Stem Cell Fate Determination Research Articles

Chromatin remodeling in tissue stem cell fate determination.

Tissue stem cells (TSCs), which reside in specialized tissues, constitute the major cell sources for tissue homeostasis and regeneration, and the contribution of transcriptional or epigenetic regulation of distinct biological processes in TSCs has been discussed in the past few decades. Meanwhile, ATP-dependent chromatin remodelers use the energy from ATP hydrolysis to remodel nucleosomes, thereby affecting chromatin dynamics and the regulation of gene expression programs in each cell type. However, the role of chromatin remodelers in tissue stem cell fate determination is less well understood. In this review, we systematically discuss recent advances in epigenetic control by chromatin remodelers of hematopoietic stem cells, intestinal epithelial stem cells, neural stem cells, and skin stem cells in their fate determination and highlight the importance of their essential role in tissue homeostasis, development, and regeneration. Moreover, the exploration of the molecular and cellular mechanisms of TSCs is crucial for advancing our understanding of tissue maintenance and for the discovery of novel therapeutic targets.

Abstract B008: Twist1 drives pancreatic ductal adenocarcinoma-mediated muscle cachexia

Abstract Cancer cachexia is a debilitating syndrome that affects the survival and quality of life of cancer patients. Up to 95% of pancreatic ductal adenocarcinoma (PDAC) patients experience muscle cachexia, yet the underlying mechanisms remain poorly understood. The transcription factor Twist1 plays essential roles in mesenchymal stem cell fate determination, and its deregulation contributes to cancer progression and metastasis. In this study, we report on the serendipitous finding that Twist1 functions in PDAC to drive muscle cachexia. We found that conditional overexpression of Twist1 in the pancreatic epithelium (pTwist1) during early development resulted in severe growth retardation, which was mainly characterized by muscle hypotrophy. This intriguing observation prompted us to inquire whether pTwist1 could function in the pancreatic epithelium to regulate muscle mass, and if so, whether this has any implication as it relates to PDAC-mediated muscle cachexia. We found that enforced expression of pTwist1 in healthy adult animals was sufficient to elicit the cardinal features of muscle cachexia, including progressive decline in body weight, physical activity, muscle force, and muscle mass. In the context of PDAC, enforced expression of pTwist1 in a KrasG12D background led to a dramatic acceleration of tumor progression, and this was associated with severe muscle cachexia. To the best of our knowledge, these findings provide the first evidence that the pro-malignant transcription factor Twist1 functions in PDAC to drive muscle cachexia, thus revealing new insights into the underlying mechanisms of muscle that could be harnessed to curb this debilitating syndrome. Citation Format: Parash Parajuli, Azeddine Atfi. Twist1 drives pancreatic ductal adenocarcinoma-mediated muscle cachexia [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B008.

Methionine Synthase 2 Represses Stem Cell Maintenance of Arabidopsis thaliana in Response to Salt Stress.

Salt stress represses the growth and development of plants that mainly depend on the continual propagation and differentiation of stem cells. WUSCHEL (WUS)/WUSCHEL-RELATED HOMEOBOX (WOX) family proteins determine stem cell fate in plants under ever-changing environments. It is not yet known how plant stem cell homeostasis is regulated under salt stress. Methionine synthase catalyzes the formation of methionine by methylating homocysteine in the one-carbon metabolism pathway. In this work, we investigated the role of Arabidopsis METHIONINE SYNTHASE 2 (AtMS2) in stem cell homeostasis under salt stress. The results showed that AtMS2 represses the stem cell maintenance of Arabidopsis in response to salt stress. Under normal growth conditions, AtMS2 is mainly localized in the cytoplasm. However, under salt stress, it exhibits significant accumulation in the nucleus. AtMS2 interacts with the WUS/WOX protein, and, together, they repress WUS/WOX expression by binding to its promoter. The mutation in AtMS2 resulted in enhanced salt tolerance. Therefore, AtMS2 might act as a key negative regulator to repress the stem cell maintenance and growth of Arabidopsis under salt stress.

