Biology subjects

Bonnet, D.

Publications and source records attributed to Bonnet, D..

15 recordsLinked to original sources

bioRxiv2026

A combined program of induced stemness and differentiation in response to interferon gamma drives acute myeloid leukaemia growth

How inflammation shapes acute myeloid leukaemia (AML) has come under scrutiny, as it may explain the disease resistance to immunotherapy approaches. IFNg has emerged as a key cytokine with paradoxical roles in suppressing and supporting AML growth, and the fundamental question of how leukemic stem cells (LSCs) respond to IFNg and whether IFNg signaling influences LSC quiescence and their capacity to regenerate disease over the long term remains unanswered. Here, we study primary human AML cells and murine models and combine bioinformatics analyses and functional assays to show that AML hierarchical heterogeneity is responsive to IFNg challenge. We uncover that IFNg triggers parallel stemness and differentiation programs; HSC/MPP-like cells enter deeper stemness, associated with quiescence and high leukaemia regeneration potential, while the surviving pool of progenitor-like cells divides faster but produces progeny that is quickly lost. Finally, with murine models we show that exposure to inflammation in vivo results in only transient impairment of leukaemia propagating capacity. This mechanism is driven by a previously unrecognised intraclonal fate bifurcation, relevant for the development of more effective therapeutic approaches.

cancer biology↗

bioRxiv2026

IBD-Derived Colonic Fibroblasts Exhibit an Osteopontin-Enriched Secretome, and Osteopontin Restrains Human Colonic Organoid Maturation

Background: Intestinal fibroblasts are extensively remodeled in inflammatory bowel disease (IBD), yet the soluble stromal signals that directly influence epithelial maturation remain incompletely understood. We examined whether fibroblasts derived from inflamed IBD colon display an osteopontin (OPN; SPP1)-enriched secretory phenotype and whether extracellular OPN directly modifies non-neoplastic human colonic epithelium. Methods: Conditioned media from 5 noninflamed-associated fibroblast (NAF) and 4 inflammatory-associated fibroblast (IAF) cultures were analyzed in the validated multi-donor cytokine-array matrix, with orthogonal SPP1 RT-qPCR validation in a complementary fibroblast cohort. Recombinant OPN was then tested in human colonic organoids from 3 donors using donor-resolved molecular and functional analyses under standard, fibroblast-conditioned, and WNT-modified culture conditions. Donor identity defined biological replication. Results: OPN showed the strongest positive rank-based separation between IAF and NAF cultures: all 4 IAF values were higher than all 5 NAF values (Cliff's delta=1.00; exact Mann-Whitney P=0.0159; median ratio=3.64; Benjamini-Hochberg q=.19). Fibroblast RT-qPCR showed approximately 10-fold higher mean SPP1 expression in IAF than NAF cultures (P<.05). In organoids, OPN consistently reduced KRT20, FABP1, CA2, and MUC2 from Day 5 to Day 9. SOX9, HES1, and NOTCH1 increased at Day 9, whereas LGR5 and ALDH provided no evidence of canonical stem-cell expansion. Organoid-area and EdU responses were modest and donor dependent. Conclusions: IBD-derived colonic fibroblasts can display an OPN-enriched secretory phenotype. In human colonic organoids, OPN is sufficient to impair epithelial maturation, whereas its effects on growth and proliferation are variable and depend on the surrounding niche.

