Differentiation Of Progenitor Cells Research Articles

An Optimized Protocol for the Generation of Alveolospheres from Wild-Type Mice

Organoid models have become an integral part of the research methodology in the lung field. These systems allow for the study of progenitor and stem cell self-renewal, self-organization, and differentiation. Distinct models of lung organoids mimicking various anatomical regions of mature lungs have emerged in parallel to the increased gain of knowledge regarding epithelial stem and progenitor cell populations and the corresponding mesenchymal cells that populate the in vivo niche. In the distal lung, type 2 alveolar epithelial cells (AEC2s) represent a stem cell population that is engaged in regenerative mechanisms in response to various insults. These cells self-renew and give rise to AEC1s that carry out gas exchange. Multiple experimental protocols allowing the generation of alveolar organoids, or alveolospheres, from murine lungs have been described. Among the drawbacks have been the requirement of transgenic mice allowing the isolation of AEC2s with high viability and purity, and the occasional emergence of bronchiolar and bronchioalveolar organoids. Here, we provide a refined gating strategy and an optimized protocol for the generation of alveolospheres from wild-type mice. Our approach not only overcomes the need for transgenic mice to generate such organoids, but also yields a pure culture of alveolospheres that is devoid of bronchiolar and bronchioalveolar organoids. Our protocol contributes to the standardization of this important research tool.

Jag1/2 maintain esophageal homeostasis and suppress foregut tumorigenesis by restricting the basal progenitor cell pool

Basal progenitor cells are crucial for maintaining foregut (the esophagus and forestomach) homeostasis. When their function is dysregulated, it can promote inflammation and tumorigenesis. However, the mechanisms underlying these processes remain largely unclear. Here, we employ genetic mouse models to reveal that Jag1/2 regulate esophageal homeostasis and foregut tumorigenesis by modulating the function of basal progenitor cells. Deletion of Jag1/2 in mice disrupts esophageal and forestomach epithelial homeostasis. Mechanistically, Jag1/2 deficiency impairs activation of Notch signaling, leading to reduced squamous epithelial differentiation and expansion of basal progenitor cells. Moreover, Jag1/2 deficiency exacerbates the deoxycholic acid (DCA)-induced squamous epithelial injury and accelerates the initiation of squamous cell carcinoma (SCC) in the forestomach. Importantly, expression levels of JAG1/2 are lower in the early stages of human esophageal squamous cell carcinoma (ESCC) carcinogenesis. Collectively, our study demonstrates that Jag1/2 are important for maintaining esophageal and forestomach homeostasis and the onset of foregut SCC.

Dysregulated cholinergic signaling inhibits oligodendrocyte maturation following demyelination.

Dysregulation of oligodendrocyte progenitor cell (OPC) recruitment and oligodendrocyte differentiation contribute to failure of remyelination in human demyelinating diseases such as multiple sclerosis (MS). Deletion of muscarinic receptor enhances OPC differentiation and remyelination. However, the role of ligand-dependent signaling versus constitutive receptor activation is unknown. We hypothesized that dysregulated acetylcholine (ACh) release upon demyelination contributes to ligand mediated activation hindering myelin repair. Following chronic cuprizone induced demyelination (male and female mice), we observed a 2.5-fold increase in ACh concentration. This increase in ACh concentration could be attributed to increased ACh synthesis or decreased acetylcholinesterase (AChE) / butyrylcholinesterase (BChE) mediated degradation. Using ChAT reporter mice, we identified increased ChAT-GFP expression following both lysolecithin and cuprizone demyelination. ChAT-GFP expression was upregulated in a subset of injured and uninjured axons following intraspinal lysolecithin induced demyelination. In cuprizone demyelinated corpus callosum, ChAT-GFP was observed in Gfap+ astrocytes and axons indicating the potential for neuronal and astrocytic ACh release. BChE expression was significantly decreased in the corpus callosum following cuprizone demyelination. This decrease was due to the loss of myelinating oligodendrocytes which were the primary source of BChE. To determine the role of ligand mediated muscarinic signaling following lysolecithin injection, we administered neostigmine, a cholinesterase inhibitor, to artificially raise ACh. We identified a dose-dependent decrease in mature oligodendrocyte density with no effect on OPC recruitment. Together, these results support a functional role of ligand mediated activation of muscarinic receptors following demyelination and suggest that dysregulation of ACh homeostasis directly contributes to failure of remyelination in MS.Significance Statement Demyelinating diseases like Multiple Sclerosis are characterized by failure of remyelination. Oligodendrocyte progenitor cell (OPC) recruitment and differentiation are crucial aspects for remyelination to occur. Here we show that increased acetylcholine (ACh) contributes to activation of muscarinic receptors that inhibit OPC differentiation. Increased choline acetyltransferase synthesis following demyelination was observed in axons and astrocytes suggestive of a potential for acetylcholine synthesis and release. The increase in ACh levels following demyelination was largely due to reduction of oligodendrocyte derived butyrylcholinesterase that modulates ACh concentration. Development of cell specific esterase stimulator to restore ACh levels may serve as an approach towards inhibiting ongoing demyelination and neurodegeneration.