Endogenous hydrogen sulfide accelerated trauma-induced heterotopic ossification through the Ca2+/ERK pathway-enhanced aberrant osteogenic activity

Unveiling of the mechanism involved in the occurrence and development of trauma-induced heterotopic ossification (tHO) is highly demanding due to current ineffective clinical treatment for it. Previous studies proposed that hydrogen sulfide (H2S) was vital for fate determination of stem cells, suggesting a potential role in the regulation of tHO development. In the current study, We found that expression of metabolic enzyme within sulfur conversion pathway was enhanced after tendon injury, leading to H2S accumulation within the tHO region. Increased production of endogenous H2S was shown to promote aberrant osteogenic activity of tendon-derived stem cells (TDSCs), which accelerated tHO formation. The inhibition of metabolic enzyme of H2S production or directly absorption of H2S could abolished osteogenic induction of TDSCs and the formation of tHO. Mechanistically, through RNA sequencing combined with rescue experiments, we demonstrated that activation of Ca2+/ERK pathway was the downstream molecular event of H2S-induced osteogenic commitment of TDSCs and tHO. For treatment strategy exploration, zine oxide nanoparticles (ZnO) as an effective H2S elimination material was validated to ideally halt the tHO formation in this study. Furthermore, in terms of chirality of nanoparticles, D-ZnO or L-ZnO nanoparticles showed superiority over R–ZnO nanoparticles in both clearing of H2S and inhibition of tHO. Our study not only revealed the mechanism of tHO through the endogenous gas signaling event from a new perspective, but also presented a applicable platform for elimination of the inordinate gas production, thus aiding the development of clinical treatment for tHO.

The Role of Lactylation in Regulating Microglial Inflammation in PTSD.

Lactylation plays an important regulatory role in neural development, neural stem cell fate determination, and the physiological function of the nervous system. Abnormal lactylation is closely related to the occurrence and development of hippocampal microglial inflammation in post-traumatic stress disorder (PTSD), so lactylation may serve as a potential therapeutic target for PTSD. This article reviewed the latest research progress on the involvement of lactylation in hippocampal microglial inflammation and its molecular mechanisms in PTSD.

Gangliosides in neural stem cell fate determination and nerve cell specification--preparation and administration.

Gangliosides are sialylated glycosphingolipids with essential but enigmatic functions in healthy and disease brains. GD3 is the predominant species in neural stem cells (NSCs) and GD3-synthase (sialyltransferase II; St8Sia1) knockout (GD3S-KO) revealed reduction of postnatal NSC pools with severe behavioral deficits including cognitive impairment, depression-like phenotypes, and olfactory dysfunction. Exogenous administration of GD3 significantly restored the NSC pools and enhanced the stemness of NSCs with multipotency and self-renewal, followed by restored neuronal functions. Our group discovered that GD3 is involved in the maintenance of NSC fate determination by interacting with epidermal growth factor receptors (EGFRs), by modulating expression of cyclin-dependent kinase (CDK) inhibitors p27 and p21, and by regulating mitochondrial dynamics via associating a mitochondrial fission protein, the dynamin-related protein-1 (Drp1). Furthermore, we discovered that nuclear GM1 promotes neuronal differentiation by an epigenetic regulatory mechanism. GM1 binds with acetylated histones on the promoter of N-acetylgalactosaminyltransferase (GalNAcT; GM2 synthase (GM2S); B4galnt1) as well as on the NeuroD1 in differentiated neurons. In addition, epigenetic activation of the GM2S gene was detected as accompanied by an apparent induction of neuronal differentiation in NSCs responding to an exogenous supplement of GM1. Interestingly, GM1 induced epigenetic activation of the tyrosine hydroxylase (TH) gene, with recruitment of Nurr1 and PITX3, dopaminergic neuron-associated transcription factors, to the TH promoter region. In this way, GM1 epigenetically regulates dopaminergic neuron specific gene expression, and it would modify Parkinson's disease. Multifunctional gangliosides significantly modulate lipid microdomains to regulate functions of important molecules on multiple sites: the plasma membrane, mitochondrial membrane, and nuclear membrane. Versatile gangliosides regulate functional neurons as well as sustain NSC functions via modulating protein and gene activities on ganglioside microdomains. Maintaining proper ganglioside microdomains benefits healthy neuronal development and millions of senior citizens with neurodegenerative diseases. Here, we introduce how to isolate GD3 and GM1 and how to administer them into the mouse brain to investigate their functions on NSC fate determination and nerve cell specification.

