Regulation Of Stem Cell Function Research Articles

The peptidase DA1 cleaves and destabilizes WUSCHEL to control shoot apical meristem size

Stem cells in plants and animals are the source of new tissues and organs. In plants, stem cells are maintained in the central zone (CZ) of multicellular meristems, and large shoot meristems with an increased stem cell population hold promise for enhancing yield. The mobile homeodomain transcription factor WUSCHEL (WUS) is a central regulator of stem cell function in plant shoot meristems. Despite its central importance, the factors that directly modulate WUS protein stability have been a long-standing question. Here, we show that the peptidase DA1 physically interacts with and cleaves the WUS protein, leading to its destabilization. Furthermore, our results reveal that cytokinin signaling represses the level of DA1 protein in the shoot apical meristem, thereby increasing the accumulation of WUS protein. Consistent with these observations, loss of DA1 function results in larger shoot apical meristems with an increased stem cell population and also influences cytokinin-induced enlargement of shoot apical meristem. Collectively, our findings uncover a previously unrecognized mechanism by which the repression of DA1 by cytokinin signaling stabilizes WUS, resulting in the enlarged shoot apical meristems with the increased stem cell number during plant growth and development.

Functional diversity of apoptotic vesicle subpopulations from bone marrow mesenchymal stem cells in tissue regeneration.

Apoptosis releases numerous apoptotic vesicles that regulate processes such as cell proliferation, immunity, and tissue regeneration and repair. Now, it has also emerged as an attractive candidate for biotherapeutics. However, apoptotic vesicles encompass a diverse range of subtypes, and it remains unclear which specific subtypes play a pivotal role. In this study, we successfully isolated different apoptotic vesicle subtypes based on their sizes and characterized them using NTA and TEM techniques, respectively. We compared the functional variances among the distinct subtypes of apoptotic vesicles in terms of stem cell proliferation, migration, and differentiation, as well as for endothelial cell and macrophage function, effectively identifying subtypes that exhibit discernible functional differences. ApoSEV (with diameter<1000nm) promoted stem cell proliferation, migration, and multi-potent differentiation, and accelerated skin wound healing of diabetes mouse model, while apoBD (with diameter>1000nm) played the opposite effect on cell function and tissue regeneration. Lastly, employing protein analysis and gene sequencing techniques, we elucidated the intrinsic mechanisms underlying these differences between different subtypes of apoEVs. Collectively, this study identified that apoptotic vesicle subtypes possessed distinct bio-functions in regulating stem cell function and behaviour and modulating tissue regeneration, which primarily attribute to the distinct profiling of protein and mRNA in different subtypes. This comprehensive analysis of specific subtypes of apoEVs would provide novel insights for potential therapeutic applications in cell biology and tissue regeneration.

Targeting the redox system for cardiovascular regeneration in aging.

Cardiovascular aging presents a formidable challenge, as the aging process can lead to reduced cardiac function and heightened susceptibility to cardiovascular diseases. Consequently, there is an escalating, unmet medical need for innovative and effective cardiovascular regeneration strategies aimed at restoring and rejuvenating aging cardiovascular tissues. Altered redox homeostasis and the accumulation of oxidative damage play a pivotal role in detrimental changes to stem cell function and cellular senescence, hampering regenerative capacity in aged cardiovascular system. A mounting body of evidence underscores the significance of targeting redox machinery to restore stem cell self-renewal and enhance their differentiation potential into youthful cardiovascular lineages. Hence, the redox machinery holds promise as a target for optimizing cardiovascular regenerative therapies. In this context, we delve into the current understanding of redox homeostasis in regulating stem cell function and reprogramming processes that impact the regenerative potential of the cardiovascular system. Furthermore, we offer insights into the recent translational and clinical implications of redox-targeting compounds aimed at enhancing current regenerative therapies for aging cardiovascular tissues.