physiology↗

bioRxiv2026

Intravital imaging uncovers remodelling of humanised bone marrow-like niches

The bone marrow haematopoietic niche is composed of a diverse array of cell types and extracellular matrix components that together support healthy haematopoiesis. However, live imaging of the bone marrow microenvironment is hampered by tissue accessibility limitations. Using intravital imaging through a titanium imaging window, we investigated the dynamics of human haematopoietic cells and mesenchymal stromal cells within an ectopically implanted humanised scaffold in an immunodeficient murine host. These cell populations expand and differentiate over time, accompanied by progressive remodelling of the scaffold. We observe migration of murine endothelial cells into the scaffold, leading to the formation of a vascular network during the initial development of the humanised niche. Subsequently, the dense collagen matrix that makes up the implanted niche is altered and larger gaps form in regions populated by mesenchymal stroma cells. Collectively, our findings demonstrate dynamic remodelling of the extracellular milieu that supports haematopoietic cell development and establish a platform for longitudinal, in vivo investigation of these processes. Altogether, we describe a novel model that aligns with the 3R guiding principles and enables real-time assessment of bone marrow cell dynamics in vivo. Summary statementRatcliffe and Mian et al. image in vivo dynamics of a bone marrow haematopoietic niche model.

cell biology↗

bioRxiv2026

A metabolically resistant spexin analogue, LIT-01-144, induces potent non-opioid peripheral antinociception in persistent pain via activation of GALR2

Chronic pain affects a significant portion of the global population and imposes substantial clinical and socioeconomic burdens. Current treatments mainly rely on opioid analgesics, which carry serious risks of dependence and misuse, underscoring the urgent need for alternative therapeutic strategies. Galanin receptors (GALR1-3) are known to be involved in modulating pain, yet their specific roles remain poorly understood due to the lack of receptor subtype-selective ligands. Recently, spexin has been identified as an endogenous peptide that selectively activates GALR2 and GALR3, offering a new scaffold for developing pharmacological tools targeting these receptor subtypes. In this study, we report the design and characterization of a modified spexin analog, LIT-01-144, engineered through N-terminal functionalization with a fluorocarbon chain to improve metabolic stability while preserving receptor selectivity. In vitro assays showed that LIT-01-144 has high potency at GALR2 and GALR3, with minimal activity at GALR1. Pharmacokinetic studies revealed a significantly longer plasma half-life compared to native spexin and no central nervous system penetration. In mice, intracerebroventricular administration of LIT-01-144 produced strong antinociceptive effects at doses ten times lower than spexin. While systemic administration showed no notable antinociception in naive animals, LIT-01-144 significantly reduced pain responses in a mouse model of persistent inflammatory pain induced by complete Freunds adjuvant (CFA). This antinociceptive activity was specifically mediated through GALR2 and was independent of opioid receptor pathways. In situ hybridization further showed an increase in Galr2-positive neurons in dorsal root ganglia of inflamed mice. Overall, these findings highlight GALR2 as a promising peripheral target for developing non-opioid analgesics and demonstrate the potential of LIT-01-144 as a valuable tool for understanding GALR2-mediated mechanisms of pain modulation.

pharmacology and toxicology↗

bioRxiv2025

DNMT3A-R882 mutations intrinsically drive dysfunctional neutropoiesis from human haematopoietic stem cells

Clonal haematopoiesis (CH) arises from the expansion of hematopoietic stem cells (HSCs) carrying leukaemia-associated somatic mutations. CH is linked to pathological immune dysregulation and a greater risk of age-related inflammatory diseases. Yet, how CH mutations impact HSC differentiation into immune effector cells remains understudied. Here, we report a single-cell resolution functional and multi-omic investigation of HSC clonal and differentiation dynamics in individuals with DNMT3A-R882 CH. DNMT3A-R882 reshapes the clonal architecture of haematopoiesis towards an aged phylogenetic structure. Functionally, DNMT3A-R882 HSCs produce decreased monocytic output but more abundant and mature neutrophil progeny compared to WT HSCs in the same individual. Whereas DNMT3A-R882 myeloid progenitors display attenuated inflammatory transcriptional programmes, DNMT3A- R882 mature neutrophils acquire proinflammatory and immunomodulatory features typical of maladaptive immunity and CH co-morbidities. Our findings, validated in humanised mice, identify aberrant DNMT3A-R882 HSC-driven neutropoiesis as a key link between CH, immune dysregulation and risk of inflammatory disease.