ARID1B maintains mesenchymal stem cell quiescence via inhibition of BCL11B-mediated non-canonical Activin signaling

ARID1B haploinsufficiency in humans causes Coffin-Siris syndrome, associated with developmental delay, facial dysmorphism, and intellectual disability. The role of ARID1B has been widely studied in neuronal development, but whether it also regulates stem cells remains unknown. Here, we employ scRNA-seq and scATAC-seq to dissect the regulatory functions and mechanisms of ARID1B within mesenchymal stem cells (MSCs) using the mouse incisor model. We reveal that loss of Arid1b in the GLI1+ MSC lineage disturbs MSCs’ quiescence and leads to their proliferation due to the ectopic activation of non-canonical Activin signaling via p-ERK. Furthermore, loss of Arid1b upregulates Bcl11b, which encodes a BAF complex subunit that modulates non-canonical Activin signaling by directly regulating the expression of activin A subunit, Inhba. Reduction of Bcl11b or non-canonical Activin signaling restores the MSC population in Arid1b mutant mice. Notably, we have identified that ARID1B suppresses Bcl11b expression via specific binding to its third intron, unveiling the direct inter-regulatory interactions among BAF subunits in MSCs. Our results demonstrate the vital role of ARID1B as an epigenetic modifier in maintaining MSC homeostasis and reveal its intricate mechanistic regulatory network in vivo, providing novel insights into the linkage between chromatin remodeling and stem cell fate determination.

METTL3 Modulates Ctsk+ Lineage Supporting Cranial Osteogenesis via Hedgehog

N6-methyladenosine (m6A) modification, a eukaryotic messenger RNA modification catalyzed by methyltransferase-like 3 (METTL3), plays a pivotal role in stem cell fate determination. Calvarial bone development and maintenance are orchestrated by the cranial sutures. Cathepsin K (CTSK)–positive calvarial stem cells (CSCs) contribute to mice calvarial ossification. However, the role of m6A modification in regulating Ctsk+ lineage cells during calvarial development remains elusive. Here, we showed that METTL3 was colocalized with cranial nonosteoclastic Ctsk+ lineage cells, which were also associated with GLI1 expression. During neonatal development, depletion of Mettl3 in the Ctsk+ lineage cells delayed suture formation and decreased mineralization. During adulthood maintenance, loss of Mettl3 in the Ctsk+ lineage cells impaired calvarial bone formation, which was featured by the increased bone porosity, enhanced bone marrow cavity, and decreased number of osteocytes with the less-developed cellular outline. The analysis of methylated RNA immunoprecipitation sequencing and RNA sequencing data indicated that loss of METTL3 reduced Hedgehog (Hh) signaling pathway. Restoration of Hh signaling pathway by crossing Sufufl/+ alleles or by local administration of SAG21 partially rescued the abnormity. Our data indicate that METTL3 modulates Ctsk+ lineage cells supporting calvarial bone formation by regulating the Hh signaling pathway, providing new insights for clinical treatment of skull vault osseous diseases.

