Tubastatin A maintains adult skeletal muscle stem cells in a quiescent state ex vivo and improves their engraftment ability in vivo.
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ABSTRACT:
SUBMITTER: Arjona M
PROVIDER: S-EPMC8758944 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Transplantation of myogenic stem cells possesses great potential for long-term repair of dystrophic muscle. However, a single donor muscle biopsy is unlikely to provide enough cells to effectively transplant the muscle mass of a patient affected by muscular dystrophy. Expansion of cells ex vivo using traditional culture techniques significantly reduces engraftment potential. We hypothesized that activation of Notch signaling during ex vivo expansion would maintain donor cell engraftment potential. In this study, we expanded freshly isolated canine muscle-derived cells on tissue culture plates coated with Delta-1(ext) -IgG to activate Notch signaling or with human IgG as a control. A model of canine-to-murine xenotransplantation was used to quantitatively compare canine muscle cell engraftment and determine whether engrafted donor cells could function as satellite cells in vivo. We show that Delta-1(ext) -IgG inhibited differentiation of canine muscle-derived cells and increased the level of genes normally expressed in myogenic precursors. Moreover, cells expanded on Delta-1(ext) -IgG resulted in a significant increase in the number of donor-derived fibers, as compared to cells expanded on human IgG, reaching engraftment levels similar to freshly isolated cells. Importantly, cells expanded on Delta-1(ext) -IgG engrafted to the recipient satellite cell niche and contributed to further regeneration. A similar strategy of expanding human muscle-derived cells on Notch ligand might facilitate engraftment and muscle regeneration for patients affected with muscular dystrophy.
| S-EPMC3448880 | biostudies-literature
Project description:We show that Tubastatin A (TubA) preserves MuSC quiescence and stem cell potency ex vivo, by inhibiting HDAC6 and, consequently, primary cilium resorption. Treatment with TubA improves MuSC engraftment potential and induces a return to quiescence in cycling MuSCs, revealing a potentially valuable approach to enhancing the therapeutic potential of MuSCs. To examine the state of quiescence preserved by TubA at the transcriptome level, we performed RNA-Seq and we found that TubA-treated MuSCs exhibit a quiescent transcriptome. The molecular mechanisms involved in the maintenance of quiescence by TubA were ribosome- and oxidative phosphorylation-related genes as well as low expression levels of cell cycle genes and Hh signaling genes.
2022-01-28 | GSE178070 | GEO
Project description:BackgroundPoor engraftment due to low cell doses restricts the usefulness of umbilical-cord-blood transplantation. We hypothesized that engraftment would be improved by transplanting cord blood that was expanded ex vivo with mesenchymal stromal cells.MethodsWe studied engraftment results in 31 adults with hematologic cancers who received transplants of 2 cord-blood units, 1 of which contained cord blood that was expanded ex vivo in cocultures with allogeneic mesenchymal stromal cells. The results in these patients were compared with those in 80 historical controls who received 2 units of unmanipulated cord blood.ResultsCoculture with mesenchymal stromal cells led to an expansion of total nucleated cells by a median factor of 12.2 and of CD34+ cells by a median factor of 30.1. With transplantation of 1 unit each of expanded and unmanipulated cord blood, patients received a median of 8.34×10(7) total nucleated cells per kilogram of body weight and 1.81×10(6) CD34+ cells per kilogram--doses higher than in our previous transplantations of 2 units of unmanipulated cord blood. In patients in whom engraftment occurred, the median time to neutrophil engraftment was 15 days in the recipients of expanded cord blood, as compared with 24 days in controls who received unmanipulated cord blood only (P<0.001); the median time to platelet engraftment was 42 days and 49 days, respectively (P=0.03). On day 26, the cumulative incidence of neutrophil engraftment was 88% with expansion versus 53% without expansion (P<0.001); on day 60, the cumulative incidence of platelet engraftment was 71% and 31%, respectively (P<0.001).ConclusionsTransplantation of cord-blood cells expanded with mesenchymal stromal cells appeared to be safe and effective. Expanded cord blood in combination with unmanipulated cord blood significantly improved engraftment, as compared with unmanipulated cord blood only. (Funded by the National Cancer Institute and others; ClinicalTrials.gov number, NCT00498316.).