cell biology↗

bioRxiv2025

Granulocyte Derived Resistin Inhibits Monocyte Maturation and Induces Immune Suppression in CMML

Chronic myelomonocytic leukaemia (CMML) is a haematological malignancy characterised by overlapping myeloid dysplasia and proliferation with persisting monocytosis. While monocytes are the cardinal malignant cell type in CMML, as a stem cell neoplasm the disease clone comprises most lineages and differentiation stages, including granulocytes. To investigate the pathogenic contribution of granulocytes in CMML maintenance and progression, we performed phenotypic, transcriptomic and functional characterization of CMML granulocytes. Compared with healthy age-matched controls, CMML granulocytes exhibit defective maturation with reduced granularity and phagocytic capacity. Transcriptome analysis revealed activation of pathways linked to proliferation, Myc activity and inflammation. Notably, RETN, which encodes the inflammatory mediator resistin, was upregulated approximately 100-fold in CMML granulocytes; but not differentially expressed in CMML PBMNCs, sorted monocytes, or stem and progenitor cells compared to their healthy counterparts. Accordingly, resistin protein levels were 10-fold higher in plasma from CMML patients and higher plasma resistin levels correlate with poor overall survival and AML-free survival. Remarkably, exposure of healthy monocytes to exogenous recombinant resistin inhibited monocyte maturation and macrophage differentiation. Transcriptome analysis of resistin treated monocytes revealed that resistin induces gene signatures related to immune suppression and myeloid-derived suppressor cell phenotype. We found SEMA4A to be a downstream target of resistin and overexpressed in CMML monocytes. Consistent with known roles for SEMA4A, CMML patients displayed higher percentage of Tregs and elevated Th2/Th1 ratio compared with healthy controls and percentage of Tregs corresponding with associated resistin levels. Furthermore, we demonstrated that resistin directly skews the Th2/Th1 ratio via binding to monocytes. In conclusion, we showed that immature granulocytes in CMML produce high levels of resistin, which contributes to defective monocyte maturation and immune suppression.

cancer biology↗

bioRxiv2025

DEK::NUP214 acts as an XPO1-dependent transcriptional activator of essential leukemia genes

The t(6;9)(p22.3;q34.1) translocation/DEK::NUP214 fusion protein defines a distinct subgroup of younger AML patients classified as a separate disease entity by the World Health Organization. DEK is a nuclear factor with multifunctional roles, including gene regulation, while its fusion partner, NUP214, plays a pivotal role in nuclear export by interacting with transport receptors such as XPO1. However, the precise mechanism by which DEK::NUP214 drives leukemia remains unclear. A comprehensive multi-omics comparison of 57 AML primary samples (including whole genome sequencing, targeted sequencing, transcriptomics, and drug screening with > 500 compounds) revealed that t(6;9) cases display a selective response to XPO1 inhibitors (Selinexor & Eltanexor) and a distinct transcriptomic signature characterized by the overexpression of FOXC1 and HOX genes that are key leukemia mediators. CUT&RUN experiments demonstrated the direct binding of DEK::NUP214 to the promoters of FOXC1 and HOXA/B clusters. Strikingly, the expression of these genes and the binding of DEK::NUP214 to their regulatory regions were selectively reduced upon XPO1 inhibition in t(6;9) cells. Altogether, these results identified a novel function of DEK::NUP214 as an XPO1-dependent transcriptional activator of key leukemia drivers and provide a rationale to explore the use of XPO1 inhibitors in this patient population.

cancer biology↗

bioRxiv2024

Unveiling mechanical interactions between cell division and extracellular matrix in human colonic epithelium organoids: A 4D study using DVC