Influence of High-Intensity Terahertz Radiation on the Differentiation of Human Neural Progenitor Cells

Influence of High-Intensity Terahertz Radiation on the Differentiation of Human Neural Progenitor Cells

Association of dietary and nutritional factors with cognitive decline, dementia, and depressive symptomatology in older individuals according to a neurogenesis-centred biological susceptibility to brain ageing.

Hippocampal neurogenesis (HN) occurs throughout the life course and is important for memory and mood. Declining with age, HN plays a pivotal role in cognitive decline (CD), dementia, and late-life depression, such that altered HN could represent a neurobiological susceptibility to these conditions. Pertinently, dietary patterns (e.g., Mediterranean diet) and/or individual nutrients (e.g., vitamin D, omega 3) can modify HN, but also modify risk for CD, dementia, and depression. Therefore, the interaction between diet/nutrition and HN may alter risk trajectories for these ageing-related brain conditions. Using a subsample (n= 371) of the Three-City cohort-where older adults provided information on diet and blood biobanking at baseline and were assessed for CD, dementia, and depressive symptomatology across 12years-we tested for interactions between food consumption, nutrient intake, and nutritional biomarker concentrations and neurogenesis-centred susceptibility status (defined by baseline readouts of hippocampal progenitor cell integrity, cell death, and differentiation) on CD, Alzheimer's disease (AD), vascular and other dementias (VoD), and depressive symptomatology, using multivariable-adjusted logistic regression models. Increased plasma lycopene concentrations (OR [95% CI] = 1.07 [1.01, 1.14]), higher red meat (OR [95% CI] = 1.10 [1.03, 1.19]), and lower poultry consumption (OR [95% CI] = 0.93 [0.87, 0.99]) were associated with an increased risk for AD in individuals with a neurogenesis-centred susceptibility. Increased vitamin D consumption (OR [95% CI] = 1.05 [1.01, 1.11]) and plasma γ-tocopherol concentrations (OR [95% CI] = 1.08 [1.01, 1.18]) were associated with increased risk for VoD and depressive symptomatology, respectively, but only in susceptible individuals. This research highlights an important role for diet/nutrition in modifying dementia and depression risk in individuals with a neurogenesis-centred susceptibility.

Features of Intracellular Signal Transduction in Neural Stem Cells under the Influence of Alkaloid Songorine, an Agonist of Fibroblast Growth Factor Receptors.

We performed a comparative in vitro study of the involvement of NF-κB, PI3K, cAMP, ERK1/2, p38, JAKs, STAT3, JNK, and p53-dependent intracellular signaling in the functioning of neural stem cells (NSC) under the influence of basic fibroblast growth factor (FGF) and FGF receptor agonist, diterpene alkaloid songorine. The significant differences in FGFR-mediated intracellular signaling in NSC were revealed for these ligands. In both cases, stimulation of progenitor cell proliferation occurs with the participation of NF-κB, PI3K, ERK1/2, JAKs, and STAT3, while JNK and p53, on the contrary, inhibit cell cycle progression. However, under the influence of songorin, cAMP- and p38-mediated cascades are additionally involved in the transmission of the NSC division-activating signal. In addition, unlike FGF, the alkaloid stimulates progenitor cell differentiation by activating ERK1/2, p38, JNK, p53, and STAT3.