Threonine dehydrogenase regulates neutrophil homeostasis but not H3K4me3 levels in zebrafish.

l-threonine dehydrogenase (Tdh) is an enzyme that links threonine metabolism to epigenetic modifications and mitochondria biogenesis. In vitro studies show that it is critical for the regulation of trimethylation of histone H3 lysine 4 (H3K4me3) levels and cell fate determination of mouse embryonic stem cells (mESCs). However, whether Tdh regulates a developmental process in vivo and, if it does, whether it also primarily regulates H3K4me3 levels in this process as it does in mESCs, remains elusive. Here, we revealed that, in zebrafish hematopoiesis, tdh is preferentially expressed in neutrophils. Knockout of tdh causes a decrease in neutrophil number and slightly suppresses their acute injury-induced migration, but, unlike the mESCs, the level of H3K4me3 is not evidently reduced in neutrophils sorted from the kidney marrow of adult tdh-null zebrafish. These phenotypes are dependent on the enzymatic activity of Tdh. Importantly, a soluble supplement of nutrients that are able to fuel the acetyl-CoA pool, such as pyruvate, glucose and branched-chain amino acids, is sufficient to rescue the reduction in neutrophils caused by tdh deletion. In summary, our study presents evidence for the functional requirement of Tdh-mediated threonine metabolism in a developmental process in vivo. It also provides an animal model for investigating the nutritional regulation of myelopoiesis and immune response, as well as a useful tool for high-throughput drug/nutrition screening.

DRP1 inhibition-mediated mitochondrial elongation abolishes cancer stemness, enhances glutaminolysis, and drives ferroptosis in oral squamous cell carcinoma.

Mitochondrial dynamics play a fundamental role in determining stem cell fate. However, the underlying mechanisms of mitochondrial dynamics in the stemness acquisition of cancer cells are incompletely understood. Metabolomic profiling of cells were analyzed by MS/MS. The genomic distribution of H3K27me3 was measured by CUT&Tag. Oral squamous cell carcinoma (OSCC) cells depended on glucose or glutamine fueling TCA cycle were monitored by 13C-isotope tracing. Organoids and tumors from patients and mice were treated with DRP1 inhibitors mdivi-1, ferroptosis inducer erastin, or combination with mdivi-1 and erastin to evaluate treatment effects. Mitochondria of OSCC stem cells own fragment mitochondrial network and DRP1 is required for maintenance of their globular morphology. Imbalanced mitochondrial dynamics induced by DRP1 knockdown suppressed stemness of OSCC cells. Elongated mitochondria increased α-ketoglutarate levels and enhanced glutaminolysis to fuel the TCA cycle by increasing glutamine transporter ASCT2 expression. α-KG promoted the demethylation of histone H3K27me3, resulting in downregulation of SNAI2 associated with stemness and EMT. Significantly, suppressing DRP1 enhanced the anticancer effects of ferroptosis. Our study reveals a novel mechanism underlying mitochondrial dynamics mediated cancer stemness acquisition and highlights the therapeutic potential of mitochondria elongation to increase the susceptibility of cancer cells to ferroptosis.

A BMP-controlled metabolic/epigenetic signaling cascade directs midfacial morphogenesis.

Craniofacial anomalies, especially midline facial defects, are among the most common birth defects in patients and are associated with increased mortality or require lifelong treatment. During mammalian embryogenesis, specific instructions arising at genetic, signaling, and metabolic levels are important for stem cell behaviors and fate determination, but how these functionally relevant mechanisms are coordinated to regulate craniofacial morphogenesis remain unknown. Here, we report that bone morphogenetic protein (BMP) signaling in cranial neural crest cells (CNCCs) is critical for glycolytic lactate production and subsequent epigenetic histone lactylation, thereby dictating craniofacial morphogenesis. Elevated BMP signaling in CNCCs through constitutively activated ACVR1 (ca-ACVR1) suppressed glycolytic activity and blocked lactate production via a p53-dependent process that resulted in severe midline facial defects. By modulating epigenetic remodeling, BMP signaling-dependent lactate generation drove histone lactylation levels to alter essential genes of Pdgfra, thus regulating CNCC behavior in vitro as well as in vivo. These findings define an axis wherein BMP signaling controls a metabolic/epigenetic cascade to direct craniofacial morphogenesis, thus providing a conceptual framework for understanding the interaction between genetic and metabolic cues operative during embryonic development. These findings indicate potential preventive strategies of congenital craniofacial birth defects via modulating metabolic-driven histone lactylation.