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Project description:Quiescence is important for sustaining neural stem cells (NSCs) in the adult brain over the lifespan. Lysosomes are digestive organelles that degrade membrane receptors after they undergo endolysosomal membrane trafficking. Enlarged lysosomes are present in quiescent NSCs (qNSCs) in the subventricular zone of the mouse brain, but it remains largely unknown how lysosomal function is involved in the quiescence. Here we show that qNSCs exhibit higher lysosomal activity and degrade activated EGF receptor by endolysosomal degradation more rapidly than proliferating NSCs. Chemical inhibition of lysosomal degradation in qNSCs prevents degradation of signaling receptors resulting in exit from quiescence. Furthermore, conditional knockout of TFEB, a lysosomal master regulator, delays NSCs quiescence in vitro and increases NSC proliferation in the dentate gyrus of mice. Taken together, our results demonstrate that enhanced lysosomal degradation is an important regulator of qNSC maintenance.
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Project description:Graft-versus-host disease (GvHD) is still the major non-relapse, life-limiting complication after hematopoietic stem cell transplantation. Modern pharmacologic immunosuppression is often insufficient and associated with significant side effects. Novel treatment strategies now include adoptive transfer of ex vivo expanded regulatory T cells (Tregs), but their efficacy in chronic GvHD is unknown. We treated three children suffering from severe, therapy-refractory GvHD with polyclonally expanded Tregs generated from the original stem cell donor. Third-line maintenance immunosuppression was tapered to cyclosporin A and low-dose steroids shortly before cell transfer. Regular follow-up included an assessment of the subjective and objective clinical development, safety parameters, and in-depth immune monitoring. All patients showed marked clinical improvement with substantially decreased GvHD activity. Laboratory follow-up showed a significant enhancement of the immunologic engraftment, including lymphocytes and dendritic cells. Monitoring the fate of Tregs by next-generation sequencing demonstrated clonal expansion. In summary, adoptive transfer of Tregs was well tolerated and able to modulate an established undesired T cell mediated allo-response. Although no signs of overimmunosuppression were detectable, the treatment of patients with invasive opportunistic infections should be undertaken with caution. Further controlled studies are necessary to confirm these encouraging effects and eventually pave the way for adoptive Treg therapy in chronic GvHD.
| S-EPMC9171381 | biostudies-literature
Project description:Within discrete regions of the developing mammalian central nervous system, small subsets of glia become specialized to function as neural stem cells. As a result of their self-renewal and neurogenic capacity, these cells later serve to replenish neurons and glia during persistent or injury-induced adult neurogenesis. SOX2, an HMG box transcription factor, plays an essential role in the maintenance of both embryonic and adult neural progenitors. It is unclear, however, which biological mechanisms regulated by SOX2 are required for neural stem cell maintenance. In this study, we address this question through genetic analysis of SOX2 function in differentiating postnatal Müller glia, a cell type that maintains neurogenic capacity in the adult retina. By utilizing molecular analysis and real-time imaging, we show that two progenitor characteristics of nascent Müller glia - their radial morphology and cell cycle quiescence - are disrupted following conditional genetic ablation of Sox2 in the mouse postnatal retina, leading to Müller cell depletion and retinal degeneration. Moreover, we demonstrate that genetic induction of the Notch signaling pathway restores Müller glial cell identity to Sox2 mutant cells, but does not secure their quiescent state. Collectively, these results uncouple the roles of SOX2 and the Notch signaling pathway in the postnatal retina, and uncover a novel role for SOX2 in preventing the depletion of postnatal Müller glia through terminal cell division.
| S-EPMC3596988 | biostudies-literature
Project description:RASA1 (also known as p120 RasGAP) is a Ras GTPase-activating protein that functions as a regulator of blood vessel growth in adult mice and humans. In humans, RASA1 mutations cause capillary malformation-arteriovenous malformation (CM-AVM); whether it also functions as a regulator of the lymphatic vasculature is unknown. We investigated this issue using mice in which Rasa1 could be inducibly deleted by administration of tamoxifen. Systemic loss of RASA1 resulted in a lymphatic vessel disorder characterized by extensive lymphatic vessel hyperplasia and leakage and early lethality caused by chylothorax (lymphatic fluid accumulation in the pleural cavity). Lymphatic vessel hyperplasia was a consequence of increased proliferation of lymphatic endothelial cells (LECs) and was also observed in mice in which induced deletion of Rasa1 was restricted to LECs. RASA1-deficient LECs showed evidence of constitutive activation of Ras in situ. Furthermore, in isolated RASA1-deficient LECs, activation of the Ras signaling pathway was prolonged and cellular proliferation was enhanced after ligand binding to different growth factor receptors, including VEGFR-3. Blockade of VEGFR-3 was sufficient to inhibit the development of lymphatic vessel hyperplasia after loss of RASA1 in vivo. These findings reveal a role for RASA1 as a physiological negative regulator of LEC growth that maintains the lymphatic vasculature in a quiescent functional state through its ability to inhibit Ras signal transduction initiated through LEC-expressed growth factor receptors such as VEGFR-3.
