Biology subjects

van Grunsven, L. A.

Publications and source records attributed to van Grunsven, L. A..

5 recordsLinked to original sources

bioRxiv2026

Increases in BCL2L1 and ID1 dosage synergistically drive fate bias and competitive advantage in human pluripotent stem cells

BackgroundGains of chromosome 20q11.21 are among the most common culture-acquired abnormalities in human pluripotent stem cells (hPSC), conferring a well-defined survival advantage while altering differentiation capacity. However, it remains unclear whether this advantage persists during differentiation, how the aneuploidy alters ectodermal and retinal pigment epithelium (RPE) lineage specification, and which genes within the minimal amplicon drive these effects. MethodsWe used three isogenic human embryonic stem cell line pairs (wild-type and 20q11.21 gain) and assessed their behaviour in two neuroectoderm differentiation systems: directed neuroectoderm induction (dual SMAD inhibition) and long-term spontaneous RPE differentiation. Competitive dynamics were measured in mixed cultures, and lineage outcomes were analysed using immunostaining, gene expression profiling and single-cell RNA sequencing. To identify driver genes, we generated BCL2L1 and ID1 overexpression lines and tested their effects under both directed and spontaneous differentiation conditions. ResultsAcross all lines and conditions, 20q cells expanded from a minor fraction to dominate mixed cultures, indicating that their competitive advantage persists beyond the undifferentiated state. Despite this dominance, pure 20q cells failed to specify to neuroectoderm or RPE. Single-cell transcriptomics revealed consistent diversion toward non-neural ectodermal and extraembryonic fates. Mechanistically, overexpression of BCL2L1 and ID1 alone or in combination impaired neuroectoderm specification, while synergistic effect of both genes promoted non-neural ectodermal outcomes under directed differentiation conditions. In spontaneous differentiation, both genes could disrupt differentiation. ConclusionsThe 20q11.21 gain couples a persistent survival advantage with a disruption of neural and RPE lineage competence, redirecting cells toward alternative ectodermal and extraembryonic fates. These effects arise from the combined action of two dosage-sensitive genes BCL2L1 and ID1 within the amplicon, illustrating how regional gene dosage can reshape developmental signalling responses in hPSC.

developmental biology↗

bioRxiv2023

Gain of 1q confers an MDM4-driven growth advantage to undifferentiated and differentiating hESC while altering their differentiation capacity

Gains of 1q are a highly recurrent chromosomal abnormality in human pluripotent stem cells. In this work, we show that gains of 1q impact the differentiation capacity to derivates of the three germ layers, leading to miss-specification to cranial placode and non-neural ectoderm during neuroectoderm differentiation and by poorer expression of lineage specific markers in hepatoblasts and cardiac progenitors. Competition assays show that the cells retain their selective advantage during differentiation, which is mediated by a higher expression of MDM4, a gene located in the common region of gain. MDM4 drives the winner phenotype of the mutant cells in both the undifferentiated and differentiating state by reducing the cells sensitivity to DNA-damage through decreased p53-mediated apoptosis. Finally, we find that cell density in culture plays a key role in promoting the competitive advantage of the cells by increasing DNA damage.

cell biology↗

bioRxiv2023

Osmolar modulation drives reversible cell cycle exit and human pluripotent cell differentiation via NF-κB and WNT signaling

Terminally differentiated cells are regarded as the most stable and common cell state in adult organisms as they reside in growth arrest and carry out their cellular function. Improving our understanding of the mechanisms involved in promoting cell cycle exit would facilitate our ability to manipulate pluripotent cells into mature tissues for both pharmacological and therapeutic use. Here, we demonstrated that a hyperosmolar environment enforced a protective p53-independent quiescent state in dedifferentiated hepatoma cells and pluripotent stem cells (PSCs)-derived models of human hepatocytes and endothelial cells, representing the endodermal and mesodermal lineages. Prolonged culture in hyperosmolar conditions stimulated transcriptional and functional cell maturation. Interestingly, hyperosmolar conditions did not only trigger cell cycle exit and cellular maturation but were also necessary to maintain this maturated state, as switching back to plasma osmolarity caused the loss of maturation markers and the gain of proliferative markers. Transcriptome analysis revealed activation of NF-{kappa}[V] and repression of WNT signaling as the two main pathways downstream of osmolarity-regulated growth arrest and cell maturation, respectively. This study revealed that increased osmolarity serves as a biochemical signal to promote long-term growth arrest, transcriptional changes, and maturation into different lineages, serving as a practical method to generate differentiated hiPSCs that resemble their mature counterpart more closely.

developmental biology↗

bioRxiv2020

A fully defined pluripotent stem cell derived multi-liver-cell model for steatohepatitis and fibrosis

Chronic liver injury, as observed in non-alcoholic steatohepatitis (NASH), progressive fibrosis, and cirrhosis, remains poorly treatable. Steatohepatitis causes hepatocyte loss in part by a direct lipotoxic insult, which is amplified by derangements in the non-parenchymal cellular (NPC) interactive network wherein hepatocytes reside, including, hepatic stellate cells, liver sinusoidal endothelial cells and liver macrophages. To create an in vitro culture model encompassing all these cells that allows studying liver steatosis, inflammation and fibrosis caused by NASH, we here developed a fully defined hydrogel microenvironment, termed hepatocyte maturation (HepMat) gel, that supports maturation and maintenance of pluripotent stem cell (PSC)-derived hepatocyte- and NPC-like cells for at least one month. The HepMat-based co-culture system modeled key molecular and functional features of TGF{beta}-induced liver fibrosis and fatty-acid induced inflammation and fibrosis better than monocultures its constituent cell populations. The novel co-culture system should open new avenues for studying mechanisms underlying liver steatosis, inflammation and fibrosis as well as for assessing drugs counteracting these effects.

bioengineering↗

bioRxiv2020

PU.1 drives specification of pluripotent stem cell-derived endothelial cells to LSEC-like cells

To date there is no representative in vitro model for liver sinusoidal endothelial cells (LSECs), as primary LSECs dedifferentiate very fast in culture and no combination of cytokines or growth factors can induce an LSEC fate in (pluripotent stem cell-derived) endothelial cells (ECs). Furthermore, the transcriptional programs driving an LSEC fate have not yet been described. Here, we first present a computational workflow (CenTFinder) that can identify transcription factors (TFs) that are crucial for modulating pathways involved in cell lineage specification. Using CenTFinder, we identified several novel LSEC-specific protein markers such as FCN2 and FCN3, which were validated by analysis of previously published single-cell RNAseq data. We also identified PU.1 (encoded by the SPI1 gene) as a major regulator of LSEC-specific immune functions. We show that SPI1 overexpression (combined with the general EC transcription factor ETV2) in human pluripotent stem cells (PSCs) induces ECs with an LSEC-like phenotype. The ETV2-SPI1-ECs display increased expression of LSEC markers such as CD32B and MRC1 as well as several of the proposed novel markers. More importantly, ETV2-SPI1-ECs acquire LSEC functions, including uptake of FSA-FITC as well as labelled IgG. In conclusion, we present the CenTFinder computational tool to identify key regulatory TFs within specific pathways, in this work pathways of lineage specification, and we demonstrate its use by the identification and validation of PU.1 as a master regulator for LSEC fating.

developmental biology↗