The miR-144/Hmgn2 regulatory axis orchestrates chromatin organization during erythropoiesis

Differentiation of stem and progenitor cells is a highly regulated process that involves the coordinated action of multiple layers of regulation. Here we show how the post-transcriptional regulatory layer instructs the level of chromatin regulation via miR-144 and its targets to orchestrate chromatin condensation during erythropoiesis. The loss of miR-144 leads to impaired chromatin condensation during erythrocyte maturation. Among the several targets of miR-144 that influence chromatin organization, the miR-144-dependent regulation of Hmgn2 is conserved from fish to humans. Our genetic probing of the miR-144/Hmgn2 regulatory axis establish that intact miR-144 target sites in the Hmgn2 3’UTR are necessary for the proper maturation of erythrocytes in both zebrafish and human iPSC-derived erythroid cells while loss of Hmgn2 rescues in part the miR-144 null phenotype. Altogether, our results uncover miR-144 and its target Hmgn2 as the backbone of the genetic regulatory circuit that controls the terminal differentiation of erythrocytes in vertebrates.

MiRNA-7145-cuedc2 axis controls hematopoiesis through JAK1/STAT3 signaling pathway

Hematopoiesis ensures tissue oxygenation, and remodeling as well as immune protection in vertebrates. During embryogenesis, hemangioblasts are the source of all blood cells. Gata1a and pu.1 are co-expressed in hemangioblasts before hemangioblasts are differentiated into blood cells. However, the genes that determine the differentiation of hemangioblasts into myeloid or erythroid cell lineages have not been fully uncovered. Here we showed that miRNA-7145, a miRNA with previously unknown function, was enriched in erythrocytes at the definitive wave, but not expressed in myeloid cells. Overexpression and loss-of-function analysis of miRNA-7145 revealed that miRNA-7145 functions as a strong inhibitor for myeloid progenitor cell differentiation while driving erythropoiesis during the primitive wave. Furthermore, we confirmed that cuedc2 is one of miRNA-7145 targeted-genes. Overexpression or knock-down of cuedc2 partially rescues the phenotype caused by miRNA-7145 overexpression or loss-of-function. As well, overexpression and loss-of-function analysis of cuedc2 showed that cuedc2 is required for myelopoiesis at the expense of erythropoiesis. Finally, we found that overexpression of zebrafish cuedc2 in 293 T cell inhibits the JAK1/STAT3 signaling pathway. Collectively, our results uncover a previously unknown miRNA-7145-cuedc2 axis, which regulate hematopoiesis through inhibiting the JAK1/STAT3 signaling pathway.

The combined administration of LNC-encapsulated retinoic acid and calcitriol stimulates oligodendrocyte progenitor cell differentiation in vitro and in vivo after intranasal administration

Intranasal administration is an efficient strategy for bypassing the BBB, favoring drug accumulation in the brain, and improving its efficiency. Lipid nanocapsules (LNC) are suitable nanocarriers for the delivery of lipophilic drugs via this route and can be used to encapsulate lipophilic molecules such as retinoic acid (RA) and calcitriol (Cal). As the hallmarks of multiple sclerosis (MS) are neuroinflammation and oligodendrocyte loss, our hypothesis was that by combining two molecules known for their pro-differentiating properties, encapsulated in LNC, and delivered by intranasal administration, we would stimulate oligodendrocyte progenitor cells (OPC) differentiation into oligodendrocytes and provide a new pro-remyelinating therapy. LNC loaded with RA (LNC-RA) and Cal (LNC-Cal) were stable for at least 8 weeks. The combination of RA and Cal was more efficient than the molecules alone, encapsulated or not, on OPC differentiation in vitro and decreased microglia cell activation in a dose-dependent manner. After the combined intranasal administration of LNC-RA and LNC-Cal in a mouse cuprizone model of demyelination, increased MBP staining was observed in the corpus callosum. In conclusion, intranasal delivery of lipophilic drugs encapsulated in LNC is a promising strategy for myelinating therapies.