Protein ligand and nanotopography separately drive the phenotype of mouse embryonic stem cells

Biochemical and biomechanical signals regulate stem cell function in the niche environments in vivo. Current in vitro culture of mouse embryonic stem cells (mESC) uses laminin (LN-511) to provide mimetic biochemical signaling (LN-521 for human systems) to maintain stemness. Alternative approaches propose topographical cues to provide biomechanical cues, however combined biochemical and topographic cues may better mimic the in vivo environment, but are largely unexplored for in vitro stem cell expansion. In this study, we directly compare in vitro signals from LN-511 and/or topographic cues to maintain stemness, using systematically-varied submicron pillar patterns or flat surfaces with or without preadsorbed LN-511. The adhesion of cells, colony formation, expression of the pluripotency marker,octamer-binding transcription factor 4 (Oct4), and transcriptome profiling were characterized. We observed that either biochemical or topographic signals could maintain stemness of mESCs in feeder-free conditions, indicated by high-level Oct4 and gene profiling by RNAseq. The combination of LN-511 with nanotopography reduced colony growth, while maintaining stemness markers, shifted the cellular phenotype indicating that the integration of biochemical and topographic signals is antagonistic. Overall, significantly faster (up to 2.5 times) colony growth was observed at nanotopographies without LN-511, suggesting for improved ESC expansion.

3D micropattern force regulates stem cell function.

3D micropattern force regulates stem cell function.

M2 macrophage-derived exosomes carry miR-142-3p to restore the differentiation balance of irradiatedBMMSCs by targeting TGF-β1.

Radiotherapy is essential to cancer treatment, while it inevitably injures surrounding normal tissues, and bone tissue is one of the most common sites prone to irradiation. Bone marrow mesenchymal stem cells (BMMSCs) are sensitive to irradiation and the irradiated dysfunction of BMMSCs may be closely related to irradiation-induced bone damage. Macropahges play important role in regulating stem cell function, bone metabolic balance and irradiation response, but the effects of macrophages on irradiated BMMSCs are still unclear. This study aimed to investigate the role of macrophages and macrophage-derived exosomes in restoring irradiated BMMSCs function. The effects of macrophage conditioned medium (CM) and macrophage-derived exosomes on osteogenic and fibrogenic differentiation capacities of irradiated BMMSCs were detected. The key microribonucleic acids (miRNAs) and targeted proteins in exosomes were also determined. The results showed that irradiation significantly inhibited the proliferation of BMMSCs, and caused differentiation imbalance of BMMSCs, with decreased osteogenic differentiation and increased fibrogenic differentiation. M2 macrophage-derived exosomes (M2D-exos) inhibited the fibrogenic differentiation and promoted the osteogenic differentiation of irradiated BMMSCs. We identified that miR-142-3p was significantly overexpressed in M2D-exos and irradiated BMMSCs treated with M2D-exos. After inhibition of miR-142-3p in M2 macrophage, the effects of M2D-exos on irradiated BMMSCs differentiation were eliminated. Furthermore, transforming growth factor beta 1 (TGF-β1), as a direct target of miR-142-3p, was significantly decreased in irradiated BMMSCs treated with M2D-exos. This study indicated that M2D-exos could carry miR-142-3p to restore the differentiation balance of irradiated BMMSCs by targeting TGF-β1. These findings pave a new way for promising and cell-free method to treat irradiation-induced bone damage.

Genetically Engineered Triple MAPT-Mutant Human-Induced Pluripotent Stem Cells (N279K, P301L, and E10+16 Mutations) Exhibit Impairments in Mitochondrial Bioenergetics and Dynamics

Pathological abnormalities in the tau protein give rise to a variety of neurodegenerative diseases, conjointly termed tauopathies. Several tau mutations have been identified in the tau-encoding gene MAPT, affecting either the physical properties of tau or resulting in altered tau splicing. At early disease stages, mitochondrial dysfunction was highlighted with mutant tau compromising almost every aspect of mitochondrial function. Additionally, mitochondria have emerged as fundamental regulators of stem cell function. Here, we show that compared to the isogenic wild-type triple MAPT-mutant human-induced pluripotent stem cells, bearing the pathogenic N279K, P301L, and E10+16 mutations, exhibit deficits in mitochondrial bioenergetics and present altered parameters linked to the metabolic regulation of mitochondria. Moreover, we demonstrate that the triple tau mutations disturb the cellular redox homeostasis and modify the mitochondrial network morphology and distribution. This study provides the first characterization of disease-associated tau-mediated mitochondrial impairments in an advanced human cellular tau pathology model at early disease stages, ranging from mitochondrial bioenergetics to dynamics. Consequently, comprehending better the influence of dysfunctional mitochondria on the development and differentiation of stem cells and their contribution to disease progression may thus assist in the potential prevention and treatment of tau-related neurodegeneration.

