Proliferation Of Progenitors Research Articles

Histone-binding protein RBBP4 is necessary to promote neurogenesis in the developing mouse neocortical progenitors.

Chromatin regulation plays a crucial role in neocortical neurogenesis, and mutations in chromatin modifiers are linked to neurodevelopmental disorders. RBBP4 is a core subunit of several chromatin-modifying complexes; however, its functional role and genome-wide occupancy profile in the neocortical primordium are unknown. To address this, we performed RBBP4 knockdown using CRISPR/Cas9 on neocortical progenitors derived from mice of both sexes at embryonic age 12.5 during deep-layer neurogenesis. Our study demonstrates that downregulation of RBBP4 in the E12.5 neocortical progenitors reduced neuronal output, specifically affecting CTIP2-expressing neurons. We demonstrate that RBBP4 plays an essential role in regulating neocortical progenitor proliferation. However, overexpression of RBBP4 alone was not sufficient to regulate neuronal fate.Genome-wide occupancy analysis revealed that RBBP4 primarily binds to distal regulatory elements, and neuron differentiation is a significant GO biological pathway of RBBP4-bound genes. Interestingly, we found that RBBP4 binds to Cdon, a receptor protein in the Shh signaling pathway, and knockdown of Cdon phenocopies RBBP4 knockdown resulting in a significant reduction in neurogenesis, particularly CTIP2-expressing neurons. CDON overexpression could rescue the phenotype caused upon loss of RBBP4 in the neocortex, thereby suggesting the functional link between RBBP4 and its target gene CDON. Our results shed light on the cellular role of RBBP4 and identify CDON as a novel regulator of deep-layer neurogenesis in the neocortical progenitors. Our findings are significant in the context of understanding how dysregulated chromatin regulation impacts cellular mechanisms in neurodevelopmental disorders.Significance Statement Chromatin modifier RBBP4 regulates chromatin structure and, thereby, gene expression. It is expressed in the dorsal telencephalon progenitors during deep-layer neurogenesis. In this study, we unveil a novel role for RBBP4 in regulating deep-layer neurogenesis in the neocortical progenitors. Our research underscores RBBP4's critical role in governing progenitor proliferation and neuronal subtype specification in the neocortex while identifying its genome-wide binding occupancy profile. Moreover, we identify Cdon as a novel binding target of RBBP4, also involved in regulating deep-layer neurogenesis. These findings illuminate the mechanisms by which chromatin modifiers influence neocortical development, offering insights into how mutations in chromatin modifiers could impact cortical development and contribute to neurodevelopmental disorders.

The Jag2/Notch1 signaling axis promotes sebaceous gland differentiation and controls progenitor proliferation.

The sebaceous gland (SG) is a vital appendage of the epidermis, and its normal homeostasis and function is crucial for effective maintenance of the skin barrier. Notch signaling is a well-known regulator of epidermal differentiation, and has also been shown to be involved in postnatal maintenance of SGs. However, the precise role of Notch signaling in regulating SG differentiation in the adult homeostatic skin remains unclear. While there is evidence to suggest that Notch1 is the primary Notch receptor involved in regulating the differentiation process, the ligand remains unknown. Using monoclonal therapeutic antibodies designed to specifically inhibit of each of the Notch ligands or receptors, we have identified the Jag2/Notch1 signaling axis as the primary regulator of sebocyte differentiation in mouse homeostatic skin. Mature sebocytes are lost upon specific inhibition of the Jag2 ligand or Notch1 receptor, resulting in the accumulation of proliferative stem/progenitor cells in the SG. Strikingly, this phenotype is reversible, as these stem/progenitor cells re-enter differentiation when the inhibition of Notch activity is lifted. Thus, Notch activity promotes correct sebocyte differentiation, and is required to restrict progenitor proliferation.