Stem Cells and Differentiation in Vascular Tissues.

Plant vascular tissues are crucial for the long-distance transport of water, nutrients, and a multitude of signal molecules throughout the plant body and, therefore, central to plant growth and development. The intricate development of vascular tissues is orchestrated by unique populations of dedicated stem cells integrating endogenous as well as environmental cues. This review summarizes our current understanding of vascular-related stem cell biology and of vascular tissue differentiation. We present an overview of the molecular and cellular mechanisms governing the maintenance and fate determination of vascular stem cells and highlight the interplay between intrinsic and external cues. In this context, we emphasize the role of transcription factors, hormonal signaling, and epigenetic modifications. We also discuss emerging technologies and the large repertoire of cell types associated with vascular tissues, which have the potential to provide unprecedented insights into cellular specialization and anatomical adaptations to distinct ecological niches.

The Role of Zinc Finger Proteins in the Fate Determination of Mesenchymal Stem Cells during Osteogenic and Adipogenic Differentiation

Zinc finger proteins (ZFPs) constitute a crucial group of transcription factors widely present in various organisms. They act as transcription factors, nucleases, and RNA-binding proteins, playing significant roles in cell differentiation, growth, and development. With extensive research on ZFPs, their roles in the determination of mesenchymal stem cells (MSCs) fate during osteogenic and adipogenic differentiation processes have become increasingly clear. ZFP521, for instance, is identified as an inhibitor of the Wnt signaling pathway and RUNX2's transcriptional activity, effectively suppressing osteogenic differentiation. Moreover, ZFP217 contributes to the inhibition of adipogenic differentiation by reducing the M6A level of the cell cycle regulator cyclin D1 (CCND1). In addition, other ZFPs can also influence the fate of mesenchymal stem cells (MSCs) during osteogenic and adipogenic differentiation through various signaling pathways, transcription factors, and epigenetic controls, participating in the subsequent differentiation and maturation of precursor cells. Given the prevalent occurrence of osteoporosis, obesity, and related metabolic disorders, a comprehensive understanding of the regulatory mechanisms balancing bone and fat metabolism is essential, with a particular focus on the fate determination of MSCs in osteogenic and adipogenic differentiation. In this review, we provide a detailed summary of how zinc finger proteins influence the osteogenic and adipogenic differentiation of MSCs through different signaling pathways, transcription factors, and epigenetic mechanisms. Additionally, we outline the regulatory mechanisms of ZFPs in controlling osteogenic and adipogenic differentiation based on various stages of MSC differentiation.

Fundamentals and Translational Applications of Stem Cells and Biomaterials in Dental, Oral and Craniofacial Regenerative Medicine.

Regenerative dental medicine continuously expands to improve treatments for prevalent clinical problems in dental and oral medicine. Stem cell based translational opportunities include regenerative therapies for tooth restoration, root canal therapy, and inflammatory processes (e.g., periodontitis). The potential of regenerative approaches relies on the biological properties of dental stem cells. These and other multipotent somatic mesenchymal stem cell (MSC) types can in principle be applied as either autologous or allogeneic sources in dental procedures. Dental stem cells have distinct developmental origins and biological markers that determine their translational utility. Dental regenerative medicine is supported by mechanistic knowledge of the molecular pathways that regulate dental stem cell growth and differentiation. Cell fate determination and lineage progression of dental stem cells is regulated by multiple cell signaling pathways (e.g., WNTs, BMPs) and epigenetic mechanisms, including DNA modifications, histone modifications, and non-coding RNAs (e.g., miRNAs and lncRNAs). This review also considers a broad range of novel approaches in which stem cells are applied in combination with biopolymers, ceramics, and composite materials, as well as small molecules (agonistic or anti-agonistic ligands) and natural compounds. Materials that mimic the microenvironment of the stem cell niche are also presented. Promising concepts in bone and dental tissue engineering continue to drive innovation in dental and non-dental restorative procedures.