The Influence of Electrical Stimulation on the Proliferation and Differentiation of Retinal Progenitor Cells

The Influence of Electrical Stimulation on the Proliferation and Differentiation of Retinal Progenitor Cells

Pharmacological modulation of inflammatory oligodendrocyte progenitor cells using three multiple sclerosis disease modifying therapies in vitro

Preclinical studies of pro-remyelinating therapies for multiple sclerosis tend to neglect the effect of the disease-relevant inflammatory milieu. Interferon-gamma (IFN-γ) is known to suppress oligodendrocyte progenitor cell (OPC) differentiation and induce a recently described immune OPC (iOPC) phenotype characterized by expression of major histocompatibility complex (MHC) molecules. We tested the effects of cladribine (CDB), dimethylfumarate (DMF), and interferon-beta (IFN-β), existing anti-inflammatory therapies for MS, on the IFN-γ-induced iOPC formation and OPC differentiation block. In line with previous reports, we demonstrate that IFN-β and DMF inhibit OPC proliferation, while CDB had no effect. None of the drugs exhibited cytotoxic effects at the physiological concentrations tested in vitro. In a differentiation assay, none of the drugs were able to promote differentiation, under inflammatory or basal conditions. To study drug effects on iOPCs, we monitored MHC expression in vitro with live cell imaging using cells isolated from MHC reporter mice. IFN-β suppressed induction of MHC class II, and DMF led to suppression of both class I and II. CDB had no effect on MHC induction. We conclude that promoting proliferation and differentiation and suppressing iOPC induction under inflammatory conditions may require separate therapeutic strategies and must be balanced for maximal repair. Our in vitro MHC screening assay can be leveraged across cell types to test the effects of drug candidates and disease-related stimuli.

Genetic Tools for Cell Lineage Tracing and Profiling Developmental Trajectories in the Skin

The epidermis is the body's first line of protection against dehydration and pathogens, continually regenerating the outermost protective skin layers throughout life. During both embryonic development and wound healing, epidermal stem and progenitor cells must respond to external stimuli and insults to build, maintain, and repair the cutaneous barrier. Recent advances in CRISPR-based methods for cell lineage tracing have remarkably expanded the potential for experiments that track stem and progenitor cell proliferation and differentiation over the course of tissue and even organismal development. Additional tools for DNA-based recording of cellular signaling cues promise to deepen our understanding of the mechanisms driving normal skin morphogenesis and response to stressors as well as the dysregulation of cell proliferation and differentiation in skin diseases and cancer. In this review, we highlight cutting-edge methods for cell lineage tracing, including in organoids and model organisms, and explore how cutaneous biology researchers might leverage these techniques to elucidate the developmental programs that support the regenerative capacity and plasticity of the skin.

Heavy Metal-Laden Particulate Dust Alters Transcriptional Regulation in Human Colonoids

Chronic heavy metals exposure in the southwestern United States co-occurs with greater incidences of metabolic and digestive diseases, particularly in the Indigenous and Hispanic populations. There is a need to understand how particulate heavy metals drive this dysbiotic state and propose a molecular explanation as a result of these exposures. Previously, our group demonstrated human colonoids to be a relevant model for investigating the physiological impact of environmental toxicants on the intestinal tract. The objective of this study is to elucidate the mechanism behind the transcriptional and cellular changes present in the intestinal epithelia upon acute exposure to non fissile uranium particulate dust. We hypothesize that non-fissile uranium exposure biases the differentiation of progenitor cells in the crypt to secretory roles through miRNA regulation. Colonoids were exposed (18 hours) to non-fissile uranium dust obtained from the Jackpile uranium mine, one of the largest open pit uranium mines, located on the Laguna Pueblo in New Mexico. Control and dust-exposed colonoids (n=3 unique donors) were digested into single cells and processed for droplet-based single cell sequencing (scRNAseq). Upon exposure, enteroendocrine cells (EEC) expanded >4 fold, along with increases in serotonin and PYY, as well as goblet cells (>3 fold) which indicates changes to the secretory lineage. Downstream pathway analysis found microRNAs miR-625-3p, miR-320a, miR-29b-3p to be differentially expressed. These results suggest that acute uranium dust exposure induces changes in differentiation pathways which are modulated by regulatory miRNAs. We would like to expand these findings by performing miRNA sequencing and comparing miRNA expression profiles between UBD exposed and control samples. Our results also provide an improved understanding of uranium dust’s impact on the gastrointestinal system, which may inform other physiological studies on the health of individuals exposed to heavy metals. This study was funded by K01DK106323, R56ES034400, P42ES025589, P20GM121176, P20GM130422, and AGA SURF. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Size-Optimized Layered Double Hydroxide Nanoparticles Promote Neural Progenitor Cells Differentiation of Embryonic Stem Cells Through the Regulation of M6A Methylation.

