 for cancer treatment, and can be repurposed as promising drug candidates for a host-directed therapy of SARS-CoV-2 infection. In conclusion, we have identified small molecule inhibitors exhibiting anti SARS-CoV-2 activity by blocking key cellular kinases, which gives insight on important mechanism of host-pathogen interaction. These compounds can be further evaluated for the treatment of COVID-19 patients following additional in vivo safety and efficacy studies. DisclosuresNone declared.

microbiology↗

bioRxiv2020

Single-cell RNA-seq analysis of human iPSC-derived motor neurons resolves early and predictive ALS signatures

Induced pluripotent stem cell (iPSC) derived neural cultures from amyotrophic lateral sclerosis (ALS) patients can reflect disease phenotypes targetable by treatments. However, widely used differentiation protocols produce mixtures of progenitors, neurons, glia, and other cells at various developmental stages and rostrocaudal neural tube segments. Here we present a methodology using single-cell RNA sequencing analysis to distinguish cell type expression in C9orf72 ALS, sporadic ALS, control, and genome-edited cultures across multiple subjects, experiments, and commercial platforms. Combinations of HOX and developmental gene expression with global clustering classified rostrocaudal, progenitor, and mantle zone fates. This demonstrated that iPSC-differentiated cells recapitulate fetal hindbrain and spinal cord development and resolved early, reproducible, and motor neuron-specific signatures of familial and sporadic ALS. This includes downregulated ELAVL3 expression, which persists into disease endstages. Single-cell analysis thus yielded predictive ALS markers in other human and mouse models which were otherwise undiscovered through bulk omics assays.

neuroscience↗

bioRxiv2020

Human iPSC-Derived Cardiomyocytes are Susceptible to SARS-CoV-2 Infection

Coronavirus disease 2019 (COVID-19) is a viral pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 is predominantly defined by respiratory symptoms, but cardiac complications including arrhythmias, heart failure, and viral myocarditis are also prevalent. Although the systemic ischemic and inflammatory responses caused by COVID-19 can detrimentally affect cardiac function, the direct impact of SARS-CoV-2 infection on human cardiomyocytes is not well-understood. We used human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) as a model system to examine the mechanisms of cardiomyocyte-specific infection by SARS-CoV-2. Microscopy and immunofluorescence demonstrated that SARS-CoV-2 can enter and replicate within hiPSC-CMs, localizing at perinuclear locations within the cytoplasm. Viral cytopathic effect induced hiPSC-CM apoptosis and cessation of beating after 72 hours of infection. These studies show that SARS-CoV-2 can infect hiPSC-CMs in vitro, establishing a model for elucidating the mechanisms of infection and potentially a cardiac-specific antiviral drug screening platform.

cell biology↗

bioRxiv2020

G4C2 repeat RNA mediates the disassembly of the nuclear pore complex in C9orf72 ALS/FTD

Nucleocytoplasmic transport, controlled by the nuclear pore complex, has recently emerged as a pathomechanism underlying neurodegenerative diseases including C9orf72 ALS/FTD. However, little is known about the underlying molecular events and the underlying biology in human neurons. Using super resolution structured illumination microscopy of twenty three nucleoporins in nuclei from C9orf72 iPSC derived neurons and postmortem human tissue we identify a unique subset of eight nucleoporins lost from human neuronal nuclei. POM121, an integral transmembrane nucleoporin, appears to coordinate the composition of the nucleoporins within human neuronal nuclei ultimately impacting nucleocytoplasmic transport, and subsequent cellular toxicity in C9orf72 iPSNs. These data suggest that POM121 is a critical nucleoporin in the maintenance of the nuclear localization of specific nucleoporins in human neurons. Moreover, loss of nuclear POM121, as a result of expanded C9orf72 ALS/FTD repeat RNA, initiates a pathological cascade affecting nucleoporin composition within neuronal nuclei, nuclear pore complex function, and overall downstream neuronal survival.

neuroscience↗

bioRxiv2020

Generation of twenty four induced pluripotent stem cell lines from twenty four members of the Lothian Birth Cohort 1936

Cognitive decline is among the most feared aspects of ageing. We have generated induced pluripotent stem cells (iPSCs) from 24 people from the Lothian Birth Cohort 1936, whose cognitive ability was tested in childhood and in older age. Peripheral blood mononuclear cells (PBMCs) were reprogrammed using non-integrating oriP/EBNA1 backbone plasmids expressing six iPSC reprogramming factors (OCT3/4 (POU5F1), SOX2, KLF4, L-Myc, shp53, Lin28, SV40LT). All lines demonstrated STR matched karyotype and pluripotency was validated by multiple methods. These iPSC lines are a valuable resource to study molecular mechanisms underlying individual differences in cognitive ageing and resilience to age-related neurodegenerative diseases.

cell biology↗