scholarly journals Improvement ofIn VitroOsteogenic Potential through Differentiation of Induced Pluripotent Stem Cells from Human Exfoliated Dental Tissue towards Mesenchymal-Like Stem Cells

2015 ◽  
Vol 2015 ◽  
pp. 1-9 ◽  
Author(s):  
Felipe Augusto Andre Ishiy ◽  
Roberto Dalto Fanganiello ◽  
Karina Griesi-Oliveira ◽  
Angela May Suzuki ◽  
Gerson Shigeru Kobayashi ◽  
...  
Keyword(s):  
Stem Cells ◽  
Dermal Fibroblasts ◽  
Deciduous Teeth ◽  
Osteogenic Potential ◽  
Dental Tissue ◽  
Self Renewal ◽  
Clinical Scenarios ◽  
Cellular Source ◽  
Induced Pluripotent

Constraints for the application of MSCs for bone reconstruction include restricted self-renewal and limited cell amounts. iPSC technology presents advantages over MSCs, providing homogeneous cellular populations with prolonged self-renewal and higher plasticity. However, it is unknown if the osteogenic potential of iPSCs differs from that of MSCs and if it depends on the iPSCs originating cellular source. Here, we compared thein vitroosteogenesis between stem cells from human deciduous teeth (SHED) and MSC-like cells from iPSCs from SHED (iPS-SHED) and from human dermal fibroblasts (iPS-FIB). MSC-like cells from iPS-SHED and iPS-FIB displayed fibroblast-like morphology, downregulation of pluripotency markers and upregulation of mesenchymal markers. Comparativein vitroosteogenesis analysis showed higher osteogenic potential in MSC-like cells from iPS-SHED followed by MSC-like cells from iPS-FIB and SHED. CD105 expression, reported to be inversely correlated with osteogenic potential in MSCs, did not display this pattern, considering that SHED presented lower CD105 expression. Higher osteogenic potential of MSC-like cells from iPS-SHED may be due to cellular homogeneity and/or to donor tissue epigenetic memory. Our findings strengthen the rationale for the use of iPSCs in bone bioengineering. Unveiling the molecular basis behind these differences is important for a thorough use of iPSCs in clinical scenarios.

scholarly journals Generation of iPSCs from Jaw Periosteal Cells Using Self-Replicating RNA

2019 ◽  
Vol 20 (7) ◽  
pp. 1648 ◽  
Author(s):  
Felix Umrath ◽  
Heidrun Steinle ◽  
Marbod Weber ◽  
Hans-Peter Wendel ◽  
Siegmar Reinert ◽  
...  
Keyword(s):  
Stem Cells ◽  
Fluorescent Protein ◽  
Osteogenic Potential ◽  
Safety Standards ◽  
Stem Cell Source ◽  
Periosteal Cells ◽  
Induced Pluripotent ◽  
Green Fluorescent ◽  
First Time

Jaw periosteal cells (JPCs) represent a suitable stem cell source for bone tissue engineering (BTE) applications. However, challenges associated with limited cell numbers, stressful cell sorting, or the occurrence of cell senescence during in vitro passaging and the associated insufficient osteogenic potential in vitro of JPCs and other mesenchymal stem/stromal cells (MSCs) are main hurdles and still need to be solved. In this study, for the first time, induced pluripotent stem cells (iPSCs) were generated from human JPCs to open up a new source of stem cells for BTE. For this purpose, a non-integrating self-replicating RNA (srRNA) encoding reprogramming factors and green fluorescent protein (GFP) as a reporter was used to obtain JPC-iPSCs with a feeder- and xeno-free reprogramming protocol to meet the highest safety standards for future clinical applications. Furthermore, to analyze the potential of these iPSCs as a source of osteogenic progenitor cells, JPC-iPSCs were differentiated into iPSC-derived mesenchymal stem/stromal like cells (iMSCs) and further differentiated to the osteogenic lineage under xeno-free conditions. The produced iMSCs displayed MSC marker expression and morphology as well as strong mineralization during osteogenic differentiation.

scholarly journals Exogenous LIN28 Is Required for the Maintenance of Self-Renewal and Pluripotency in Presumptive Porcine-Induced Pluripotent Stem Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Warunya Chakritbudsabong ◽  
Somjit Chaiwattanarungruengpaisan ◽  
Ladawan Sariya ◽  
Sirikron Pamonsupornvichit ◽  
Joao N. Ferreira ◽  
...  
Keyword(s):  
Stem Cells ◽  
Alkaline Phosphatase ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Embryoid Bodies ◽  
Self Renewal ◽  
Fetal Fibroblasts ◽  
Induced Pluripotent