The committed differentiation fate regulation has been a difficult problem in the fields of stem cell research, evidence showed that nanomaterials could promote the differentiation of stem cells into specific cell types. Layered double hydroxide (LDH) nanoparticles possess the regulation function of stem cell fate, while the underlying mechanism needs to be investigated. In this study, the process of embryonic stem cells (ESCs) differentiate to neural progenitor cells (NPCs) by magnesium aluminum LDH (MgAl-LDH) was investigated. MgAl-LDH with diameters of 30, 50, and 100 nm were synthesized and characterized, and their effects on the cytotoxicity and differentiation of NPCs were detected in vitro. Dot blot and MeRIP-qPCR were performed to detect the level of m6A RNA methylation in nanoparticles-treated cells. Our work displayed that LDH nanoparticles of three different sizes were biocompatible with NPCs, and the addition of MgAl-LDH could significantly promote the process of ESCs differentiate to NPCs. 100 nm LDH has a stronger effect on promoting NPCs differentiation compared to 30 nm and 50 nm LDH. In addition, dot blot results indicated that the enhanced NPCs differentiation by MgAl-LDH was closely related to m6A RNA methylation process, and the major modification enzyme in LDH controlled NPCs differentiation may be the m6A RNA methyltransferase METTL3. The upregulated METTL3 by LDH increased the m6A level of Sox1 mRNA, enhancing its stability. This work reveals that MgAl-LDH nanoparticles can regulate the differentiation of ESCs into NPCs by increasing m6A RNA methylation modification of Sox1.

Triptolide promotes differentiation of human monocytes into immunosuppressive MDSCs

BackgroundMyeloid-derived suppressor cells (MDSCs) negatively modulate immune activity. Prior investigations have shown much promise in using MDSCs-assisted immunotherapy for organ transplantation patients. Additionally, owing to its immunosuppressive activity, MDSCs can also be used to manage immune-associated disorders. MethodsGranulocyte-macrophage colony-stimulating factor (GM-CSF) was employed to stimulate myeloid progenitor cell differentiation. Triptolide (PG490) was introduced toward the later phases of in vitro MDSCs induction. Lastly, real-time PCR (RT-PCR) and flow cytometry were used to assess transcript expression and cell phenotype, and a mouse skin transplantation model was established to evaluate the MDSCs-mediated immune suppression in vivo. ResultsCo-stimulation with PG490 and GM-CSF potently induced myeloid-derived monocytes to form MDSCs, with remarkable immune-suppressive activity. The underlying mechanism involved downregulation of T cell proliferation, activation, enhancement of inflammatory cytokine release, as well as T cell conversion to Treg cells. PG490 strongly enhanced iNOS expression in MDSCs, and iNOS inhibition successfully reversed the immune-suppression. The PG490- and GM-CSF-induced MDSCs substantially extended survival duration of murine skin grafts, thereby validating their strong immune-suppressive activity in vivo. ConclusionsHerein, we presented a new approach involving MDSCs-based immunosuppression in vitro. PG490 and GM-CSF co-treatment strongly induced immuno-suppressive activity in MDSCs both in vitro and in vivo. Our findings highlight the promise of applying MDSCs-based therapy in clinical organ transplantation treatment.

Linking Altered Nephrogenesis with Hypertension using Single Nuclei RNA Sequencing of Fetal Kidney at E15.5, E16.5 and E17.5 in a Novel Nephron Deficient Rat Model

