Showing papers in "Stem Cells in 2020"
TL;DR: The global trends associated with preclinical development of MSC‐derived exosome‐based therapies, including immunogenicity, source of exosomes, isolation methods, biodistribution, and disease categories tested to date are reviewed.
Abstract: Exosomes are nanovesicles secreted by virtually all cells. Exosomes mediate the horizontal transfer of various macromolecules previously believed to be cell-autonomous in nature, including nonsecretory proteins, various classes of RNA, metabolites, and lipid membrane-associated factors. Exosomes derived from mesenchymal stem/stromal cells (MSCs) appear to be particularly beneficial for enhancing recovery in various models of disease. To date, there have been more than 200 preclinical studies of exosome-based therapies in a number of different animal models. Despite a growing number of studies reporting the therapeutic properties of MSC-derived exosomes, their underlying mechanism of action, pharmacokinetics, and scalable manufacturing remain largely outstanding questions. Here, we review the global trends associated with preclinical development of MSC-derived exosome-based therapies, including immunogenicity, source of exosomes, isolation methods, biodistribution, and disease categories tested to date. Although the in vivo data assessing the therapeutic properties of MSC-exosomes published to date are promising, several outstanding questions remain to be answered that warrant further preclinical investigation.
137 citations
TL;DR: In this review, the process of HSPC and MSC homing is examined, as are methods to enhance this process.
Abstract: Stem cell homing is a multistep endogenous physiologic process that is also used by exogenously administered hematopoietic stem and progenitor cells (HSPCs). This multistep process involves cell migration and is essential for hematopoietic stem cell transplantation. The process can be manipulated to enhance ultimate engraftment potential, and understanding stem cell homing is also important to the understanding of stem cell mobilization. Homing is also of potential importance in the recruitment of marrow mesenchymal stem and stromal cells (MSCs) to sites of injury and regeneration. This process is less understood but assumes importance when these cells are used for repair purposes. In this review, the process of HSPC and MSC homing is examined, as are methods to enhance this process.
104 citations
TL;DR: The tripartite-motif (TRIM) family of proteins represents one of the largest classes of putative single protein RING-finger E3 ubiquitin ligases as discussed by the authors.
Abstract: The tripartite-motif (TRIM) family of proteins represents one of the largest classes of putative single protein RING-finger E3 ubiquitin ligases. The members of this family are characterized by an N-terminal TRIM motif containing one RING-finger domain, one or two zinc-finger domains called B boxes (B1 box and B2 box), and a coiled-coil region. The TRIM motif can be found in isolation or in combination with a variety of C-terminal domains, and based on C-terminus, TRIM proteins are classified into 11 distinct groups. Because of the complex nature of TRIM proteins, they are implicated in a variety of cellular functions and biological processes, including regulation of cell proliferation, cell division and developmental processes, cancer transformation, regulation of cell metabolism, autophagocytosis, modification of chromatin status, regulation of gene transcription, post-translational modifications, and interactions with pathogens. Here, we demonstrate the specific activities of TRIM family proteins that contribute to the cancer stem cell phenotype. A growing body of evidence demonstrates that several TRIM members guarantee the acquisition of stem cell properties and the ability to sustain stem-like phenotype by cancer cells using distinct mechanisms. For other members, further work is needed to understand their full contribution to stem cell self-renewal. Identification of TRIM proteins that possess the potential to serve as therapeutic targets may result in the development of new therapeutic strategies. Finally, these strategies may result in the disruption of the machinery of stemness acquisition, which may prevent tumor growth, progression, and overcome the resistance to anticancer therapies.
69 citations
TL;DR: The current status and future potential of combining retinal organoids as human models with recent technologies to advance the development of gene, cell, and drug therapies for retinopathies are discussed.
Abstract: Retinal diseases constitute a genetically and phenotypically diverse group of clinical conditions leading to vision impairment or blindness with limited treatment options. Advances in reprogramming of somatic cells to induced pluripotent stem cells and generation of three-dimensional organoids resembling the native retina offer promising tools to interrogate disease mechanisms and evaluate potential therapies for currently incurable retinal neurodegeneration. Next-generation sequencing, single-cell analysis, advanced electrophysiology, and high-throughput screening approaches are expected to greatly expand the utility of stem cell-derived retinal cells and organoids for developing personalized treatments. In this review, we discuss the current status and future potential of combining retinal organoids as human models with recent technologies to advance the development of gene, cell, and drug therapies for retinopathies.
67 citations
TL;DR: A comprehensive review of plausible metabolic switches in response to implantation and of the various strategies currently used to leverage MSC metabolism to improve stem cell‐based therapeutics is provided.
Abstract: In tissue engineering and regenerative medicine, stem cell-specifically, mesenchymal stromal/stem cells (MSCs)-therapies have fallen short of their initial promise and hype. The observed marginal, to no benefit, success in several applications has been attributed primarily to poor cell survival and engraftment at transplantation sites. MSCs have a metabolism that is flexible enough to enable them to fulfill their various cellular functions and remarkably sensitive to different cellular and environmental cues. At the transplantation sites, MSCs experience hostile environments devoid or, at the very least, severely depleted of oxygen and nutrients. The impact of this particular setting on MSC metabolism ultimately affects their survival and function. In order to develop the next generation of cell-delivery materials and methods, scientists must have a better understanding of the metabolic switches MSCs experience upon transplantation. By designing treatment strategies with cell metabolism in mind, scientists may improve survival and the overall therapeutic potential of MSCs. Here, we provide a comprehensive review of plausible metabolic switches in response to implantation and of the various strategies currently used to leverage MSC metabolism to improve stem cell-based therapeutics.
64 citations
TL;DR: In this paper, the aberrant lipid metabolism that CSCs exploit to boost their survival, which comprises upregulation in de novo lipogenesis, lipid droplet synthesis, lipid desaturation, and β-oxidation.