Cell division is a major event in tissue homeostasis, enabling renewal and regeneration. Stem cells, in particular, play an important role in this homeostasis, thanks to their ability to perform symmetric or asymmetric cell divisions. To study cell division, the human colon epithelium represents a model of choice due to its rapid renewal and therefore high proliferative potential. Currently, studying the live mechanical interactions between the epithelium and its matrix in vivo is challenging due to the lack of suitable methods. 3D human colon organoids seeded in Matrigel(R) are good models for this purpose as, from isolated stem cells, they recapitulate the tissue architecture organization and properties. This culture set-up also allows to study the matrix displacements around the organoid. Here, we studied the impact of cell division within the colonic epithelium on the extracellular matrix. We performed and validated an original experimental and analytical process with a 3D time-lapse confocal microscopy to follow cell mitosis and matrix movements on which we performed Digital Volume Correlation. We showed that these two different types of cell division impact the matrix differently with the asymmetric divisions causing a mainly uniaxial displacement, whereas symmetric ones involved a multiaxial and more important one.

cell biology↗

bioRxiv2024

Acute, chronic and conditioned effects of intranasal oxytocin in the mu opioid receptor knockout mouse model of autism: social context matters

Autism Spectrum Disorders (ASD) are neurodevelopmental disorders whose diagnosis relies on deficient social interaction and communication together with repetitive behaviours. Multiple studies have highlighted the potential of oxytocin (OT) to ameliorate behavioural abnormalities in animal models and subjects with ASD. Clinical trials, however, yielded disappointing results. Our study aimed at challenging hypotheses accounting for such negative results by assessing the behavioural effects of different regimens of OT administration in the Oprm1 null mouse model of ASD. We assessed the effects of intranasal OT injected once at different doses and time points following administration, or chronically, on ASD-related behaviours in Oprm1+/+and Oprm1-/- mice. We then tested whether pairing intranasal OT injection with social experience would influence its outcome on ASD-like core symptoms, and measured gene expression in several regions of the reward/social circuit. Acute intranasal OT improved social behaviour in Oprm1-/- mice at a moderate dose (0.3 IU) shortly after administration (5 min). Effects on non-social behaviours were limited. Chronic OT at this dose maintained beneficial effects in Oprm1 null mice but was deleterious in wild-type mice. Finally, improvements in the social behaviour of Oprm1-/- mice were greater and longer lasting when OT was administered in a social context, while the expression of OT and vasopressin receptor genes, as well as marker genes of striatal projection neurons, was suppressed. Our results highlight the importance of considering dosage and social context when evaluating the effects of OT treatment in ASD. HighlightsO_LIAcute intranasal oxytocin improved social behaviour shortly after administration C_LIO_LIEffects of oxytocin on non-social behaviours were limited C_LIO_LIChronic oxytocin maintained beneficial effects in mutant mice but was deleterious in wild-type mice C_LIO_LIOxytocin triggers greater and longer-lasting improvements in social behaviour when administered within a social context C_LIO_LIChronic oxytocin suppresses the expression of its own receptor gene and key striatal neuron gene markers C_LI

animal behavior and cognition↗

bioRxiv2023

Biased activation of the vasopressin V2 receptor probed by NMR, paramagnetic ligands, and molecular dynamics simulations

G protein-coupled receptors (GPCRs) control critical intercellular communications by responding to extracellular stimuli and undertaking conformational changes to convey signals to intracellular effectors. We combined NMR, molecular pharmacology, and molecular dynamics (MD) simulations to study the conformational diversity of the vasopressin V2 GPCR subtype (V2R) bound to different types of ligands: the antagonist tolvaptan, the endogenous unbiased agonist arginine-vasopressin, and MCF14, a Gs-protein biased agonist. We developed a double-labeling NMR scheme to study the conformational dynamics: V2R was subjected to lysine 13CH3 methylation, whereas the agonists were tagged with a paramagnetic probe. Paramagnetic relaxation enhancements were used to validate the ligand binding poses in the MD simulations. We found that the bias for the Gs protein over the {beta}-arrestin pathway involves interactions between the conserved NPxxY motif in the transmembrane helix (TM) 7 and a central hydrophobic patch in TM3, which constrains TM7 and likely inhibits {beta}-arrestin signaling. A similar mechanism was observed for the pathogenic mutation, I1303.43N, which constitutively activates the Gs protein without concomitant {beta}-arrestin recruitment. This mechanism resembles to opioid receptors findings indicating common patterns in class A GPCRs.