High levels of Wnt signaling reduce proliferation and promote deep glandular differentiation in human gastric corpus-derived organoids

Introduction: The Wnt signaling pathway regulates gastrointestinal stem cell identity and function. Throughout the stomach, a gradient of Wnt signaling is established across the epithelium with highest levels at the gland base. However, the proximal (corpus) and distal (antrum) regions of the stomach have strikingly different epithelial architectures, with antral stem cells and proliferation localized to the gland base while corpus proliferation is localized higher-up in the glands where Wnt levels are diminished. It is unknown how Wnt may differentially regulate cell function and identity within these regions. Furthermore, activation of Wnt signaling through genetic diseases, such as familial adenomatous polyposis (FAP), results in abundant polyposis of the corpus while the antrum is mostly spared. This study investigated how activation of Wnt signaling in human gastric corpus- and antral-derived organoids regulates stem cell function, with the hypothesis that corpus stem/progenitor cells have a lower threshold for optimal Wnt signaling. Methods: Human patient-matched corpus and antral organoids from two individuals were grown in the presence of the Wnt agonist CHIR99021 to assess Wnt sensitivity in vitro­. Corpus organoids were then grown in high levels of CHIR99021 to assess how Wnt activation affected growth, proliferation, differentiation, and stem cell maintenance. Results: Treatment with CHIR99021 in media free of exogenous Wnt/R-spondin stimulated a maximal growth response in corpus organoids at lower concentrations than their respective patient-matched antral organoids. Increased Wnt signaling beyond the maximal growth concentration also suppressed proliferation at a rapid rate in corpus organoids. Chronic treatment with increasing levels of CHIR99021 reduced organoid size, suppressed surface cell differentiation, and promoted expression of markers for differentiated deep glandular cells, including neck and chief cells. Surprisingly, organoids retained in high levels of CHIR99021 had enhanced organoid formation ability when passaged, indicating that progenitor cell status was maintained in these non-proliferative, deep glandular cell-enriched organoids. Passaging these quiescent organoids into low Wnt conditions led to a rapid rescue of normal growth, organoid morphology, and surface cell differentiation. Conclusion: Our findings suggest that progenitor cells in human corpus organoids have a lower threshold for optimal Wnt signaling than progenitor cells in the antrum. We demonstrate that Wnt signaling in the corpus plays a central role in defining a bimodal axis of differentiation, and that high activation of Wnt signaling promotes stemness while simultaneously suppressing proliferation. Funding support for this research was provided through T32DK094775 (KPM), R25GM086262 (ED), and R01DK126451 (LCS). This is the full abstract presented at the American Physiology Summit 2023 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.

MY-1-Loaded Nano-Hydroxyapatite Accelerated Bone Regeneration by Increasing Type III Collagen Deposition in Early-Stage ECM via a Hsp47-Dependent Mechanism.

The extracellular matrix (ECM) plays a crucial part in regulating stem cellfunction through its distinctive mechanical and chemical effect. Therefore, it is worth studying how to activate the driving force of osteoblast cells by dynamic changing of ECM and accelerate the bone regeneration. In this research, a novel peptide MY-1is designed and synthesized. To achieve its sustained releasing, the nano-hydroxyapatite (nHA) is chosen as the carrier of MY-1by mixed adsorption. The results reveal that the sustainable releasing of MY-1regulates the synthesis and secretion of ECM from rat bone marrow mesenchymal stem cells (rBMSCs), which promotes the cell migration and osteogenic differentiation in the early stage of bone regeneration. Further analyses demonstrate that MY-1increases the expression and nuclear translocation of β-catenin, and then upregulates the level of heat shock protein 47 (Hsp47), thereby accelerating the synthesis and secretion of type IIIcollagen (Col III) at the early stage. Finally, the promoted rapid transformation of Col III to Col I at late stage benefits the bone regeneration. Hence, this study can provide a theoretical basis for the local application of MY-1in bone regeneration.