Abstract: Emerging evidence in cancer metabolomics has identified reprogrammed metabolic pathways to be a major hallmark of cancer, among which deregulated lipid metabolism is a prominent field receiving increasing attention. Cancer stem cells (CSCs) comprise <0.1% of the tumor bulk and possess high self-renewal, tumor-initiating properties, and are responsible for therapeutic resistance, disease recurrence, and tumor metastasis. Hence, it is imperative to understand the metabolic rewiring occurring in CSCs, especially their lipid metabolism, on which there have been recent reports. CSCs rely highly upon lipid metabolism for maintaining their stemness properties and fulfilling their biomass and energy demands, ultimately leading to cancer growth and invasion. Hence, in this review we will shed light on the aberrant lipid metabolism that CSCs exploit to boost their survival, which comprises upregulation in de novo lipogenesis, lipid droplet synthesis, lipid desaturation, and β-oxidation. Furthermore, the metabolic regulators involved in the process, such as key lipogenic enzymes, are also highlighted. Finally, we also summarize the therapeutic strategies targeting the key regulators involved in CSCs' lipid metabolism, which thereby demonstrates the potential to develop powerful and novel therapeutics against the CSC lipid metabolome.
57 citations
TL;DR: Activation of the Piezo1 channel stimulates MSC migration via inducing ATP release and subsequent activation of the P2 receptor purinergic signaling and downstream PYK2 and MEK/ERK signaling pathways, thus revealing novel insights into the molecular and signaling mechanisms regulating MSC Migration.
Abstract: In this study, we examined the Ca2+ -permeable Piezo1 channel, a newly identified mechanosensing ion channel, in human dental pulp-derived mesenchymal stem cells (MSCs) and hypothesized that activation of the Piezo1 channel regulates MSC migration via inducing ATP release and activation of the P2 receptor purinergic signaling. The Piezo1 mRNA and protein were readily detected in hDP-MSCs from multiple donors and, consistently, brief exposure to Yoda1, the Piezo1 channel-specific activator, elevated intracellular Ca2+ concentration. Yoda1-induced Ca2+ response was inhibited by ruthenium red or GsMTx4, two Piezo1 channel inhibitors, and also by Piezo1-specific siRNA. Brief exposure to Yoda1 also induced ATP release. Persistent exposure to Yoda1 stimulated MSC migration, which was suppressed by Piezo1-specific siRNA, and also prevented by apyrase, an ATP scavenger, or PPADS, a P2 generic antagonist. Furthermore, stimulation of MSC migration induced by Yoda1 as well as ATP was suppressed by PF431396, a PYK2 kinase inhibitor, or U0126, an inhibitor of the mitogen-activated protein kinase MEK/ERK signaling pathway. Collectively, these results suggest that activation of the Piezo1 channel stimulates MSC migration via inducing ATP release and subsequent activation of the P2 receptor purinergic signaling and downstream PYK2 and MEK/ERK signaling pathways, thus revealing novel insights into the molecular and signaling mechanisms regulating MSC migration. Such findings provide useful information for evolving a full understanding of MSC migration and homing and developing strategies to improve MSC-based translational applications.
53 citations
TL;DR: This review critically assess recent translational research related to the outcomes and mechanistic basis of MSC effects on T‐reg and provides a perspective on the potential for this knowledge base to be further exploited for the treatment of autoimmune disorders and transplants.
Abstract: The immunomodulatory potential of mesenchymal stromal cells (MSCs) and regulatory T cells (T-reg) is well recognized by translational scientists in the field of regenerative medicine and cellular therapies. A wide range of preclinical studies as well as a limited number of human clinical trials of MSC therapies have not only shown promising safety and efficacy profiles but have also revealed changes in T-reg frequency and function. However, the mechanisms underlying this potentially important observation are not well understood and, consequently, the optimal strategies for harnessing MSC/T-reg cross-talk remain elusive. Cell-to-cell contact, production of soluble factors, reprogramming of antigen presenting cells to tolerogenic phenotypes, and induction of extracellular vesicles ("exosomes") have emerged as possible mechanisms by which MSCs produce an immune-modulatory milieu for T-reg expansion. Additionally, these two cell types have the potential to complement each other's immunoregulatory functions, and a combinatorial approach may exert synergistic effects for the treatment of immunological diseases. In this review, we critically assess recent translational research related to the outcomes and mechanistic basis of MSC effects on T-reg and provide a perspective on the potential for this knowledge base to be further exploited for the treatment of autoimmune disorders and transplants.
51 citations
TL;DR: The involvement of m6A modification in CSCs provides a new direction for exploring cancer pathogenesis and inspires the development of effective strategies to fully eliminate both cancer cells and C SCs.
Abstract: Cancer stem cells (CSCs), a unique subset of undifferentiated cells with stem cell-like properties, have emerged as driving forces in mediating tumor growth, metastasis, and therapeutic resistance. Recent advances have highlighted that N6-methyladenosine (m6A) RNA modification plays an important role in cancer biology and CSCs. Dynamic m6A decoration has been demonstrated to be involved in CSC generation and maintenance, governing cancer progression and therapeutic resistance. In this review, we provide the first overview of the current knowledge of m6A modification implicated in CSCs and their impact on CSC properties, tumor progression, and responses to treatment. Finally, we also highlight the potential of m6A machinery as novel targets for cancer therapeutics. The involvement of m6A modification in CSCs provides a new direction for exploring cancer pathogenesis and inspires the development of effective strategies to fully eliminate both cancer cells and CSCs.