Porcine species have been used in preclinical transplantation models for assessing the efficiency and safety of transplants before their application in human trials. Porcine-induced pluripotent stem cells (piPSCs) are traditionally established using four transcription factors (4TF): OCT4, SOX2, KLF4, and C-MYC. However, the inefficiencies in the reprogramming of piPSCs and the maintenance of their self-renewal and pluripotency remain challenges to be resolved. LIN28 was demonstrated to play a vital role in the induction of pluripotency in humans. To investigate whether this factor is similarly required by piPSCs, the effects of adding LIN28 to the 4TF induction method (5F approach) on the efficiency of piPSC reprogramming and maintenance of self-renewal and pluripotency were examined. Using a retroviral vector, porcine fetal fibroblasts were transfected with human OCT4, SOX2, KLF4, and C-MYC with or without LIN28. The colony morphology and chromosomal stability of these piPSC lines were examined and their pluripotency properties were characterized by investigating both their expression of pluripotency-associated genes and proteins and in vitro and in vivo differentiation capabilities. Alkaline phosphatase assay revealed the reprogramming efficiencies to be 0.33 and 0.17% for the 4TF and 5TF approaches, respectively, but the maintenance of self-renewal and pluripotency until passage 40 was 6.67 and 100%, respectively. Most of the 4TF-piPSC colonies were flat in shape, showed weak positivity for alkaline phosphatase, and expressed a significantly high level of SSEA-4 protein, except for one cell line (VSMUi001-A) whose properties were similar to those of the 5TF-piPSCs; that is, tightly packed and dome-like in shape, markedly positive for alkaline phosphatase, and expressing endogenous pluripotency genes (pOCT4, pSOX2, pNANOG, and pLIN28), significantly high levels of pluripotent proteins (OCT4, SOX2, NANOG, LIN28, and SSEA-1), and a significantly low level of SSEA-4 protein. VSMUi001-A and all 5F-piPSC lines formed embryoid bodies, underwent spontaneous cardiogenic differentiation with cardiac beating, expressed cardiomyocyte markers, and developed teratomas. In conclusion, in addition to the 4TF, LIN28 is required for the effective induction of piPSCs and the maintenance of their long-term self-renewal and pluripotency toward the development of all germ layers. These piPSCs have the potential applicability for veterinary science.

scholarly journals Comparative Analysis of Proliferation and Differentiation Potentials of Stem Cells from Inflamed Pulp of Deciduous Teeth and Stem Cells from Exfoliated Deciduous Teeth

2014 ◽  
Vol 2014 ◽  
pp. 1-12 ◽  
Author(s):  
Shi Yu ◽  
Shu Diao ◽  
Jinsong Wang ◽  
Gang Ding ◽  
Dongmei Yang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Tissue Regeneration ◽  
Deciduous Teeth ◽  
Inflammatory Factors ◽  
Dental Tissue ◽  
Significant Difference ◽  
Proliferation And Differentiation ◽  
Cell Sources ◽  
Irreversible Pulpitis

Stem cells isolated from exfoliated deciduous teeth (SHEDs) are highly capable of proliferation and differentiation, and they represent good cell sources for mesenchymal stem cell- (MSC-) mediated dental tissue regeneration, but the supply of SHEDs is limited. A previous study found that stem cells could be isolated from inflamed tissues, but it is unknown whether primary dental pulp diagnosed with irreversible pulpitis might contain stem cells with appropriate tissue regeneration capacity. In this study, we aimed to isolate stem cells from both inflamed pulps of deciduous teeth (SCIDs) and SHEDs from Chinese children and to compare their proliferation and differentiation potentials. Our results showed that SCIDs were positive for cell surface markers, including CD105, CD90, and CD146, and they had high proliferation ability and osteogenic, adipogenic, and chondrogenic differentiation potentials. There was no significant difference in proliferation and differentiation potentials between SCIDs and SHEDs. The mRNA of inflammatory factors, including IL-1β, IL-6, and TNF-α, was expressed at similar levels in SCIDs and SHEDs, but SCIDs secreted more TNF-αprotein. In conclusion, ourin vitroresults showed that SCIDs have proliferation and differentiation potentials similar to those of SHEDs. Thus, SCIDs represent a new potentially applicable source for MSC mediated tissue regeneration.