biophysics↗

bioRxiv2023

Torsion of the heart tube by shortage of progenitor cells : identification of Greb1l as a genetic determinant of criss-cross heart in mice

Despite their burden and impact, most congenital defects remain poorly understood by lack of knowledge of the embryological mechanisms. Here, we identify Greb1l mutants as the first mouse model of criss-cross heart. Based on 3D quantifications of shape changes, we demonstrate that torsion of the atrioventricular canal occurs together with supero-inferior ventricles at E10.5, after heart looping. Mutants phenocopy specific features of partial deficiency in retinoic acid signalling, suggesting that GREB1L is a novel modulator of this signalling. Spatio-temporal gene mapping and cross-correlated transcriptomic analyses further reveal the role of Greb1l in maintaining a pool of precursor cells during heart tube elongation, by controlling ribosome biogenesis and cell differentiation. Growth arrest and malposition of the outflow tract are predictive of abnormal tube remodelling in mutants. Our work on a rare cardiac malformation opens novel perspectives on the origin of a broader spectrum of congenital defects associated with GREB1L in humans. HighlightsO_LIGreb1l inactivation is the first model of criss-cross heart C_LIO_LIGrowth arrest of the outflow tract and reduced pole distance are predictive of the torsion of the atrioventricular canal, and also account for associated defects of supero-inferior ventricles and malposition of the great vessels C_LIO_LIVentricle position needs to be maintained after heart looping C_LIO_LIGREB1L, which is associated in humans with a spectrum of congenital defects, is required to maintain precursor cells, by promoting ribosome biogenesis and restricting cell differentiation. C_LIO_LIGREB1L is a novel factor involved in retinoic acid signalling. C_LI In BriefGREB1L is associated with a spectrum of congenital defects in humans. Bernheim et al now uncover its function in maintaining a reservoir of precursor cells. Inactivation of Greb1l in the mouse impairs the elongation of the heart tube leading to criss-cross heart with supero-inferior ventricles.

developmental biology↗

bioRxiv2021

A monocyte/dendritic cell molecular signature of SARS-CoV2-related multisystem inflammatory syndrome in children (MIS-C) with severe myocarditis

SARS-CoV-2 infection in children is generally milder than in adults, yet a proportion of cases result in hyperinflammatory conditions often including myocarditis. To better understand these cases, we applied a multi-parametric approach to the study of blood cells of 56 children hospitalized with suspicion of SARS-CoV-2 infection. The most severe forms of MIS-C (multisystem inflammatory syndrome in children related to SARS-CoV-2), that resulted in myocarditis, were characterized by elevated levels of pro-angiogenesis cytokines and several chemokines. Single-cell transcriptomic analyses identified a unique monocyte/dendritic cell gene signature that correlated with the occurrence of severe myocarditis, characterized by sustained NF-{kappa}B activity, TNF- signaling, associated with decreased gene expression of NF-{kappa}B inhibitors. We also found a weak response to type-I and type-II interferons, hyperinflammation and response to oxidative stress related to increased HIF-1 and VEGF signaling. These results provide potential for a better understanding of disease pathophysiology.

immunology↗

bioRxiv2021

Neural stem cells alter nucleocytoplasmic partitioning and accumulate nuclear polyadenylated transcripts during quiescence