51 citations
TL;DR: The results illustrated that hUMSC‐Exos restored ovarian phenotype and function in a POI mouse model, promoted proliferation of CTX‐damaged hGCs and ovarian cells, and alleviated ROS accumulation by delivering exosomal miR‐17‐5P and inhibiting SIRT7 expression.
Abstract: Premature ovarian insufficiency (POI) is clinically irreversible in women aged over 40 years. Although numerous studies have demonstrated satisfactory outcomes of mesenchymal stem cell therapy, the underlying therapeutic mechanism remains unclear. Exosomes were collected from the culture medium of human umbilical cord mesenchymal stem cells (hUMSCs) and assessed by electron microscopy and Western blot (WB) analysis. Then, exosomes were added to the culture medium of cyclophosphamide (CTX)-damaged human granulosa cells (hGCs), and the mixture was injected into the ovaries of CTX-induced POI model mice before detection of antiapoptotic and apoptotic gene expression. Next, the microRNA expression profiles of hUMSC-derived exosomes (hUMSC-Exos) were detected by small RNA sequencing. The ameliorative effect of exosomal microRNA-17-5P (miR-17-5P) was demonstrated by miR-17-5P knockdown before assessment of ovarian phenotype and function, reactive oxygen species (ROS) levels and SIRT7 expression. Finally, SIRT7 was inhibited or overexpressed by RNA interference or retrovirus transduction, and the protein expression of PARP1, γH2AX, and XRCC6 was analyzed. The ameliorative effect of hUMSC-Exos on POI was validated. Our results illustrated that hUMSC-Exos restored ovarian phenotype and function in a POI mouse model, promoted proliferation of CTX-damaged hGCs and ovarian cells, and alleviated ROS accumulation by delivering exosomal miR-17-5P and inhibiting SIRT7 expression. Moreover, our findings elucidated that miR-17-5P repressed PARP1, γH2AX, and XRCC6 by inhibiting SIRT7. Our findings suggest a critical role for exosomal miR-17-5P and its downstream target mRNA SIRT7 in hUMSC transplantation therapy. This study indicates the promise of exosome-based therapy for POI treatment.
48 citations
TL;DR: The robust generation of MOs with homogeneous distribution of midbrain dopaminergic (mDA) neurons is described, indicating that MOs could be a proper human model system for studying the in vivo pathology of Parkinson's disease (PD).
Abstract: Recent studies have demonstrated the generation of midbrain-like organoids (MOs) from human pluripotent stem cells. However, the low efficiency of MO generation and the relatively immature and heterogeneous structures of the MOs hinder the translation of these organoids from the bench to the clinic. Here we describe the robust generation of MOs with homogeneous distribution of midbrain dopaminergic (mDA) neurons. Our MOs contain not only mDA neurons but also other neuronal subtypes as well as functional glial cells, including astrocytes and oligodendrocytes. Furthermore, our MOs exhibit mDA neuron-specific cell death upon treatment with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, indicating that MOs could be a proper human model system for studying the in vivo pathology of Parkinson's disease (PD). Our optimized conditions for producing homogeneous and mature MOs might provide an advanced patient-specific platform for in vitro disease modeling as well as for drug screening for PD.
TL;DR: The Wnt/β‐catenin pathway is identified as a critical determinant of cardiac myocyte subtype commitment during ESC differentiation: endogenous Wnt signaling favors the pacemaker lineage, whereas its suppression promotes the chamber cardiomyocyte lineage.
Abstract: Cardiac differentiation of embryonic stem cells (ESCs) can give rise to de novo chamber cardiomyocytes and nodal pacemaker cells. Compared with our understanding of direct differentiation toward atrial and ventricular myocytes, the mechanisms for nodal pacemaker cell commitment are not well understood. Taking a cue from the prominence of canonical Wnt signaling during cardiac pacemaker tissue development in chick embryos, we asked if modulations of Wnt signaling influence cardiac progenitors to bifurcate to either chamber cardiomyocytes or pacemaker cells. Omitting an exogenous Wnt inhibitor, which is routinely added to maximize cardiac myocyte yield during differentiation of mouse and human ESCs, led to increased yield of spontaneously beating cardiomyocytes with action potential properties similar to those of native sinoatrial node pacemaker cells. The pacemaker phenotype was accompanied by enhanced expression of genes and gene products that mark nodal pacemaker cells such as Hcn4, Tbx18, Tbx3, and Shox2. Addition of exogenous Wnt3a ligand, which activates canonical Wnt/β-catenin signaling, increased the yield of pacemaker-like myocytes while reducing cTNT-positive pan-cardiac differentiation. Conversely, addition of inhibitors of Wnt/β-catenin signaling led to increased chamber myocyte lineage development at the expense of pacemaker cell specification. The positive impact of canonical Wnt signaling on nodal pacemaker cell differentiation was evidenced in direct differentiation of two human ESC lines and human induced pluripotent stem cells. Our data identify the Wnt/β-catenin pathway as a critical determinant of cardiac myocyte subtype commitment during ESC differentiation: endogenous Wnt signaling favors the pacemaker lineage, whereas its suppression promotes the chamber cardiomyocyte lineage.
TL;DR: High‐content, reproducible evidence suggests that the CD146+ (POS) MSC subpopulation are the mediators of the beneficial effects achieved using crude BM‐MSCs, leading to translational implications for improving cell therapy and manufacturing.