The Jag2/Notch1 signaling axis promotes sebaceous gland differentiation and controls progenitor proliferation

The sebaceous gland (SG) is a vital appendage of the epidermis, and its normal homeostasis and function is crucial for effective maintenance of the skin barrier. Notch signaling is a well-known regulator of epidermal differentiation, and has also been shown to be involved in postnatal maintenance of SGs. However, the precise role of Notch signaling in regulating SG differentiation in the adult homeostatic skin remains unclear. While there is evidence to suggest that Notch1 is the primary Notch receptor involved in regulating the differentiation process, the ligand remains unknown. Using monoclonal therapeutic antibodies designed to specifically inhibit of each of the Notch ligands or receptors, we have identified the Jag2/Notch1 signaling axis as the primary regulator of sebocyte differentiation in mouse homeostatic skin. Mature sebocytes are lost upon specific inhibition of the Jag2 ligand or Notch1 receptor, resulting in the accumulation of proliferative stem/progenitor cells in the SG. Strikingly, this phenotype is reversible, as these stem/progenitor cells re-enter differentiation when the inhibition of Notch activity is lifted. Thus, Notch activity promotes correct sebocyte differentiation, and is required to restrict progenitor proliferation.

High CD44 expression and enhanced E-selectin binding identified as biomarkers of chemoresistant leukemic cells in human T-ALL.

T-cell acute lymphoblastic leukemia (T-ALL) is a hematopoietic malignancy characterized by increased proliferation and incomplete maturation of T-cell progenitors, for which relapse is often of poor prognosis. To improve patient outcomes, it is critical to understand the chemoresistance mechanisms arising from cell plasticity induced by the bone marrow (BM) microenvironment. Single-cell RNA sequencing of human T-ALL cells from adipocyte-rich and adipocyte-poor BM revealed a distinct leukemic cell population defined by quiescence and high CD44 expression (Ki67neg/lowCD44high). During in vivo treatment, these cells evaded chemotherapy, and were further called Chemotherapy-resistant Leukemic Cells (CLCs). Patient sample analysis revealed Ki67neg/lowCD44high CLCs at diagnosis and during relapse, with each displaying a specific transcriptomic signature. Interestingly, CD44high expression in T-ALL Ki67neg/low CLCs was associated with E-selectin binding. Analysis of 39 human T-ALL samples revealed significantly enhanced E-selectin binding activity in relapse/refractory samples compared with drug-sensitive samples. These characteristics of chemoresistant T-ALL CLCs provide key insights for prognostic stratification and novel therapeutic options.

Evolving cell states and oncogenic drivers during the progression of IDH-mutant gliomas.

Isocitrate dehydrogenase (IDH) mutants define a class of gliomas that are initially slow-growing but inevitably progress to fatal disease. To characterize their malignant cell hierarchy, we profiled chromatin accessibility and gene expression across single cells from low-grade and high-grade IDH-mutant gliomas and ascertained their developmental states through a comparison to normal brain cells. We provide evidence that these tumors are initially fueled by slow-cycling oligodendrocyte progenitor cell-like cells. During progression, a more proliferative neural progenitor cell-like population expands, potentially through partial reprogramming of 'permissive' chromatin in progenitors. This transition is accompanied by a switch from methylation-based drivers to genetic ones. In low-grade IDH-mutant tumors or organoids, DNA hypermethylation appears to suppress interferon (IFN) signaling, which is induced by IDH or DNA methyltransferase 1 inhibitors. High-grade tumors frequently lose this hypermethylation and instead acquire genetic alterations that disrupt IFN and other tumor-suppressive programs. Our findings explain how these slow-growing tumors may progress to lethal malignancies and have implications for therapies that target their epigenetic underpinnings.

KAT6B is required for histone 3 lysine 9 acetylation and SOX gene expression in the developing brain.

Heterozygous mutations in the histone lysine acetyltransferase gene KAT6B (MYST4/MORF/QKF) underlie neurodevelopmental disorders, but the mechanistic roles of KAT6B remain poorly understood. Here, we show that loss of KAT6B in embryonic neural stem and progenitor cells (NSPCs) impaired cell proliferation, neuronal differentiation, and neurite outgrowth. Mechanistically, loss of KAT6B resulted in reduced acetylation at histone H3 lysine 9 and reduced expression of key nervous system development genes in NSPCs and the developing cortex, including the SOX gene family, in particular Sox2, which is a key driver of neural progenitor proliferation, multipotency and brain development. In the fetal cortex, KAT6B occupied the Sox2 locus. Loss of KAT6B caused a reduction in Sox2 promoter activity in NSPCs. Sox2 overexpression partially rescued the proliferative defect of Kat6b -/- NSPCs. Collectively, these results elucidate molecular requirements for KAT6B in brain development and identify key KAT6B targets in neural precursor cells and the developing brain.