| S-EPMC3266774 | biostudies-literature
Project description:Skeletal myogenesis is potently regulated by the extracellular milieu of growth factors and cytokines. We observed that cardiotrophin-1 (CT-1), a member of the interleukin-6 (IL-6) family of cytokines, is a potent regulator of skeletal muscle differentiation. The normal up-regulation of myogenic marker genes, myosin heavy chain (MyHC), myogenic regulatory factors (MRFs), and myocyte enhancer factor 2s (MEF2s) were inhibited by CT-1 treatment. CT-1 also represses myogenin (MyoG) promoter activation. CT-1 activated two signaling pathways: signal transducer and activator of transcription 3 (STAT3), and mitogen-activated protein kinase kinase (MEK), a component of the extracellular signal-regulated MAPK (ERK) pathway. In view of the known connection between CT-1 and STAT3 activation, we surprisingly found that pharmacological blockade of STAT3 activity had no effect on the inhibition of myogenesis by CT-1 suggesting that STAT3 signaling is dispensable for myogenic repression. Conversely, MEK inhibition potently reversed the inhibition of myotube formation and attenuated the repression of MRF transcriptional activity mediated by CT-1. Taken together, these data indicate that CT-1 represses skeletal myogenesis through interference with MRF activity by activation of MEK/ERK signaling. In agreement with these in vitro observations, exogenous systemic expression of CT-1 mediated by adenoviral vector delivery increased the number of myonuclei in normal post-natal mouse skeletal muscle and also delayed skeletal muscle regeneration induced by cardiotoxin injection. The expression pattern of CT-1 in embryonic and post-natal skeletal muscle and in vivo effects of CT-1 on myogenesis implicate CT-1 in the maintenance of the undifferentiated state in muscle progenitor cells.
| S-EPMC2740593 | biostudies-literature
Project description:ContextType 2 diabetes (T2D) has features of disordered lipid and glucose metabolism, due in part to reduced mitochondrial content.ObjectiveOur objective was to investigate effects of different types of exercise on mitochondrial content and substrate oxidation in individuals with T2D (ancillary study of the randomized controlled trial Health Benefits of Aerobic and Resistance Training in Individuals with Type 2 Diabetes, HART-D).InterventionT2D individuals were randomized to aerobic training (AT, n = 12), resistance training (RT, n = 18), combination training (ATRT, n = 12), or nonexercise control (n = 10). Blood draws, peak oxygen consumption tests, dual-energy x-ray absorptiometry scans and muscle biopsies of vastus lateralis were performed before and after 9 months. Ex vivo substrate oxidations ((14)CO2), mitochondrial content, and enzyme activities were measured. Glycated hemoglobin A1c and free fatty acids were also determined.ResultsMitochondrial content increased after RT and ATRT. Octanoate oxidation increased after AT and ATRT, whereas palmitate, pyruvate, and acetate oxidations increased in all exercise groups. Exercise-induced responses in mitochondrial DNA were associated with improvements in peak oxygen consumption, β-hydroxyacyl-coenzyme A dehydrogenase activity, and palmitate oxidation.ConclusionsNine months of AT and RT significantly improved most aspects of skeletal muscle mitochondrial content and substrate oxidation, whereas the combination improved all aspects. These exercise responses were associated with clinical improvements, indicating that long-term training, especially combination, is an effective lifestyle therapy for individuals with T2D by way of improving muscle substrate metabolism.
| S-EPMC3615199 | biostudies-literature
Project description:Due to their unique longevity and capacity to secrete high levels of protein, plasma B cells have the potential to be used as a cell therapy for protein replacement. Here, we show that ex vivo engineered human plasma cells exhibit single-cell RNA profiles, scanning electron micrograph ultrastructural features, and in vivo homing capacity of long-lived plasma cells. After transferring human plasma cells to immunodeficient mice in the presence of the human cytokines BAFF and IL-6, we observe increases in retention of plasma cells in the bone marrow, with engraftment exceeding a year. The most profound in vivo effects of human IL-6 are observed within 20 days of transfer and could be explained by decreased apoptosis in newly differentiated plasma cells. Collectively, these results show that ex vivo engineered and differentiated human plasma cells have the potential for long-lived in vivo protein secretion, which can be modeled in small animals.
| S-EPMC9573882 | biostudies-literature