Abstract: CD146+ bone marrow-derived mesenchymal stem/stromal cells (BM-MSCs) play key roles in the perivascular niche, skeletogenesis, and hematopoietic support; however, comprehensive evaluation of therapeutic potency has yet to be determined. In this study, in vitro inflammatory priming to crude human BM-MSCs (n = 8) captured a baseline of signature responses, including enriched CD146+ with coexpression of CD107aHigh , CXCR4High , and LepRHigh , transcriptional profile, enhanced secretory capacity, and robust immunomodulatory secretome and function, including immunopotency assays (IPAs) with stimulated immune cells. These signatures were significantly more pronounced in CD146+ (POS)-sorted subpopulation than in the CD146- (NEG). Mechanistically, POS BM-MSCs showed a markedly higher secretory capacity with significantly greater immunomodulatory and anti-inflammatory protein production upon inflammatory priming compared with the NEG BM-MSCs. Moreover, IPAs with stimulated peripheral blood mononuclear cells and T lymphocytes demonstrated robust immunosuppression mediated by POS BM-MSC while inducing significant frequencies of regulatory T cells. in vivo evidence showed that POS BM-MSC treatment promoted pronounced M1-to-M2 macrophage polarization, ameliorating inflammation/fibrosis of knee synovium and fat pad, unlike treatment with NEG BM-MSCs. These data correlate the expression of CD146 with innately higher immunomodulatory and secretory capacity, and thus therapeutic potency. This high-content, reproducible evidence suggests that the CD146+ (POS) MSC subpopulation are the mediators of the beneficial effects achieved using crude BM-MSCs, leading to translational implications for improving cell therapy and manufacturing.
TL;DR: A novel mechanism of lung epithelial progenitor cell activation in homeostasis and emphysema is revealed, which largely increased the number of alveolar organoids.
Abstract: Wnt/β-catenin signaling regulates progenitor cell fate decisions during lung development and in various adult tissues. Ectopic activation of Wnt/β-catenin signaling promotes tissue repair in emphysema, a devastating lung disease with progressive loss of parenchymal lung tissue. The identity of Wnt/β-catenin responsive progenitor cells and the potential impact of Wnt/β-catenin signaling on adult distal lung epithelial progenitor cell function in emphysema are poorly understood. Here, we used a TCF/Lef:H2B/GFP reporter mice to investigate the role of Wnt/β-catenin signaling in lung organoid formation. We identified an organoid-forming adult distal lung epithelial progenitor cell population characterized by a low Wnt/β-catenin activity, which was enriched in club and alveolar epithelial type (AT)II cells. Endogenous Wnt/β-catenin activity was required for the initiation of multiple subtypes of distal lung organoids derived from the Wntlow epithelial progenitors. Further ectopic Wnt/β-catenin activation specifically led to an increase in alveolar organoid number; however, the subsequent proliferation of alveolar epithelial cells in the organoids did not require constitutive Wnt/β-catenin signaling. Distal lung epithelial progenitor cells derived from the mouse model of elastase-induced emphysema exhibited reduced organoid forming capacity. This was rescued by Wnt/β-catenin signal activation, which largely increased the number of alveolar organoids. Together, our study reveals a novel mechanism of lung epithelial progenitor cell activation in homeostasis and emphysema.
TL;DR: As a whole, ASCs exhibited an immune profile consistent with a stronger inhibition of immune response and a lower immunogenicity, supporting the use of adipose tissue as a valuable source for clinical applications.
Abstract: Clinical-grade mesenchymal stromal cells (MSCs) can be expanded from bone marrow and adipose tissue to treat inflammatory diseases and degenerative disorders. However, the influence of their tissue of origin on their functional properties, including their immunosuppressive activity, remains unsolved. In this study, we produced paired bone marrow-derived mesenchymal stromal cell (BM-MSC) and adipose-derived stromal cell (ASC) batches from 14 healthy donors. We then compared them using transcriptomic, phenotypic, and functional analyses and validated our results on purified native MSCs to infer which differences were really endowed by tissue of origin. Cultured MSCs segregated together owing to their tissue of origin based on their gene expression profile analyzed using differential expression and weighted gene coexpression network analysis. This translated into distinct immune-related gene signatures, phenotypes, and functional cell interactions. Importantly, sorted native BM-MSCs and ASCs essentially displayed the same distinctive patterns than their in vitro-expanded counterparts. As a whole, ASCs exhibited an immune profile consistent with a stronger inhibition of immune response and a lower immunogenicity, supporting the use of adipose tissue as a valuable source for clinical applications.
TL;DR: The results indicate Wnt5a as a niche factor in the adult hippocampus that promotes neuronal differentiation and development through activation of noncanonical Wnt signaling pathways.
Abstract: In the adult hippocampus, new neurons are generated in the dentate gyrus. The Wnt signaling pathway regulates this process, but little is known about the endogenous Wnt ligands involved. We investigated the role of Wnt5a on adult hippocampal neurogenesis. Wnt5a regulates neuronal morphogenesis during embryonic development, and maintains dendritic architecture of pyramidal neurons in the adult hippocampus. Here, we determined that Wnt5a knockdown in the mouse dentate gyrus by lentivirus-mediated shRNA impaired neuronal differentiation of progenitor cells, and reduced dendritic development of adult-born neurons. In cultured adult hippocampal progenitors (AHPs), Wnt5a knockdown reduced neuronal differentiation and morphological development of AHP-derived neurons, whereas treatment with Wnt5a had the opposite effect. Interestingly, no changes in astrocytic differentiation were observed in vivo or in vitro, suggesting that Wnt5a does not affect fate-commitment. By using specific inhibitors, we determined that Wnt5a signals through CaMKII to induce neurogenesis, and promotes dendritic development of newborn neurons through activating Wnt/JNK and Wnt/CaMKII signaling. Our results indicate Wnt5a as a niche factor in the adult hippocampus that promotes neuronal differentiation and development through activation of noncanonical Wnt signaling pathways.
TL;DR: H19 was the most highly upregulated lncRNA in neural stem cells (NSCs) of the subventricular zone (SVZ) of rats subjected to focal cerebral ischemia and regulated neurogenesis‐related miRNAs, providing novel insights into the epigenetic regulation of lncRNAs in stroke‐induced Neurogenesis.