Cancellation of c-MYC Silencing in Human Induced Pluripotent Stem Cells Contributes to the Efficient in Vitro Production of Platelets with the Ability of Hemostasis In Vivo.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1488-1488 ◽  
Author(s):  
Naoya Takayama ◽  
Sou Nakamura ◽  
Satoshi Nishimura ◽  
Ryoko Ohnishi ◽  
Kazutoshi Takahashi ◽  
...  
Keyword(s):  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Blood Platelets ◽  
Dermal Fibroblasts ◽  
Efficient Production ◽  
Nog Mice ◽  
Induced Pluripotent

Abstract Abstract 1488 Poster Board I-511 Human induced pluripotent stem cells (hiPSCs) generated from somatic cells by introduction of OCT3/4, SOX2, KLF4 and c-MYC represent a potential source of hematopoietic cells for transfusion without the risk of immune rejection. We recently established an in vitro culture system with which hiPSCs could be differentiated into the unique structure of an “in vitro hematopoietic niche” containing hematopoietic progenitors. Upon further cultivation under appropriate conditions, these hematopoietic progenitor cells differentiated into megakaryocytes, which could then generate platelets with morphologies indistinguishable from peripheral blood platelets regardless of either 4-factor iPSCs (n=8, 4-factor hiPSC clones generated from adult dermal fibroblasts through induction with c-MYC) or 3-factor iPSC clones (without c-MYC, n=3). It is well known that iPSC differentiation yields a heterogeneous population of clones. To select the best hiPSC clone for platelet production, we quantified thrombopoiesis with 11 independent hiPSC clones by comparison with human embryonic stem cells (hESCs) evaluated previously (Takayama et al., Blood, 2008) as a reference. Particularly noteworthy is our finding that 4-factor iPSCs have an advantage over 3-factor iPSCs or hESCs (P<0.01) that is mediated through cancellation of c-MYC silencing (re-activation) over the course of differentiation evidenced by RT-PCR studies. Indeed, ectopic expression of c-MYC, but not OCT3/4, SOX2 or KLF4, using a retroviral vector in hESC-derived progenitors accelerated both megakaryopoiesis and thrombopoiesis. By contrast, the platelet activation statuses (i.e., PAC-1 ligation with activated integrin αIIbβ3 following agonist stimulation) were comparable for platelets obtained from 4-factor hiPSCs and hESCs, though levels of c-MYC clearly differed, indicating that at least integrin activation is independent of c-MYC. To further estimate the in vivo functionality of iPSC-derived platelets, we developed a mouse model for transfusion. Irradiation (2.0 Gy, 9 days beforehand) induced thrombocytopenia in NOG (nod-scid/IL-2 γc-null) mice. Subsequent flow cytometry showed that 2 hrs after transfusion (1.0∼1.2×107 platelets per a mouse) of NOG mice via the tail vein, the circulating levels of selected 4-factor iPSC-derived platelets were similar to those of human adult platelets (platelet chimerism of human CD41/mouse CD41; 4∼10%). Moreover, by using our recently established in vivo imaging system, which enables observation of single platelet behavior, we observed that 4-factor iPSC-derived platelets circulate in NOG mice and contribute to the development thrombi within their vessels, suggesting the in vivo functionality of iPSC-derived platelets is intact. A number of studies have suggested that c-MYC can have deleterious effects leading to in vivo oncogeneity after transplantation in vivo. By contrast, our data strongly indicate the importance of c-MYC for platelet generation from hiPSCs and hESCs. Given that anucleate platelets are routinely irradiated before transfusion in clinical settings, use of c-MYC for hiPSCs generation may contribute to the efficient production of HLA-matched platelet concentrates for those requiring repeated transfusion. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Exosomes Secreted from Induced Pluripotent Stem Cell-Derived Mesenchymal Stem Cells Accelerate Skin Cell Proliferation

2018 ◽  
Vol 19 (10) ◽  
pp. 3119 ◽  
Author(s):  
Soo Kim ◽  
Seul Lee ◽  
Hyunjung Kim ◽  
Tae Kim
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Dermal Fibroblasts ◽  
Skin Cells ◽  
Significant Difference ◽  
Induced Pluripotent

