Effects of erythropoietin in murine-induced pluripotent cell-derived panneural progenitor cells.
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SUBMITTER: Offen N
PROVIDER: S-EPMC3883967 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Resolving the immunogenicity of cells derived from induced pluripotent stem cells (iPSCs) remains an important challenge for cell transplant strategies that use banked allogeneic cells. Thus, we evaluated the immunogenicity of mouse fetal neural stem/progenitor cells (fetus-NSPCs) and iPSC-derived neural stem/progenitor cells (iPSC-NSPCs) both in vitro and in vivo. Flow cytometry revealed the low expression of immunological surface antigens, and these cells survived in all mice when transplanted syngeneically into subcutaneous tissue and the spinal cord. In contrast, an allogeneic transplantation into subcutaneous tissue was rejected in all mice, and allogeneic cells transplanted into intact and injured spinal cords survived for 3 months in approximately 20% of mice. In addition, cell survival was increased after co-treatment with an immunosuppressive agent. Thus, the immunogenicity and post-transplantation immunological dynamics of iPSC-NSPCs resemble those of fetus-NSPCs.
| S-EPMC5636829 | biostudies-other
Project description:BackgroundCardiovascular progenitor cells (CPCs) have been identified within the developing mouse heart and differentiating pluripotent stem cells by intracellular transcription factors Nkx2.5 and Islet 1 (Isl1). Study of endogenous and induced pluripotent stem cell (iPSC)-derived CPCs has been limited due to the lack of specific cell surface markers to isolate them and conditions for their in vitro expansion that maintain their multipotency.Methodology/principal findingsWe sought to identify specific cell surface markers that label endogenous embryonic CPCs and validated these markers in iPSC-derived Isl1(+)/Nkx2.5(+) CPCs. We developed conditions that allow propagation and characterization of endogenous and iPSC-derived Isl1(+)/Nkx2.5(+) CPCs and protocols for their clonal expansion in vitro and transplantation in vivo. Transcriptome analysis of CPCs from differentiating mouse embryonic stem cells identified a panel of surface markers. Comparison of these markers as well as previously described surface markers revealed the combination of Flt1(+)/Flt4(+) best identified and facilitated enrichment for Isl1(+)/Nkx2.5(+) CPCs from embryonic hearts and differentiating iPSCs. Endogenous mouse and iPSC-derived Flt1(+)/Flt4(+) CPCs differentiated into all three cardiovascular lineages in vitro. Flt1(+)/Flt4(+) CPCs transplanted into left ventricles demonstrated robust engraftment and differentiation into mature cardiomyocytes (CMs).Conclusion/significanceThe cell surface marker combination of Flt1 and Flt4 specifically identify and enrich for an endogenous and iPSC-derived Isl1(+)/Nkx2.5(+) CPC with trilineage cardiovascular potential in vitro and robust ability for engraftment and differentiation into morphologically and electrophysiologically mature adult CMs in vivo post transplantation into adult hearts.
| S-EPMC3467279 | biostudies-literature
Project description:Glucocorticoids (GCs) are frequently used for treating and preventing chronic lung disease and circulatory dysfunction in premature infants. However, there is growing concern about the detrimental effects of systemic GC administration on neurodevelopment. The first choice of GCs to minimize the adverse effects on the developing brain is still under debate. We investigated the effect of commonly used GCs such as dexamethasone (DEX), betamethasone (BET) and hydrocortisone (HDC) on the proliferation of human-induced pluripotent stem cell (iPSC)-derived neuronal progenitor cells (NPCs). In this study, NPCs were treated with various concentrations of GCs and subjected to cell proliferation assays. Furthermore, we quantified the number of microtubule-associated protein 2 (MAP2) positive neurons in NPCs by immunostaining. All GCs promoted NPC proliferation in a dose-dependent manner. We also confirmed that MAP2-positive neurons in NPCs increased upon GC treatment. However, differential effects of GCs on MAP2 positive neurons were observed when we treated NPCs with H2O2. The total numbers of NPCs increased upon any GC treatment even under oxidative conditions but the numbers of MAP2 positive neurons increased only by HDC treatment. GCs promoted human iPSCsâ€"derived NPC proliferation and the differential effects of GCs became apparent under oxidative stress. Our results may support HDC as the preferred choice over DEX and BET to prevent adverse effects on the developing human brain.