Rejuvenating muscle stem cells with the glutathione system

Aging results from the combination of complex processes still largely undefined. In this issue, Benjamin etal. use multiomic analysis to reveal a causative role of altered glutathione (GSH) synthesis and metabolism in age-dependent muscle stem cell (MuSC) dysfunction, casting light on novel mechanisms regulating stem cell function and on therapeutic approaches to improve defective regeneration in the aged muscle.

FRUCTOSE INSENSITIVE1 regulates stem cell function in Arabidopsis in response to fructose signalling.

Stem cell function in different meristems of Arabidopsis is mainly defined by WUSCHEL (WUS)/WUSCHEL-RELATED HOMEOBOX (WOX) family of proteins. Sugars have also been demonstrated to play pivotal roles in stem cell function and development of plants. As a cytosolic fructose 1,6-bisphosphatase, FRUCTOSE INSENSITIVE1 (FINS1) has been demonstrated to regulate plant growth in response to fructose signalling. However, it remains to be elucidated how stem cell function is regulated in response to fructose signalling. Our work showed that FINS1 interacts with WUS/WOX protein as a complex, which further binds to the promoter of WUS/WOX and regulates its expression in response to fructose signalling. FINS1 might act as a bifunctional factor that promotes WUS/WOX expression in the presence of low concentrations of fructose, and represses WUS/WOX expression in the presence of high concentrations of fructose. Therefore, FINS1 regulates stem cell function in response to fructose signalling during the growth and development of Arabidopsis.

Dissecting Physical and Biochemical Effects in Nanotopographical Regulation of Cell Behavior.

Regulating cell behavior using nanotopography has been widely implemented. To facilitate cell adhesion, physical nanotopography is usually coated with adhesive proteins such as fibronectin (FN). However, the confounding effects of physical and biochemical cues of nanotopography hinder the understanding of nanotopography in regulating cell behavior, which ultimately limits the biomedical applications of nanotopography. To delineate the roles of the physical and biochemical cues in cell regulation, we fabricate substrates that have either the same physical nanotopography but different biochemical (FN) nanopatterns or identical FN nanopatterns but different physical nanotopographies. We then examine the influences of physical and biochemical cues of nanotopography on spreading, nuclear deformation, mechanotransduction, and function of human mesenchymal stem cells (hMSCs). Our results reveal that physical topographies, especially nanogratings, dominantly control cell spreading, YAP localization, proliferation, and differentiation of hMSCs. However, biochemical FN nanopatterns affect hMSC elongation, YAP intracellular localization, and lamin a/c (LAMAC) expression. Furthermore, we find that physical nanogratings induce nanoscale curvature of nuclei at the basal side, which attenuates the osteogenic differentiation of hMSCs. Collectively, our study highlights the dominant effect of physical nanotopography in regulating stem cell functions, while suggesting that fine-tuning of cell behavior can be achieved through altering the presentation of biochemical cues on substrate surfaces.

Measuring Oxygen Consumption Rate (OCR) and Extracellular Acidification Rate (ECAR) in Muscle Stem Cells Using a Seahorse Analyzer: Applicability for Aging Studies.

In recent years, evidence showing metabolism as a fundamental regulator of stem cell functions has emerged. In skeletal muscle, its stem cells (satellite cells) sustain muscle regeneration, although they lose their regenerative potential with aging, and this has been attributed, at least in part, to changes in their metabolism. In this chapter, we describe a protocol to analyze the metabolism of satellite cells using the Seahorse technology, which can be applied to aging mice.

Wnt signalling in cell division: from mechanisms to tissue engineering.