This study sought to elucidate molecular mechanisms linking altered nephrogenesis in the HSRA rat, a novel model of nephron deficiency, to the development of hypertension and chronic kidney disease (CKD) later in life. The HSRA model gives rise to offspring born with a single kidney (-S) 50-75% of the time. The single kidneys from HSRA-S animals exhibit reduced nephron endowment compared to either kidney from two-kidney offspring (-C). We hypothesize that early molecular factors during development contribute to the observed reduction in nephron number in -S animals. Timed breeding and embryonic kidney isolation from -S and -C animals were performed at gestational days (GD) 15.5, 16.5, or 17.5. Single nuclei RNA sequencing (snRNA-seq) was conducted using n=3 -S kidneys and n=3 -C kidneys for each timepoint. The library was prepared using a 10x Chromium Single Cell 3’ kit, followed by sequencing on an Illumina NextSeq2000. The average number of cells captured per sample was 16,997, yielding an average of 25,335 reads/cell, with a median of 1,512 genes/cell identified. Bioinformatic analysis using Cell Ranger and Seurat (v4) revealed various renal progenitor cell types including metanephric mesenchymal (MM) and ureteric bud cells, alongside tubular, stromal, and glomerular endothelial progenitors. Podocytes emerged by GD17.5, originating from the MM cell population, as evidenced by their extension from this cluster. Global differential expression analysis revealed minimal transcriptional differences between -S and -C fetal kidneys at GD 16.5 and 17.5. However, a down-regulation of ribosomal genes was observed on GD15.5 in the -S group, suggesting premature cellular differentiation contributing to the diminished nephron formation. Markedly, cell type-specific gene expression differences were observed at GD 16.5 and 17.5, highlighting the superior resolution that snRNAseq provides compared to bulk RNA sequencing methods. We are currently performing trajectory analysis using Monocle3 to investigate cellular fate pathways and uncover genes involved during nephron progenitor cell differentiation. These preliminary findings shed light on the cellular and molecular landscape during critical developmental stages in the kidney of the HSRA rat that could link developmental factors affecting nephron number to hypertension and CKD pathologies later in life. This research was generously supported by the Graduate School of Health Sciences at UMMC. MRG is supported by NIH R01HL137673. The UMMC Molecular and Genomics Facility is supported, in part, by funds from the NIGMS, including Molecular Center of Health and Disease-COBRE (P20GM144041-Garrett), Mississippi INBRE (P20GM103476-Flynt), and Cardiorenal and Metabolic Diseases-COBRE (P20GM104357-Hall). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Distinct Treatment Outcomes between MVID Patient-Modeled Mice Suggest Epithelial Cell Maturation Is Influenced by MYO5B Functional State

Background and aims: Microvillus inclusion disease (MVID) is a devastating congenital diarrhea and malnutrition disorder caused by different point mutations in Myosin Vb (MYO5B). Specific genotype-dependent epithelial phenotypes or treatment options have not been fully investigated. Using tamoxifen-inducible epithelial-specific MYO5B-deficient (Myo5bΔIEC) mice, we have reported that MYO5B loss disrupts progenitor cell differentiation and that Lyso-Phosphatidic Acid (LPA) signaling and Notch inhibition can ameliorate the differentiation defects. We recently generated another MVID patient-modeled mouse strain, Vil1-CreERT2;Myo5bflox/G519R (Myo5b(G519R)). This study compared the epithelial pathophysiology and the effects of drug treatment between Myo5b(G519R) and Myo5bΔIEC mice. Methods: The MVID model mice and littermate controls received a single dose of tamoxifen, and their intestinal tissues were harvested on day 4 post-tamoxifen. For treatment: the small chemical LPAR5 agonist, Compound-1, or vehicle were given intraperitoneally once a day. Another set of mice received the Notch2-targeted inhibitor, Nirogacestat, on days 1 and 3. Digital image analysis scripts were developed for segmentation of immunostained tissues. Staining intensities and localization (membrane vs. cytoplasm) of markers were assessed in >300,000 epithelial cells per section. Results: Epithelial transcriptional signatures were analyzed by RNA-sequencing of enteroids generated from jejunal crypts of Myo5b(G519R) and Myo5bΔIEC mice. Enteroids from both mice showed significant upregulation of Notch signaling mediators and the downregulation of tuft cell markers and mitochondrial metabolic genes compared to control enteroids. Immunostaining of tissue sections consistently demonstrated that functional Myo5b loss decreases tuft cell numbers and mitochondrial densities in crypt and villus epithelial cells. In the vehicle treated MVID mouse tissues, sodium transporters of enterocytes, such as Na+/H+ exchanger (NHE)3 and Na+-dependent glucose transporter (SGLT)1, were mislocalized from the apical membrane and colocalized with lysosomal markers similar to what is observed in MVID patient tissues. Compound-1 treatment increased the apical localization of NHE3 and SGLT1 both in Myo5b(G519R) and Myo5bΔIEC mouse small intestines, whereas Nirogacestat improved localizations only in Myo5bΔIEC but not Myo5b(G519R) mouse tissues. Conclusions: Functional MYO5B loss, caused by the point mutation at G519R or total knockout directly disrupts Notch signaling and energy metabolic pathways in the epithelial cells, resulting in differentiation defects and malnutrition. The distinct treatment effcacies between the two MVID mouse models suggest that the total loss and impaired function of MYO5B differentially affect epithelial cell functions. NIH R01DK128190, RC2DK118640. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Single-cell analyses reveal transient retinal progenitor cells in the ciliary margin of developing human retina