TMOD-17. FOXR2-ACTIVATED CNS NEUROBLASTOMA ORIGINATES FROM THE MEDIAL GANGLIONIC EMINENCE LINEAGE

Abstract FOXR2-activated CNS neuroblastoma is a rare subtype of pediatric brain tumor characterized by the elevated expression of the transcription factor FOXR2 due to genomic rearrangement. However, the precise pathogenic mechanisms, including the cell type of origin and the function of FOXR2, are not fully understood. An expression profile analysis of patient tumors showed increased expression of the marker genes associated with the Medial Ganglionic Eminence (MGE) in the ventral forebrain such as NKX2.1 and SOX6. Based on this result, we hypothesized that FOXR2-activated CNS neuroblastoma originates from the MGE lineage. To test this hypothesis, we overexpressed FOXR2 in human embryonic stem cell-derived MGE progenitors and, as a control cell type, cortical progenitors. Our data showed that FOXR2 overexpression resulted in a significantly increased proliferation in the MGE progenitors but not in the cortical progenitors, indicating a cell-type-specific effect of FOXR2. When injected into the brain of immunodeficient mice, FOXR2-expressing MGE progenitors formed tumors that molecularly resemble patient tumors, while FOXR2-expressing cortical progenitors did not. To further dissect the underlying molecular mechanisms, we conducted an RNA-seq experiment and found that FOXR2 overexpression activated the MEK/ERK signaling pathway through a suppression of the endogenous RAS inhibitor DIRAS3. Moreover, the FDA-approved MEK inhibitor trametinib significantly suppressed the proliferation and induced cell death in FOXR2-expressing MGE progenitors in vitro with the IC50 value of 30 nM. Collectively, our study has revealed the cell type of origin of FOXR2-activated CNS neuroblastoma and identified the MEK/ERK pathway as a critical downstream of FOXR2 that can be targeted therapeutically.

Enhancing Ex Vivo Limbal Epithelial Cell Expansion on Amniotic Membrane: A Comparative Study of Monolayer (2D) Versus Sandwich (3D) Culture Configurations.

This study compared two-dimensional (monolayer) and three-dimensional (sandwich) systems for expanding ex vivo limbal epithelial cells on amniotic membrane and evaluated the outcomes after transplantation into rabbits with experimentally induced limbal stem cell deficiency. Evaluations included markers for progenitor cells, proliferation, apoptosis, and clinical monitoring for up to 63 days. In the monolayer culture, epithelial cells derived from limbal explants were expanded on amniotic membrane as the substrate. In the sandwich culture, the cells were cultured between 2 layers of amniotic membrane. Evaluations included markers for progenitor cells, proliferation, and apoptosis, along with clinical monitoring for up to 63 days. Sandwich cultures demonstrated increased cellular proliferation and fewer progenitor cells compared with monolayer cultures. In treating limbal stem cell deficiency, the group receiving transplantation from sandwich cultures exhibited reduced neovascularization and decreased corneal ulceration compared with those treated with monolayer cultures, with similar clinical outcomes in corneal opacity. The configuration of the culture system did not affect the presence of apoptotic cells. Corneas treated with sandwich cultures showed a higher presence of progenitor cells compared with the monolayer group, suggesting a potential long-term viability advantage for these transplants. In conclusion, although the sandwich culture system enhanced cellular proliferation, it also resulted in a decrease in progenitor cells within the cultures. Nevertheless, both systems demonstrated comparable therapeutic efficacy in treating limbal stem cell deficiency, with the sandwich approach potentially offering long-term benefits because of the increased presence of progenitor cells in the transplanted cornea.