Wnt signalling is an essential player in tissue formation, notably in the regulation of stem cell function. Wnt signalling is best known for its roles in G1/S progression. However, a complex Wnt programme that also mediates mitotic progression and asymmetric cell division (ACD) is emerging. Recent developments in this area have provided mechanistic insights as well as tools to engineer or target Wnt signalling for translational and therapeutic purposes. Here, we discuss the bidirectional relationship between Wnt activity and mitosis. We emphasise how various Wnt-dependent mechanisms control spindle dynamics, chromosome segregation, and ACD. Finally, we illustrate how knowledge about these mechanisms has been successfully employed in tissue engineering for regenerative medicine applications.

SZT2 maintains hematopoietic stem cell homeostasis via nutrient-mediated mTORC1 regulation.

The mTORC1 pathway coordinates nutrient and growth factor signals to maintain organismal homeostasis. Whether nutrient signaling to mTORC1 regulates stem cell function remains unknown. Here, we show that SZT2 — a protein required for mTORC1 downregulation upon nutrient deprivation — is critical for hematopoietic stem cell (HSC) homeostasis. Ablation of SZT2 in HSCs decreased the reserve and impaired the repopulating capacity of HSCs. Furthermore, ablation of both SZT2 and TSC1 — 2 repressors of mTORC1 on the nutrient and growth factor arms, respectively — led to rapid HSC depletion, pancytopenia, and premature death of the mice. Mechanistically, loss of either SZT2 or TSC1 in HSCs led to only mild elevation of mTORC1 activity and reactive oxygen species (ROS) production. Loss of both SZT2 and TSC1, on the other hand, simultaneously produced a dramatic synergistic effect, with an approximately 10-fold increase of mTORC1 activity and approximately 100-fold increase of ROS production, which rapidly depleted HSCs. These data demonstrate a critical role of nutrient mTORC1 signaling in HSC homeostasis and uncover a strong synergistic effect between nutrient- and growth factor–mediated mTORC1 regulation in stem cells.

Interplay between mechanics and signalling in regulating cell fate.

Mechanical signalling affects multiple biological processes during development and in adult organisms, including cell fate transitions, cell migration, morphogenesis and immune responses. Here, we review recent insights into the mechanisms and functions of two main routes of mechanical signalling: outside-in mechanical signalling, such as mechanosensing of substrate properties or shear stresses; and mechanical signalling regulated by the physical properties of the cell surface itself. We discuss examples of how these two classes of mechanical signalling regulate stem cell function, as well as developmental processes in vivo. We also discuss how cell surface mechanics affects intracellular signalling and, in turn, how intracellular signalling controls cell surface mechanics, generating feedback into the regulation of mechanosensing. The cooperation between mechanosensing, intracellular signalling and cell surface mechanics has a profound impact on biological processes. We discuss here our understanding of how these three elements interact to regulate stem cell fate and development.

Upregulation of BMI1-suppressor miRNAs (miR-200c, miR-203) during terminal differentiation of colon epithelial cells.

MicroRNAs (miRNAs) are key regulators of stem cell functions, including self-renewal and differentiation. In this study, we aimed to identify miRNAs that are upregulated during terminal differentiation in the human colon epithelium, and elucidate their role in the mechanistic control of stem cell properties. "Bottom-of-the-crypt" (EPCAM+/CD44+/CD66alow) and "top-of-the-crypt" (EPCAM+/CD44neg/CD66ahigh) epithelial cells from 8 primary colon specimens (6 human, 2 murine) were purified by flow cytometry and analyzed for differential expression of 335 miRNAs. The miRNAs displaying the highest upregulation in "top-of-the-crypt" (terminally differentiated) epithelial cells were tested for positive correlation and association with survival outcomes in a colon cancer RNA-seq database (n = 439 patients). The two miRNAs with the strongest "top-of-the-crypt" expression profile were evaluated for capacity to downregulate self-renewal effectors and inhibit in vitro proliferation of colon cancer cells, in vitro organoid formation by normal colon epithelial cells and in vivo tumorigenicity by patient-derived xenografts (PDX). Six miRNAs (miR-200a, miR-200b, miR-200c, miR-203, miR-210, miR-345) were upregulated in "top-of-the-crypt" cells and positively correlated in expression among colon carcinomas. Overexpression of the three miRNAs with the highest inter-correlation coefficients (miR-200a, miR-200b, miR-200c) associated with improved survival. The top two over-expressed miRNAs (miR-200c, miR-203) cooperated synergistically in suppressing expression of BMI1, a key regulator of self-renewal in stem cell populations, and in inhibiting proliferation, organoid-formation and tumorigenicity of colon epithelial cells. In the colon epithelium, terminal differentiation associates with the coordinated upregulation of miR-200c and miR-203, which cooperate to suppress BMI1 and disable the expansion capacity of epithelial cells.