Abstract: Neurogenesis contributes to poststroke recovery. Long noncoding RNAs (lncRNAs) participate in the regulation of stem cell self-renewal and differentiation. However, the role of lncRNAs in stroke-induced neurogenesis remains unknown. In this study, we found that H19 was the most highly upregulated lncRNA in neural stem cells (NSCs) of the subventricular zone (SVZ) of rats subjected to focal cerebral ischemia. Deletion of H19 suppressed cell proliferation, promoted cell death, and blocked NSC differentiation. RNA sequencing analysis revealed that genes deregulated by H19 knockdown were those that are involved in transcription, apoptosis, proliferation, cell cycle, and response to hypoxia. H19 knockdown significantly increased the transcription of cell cycle-related genes including p27, whereas overexpression of H19 substantially reduced expression of these genes through the interaction with chromatin remodeling proteins EZH2 and SUZ12. Moreover, H19 regulated neurogenesis-related miRNAs. Inactivation of H19 in NSCs of ischemic rats attenuated spontaneous functional recovery after stroke. Collectively, our data provide novel insights into the epigenetic regulation of lncRNAs in stroke-induced neurogenesis.
TL;DR: It is suggested that SDF‐1‐CXCR4 signaling in the stromal microenvironment cells plays a crucial role in maintenance of HSPCs during homeostasis, and promotes niche regeneration and early hematopoietic reconstitution after transplantation.
Abstract: The bone marrow microenvironment/niche plays a key role in regulating hematopoietic stem and progenitor cell (HSPC) activities, however mechanisms regulating niche cell function are not well understood. In this study, we show that niche intrinsic expression of the CXCR4 chemokine receptor critically regulates HSPC maintenance during study-state, and promotes early hematopoietic regeneration after myeloablative irradiation. At steady-state, chimeric mice with wild-type (WT) HSPC and marrow stroma that lack CXCR4 show decreased HSPC quiescence, and their repopulation capacity was markedly reduced. Mesenchymal stromal cells (MSC) were significantly reduced in the bone marrow (BM) of CXCR4 deficient mice, which was accompanied by decreased levels of the HSPC supporting factors stromal cell-derived factor-1 (SDF-1) and stem cell factor (SCF). CXCR4 also plays a crucial role in survival and restoration of BM stromal cells after myeloablative irradiation, where loss of BM stromal cells was more severe in CXCR4 deficient mice compared to WT mice. In addition, transplantation of WT donor HSPC into CXCR4 deficient recipient mice demonstrated reduced HSPC homing and early hematopoietic reconstitution. We found that CXCR4 signaling attenuates irradiation-induced BM stromal cell loss by up-regulating the expression of the anti-apoptotic protein Survivin via the PI3K pathway. Our study suggests that SDF-1-CXCR4 signaling in the stromal microenvironment cells plays a crucial role in maintenance of HSPCs during homeostasis, and promotes niche regeneration and early hematopoietic reconstitution after transplantation. Modulation of CXCR4 signaling in the HSPC microenvironment could be a means to enhance hematopoietic recovery after clinical hematopoietic cell transplantation.
TL;DR: It is confirmed that EVs may represent an alternative to whole MSCs for aGvHD prevention, once the effective dose is reproducibly identified according to EV‐IFU and EV‐TD definition.
Abstract: Graft-vs-host-disease (GvHD) is currently the main complication of allogeneic hematopoietic stem cell transplantation. Mortality and morbidity rates are particularly high, especially in steroid-refractory acute GvHD (aGvHD). Immune regulatory human bone marrow mesenchymal stromal cells (hMB-MSCs) represent a therapeutic approach to address this issue. Unfortunately, their effect is hardly predictable in vivo due to several variables, that is, MSC tissue origin, concentration, dose number, administration route and timing, and inflammatory status of the recipient. Interestingly, human bone marrow MSC-derived extracellular vesicles (hBM-MSC-EVs) display many of the hBM-MSC immunoregulatory properties due to their content in paracrine factors that greatly varies according to the collection method. In this study, we focused on the immunological characterization of hBM-MSC-EVs on their capability of inducing regulatory T-cells (T-regs) both in vitro and in a xenograft mouse model of aGvHD. We correlated these data with the aGvHD incidence and degree following hBM-MSC-EV intravenous administration. Thus, we first quantified the EV immunomodulation in vitro in terms of EV immunomodulatory functional unit (EV-IFU), that is, the lowest concentration of EVs leading in vitro to at least threefold increase of the T-regs compared with controls. Second, we established the EV therapeutic dose in vivo (EV-TD) corresponding to 10-fold the in vitro EV-IFU. According to this approach, we observed a significant improvement of both mouse survival and control of aGvHD onset and progression. This study confirms that EVs may represent an alternative to whole MSCs for aGvHD prevention, once the effective dose is reproducibly identified according to EV-IFU and EV-TD definition.
TL;DR: The results indicate that addition of RA + T3 during days 90 to 120 of differentiation enhanced the generation of rod and S‐cone photoreceptor formation, while the combined addition of DAPT from days 28 to 42 with RA during days 30 to120 of differentiation led to enhanced generation of L/M‐cones at the expense of rods.