TLX3 regulates CGN progenitor proliferation during cerebellum development and its dysfunction can lead to autism

TLX3 regulates CGN progenitor proliferation during cerebellum development and its dysfunction can lead to autism

Wnt2bb signaling promotes pharyngeal chondrogenic precursor proliferation and chondrocyte maturation by activating Yap expression in zebrafish

Pharyngeal cartilage morphogenesis is crucial for the formation of craniofacial structures. Cranial neural crest cells are specified at the neural plate border, migrate to pharyngeal arches, and differentiate into pharyngeal chondrocytes, which subsequently flatten, elongate, and stack like coins during maturation. Although the developmental processes prior to chondrocyte maturation have been extensively studied, their subsequent changes in morphology and organization remain largely elusive. Here, we show that wnt2bb is expressed in the pharyngeal ectoderm adjacent to the chondrogenic precursor cells in zebrafish. Inactivation of Wnt2bb leads to a reduction in nuclear β-catenin, which impairs chondrogenic precursor proliferation and disrupts chondrocyte morphogenesis and organization, eventually causing a severe shrinkage of pharyngeal cartilages. Moreover, the decrease of β-catenin in wnt2bb−/− mutants is accompanied by the reduction of Yap expression. Reactivation of Yap can restore the proliferation of chondrocyte progenitors as well as the proper size, shape, and stacking of pharyngeal chondrocytes. Our findings suggest that Wnt/β-catenin signaling promotes Yap expression to regulate pharyngeal cartilage formation in zebrafish.

Regulating the formation of Müller glia-derived progenitor cells in the retina.

We summarize recent findings in different animal models regarding the different cell-signaling pathways and gene networks that influence the reprogramming of Müller glia into proliferating, neurogenic progenitor cells in the retina. Not surprisingly, most of the cell-signaling pathways that guide the proliferation and differentiation of embryonic retinal progenitors also influence the ability of Müller glia to become proliferating Müller glia-derived progenitor cells (MGPCs). Further, the neuronal differentiation of MGPC progeny is potently inhibited by networks of neurogenesis-suppressing genes in chick and mouse models but occurs freely in zebrafish. There are important differences between the model systems, particularly pro-inflammatory signals that are active in mature Müller glia in damaged rodent and chick retinas, but less so in fish retinas. These pro-inflammatory signals are required to initiate the process of reprogramming, but if sustained suppress the potential of Müller glia to become neurogenic MGPCs. Further, there are important differences in how activated Müller glia up- or downregulate pro-glial transcription factors in the different model systems. We review recent findings regarding regulatory cell signaling and gene networks that influence the activation of Müller glia and the transition of these glia into proliferating progenitor cells with neurogenic potential in fish, chick, and mouse model systems.

Stem and progenitor cell proliferation are independently regulated by cell type-specific cyclinD genes.

Regeneration and homeostatic turnover of solid tissues depend on the proliferation of symmetrically dividing adult stem cells, which either remain stem cells or differentiate based on their niche position. Here we demonstrate that in zebrafish lateral line sensory organs, stem and progenitor cell proliferation are independently regulated by two cyclinD genes. Loss of ccnd2a impairs stem cell proliferation during development, while loss of ccndx disrupts hair cell progenitor proliferation but allows normal differentiation. Notably, ccnd2a can functionally replace ccndx, indicating that the respective effects of these Cyclins on proliferation are due to cell type-specific expression. However, even though hair cell progenitors differentiate normally in ccndx mutants, they are mispolarized due to hes2 and Emx2 downregulation. Thus, regulated proliferation ensures that equal numbers of hair cells are polarized in opposite directions. Our study reveals cell type-specific roles for cyclinD genes in regulating the different populations of symmetrically dividing cells governing organ development and regeneration, with implications for regenerative medicine and disease.

Microglia regulate GABAergic neurogenesis in prenatal human brain through IGF1.

GABAergic neurons are an essential cellular component of neural circuits. Their abundance and diversity have enlarged significantly in the human brain, contributing to the expanded cognitive capacity of humans. However, the developmental mechanism of the extended production of GABAergic neurons in the human brain remains elusive. Here, we use single-cell transcriptomics, bioinformatics, and histological analyses to uncover microglial regulation of the sustained proliferation of GABAergic progenitors and neuroblasts in the human medial ganglionic eminence (hMGE). We show that insulin-like growth factor 1 (IGF1) and its receptor IGR1R as the top ligand-receptor pair underlying microglia-progenitor communication in the prenatal human brain. Using our newly developed neuroimmune hMGE organoids, which mimics hMGE cytoarchitecture and developmental trajectory, we demonstrate that microglia-derived IGF1 promotes progenitor proliferation and the production of GABAergic neurons. Conversely, IGF1-neutralizing antibodies and IGF1 knockout human embryonic stem cells (hESC)-induced microglia (iMG) completely abolished iMG-mediated progenitor proliferation. Together, these findings reveal a previously unappreciated role of microglia-derived IGF1 in promoting proliferation of neural progenitors and the development of GABAergic neurons.