Quiescence is a cellular state characterised by reversible cell-cycle arrest and diminished biosynthetic activity that protects against environmental insults, replicative exhaustion and proliferation-induced mutations1. Entry into and exit from this state controls development, maintenance and repair of tissues plus, in the adult central nervous system, generation of new neurons and thus cognition and mood2-4. Cancer stem cells too can undergo quiescence, which confers them resistance to current therapies5, 6. Despite clinical relevance, quiescence is poorly understood and is defined functionally given lack of molecular markers. Decrease of the most resource-intensive cellular process of protein synthesis is a feature of quiescence, controlled across species and cell types by inhibition of the Target of Rapamycin (TOR) pathway1, 7. Here, we combine Drosophila genetics and a mammalian model to show that altered nucleocytoplasmic partitioning and nuclear accumulation of polyadenylated RNAs are novel evolutionarily conserved hallmarks of quiescence regulation. Furthermore, nuclear accumulation of messenger RNA (mRNA) in quiescent NSCs (qNSCs) largely predicts protein downregulation, accounting for uncoupling between transcriptome and proteome in quiescence. These mechanisms provide a previously unappreciated regulatory layer to reducing protein synthesis in quiescent cells, whilst priming them for reactivation in response to appropriate cues.

developmental biology↗

bioRxiv2020

CKS1-dependent proteostatic regulation has dual roles combating acute myeloid leukemia whilst protecting normal hematopoiesis

Acute myeloid leukemia (AML) is an aggressive hematological disorder comprising a hierarchy of quiescent leukemic stem cells (LSCs) and proliferating blasts with limited self-renewal ability. AML has a dismal prognosis, with extremely low two-year survival rates in the poorest cytogenetic risk patients, primarily due to the failure of intensive chemotherapy protocols unable to deplete LSCs, which reconstitute the disease in vivo, and the significant toxicity towards healthy hematopoietic cells. Whilst much work has been done to identify genetic and epigenetic vulnerabilities in AML LSCs, little is known about protein dynamics and the role of protein degradation in drug resistance and relapse. Here, using a highly specific inhibitor of the SCFSKP2-CKS1 complex, we report a dual role for CKS1-dependent protein degradation in reducing AML blasts in vivo, and importantly depleting LSCs. Whilst many AML LSC targeted therapies show significant toxicity to healthy hematopoiesis, inhibition of CKS1-dependent protein degradation has the opposite effect, protecting normal hematopoietic cells from chemotherapeutic toxicity. Together these findings demonstrate CKS1-dependent proteostasis is key for normal and malignant hematopoiesis. SignificanceCKS1-dependent protein degradation is a specific vulnerability in AML LSCs. Specific inhibition of SCFSKP2-CKS1 is lethal to CKS1Bhigh AML blasts and all AML LSCs. Normal hematopoiesis is protected from chemotherapeutic toxicity by inhibition of CKS1-dependent protein degradation, substantiating a dual role for CKS1-dependent protein degradation in clinical treatment of AML.

cancer biology↗

bioRxiv2020

Dynamic regulation of hierarchical heterogeneity in Acute Myeloid Leukemia serves as a tumor immunoevasion mechanism

Acute Myeloid Leukemia, a hematological malignancy with poor clinical outcome, is composed of hierarchically heterogeneous cells. We examine the contribution of this heterogeneity to disease progression in the context of anti-tumor immune responses and investigate whether these responses regulate the balance between stemness and differentiation in AML. Combining phenotypic analysis with proliferation dynamics and fate-mapping of AML cells in a murine AML model, we demonstrate the presence of a terminally differentiated, chemoresistant population expressing high levels of PDL1. We show that PDL1 upregulation in AML cells, following exposure to IFN{gamma} from activated T cells, is coupled with AML differentiation and the dynamic balance between proliferation, versus differentiation and immunosuppression, facilitates disease progression in the presence of immune responses. This microenvironment-responsive hierarchical heterogeneity in AML may be key in facilitating disease growth at the population level at multiple stages of disease, including following bone marrow transplantation and immunotherapy.

cancer biology↗