| S-EPMC4174547 | biostudies-literature
Project description:BackgroundSeveral pieces of evidence from in vitro studies showed that brain-derived neurotrophic factor (BDNF) promotes proliferation and differentiation of neural stem/progenitor cells (NSCs) into neurons. Moreover, the JAK2 pathway was proposed to be associated with mouse NSC proliferation. BDNF could activate the STAT-3 pathway and induce proliferation in mouse NSCs. However, its effects on proliferation are not fully understood and JAK/STAT pathway was proposed to play a role in this activity.MethodsIn the present study, the effects of BDNF on cell proliferation and neurite outgrowth of Alzheimer's disease (AD) induced pluripotent stem cells (iPSCs)-derived human neural progenitor cells (hNPCs) were examined. Moreover, a specific signal transduction pathway important in cell proliferation was investigated using a JAK2 inhibitor (AG490) to clarify the role of that pathway.ResultsThe proliferative effect of BDNF was remarkably observed as an increase in Ki-67 positive cells. The cell number of hNPCs was significantly increased after BDNF treatment represented by cellular metabolic activity of the cells measured by MTT assay. This noticeable effect was statistically shown at 20 ng/ml of BDNF treatment. BDNF, however, did not promote neurite outgrowth but increased neuronal cell number. It was found that AG490 suppressed hNPCs proliferation. However, this inhibitor partially decreased BDNF-induced hNPCs proliferation. These results demonstrated the potential role of BDNF for the amelioration of AD through the increase of AD-derived hNPCs number.
| S-EPMC8123227 | biostudies-literature
Project description:BackgroundCurrently, there is no regenerative therapy for patients with neurological and neurodegenerative disorders. Cell-therapies have emerged as a potential treatment for numerous brain diseases. Despite recent advances in stem cell technology, major concerns have been raised regarding the feasibility and safety of cell therapies for clinical applications.MethodsWe generated good manufacturing practice (GMP)-compatible neural progenitor cells (NPCs) from transgene- and xeno-free induced pluripotent stem cells (iPSCs) that can be smoothly adapted for clinical applications. NPCs were characterized in vitro for their differentiation potential and in vivo after transplantation into wild type as well as genetically immunosuppressed mice.ResultsGenerated NPCs had a stable gene-expression over at least 15 passages and could be scaled for up to 1018 cells per initially seeded 106 cells. After withdrawal of growth factors in vitro, cells adapted a neural fate and mainly differentiated into active neurons. To ensure a pure NPC population for in vivo applications, we reduced the risk of iPSC contamination by applying micro RNA-switch technology as a safety checkpoint. Using lentiviral transduction with a fluorescent and bioluminescent dual-reporter construct, combined with non-invasive in vivo bioluminescent imaging, we longitudinally tracked the grafted cells in healthy wild-type and genetically immunosuppressed mice as well as in a mouse model of ischemic stroke. Long term in-depth characterization revealed that transplanted NPCs have the capability to survive and spontaneously differentiate into functional and mature neurons throughout a time course of a month, while no residual pluripotent cells were detectable.ConclusionWe describe the generation of transgene- and xeno-free NPCs. This simple differentiation protocol combined with the ability of in vivo cell tracking presents a valuable tool to develop safe and effective cell therapies for various brain injuries.