Arginine methylation of the p30 C/EBPα oncoprotein regulates progenitor proliferation and myeloid differentiation

Arginine methylation of the p30 C/EBPα oncoprotein regulates progenitor proliferation and myeloid differentiation

Negative cell cycle regulation by calcineurin is necessary for proper beta cell regeneration in zebrafish.

Stimulation of pancreatic beta cell regeneration could be a therapeutic lead to treat diabetes. Unlike humans, the zebrafish can efficiently regenerate beta cells, notably from ductal pancreatic progenitors. To gain insight into the molecular pathways involved in this process, we established the transcriptomic profile of the ductal cells after beta cell ablation in the adult zebrafish. These data highlighted the protein phosphatase calcineurin (CaN) as a new potential modulator of beta cell regeneration. We showed that CaN overexpression abolished the regenerative response, leading to glycemia dysregulation. On the opposite, CaN inhibition increased ductal cell proliferation and subsequent beta cell regeneration. Interestingly, the enhanced proliferation of the progenitors was paradoxically coupled with their exhaustion. This suggests that the proliferating progenitors are next entering in differentiation. CaN appears as a guardian which prevents an excessive progenitor proliferation to preserve the pool of progenitors. Altogether, our findings reveal CaN as a key player in the balance between proliferation and differentiation to enable a proper beta cell regeneration.

The transcription elongation factors Spt4 and Spt5 control neural progenitor proliferation and are implicated in neuronal remodeling during Drosophila mushroom body development.

Spt4 and Spt5 form the DRB sensitivity inducing factor (DSIF) complex that regulates transcription elongation at multiple steps including promotor-proximal pausing, processivity and termination. Although this implicated a general role in transcription, several studies pointed to smaller sets of target genes and indicated a more specific requirement in certain cellular contexts. To unravel common or distinct functions of Spt4 and Spt5 in vivo, we generated knock-out alleles for both genes in Drosophila melanogaster. Using the development of the mushroom bodies as a model, we provided evidence for two common functions of Spt4 and Spt5 during mushroom body development, namely control of cell proliferation of neural progenitor cells and remodeling of axonal projections of certain mushroom body neurons. This latter function is not due to a general requirement of Spt4 and Spt5 for axon pathfinding of mushroom body neurons, but due to distinct effects on the expression of genes controlling remodeling.

Negative cell cycle regulation by calcineurin is necessary for proper beta cell regeneration in zebrafish

Stimulation of pancreatic beta cell regeneration could be a therapeutic lead to treat diabetes. Unlike humans, the zebrafish can efficiently regenerate beta cells, notably from ductal pancreatic progenitors. To gain insight into the molecular pathways involved in this process, we established the transcriptomic profile of the ductal cells after beta cell ablation in the adult zebrafish. These data highlighted the protein phosphatase calcineurin (CaN) as a new potential modulator of beta cell regeneration. We showed that CaN overexpression abolished the regenerative response, leading to glycemia dysregulation. On the opposite, CaN inhibition increased ductal cell proliferation and subsequent beta cell regeneration. Interestingly, the enhanced proliferation of the progenitors was paradoxically coupled with their exhaustion. This suggests that the proliferating progenitors are next entering in differentiation. CaN appears as a guardian which prevents an excessive progenitor proliferation to preserve the pool of progenitors. Altogether, our findings reveal CaN as a key player in the balance between proliferation and differentiation to enable a proper beta cell regeneration.