Elevated Id2 expression causes defective meiosis and spermatogenesis in mice.

Inhibitors of DNA binding (ID) proteins mainly inhibit gene expression and regulate cell fate decisions by interacting with E-proteins. All four ID proteins (ID1-4) are present in the testis, and ID4 has a particularly important role in spermatogonial stem cell fate determination. Several lines of evidence indicate that ID proteins are involved in meiosis; however, functional experiments have not been conducted to validate this observation. In this study, we report that ID2 is enriched in spermatocytes and that forced ID2 expression in germ cells causes defects in spermatogenesis. A detailed analysis demonstrated that Id2 overexpression (Id2 OE) decreased the total number of spermatogonia and changed the dynamics of meiosis progression. Specifically, spermatocytes were enriched in the zygotene stage, and the proportion of pachytene spermatocytes was significantly decreased, indicating defects in the zygotene-pachytene transition. The number of MLH1-positive foci per cell was decreased in pachytene spermatocytes from Id2 OE testes, suggesting abnormalities in recombination. Transcriptome analysis revealed that forced Id2 expression changed the expression of a list of genes mainly associated with meiosis and spermatid development. ID2 protein is expressed in spermatocytes, and its genetic ablation in the germline does not affect spermatogenesis, likely due to genetic compensation of its family members. However, forced Id2 expression changes meiosis progression and causes defects in spermiogenesis. These data provide important evidence that ID proteins play pivotal roles in male meiosis and spermatid development.

Transcription Factor EB-Mediated Lysosomal Function Regulation for Determining Stem Cell Fate under Metabolic Stress.

Stem cells require high amounts of energy to replicate their genome and organelles and differentiate into numerous cell types. Therefore, metabolic stress has a major impact on stem cell fate determination, including self-renewal, quiescence, and differentiation. Lysosomes are catabolic organelles that influence stem cell function and fate by regulating the degradation of intracellular components and maintaining cellular homeostasis in response to metabolic stress. Lysosomal functions altered by metabolic stress are tightly regulated by the transcription factor EB (TFEB) and TFE3, critical regulators of lysosomal gene expression. Therefore, understanding the regulatory mechanism of TFEB-mediated lysosomal function may provide some insight into stem cell fate determination under metabolic stress. In this review, we summarize the molecular mechanism of TFEB/TFE3 in modulating stem cell lysosomal function and then elucidate the role of TFEB/TFE3-mediated transcriptional activity in the determination of stem cell fate under metabolic stress.

Understanding the Functional Differences between PRC2.1 and PRC2.2 in Hematopoiesis