| S-EPMC9482172 | biostudies-literature
Project description:PURPOSE: Glial cell line-derived neurotrophic factor (GDNF) is neuroprotective of retinal neurons, and transduced retinal progenitor cells (RPCs) can deliver this cytokine for the treatment of retinal diseases, yet the potential effects of GDNF on RPCs have received little attention. METHODS: Murine RPCs were assessed under multiple conditions in the presence or absence of epidermal growth factor (EGF, 20 ng/ml) and/or GDNF (10 ng/ml) using a variety of techniques, including live-cell imaging, caspase-3 activity assay, whole genome microarray, quantitative polymerase chain reaction (qPCR), and western blotting. RESULTS: Live monitoring revealed that formation of initial aggregates resulted largely from the collision and adherence of dissociated RPCs, as opposed to clonal proliferation. Spheres enlarged in size and number, with more reaching the threshold criteria for cross-sectional areas in the EGF+GDNF condition. Proliferation was measurably augmented in association with EGF+GDNF, and K(i)-67 expression was modestly increased (1.07 fold), as were hairy and enhancer of split 5 (Hes5), mammalian achaete-scute homolog 1 (Mash1), and Vimentin. However, global gene expression did not reveal a notable treatment-related response, and the expression of the majority of progenitor and lineage markers examined remained stable. GDNF reduced RPC apoptosis, compared to complete growth-factor withdrawal, although it could not by itself sustain mitotic activity. CONCLUSIONS: These data support the feasibility of developing GDNF-transduced RPCs as potential therapeutic agents for use in retinal diseases.
| S-EPMC3012652 | biostudies-literature
Project description:Mucopolysaccharidosis type IIIA (MPS IIIA) is characterised by a progressive neurological decline leading to early death. It is caused by bi-allelic loss-of-function mutations in SGSH encoding sulphamidase, a lysosomal enzyme required for heparan sulphate glycosaminoglycan (HS GAG) degradation, that results in the progressive build-up of HS GAGs in multiple tissues most notably the central nervous system (CNS). Skin fibroblasts from two MPS IIIA patients who presented with an intermediate and a severe clinical phenotype, respectively, were reprogrammed into induced pluripotent stem cells (iPSCs). The intermediate MPS IIIA iPSCs were then differentiated into neural progenitor cells (NPCs) and subsequently neurons. The patient derived fibroblasts, iPSCs, NPCs and neurons all displayed hallmark biochemical characteristics of MPS IIIA including reduced sulphamidase activity and increased accumulation of an MPS IIIA HS GAG biomarker. Proliferation of MPS IIIA iPSC-derived NPCs was reduced compared to control, but could be partially rescued by reintroducing functional sulphamidase enzyme, or by doubling the concentration of the mitogen fibroblast growth factor 2 (FGF2). Whilst both control heparin, and MPS IIIA HS GAGs had a similar binding affinity for FGF2, only the latter inhibited FGF signalling, suggesting accumulated MPS IIIA HS GAGs disrupt the FGF2:FGF2 receptor:HS signalling complex. Neuronal differentiation of MPS IIIA iPSC-derived NPCs was associated with a reduction in the expression of neuronal cell marker genes βIII-TUBULIN, NF-H and NSE, revealing reduced neurogenesis compared to control. A similar result was achieved by adding MPS IIIA HS GAGs to the culture medium during neuronal differentiation of control iPSC-derived NPCs. This study demonstrates the generation of MPS IIIA iPSCs, and NPCs, the latter of which display reduced proliferation and neurogenic capacity. Reduced NPC proliferation can be explained by a model in which soluble MPS IIIA HS GAGs compete with cell surface HS for FGF2 binding. The mechanism driving reduced neurogenesis remains to be determined but appears downstream of MPS IIIA HS GAG accumulation. Highlights • Induced pluripotent stem cells (iPSCs) were generated from MPS IIIA skin fibroblasts.• MPS IIIA iPSCs were successfully differentiated into neural progenitor cells (NPCs).• MPS IIIA NPC proliferation was reduced due to interference with FGF2 signalling.• Neurogenesis from MPS IIIA iPSC-derived NPCs is reduced via an unknown mechanism.