The emergence of retinal progenitor cells and differentiation to various retinal cell types represent fundamental processes during retinal development. Herein, we provide a comprehensive single cell characterisation of transcriptional and chromatin accessibility changes that underline retinal progenitor cell specification and differentiation over the course of human retinal development up to midgestation. Our lineage trajectory data demonstrate the presence of early retinal progenitors, which transit to late, and further to transient neurogenic progenitors, that give rise to all the retinal neurons. Combining single cell RNA-Seq with spatial transcriptomics of early eye samples, we demonstrate the transient presence of early retinal progenitors in the ciliary margin zone with decreasing occurrence from 8 post-conception week of human development. In retinal progenitor cells, we identified a significant enrichment for transcriptional enhanced associate domain transcription factor binding motifs, which when inhibited led to loss of cycling progenitors and retinal identity in pluripotent stem cell derived organoids.

YAP and TAZ differentially regulate postnatal cortical progenitor proliferation and astrocyte differentiation.

WW domain-containing transcription regulator 1 (TAZ) and Yes-associated protein (YAP) are transcriptional co-activators traditionally studied together as a part of the Hippo pathway and best known for their roles in stem cell proliferation and differentiation. Despite their similarities, TAZ and YAP can exert divergent cellular effects by differentially interacting with other signaling pathways that regulate stem cell maintenance or differentiation. In this study, we show that TAZ regulates astrocytic differentiation and maturation of postnatal neural stem and progenitor cells (NPCs), and that TAZ mediates some but not all of the effects of bone morphogenetic protein (BMP) signaling on astrocytic development. By contrast, both TAZ and YAP mediate effects on NPC fate of β1-integrin and integrin-linked kinase (ILK) signaling, and these effects are dependent on extracellular matrix (ECM) cues. These findings demonstrate that TAZ and YAP perform divergent functions in the regulation of astrocyte differentiation, where YAP regulates cell cycle states of astrocytic progenitors and TAZ regulates differentiation and maturation from astrocytic progenitors into astrocytes.

Temporal regulation of the Mediator complex during muscle proliferation, differentiation, regeneration, aging, and disease.

Genesis of skeletal muscle relies on the differentiation and fusion of mono-nucleated muscle progenitor cells into the multi-nucleated muscle fiber syncytium. The temporally-controlled cellular and morphogenetic changes underlying this process are initiated by a series of highly coordinated transcription programs. At the core, the myogenic differentiation cascade is driven by muscle-specific transcription factors, i.e., the Myogenic Regulatory Factors (MRFs). Despite extensive knowledge on the function of individual MRFs, very little is known about how they are coordinated. Ultimately, highly specific coordination of these transcription programs is critical for their masterfully timed transitions, which in turn facilitates the intricate generation of skeletal muscle fibers from a naïve pool of progenitor cells. The Mediator complex links basal transcriptional machinery and transcription factors to regulate transcription and could be the integral component that coordinates transcription factor function during muscle differentiation, growth, and maturation. In this study, we systematically deciphered the changes in Mediator complex subunit expression in skeletal muscle development, regeneration, aging, and disease. We incorporated our in vitro and in vivo experimental results with analysis of publicly available RNA-seq and single nuclei RNA-seq datasets and uncovered the regulation of Mediator subunits in different physiological and temporal contexts. Our experimental results revealed that Mediator subunit expression during myogenesis is highly dynamic. We also discovered unique temporal patterns of Mediator expression in muscle stem cells after injury and during the early regeneration period, suggesting that Mediator subunits may have unique contributions to directing muscle stem cell fate. Although we observed few changes in Mediator subunit expression in aging muscles compared to younger muscles, we uncovered extensive heterogeneity of Mediator subunit expression in dystrophic muscle nuclei, characteristic of chronic muscle degeneration and regeneration cycles. Taken together, our study provides a glimpse of the complex regulation of Mediator subunit expression in the skeletal muscle cell lineage and serves as a springboard for mechanistic studies into the function of individual Mediator subunits in skeletal muscle.