MITOCHONDRIA: Mitochondrial dynamics in the regulation of stem cells.

Mitochondria are considered the metabolic hubs within a cell. These organelles are highly dynamic and continuously undergo cycles of fission and fusion events. The balance in the dynamic state of mitochondria is critical for maintaining key physiological events within cells. Here we discuss the emerging role of mitochondrial dynamics in regulating stem cell function and highlight the crosstalk between mitochondrial shape and intracellular signaling cascades within the context of stem cells.

CLAVATA modulates auxin homeostasis and transport to regulate stem cell identity and plant shape in a moss.

Summary The CLAVATA pathway is a key regulator of stem cell function in the multicellular shoot tips of Arabidopsis, where it acts via the WUSCHEL transcription factor to modulate hormone homeostasis. Broad‐scale evolutionary comparisons have shown that CLAVATA is a conserved regulator of land plant stem cell function, but CLAVATA acts independently of WUSCHEL‐like (WOX) proteins in bryophytes. The relationship between CLAVATA, hormone homeostasis and the evolution of land plant stem cell functions is unknown.Here we show that in the moss, Physcomitrella (Physcomitrium patens), CLAVATA affects stem cell activity by modulating hormone homeostasis. CLAVATA pathway genes are expressed in the tip cells of filamentous tissues, regulating cell identity, filament branching, plant spread and auxin synthesis. The receptor‐like kinase PpRPK2 plays the major role, and Pprpk2 mutants have abnormal responses to cytokinin, auxin and auxin transport inhibition, and show reduced expression of PIN auxin transporters.We propose a model whereby PpRPK2 modulates auxin gradients in filaments to determine stem cell identity and overall plant form.Our data indicate that CLAVATA‐mediated auxin homeostasis is a fundamental property of plant stem cell function, probably exhibited by the last shared common ancestor of land plants.

Construction and Evaluation of Recombinant Chimeric Fibrillin and Elastin Fragment in Human Mesenchymal Stem Cells.

Diverse extracellular matrix (ECM) proteins physically interact with stem cells and regulate stem cell function. However, the large molecular weight of the natural ECM renders large-scale fabrication of a similar functional structure challenging. The objective of this study was to construct a low molecular weight and multifunctional chimeric form of recombinant ECM to stimulate mesenchymal stem cell (MSC) for tissue repair. We engineered Fibrillin-1PF14 fused to an elastin-like polypeptide to develop a new biomimetic ECM for stem cell differentiation and investigated whether this recombinant chimeric Fibrillin-Elastin fragment (rcFE) was effective on human nasal inferior turbinate-derived mesenchymal stem cells (hTMSCs). hTMSCs were grown in the medium supplemented with rcFE, then the effect of the protein was confirmed through cell adhesion assay, proliferation assay, and real-time PCR. rcFE enhanced the adhesion activity of hTMSCs by 2.7-fold at the optimal concentration, and the proliferation activity was 2.6-fold higher than that of the control group (non-treatment rcFE). In addition, when smooth muscle cell differentiation markers were identified by real-time PCR, Calponin increased about 6-fold, α-actin about 9-fold, and MYH11 about 10-fold compared to the control group. Chimeric rcFE enhanced cellular functions such as cell adhesion, proliferation, and smooth muscle differentiation of hTMSCs, suggesting that the rcFE can facilitate the induction of tissue regeneration.