Epigenetic mechanisms play an important role in the determination of hematopoietic stem and progenitor cell (HSPC) fate through the modification of chromatin architecture. Polycomb repressive complex 2 (PRC2) is an indispensable epigenetic modifier that catalyzes histone 3 lysine 27 tri-methylation (H3K27me3) repressive mark through its catalytic subunit EZH2. PRC2 is divided into two independent complexes, PRC2.1 and PRC2.2, based on the specific interacting accessory proteins. PRC2.1 comprises one of the three polycomb-like proteins (PCL1/PHF1, PCL2/MTF2, or PCL3/PHF19), as well as one of the two recently identified accessory proteins, EPOP or PALI1. However, PRC2.2 contains either JARID2 or AEBP2 as co-factors. These accessory proteins aid in the recruitment of the PRC2 complexes to different sites in the genome to catalyze H3K27me3. Recent studies performed on mouse embryonic stem cells showed that PRC2.1 and PRC2.2 are involved in distinct lineage specification and commitment through their specific co-factors. However, the functional differences between PRC2.1 and PRC2.2 in HSPC differentiation and lineage commitment has not been thoroughly characterized. To compare the complete loss of PRC2 versus PRC2.2-specific loss of function in normal hematopoiesis, we generated Mx1-Cre: Jarid2fl/fl and Mx1-Cre: Ezh2fl/fl mouse strains. Two-month old Mx1-Cre(+) control, Mx1-Cre(+): Jarid2fl/fl, and Mx1-Cre(+): Ezh2fl/fl mice were injected with six doses of polyinosinic:polycytidylic acid (pIpC) to promote conditional deletion of Jarid2 and Ezh2. Two months post-pIpC injection, whole bone marrow (WBM) cells were used to perform competitive transplantation (5x10 5 CD45.2 test WBM plus 5x10 5 CD45.1 competitor WBM cells). Jarid2-KO primary recipients showed the highest CD45.2 peripheral blood (PB) engraftment (~80%), and the highest tri-lineage (myeloid 80%, B cell 70%, T cell 80%) chimerism compared to the control (myeloid 50%, B cell 45%, T cell 25%) and Ezh2-KO (myeloid 65%, B cell 5%, T cell 5%) recipients. Ezh2-KO recipients showed the lowest CD45.2 PB engraftment (~20%), and were deficient for PB lymphoid cell production. Jarid2-KO recipients showed the highest bone marrow (BM) engraftment (~ 90%), while Ezh2-KO recipients showed significant BM lymphoid engraftment (B cell 55% and T cell 40%) inverse to what was observed in the PB (B cell 5% and T cell 5%). This suggested that complete loss of PRC2 function promotes blockage in the migration of mature lymphoid cells from BM to the PB. Jarid2-KO and Ezh2-KO recipients showed significant reductions in phenotypically defined HSC cell number and frequency compared to control recipients suggesting that PRC2.2 function through its co-factor Jarid2 to maintain HSC pool. To further investigate the functional effect of both genes' loss of function at the molecular level, we performed single cell transcriptomic analysis from KSL (lineage-, c-Kit+, Sca1+, CD45.2+) cells sorted from the aforementioned recipients. Based on transcriptomic analysis, Ezh2 loss of function promoted an increase in MPP4 and precursor B cell populations in the BM of recipient mice. However, no significant changes was observed between control and Jarid2-KO clusters. Analysis showed that the Mtf2 co-factor has the highest expression level throughout the whole KSL clusters compared to other PRC2.1 and PRC2.2 accessory proteins. These results suggested that Mtf2-associated PRC2.1 is necessary to restrict MPP4 self-renewal and to repress B cell priming in HSPCs. Although no significant change was observed in the HSC cluster between Jarid2-KO and Ezh2-KO genotypes, Jarid2 and Ezh2 loss of functions showed significant differential gene expression in the HSC population when compared to each other and to the control suggesting that Jarid2-associated PRC2.2 regulates a unique molecular program in HSCs independently from PRC2.1. Combined with our functional data, these results suggested that PRC2.2 restricts HSC-specific gene expression in non-self-renewing progenitor cells through its co-factor Jarid2; while PRC2.1 restricts MPP4 fate and B cell priming through its co-factor Mtf2. Ongoing studies are aimed to delineate the distinct molecular programs regulated by PRC2.1 and PRC2.2 in HSPCs using CRISPR-Cas9 sgRNA lentiviral system targeting each of the PRC2.1 and PRC2.2 accessory proteins independently and validation in lineage-specific functional assays.

StemDriver:a knowledgebase of gene functions for hematopoietic stem cell fate determination.