Abstract: Cell replacement therapy is a promising treatment for irreversible retinal cell death in diverse diseases such as Stargardt's disease, age-related macular degeneration, and retinitis pigmentosa. The final impact of all retinal dystrophies is the loss of photoreceptors; hence, there is a pressing need for research into replacement. Seminal work has shown that a simple three-dimensional culture system enables differentiation of human pluripotent stem cells to retinal organoids containing large numbers of photoreceptors developing alongside retinal neurons and Muller glia cells in a laminated structure that resembles the native retina. Despite these promising developments, current protocols show different efficiencies across pluripotent stem cells and result in retinal organoids with a mixture of photoreceptor cells at varying maturation states, along with nonphotoreceptor cell types. In this study, we investigated the impact of stage-specific addition of retinoic acid (RA), 9-cis-retinal, 11-cis-retinal, levodopa (l-DOPA), triiodothyronine (T3), and γ-secretase inhibitor ((2S)-N-[(3,5-Difluorophenyl)acetyl]-l-alanyl-2-phenyl]glycine1,1-dimethylethyl ester2L [DAPT]) in the generation of cone and rod photoreceptors. Our results indicate that addition of RA + T3 during days 90 to 120 of differentiation enhanced the generation of rod and S-cone photoreceptor formation, while the combined addition of DAPT from days 28 to 42 with RA during days 30 to 120 of differentiation led to enhanced generation of L/M-cones at the expense of rods. l-DOPA when added together with RA during days 90 to 120 of differentiation also promoted the emergence of S-cones at the expense of rod photoreceptors. Collectively, these data represent an advance in our ability to direct generation of rod and cone photoreceptors in vitro.
TL;DR: How the EVs produced by stem cells are being aggressively pursued for clinical applications, including their potential use as therapeutic delivery systems and for their regenerative capabilities are highlighted.
Abstract: Stem cells use a variety of mechanisms to help maintain their pluripotency and promote self-renewal, as well as, at the appropriate time, to differentiate into specialized cells. One such mechanism that is attracting significant attention from the stem cell, development, and regenerative medicine research communities involves a form of intercellular communication, specifically, the ability of cells to form and release nontraditional membrane-enclosed structures, referred to as extracellular vesicles (EVs). There are two major classes of EVs, microvesicles (MVs), which are generated through the outward budding and fission of the plasma membrane, and exosomes, which are formed as multivesicular bodies (MVBs) in the endo-lysosomal pathway that fuse with the cell surface to release their contents. Although they differ in how they are formed, both MVs and exosomes have been shown to contain a diverse array of bioactive cargo, such as proteins, RNA transcripts, microRNAs, and even DNA, which can be transferred to other cells and promote phenotypic changes. Here, we will describe what is currently known regarding EVs and the roles they play in stem cell biology and different aspects of early development. We will also highlight how the EVs produced by stem cells are being aggressively pursued for clinical applications, including their potential use as therapeutic delivery systems and for their regenerative capabilities.
TL;DR: It is shown that intestinal fibroblast‐derived EVs are involved in forming the ISC niche by transmitting Wnt and epidermal growth factor (EGF) activity and it is proved that loss in ISC number in the absence of EGF is prevented by fibro Blast EVs.
Abstract: Extracellular vesicles (EV) are membrane-surrounded vesicles that represent a novel way of intercellular communication by carrying biologically important molecules in a concentrated and protected form. The intestinal epithelium is continuously renewed by a small proliferating intestinal stem cell (ISC) population, residing at the bottom of the intestinal crypts in a specific microenvironment, the stem cell niche. By using 3D mouse and human intestinal organoids, we show that intestinal fibroblast-derived EVs are involved in forming the ISC niche by transmitting Wnt and epidermal growth factor (EGF) activity. With a mouse model that expresses EGFP in the Lgr5+ ISCs, we prove that loss in ISC number in the absence of EGF is prevented by fibroblast-derived EVs. Furthermore, we demonstrate that intestinal fibroblast-derived EVs carry EGF family members, such as amphiregulin. Mechanistically, blocking EV-bound amphiregulin inhibited the EV-induced survival of organoids. In contrast, EVs have no role in transporting R-Spondin, a critical niche factor amplifying Wnt signaling. Collectively, we prove the important role of fibroblast-derived EVs as a novel transmission mechanism of factors in the normal ISC niche.
TL;DR: In this paper, Mesenchymal stem cells (MSCs) have been reported to prevent renal injuries via immune regulation in diabetic models, but whether these benefits are owing to the regulation of Mφ, and the underlying signaling pathways are unknown.
Abstract: Diabetic nephropathy (DN) is a leading cause of end-stage renal disease. Chronic inflammation is recognized as a key causal factor in the development and progression of DN, and the imbalance of M1/M2 macrophages (Mφ) contributes to this process. Mesenchymal stem cells (MSCs) have been reported to prevent renal injuries via immune regulation in diabetic models, but whether these benefits are owing to the regulation of Mφ, and the underlying signaling pathways are unknown. Here, we showed that MSCs elicited Mφ into M2 phenotype and prevented renal injuries in DN mice, but these effects were abolished when the Mφ were depleted by clodronate liposomes (Lipo-Clod), suggesting that Mφ were necessary for renal protection of MSCs in DN mice. Moreover, the MSCs promoted M2 polarization was attributable to the activation of transcription factor EB (TFEB) and subsequent restore of lysosomal function and autophagy activity in Mφ. Furthermore, in vivo adoptive transfer of Mφin vivo (Mφ from DN + MSCs mice) or MφMSCs (Mφ cocultured with MSCs in vitro) to DN mice improved renal function. While, TFEB knockdown in Mφ significantly abolished the protective role of MφMSCs . Altogether, these findings revealed that MSCs suppress inflammatory response and alleviate renal injuries in DN mice via TFEB-dependent Mφ switch.
TL;DR: The data suggest that Treg enrichment by MSCs results from Tcon conversion into Treg‐like cells, rather than to expansion of natural Treg, possibly through mechanisms involving TGF‐β and/or PD‐1/PDL‐1 expression.