| S-EPMC8531667 | biostudies-literature
Project description:Induced pluripotent stem cell (iPSC)-derived dopaminergic (DA) neurons are an expected source for cell-based therapies for Parkinson's disease (PD). The regulatory criteria for the clinical application of these therapies, however, have not been established. Here we show the results of our pre-clinical study, in which we evaluate the safety and efficacy of dopaminergic progenitors (DAPs) derived from a clinical-grade human iPSC line. We confirm the characteristics of DAPs by in vitro analyses. We also verify that the DAP population include no residual undifferentiated iPSCs or early neural stem cells and have no genetic aberration in cancer-related genes. Furthermore, in vivo studies using immunodeficient mice reveal no tumorigenicity or toxicity of the cells. When the DAPs are transplanted into the striatum of 6-OHDA-lesioned rats, the animals show behavioral improvement. Based on these results, we started a clinical trial to treat PD patients in 2018.
| S-EPMC7338530 | biostudies-literature
Project description:BACKGROUND:Renal ischemia-reperfusion (I/R) injury is a major cause of acute kidney injury (AKI), which is associated with high morbidity and mortality. AKI is a serious and costly medical condition. Effective therapy for AKI is an unmet clinical need, and molecular mechanisms underlying the interactions between an injured kidney and distant organs remain unclear. Therefore, novel therapeutic strategies should be developed. METHODS:We directed the differentiation of human induced pluripotent stem (iPS) cells into endothelial progenitor cells (iEPCs), which were then applied for treating mouse AKI. The mouse model of AKI was induced by I/R injury. RESULTS:We discovered that intravenously infused iEPCs were recruited to the injured kidney, expressed the mature endothelial cell marker CD31, and replaced injured endothelial cells. Moreover, infused iEPCs produced abundant proangiogenic proteins, which entered into circulation. In AKI mice, blood urea nitrogen and plasma creatinine levels increased 2?days after I/R injury and reduced after the infusion of iEPCs. Tubular injury, cell apoptosis, and peritubular capillary rarefaction in injured kidneys were attenuated accordingly. In the AKI mice, iEPC therapy also ameliorated apoptosis of cardiomyocytes and cardiac dysfunction, as indicated by echocardiography. The therapy also ameliorated an increase in serum brain natriuretic peptide. Regarding the relevant mechanisms, indoxyl sulfate and interleukin-1? synergistically induced apoptosis of cardiomyocytes. Systemic iEPC therapy downregulated the proapoptotic protein caspase-3 and upregulated the anti-apoptotic protein Bcl-2 in the hearts of the AKI mice, possibly through the reduction of indoxyl sulfate and interleukin-1?. CONCLUSIONS:Therapy using human iPS cell-derived iEPCs provided a protective effect against ischemic AKI and remote cardiac dysfunction through the repair of endothelial cells and the attenuation of cardiomyocyte apoptosis.
| S-EPMC6288873 | biostudies-literature
Project description:Human embryonic stem (ES) cells and induced pluripotent stem (iPS) cells represent a potential alternative source for red blood cell transfusion. However, when using traditional methods with embryoid bodies, ES cell-derived erythroid cells predominantly express embryonic type ?-globin, with lesser fetal type ?-globin and very little adult type ?-globin. Furthermore, no ?-globin expression is detected in iPS cell-derived erythroid cells. ES cell-derived sacs (ES sacs) have been recently used to generate functional platelets. Due to its unique structure, we hypothesized that ES sacs serve as hemangioblast-like progenitors capable to generate definitive erythroid cells that express ?-globin. With our ES sac-derived erythroid differentiation protocol, we obtained ?120 erythroid cells per single ES cell. Both primitive (?-globin expressing) and definitive (?- and ?-globin expressing) erythroid cells were generated from not only ES cells but also iPS cells. Primitive erythropoiesis is gradually switched to definitive erythropoiesis during prolonged ES sac maturation, concurrent with the emergence of hematopoietic progenitor cells. Primitive and definitive erythroid progenitor cells were selected on the basis of glycophorin A or CD34 expression from cells within the ES sacs before erythroid differentiation. This selection and differentiation strategy represents an important step toward the development of in vitro erythroid cell production systems from pluripotent stem cells. Further optimization to improve expansion should be required for clinical application. Stem Cells 2016;34:1541-1552.
| S-EPMC4892957 | biostudies-literature