StemDriver is a comprehensive knowledgebase dedicated to the functional annotation of genes participating in the determination of hematopoietic stem cell fate, available at http://biomedbdc.wchscu.cn/StemDriver/. By utilizing single-cell RNA sequencing data, StemDriver has successfully assembled a comprehensive lineage map of hematopoiesis, capturing the entire continuum from the initial formation of hematopoietic stem cells to the fully developed mature cells. Extensive exploration and characterization were conducted on gene expression features corresponding to each lineage commitment. At the current version, StemDriver integrates data from 42 studies, encompassing a diverse range of 14 tissue types spanning from the embryonic phase to adulthood. In order to ensure uniformity and reliability, all data undergo a standardized pipeline, which includes quality data pre-processing, cell type annotation, differential gene expression analysis, identification of gene categories correlated with differentiation, analysis of highly variable genes along pseudo-time, and exploration of gene expression regulatory networks. In total, StemDriver assessed the function of 23 839 genes for human samples and 29 533 genes for mouse samples. Simultaneously, StemDriver also provided users with reference datasets and models for cell annotation. We believe that StemDriver will offer valuable assistance to research focused on cellular development and hematopoiesis.

Combined 3D bioprinting and tissue-specific ECM system reveals the influence of brain matrix on stem cell differentiation.

We have previously shown that human and murine breast extracellular matrix (ECM) can significantly impact cellular behavior, including stem cell fate determination. It has been established that tissue-specific extracellular matrix from the central nervous system has the capacity to support neuronal survival. However, the characterization of its influence on stem cell differentiation and its adaptation to robust 3D culture models is underdeveloped. To address these issues, we combined our 3D bioprinter with hydrogels containing porcine brain extracellular matrix (BMX) to test the influence of the extracellular matrix on stem cell differentiation. Our 3D bioprinting system generated reproducible 3D neural structures derived from mouse embryonic stem cells (mESCs). We demonstrate that the addition of BMX preferentially influences 3D bioprinted mESCs towards neural lineages compared to standard basement membrane (Geltrex/Matrigel) hydrogels alone. Furthermore, we demonstrate that we can transplant these 3D bioprinted neural cellular structures into a mouse's cleared mammary fat pad, where they continue to grow into larger neural outgrowths. Finally, we demonstrate that direct injection of human induced pluripotent stem cells (hiPSCS) and neural stem cells (NSCs) suspended in pure BMX formed neural structures in vivo. Combined, these findings describe a unique system for studying brain ECM/stem cell interactions and demonstrate that BMX can direct pluripotent stem cells to differentiate down a neural cellular lineage without any additional specific differentiation stimuli.

Resveratrol Mediated Regulation of Hippocampal Neuroregenerative Plasticity via SIRT1 Pathway in Synergy with Wnt Signaling: Neurotherapeutic Implications to Mitigate Memory Loss in Alzheimer's Disease.

Alzheimer's disease (AD) is a major form of dementia. Abnormal amyloidogenic event-mediated degeneration of cholinergic neurons in the cognitive centers of the brain has been attributed to neuropathological sequelae and behavioral deficits in AD. Besides, impaired adult neurogenesis in the hippocampus has experimentally been realized as an underlying cause of dementia regardless of neurodegeneration. Therefore, nourishing the neurogenic process in the hippocampus has been considered an effective therapeutic strategy to mitigate memory loss. In the physiological state, the Wnt pathway has been identified as a potent mitogenic generator in the hippocampal stem cell niche. However, downstream components of Wnt signaling have been noticed to be downregulated in AD brains. Resveratrol (RSV) is a potent Sirtuin1 (SIRT1) enhancer that facilitates neuroprotection and promotes neurogenesis in the hippocampus of the adult brain. While SIRT1 is an important positive regulator of Wnt signaling, ample reports indicate that RSV treatment strongly mediates the fate determination of stem cells through Wnt signaling. However, the possible therapeutic roles of RSV-mediated SIRT1 enhancement on the regulation of hippocampal neurogenesis and reversal of memory loss through the Wnt signaling pathway have not been addressed yet. Taken together, this review describes RSV-mediated effects on the regulation of hippocampal neurogenesis via the activation of SIRT1 in synergy with the Wnt signaling. Further, the article emphasizes a hypothesis that RSV treatment can provoke the activation of quiescent neural stem cells and prime their neurogenic capacity in the hippocampus via Wnt signaling in AD.