Type V collagen exhibits distinct regulatory activities in TMJ articular disc versus condylar cartilage during postnatal growth and remodeling

Understanding matrix molecular activities that regulate the postnatal growth and remodeling of the temporomandibular joint (TMJ) articular disc and condylar cartilage will enable the development of effective regenerative strategies targeting TMJ disorders. This study elucidated the distinct roles of type V collagen (collagen V) in regulating these two units. Studying the TMJ of young adult Col5a1+/− mice, we found that loss of collagen V resulted in substantial changes in the proliferation, clustering and density of progenitors in condylar cartilage, but did not have a major impact on disc cells that are more fibroblast-like. Although loss of collagen V led to thickened collagen fibrils with increased heterogeneity in the disc, there were no significant changes in local micromodulus, except for a reduction at the posterior end of the inferior side. Following the induction of aberrant occlusal loading by the unilateral anterior crossbite (UAC) procedure, both wild-type (WT) and Col5a1+/− condylar cartilage exhibited salient remodeling, and Col5a1+/− condyle developed more pronounced degeneration and tissue hypertrophy at the posterior end than the WT. In contrast, neither UAC nor collagen V deficiency induced marked changes in the morphology or biomechanical properties of the disc. Together, our findings highlight the distinct roles of collagen V in regulating these two units during postnatal growth and remodeling, emphasizing its more crucial role in condylar cartilage due to its impact on the highly mechanosensitive progenitors. These results provide the foundation for using collagen V to improve the regeneration of TMJ and the care of patients with TMJ disorders. Statement of significanceSuccessful regeneration of the temporomandibular joint (TMJ) articular disc and condylar cartilage remains a significant challenge due to the limited understanding of matrix molecular activities that regulate the formation and remodeling of these tissues. This study demonstrates that collagen V plays distinct and critical roles in these processes. In condylar cartilage, collagen V is essential for regulating progenitor cell fate and maintaining matrix integrity. In the disc, collagen V also regulates fibril structure and local micromechanics, but has a limited impact on cell phenotype or its remodeling response. Our findings establish collagen V as a key component in maintaining the integrity of these two units, with a more crucial role in condylar cartilage due to its impact on progenitor cell activities.

Characterization of Age-Dependent Changes in Skeletal Muscle Repair and Regeneration Using a Mouse Model of Acute Muscle Injury.

Acute skeletal muscle injury initiates a process of necrosis, debris clearance, and ultimately tissue regeneration via myogenesis. While skeletal muscle stem cells (MuSCs) are responsible for populating the proliferative myogenic progenitor pool to fuel muscle repair, recruited and resident immune cells have a central role in the regulation of muscle regeneration via the execution of phagocytosis and release of soluble factors that act directly on MuSCs to regulate myogenic differentiation. Therefore, the timing of MuSC proliferation and differentiation is closely linked to the populations and behaviors of immune cells present within skeletal muscle. This has important implications for aging andmuscle repair, as systemic changes in immune system function contribute to a decline in muscle regenerative capacity. Here, we present adapted protocols for the isolation of mononuclear cells from skeletal muscles for the quantification of immune cell populations using flow cytometry. We also describe a cardiotoxin skeletal muscle injury protocol and detail the expected outcomes including immune cell infiltration to the injured sites and formation of new myocytes. As immune cell function is substantially influenced by aging, we extend these approaches and outcomes to aged mice.

Comparative single-cell multiome identifies evolutionary changes in neural progenitor cells during primate brain development.

Understanding the cellular and genetic mechanisms driving human-specific features of cortical development remains a challenge. We generated a cell-type resolved atlas of transcriptome and chromatin accessibility in the developing macaque and mouse prefrontal cortex (PFC). Comparing with published human data, our findings demonstrate that although the cortex cellular composition is overall conserved across species, progenitor cells show significant evolutionary divergence in cellular properties. Specifically, human neural progenitors exhibit extensive transcriptional rewiring in growth factor and extracellular matrix (ECM) pathways. Expression of the human-specific progenitor marker ITGA2 in the fetal mouse cortex increases the progenitor proliferation and the proportion of upper-layer neurons. These transcriptional divergences are primarily driven by altered activity in the distal regulatory elements. The chromatin regions with human-gained accessibility are enriched with human-specific sequence changes and polymorphisms linked to intelligence and neuropsychiatric disorders. Our results identify evolutionary changes in neural progenitors and putative gene regulatory mechanisms shaping primate brain evolution.