Abstract: Regulatory T cells (Treg) play a critical role in immune tolerance. The scarcity of Treg therapy clinical trials in humans has been largely due to the difficulty in obtaining sufficient Treg numbers. We performed a preclinical investigation on the potential of mesenchymal stromal cells (MSCs) to expand Treg in vitro to support future clinical trials. Human peripheral blood mononuclear cells from healthy donors were cocultured with allogeneic bone marrow-derived MSCs expanded under xenogeneic-free conditions. Our data show an increase in the counts and frequency of CD4+ CD25high Foxp3+ CD127low Treg cells (4- and 6-fold, respectively) after a 14-day coculture. However, natural Treg do not proliferate in coculture with MSCs. When purified conventional CD4 T cells (Tcon) are cocultured with MSCs, only cells that acquire a Treg-like phenotype proliferate. These MSC-induced Treg-like cells also resemble Treg functionally, since they suppress autologous Tcon proliferation. Importantly, the DNA methylation profile of MSC-induced Treg-like cells more closely resembles that of natural Treg than of Tcon, indicating that this population is stable. The expression of PD-1 is higher in Treg-like cells than in Tcon, whereas the frequency of PDL-1 increases in MSCs after coculture. TGF-β levels are also significantly increased MSC cocultures. Overall, our data suggest that Treg enrichment by MSCs results from Tcon conversion into Treg-like cells, rather than to expansion of natural Treg, possibly through mechanisms involving TGF-β and/or PD-1/PDL-1 expression. This MSC-induced Treg population closely resembles natural Treg in terms of phenotype, suppressive ability, and methylation profile.
TL;DR: The therapeutic benefit of exosomes secreted by stem and progenitor cells in preclinical models of ischemic heart disease is discussed.
Abstract: The adult human heart has limited regenerative capacity; hence, stem cell therapy has been investigated as a potential approach for cardiac repair. However, a large part of the benefit of the injection of stem and progenitor cells into injured hearts is mediated by secreted factors. Exosomes-nano-sized secreted extracellular vesicles of endosomal origin-have emerged as key signaling organelles in intercellular communication, and are now viewed as the key regenerative constituent of the secretome of stem and progenitor cells. Exosomes released from mesenchymal stem cells, cardiac-derived progenitor cells, embryonic stem cells, induced pluripotent stem cells (iPSCs), and iPSC-derived cardiomyocytes exhibit cardioprotective, immunomodulatory, and reparative abilities. This concise review discusses the therapeutic benefit of exosomes secreted by stem and progenitor cells in preclinical models of ischemic heart disease.
TL;DR: It is shown that ectopic expression of miR‐137 in hiNSCs reduces proliferation and accelerates neuronal differentiation and migration and that reduced levels of MEF2A reduce the transcription of PGC1α, which in turn impacts mitochondrial dynamics.
Abstract: The role of miRNAs in determining human neural stem cell (NSC) fate remains elusive despite their high expression in the developing nervous system. In this study, we investigate the role of miR-137, a brain-enriched miRNA, in determining the fate of human induced pluripotent stem cells-derived NSCs (hiNSCs). We show that ectopic expression of miR-137 in hiNSCs reduces proliferation and accelerates neuronal differentiation and migration. TargetScan and MicroT-CDS predict myocyte enhancer factor-2A (MEF2A), a transcription factor that regulates peroxisome proliferator-activated receptor-gamma coactivator (PGC1α) transcription, as a target of miR-137. Using a reporter assay, we validate MEF2A as a downstream target of miR-137. Our results indicate that reduced levels of MEF2A reduce the transcription of PGC1α, which in turn impacts mitochondrial dynamics. Notably, miR-137 accelerates mitochondrial biogenesis in a PGC1α independent manner by upregulating nuclear factor erythroid 2 (NFE2)-related factor 2 (NRF2) and transcription factor A of mitochondria (TFAM). In addition, miR-137 modulates mitochondrial dynamics by inducing mitochondrial fusion and fission events, resulting in increased mitochondrial content and activation of oxidative phosphorylation (OXPHOS) and oxygen consumption rate. Pluripotency transcription factors OCT4 and SOX2 are known to have binding sites in the promoter region of miR-137 gene. Ectopic expression of miR-137 elevates the expression levels of OCT4 and SOX2 in hiNSCs which establishes a feed-forward self-regulatory loop between miR-137 and OCT4/SOX2. Our study provides novel molecular insights into NSC fate determination by miR-137.
TL;DR: The observations of increased MSC‐mediated mitochondria transfer to hypoxia‐exposed mouse islets are consistent with this and suggest that MSCs are most effective in supporting the secretory function of compromised β‐cells, thus enabling the more widespread application of clinical islet transplantation.
Abstract: Pretransplant islet culture is associated with the loss of islet cell mass and insulin secretory function. Insulin secretion from islet β-cells is primarily controlled by mitochondrial ATP generation in response to elevations in extracellular glucose. Coculture of islets with mesenchymal stromal cells (MSCs) improves islet insulin secretory function in vitro, which correlates with superior islet graft function in vivo. This study aimed to determine whether the improved islet function is associated with mitochondrial transfer from MSCs to cocultured islets. We have demonstrated mitochondrial transfer from human adipose MSCs to human islet β-cells in coculture. Fluorescence imaging showed that mitochondrial transfer occurs, at least partially, through tunneling nanotube (TNT)-like structures. The extent of mitochondrial transfer to clinically relevant human islets was greater than that to experimental mouse islets. Human islets are subjected to more extreme cellular stressors than mouse islets, which may induce "danger signals" for MSCs, initiating the donation of MSC-derived mitochondria to human islet β-cells. Our observations of increased MSC-mediated mitochondria transfer to hypoxia-exposed mouse islets are consistent with this and suggest that MSCs are most effective in supporting the secretory function of compromised β-cells. Ensuring optimal MSC-derived mitochondria transfer in preculture and/or cotransplantation strategies could be used to maximize the therapeutic efficacy of MSCs, thus enabling the more widespread application of clinical islet transplantation.
TL;DR: Wnt7b can enhance both self‐renewal and osteogenic differentiation of BMSCs via inducing Sox11, which presents a new crosstalk between Wnt and Sox signaling in B MSCs.
Abstract: As a profoundly anabolic regulator of bone, Wnt7b is well acknowledged to enhance osteoblast activities. Here, we report that bone marrow mesenchymal stem cells (BMSCs) are another important population responding to Wnt7b. In this study, we systematically investigated the in vivo role of Wnt7b in BMSCs using transgenic mice, high-throughput RNA-seq, immunohistochemistry, RT-qPCR, and in situ hybridization. These methods led us to uncover that Sox11 is induced via Wnt7b in BMSCs. Colony formation assay, flow cytometry, EdU incorporation labeling, RT-qPCR, and Western blot were conducted to detect the self-renewal capacity of BMSCs. Alkaline phosphatase staining, alizarin red staining, and ex vivo BMSCs transplantation were utilized to detect the osteogenic ability of BMSCs. ChIP-qPCR, shRNAs, and immunofluorescence staining were utilized to investigate the underlying mechanisms. Consequently, bone-derived Wnt7b was found to decrease in osteoporosis and elevate in bone fracture healing. During bone fracture healing, Wnt7b was particularly expressed in the mesenchymal cells residing within healing frontiers. RNA-seq data of Wnt7b-overexpressed bones uncovered the significant upregulation of Sox11. Histological results further unveiled that Sox11 is specifically increased in BMSCs. Wnt7b-induced Sox11 was demonstrated to reinforce both self-renewal and osteogenic differentiation of BMSCs. Mechanistically, Wnt7b activates the Ca2+ -dependent Nfatc1 signaling to directly induce Sox11 transcription, which in turn activates the transcriptions of both proliferation-related transcription factors (Ccnb1 and Sox2) and osteogenesis-related factors (Runx2, Sp7) in BMSCs. It is intriguing that this Wnt7b-Sox11 signaling in BMSCs is β-Catenin-independent. Overall, this study provides brand new insights of Wnt7b in bone formation, namely, Wnt7b can enhance both self-renewal and osteogenic differentiation of BMSCs via inducing Sox11. These findings present a new crosstalk between Wnt and Sox signaling in BMSCs.
TL;DR: How hiPSCs could contribute to improving the diagnosis, prognosis, and treatment of an individual with a suspected genetic cardiac disease is discussed, thereby developing better risk stratification and clinical management strategies for these potentially lethal but treatable disorders.
Abstract: Research on mechanisms underlying monogenic cardiac diseases such as primary arrhythmias and cardiomyopathies has until recently been hampered by inherent limitations of heterologous cell systems, where mutant genes are expressed in noncardiac cells, and physiological differences between humans and experimental animals. Human-induced pluripotent stem cells (hiPSCs) have proven to be a game changer by providing new opportunities for studying the disease in the specific cell type affected, namely the cardiomyocyte. hiPSCs are particularly valuable because not only can they be differentiated into unlimited numbers of these cells, but they also genetically match the individual from whom they were derived. The decade following their discovery showed the potential of hiPSCs for advancing our understanding of cardiovascular diseases, with key pathophysiological features of the patient being reflected in their corresponding hiPSC-derived cardiomyocytes (the past). Now, recent advances in genome editing for repairing or introducing genetic mutations efficiently have enabled the disease etiology and pathogenesis of a particular genotype to be investigated (the present). Finally, we are beginning to witness the promise of hiPSC in personalized therapies for individual patients, as well as their application in identifying genetic variants responsible for or modifying the disease phenotype (the future). In this review, we discuss how hiPSCs could contribute to improving the diagnosis, prognosis, and treatment of an individual with a suspected genetic cardiac disease, thereby developing better risk stratification and clinical management strategies for these potentially lethal but treatable disorders.
TL;DR: PDGFRα marks distinct perivascular osteoprogenitor cell subpopulations within adipose tissue, which may contribute to the improved understanding of pathologic mineralization/ossification.
Abstract: The perivascular niche within adipose tissue is known to house multipotent cells, including osteoblast precursors. However, the identity of perivascular subpopulations that may mineralize or ossify most readily is not known. Here, we utilize inducible PDGFRα (platelet-derived growth factor alpha) reporter animals to identify subpopulations of perivascular progenitor cells. Results showed that PDGFRα-expressing cells are present in four histologic niches within inguinal fat, including two perivascular locations. PDGFRα+ cells are most frequent within the tunica adventitia of arteries and veins, where PDGFRα+ cells populate the inner aspects of the adventitial layer. Although both PDGFRα+ and PDGFRα- fractions are multipotent progenitor cells, adipose tissue-derived PDGFRα+ stromal cells proliferate faster and mineralize to a greater degree than their PDGFRα- counterparts. Likewise, PDGFRα+ ectopic implants reconstitute the perivascular niche and ossify to a greater degree than PDGFRα- cell fractions. Adventicytes can be further grouped into three distinct groups based on expression of PDGFRα and/or CD34. When further partitioned, adventicytes co-expressing PDGFRα and CD34 represented a cell fraction with the highest mineralization potential. Long-term tracing studies showed that PDGFRα-expressing adventicytes give rise to adipocytes, but not to other cells within the vessel wall under homeostatic conditions. However, upon bone morphogenetic protein 2 (BMP2)-induced ossicle formation, descendants of PDGFRα+ cells gave rise to osteoblasts, adipocytes, and "pericyte-like" cells within the ossicle. In sum, PDGFRα marks distinct perivascular osteoprogenitor cell subpopulations within adipose tissue. The identification of perivascular osteoprogenitors may contribute to our improved understanding of pathologic mineralization/ossification.