Induced pluripotent stem cell (iPSC)-derived mesenchymal stem cells (iMSCs) serve as a unique source for cell therapy. We investigated whether exosomes from iMSCs promote the proliferation of human keratinocytes (HaCaT) and human dermal fibroblasts (HDFs). iPSCs were established from human Wharton’s jelly MSCs and were allowed to differentiate into iMSCs. Exosomes were collected from the culture supernatant of MSCs (MSC-exo) and iMSCs (iMSC-exo), and their characteristics were investigated. Both exosome types possessed basic characteristics of exosomes and were taken up by skin cells in vitro and in vivo. A significant increase in HaCaT proliferation was observed with iMSC-exo, although both exosomes increased the viability and cell cycle progression in HaCaT and HDFs. No significant difference was observed in the closure of wound scratch and the expression of reparative genes between cells treated with the two exosome types. Both exosomes enhanced the secretion of collagen in HaCaT and HDFs; however, an increase in fibronectin level was observed only in HaCaT, and this effect was better with iMSC-exo treatment. Only iMSC-exo increased the phosphorylation of extracellular signal-regulated kinase (ERK)-1/2. Our results indicate that iMSC-exo promote the proliferation of skin cells by stimulating ERK1/2 and highlight the application of iMSCs for producing exosomes.

Derivation of Hematopoietic Progenitor Cells From Vector-Free Human Induced Pluripotent Stem Cells

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4746-4746
Author(s):  
Friedrich Schuening ◽  
Michail Zaboikin ◽  
Tatiana Zaboikina ◽  
Narasimhachar Srinivasakumar
Keyword(s):  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Conflicts Of Interest ◽  
Dermal Fibroblasts ◽  
Embryoid Bodies ◽  
Chromosomal Anomalies ◽  
Hematopoietic Stem ◽  
Induced Pluripotent

Abstract Abstract 4746 Induced pluripotent stem cells (iPSCs), due to their self-renewal and differentiation capability, have tremendous potential in regenerative medicine. Differentiation of IPSCs in vitro to obtain sufficient number of hematopoietic stem cells (HSCs) and their progenitors (HPCs) from iPSCs for therapeutic purposes is a holy grail of cellular therapy. To this end, we are comparing different in vitro differentiation approaches for generation of HSCs/HPCs from IPSCs. We have generated iPSCs from human adult dermal fibroblasts using two different reprogramming methods: 1) Transduction with retroviral vectors encoding human Klf4, Oct3/4, Sox2 and cMyc or 2) Electroporation with Epstein–Barr virus (EBV) based episomal plasmid vectors encoding Klf4, Oct3/4, Sox2, L-Myc and p53 targeting shRNA. The transduced/electroporated cells were reprogrammed on SNL5 mouse feeder cells. Putative iPSC-like colonies were cloned and adapted to grow under feeder-free conditions on Matrigel (BD) in mTeSR1 (Stem Cell Technologies) medium. From over 30 individual clones isolated, six were further characterized for: 1) expression of pluripotency markers (Tra-1–60, SSEA-3, SSEA-4, Nanog and Oct3/4) by immunofluorescence; 2) endogenous and total mRNA expression by quantitative real-time reverse-transcriptase PCR (RT-qPCR) for Klf4, Oct3/4, Sox2 and cMyc to distinguish between cellular and vector derived expression of reprogramming factors; 3) RT-qPCR to determine expression of other markers of pluripotency such as Nanog and DNA methyl transferease; 4) karyotype analysis to determine chromosomal anomalies. The vector-free IPSC clones were also tested for residual integrated EBV plasmid DNA by qPCR. Trilineage differentiation ability of the clones was determined through embryoid body formation in suspension cultures, and subsequent staining of resulting embryoid bodies after adherence to gelatin coated dishes for makers of ectoderm, mesoderm and endoderm. HSCs/HPCs were obtained from IPSCs by 1) coculture with OP9 stromal cells, or 2) step-wise differentiation in feeder-free conditions on Matrigel under defined conditions in the presence of appropriate growth factors [Niwa A et al. PLoS One. (2011); 6(7):e22261.]. The resultant HSCs/HPCs were subjected to colony forming assays in semi-solid medium containing hematopoietic cytokines. Both erythroid and myelomonocytic colonies could be readily identified. The influence of ambient oxygen concentration on the HSC/HPC derivation procedure is being investigated. The results of these studies will be presented. Disclosures: No relevant conflicts of interest to declare.

In vitro effect of prolactin on the osteogenic potential of bone marrow mesenchymal stem cells of rats

2013 ◽  
Author(s):  
Melo Ocarino Natalia de ◽  
Silvia Silva Santos ◽  
Lorena Rocha ◽  
Juneo Freitas ◽  
Reis Amanda Maria Sena ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Potential ◽  
Marrow Mesenchymal Stem Cells ◽  
Vitro Effect

Stem Cells Derived from Human Exfoliated Deciduous teeth [shed] in Neuronal Disorders: A Review

2020 ◽  
Vol 16 ◽  
Author(s):  
Minu Anoop ◽  
Indrani Datta
Keyword(s):  
Stem Cells ◽  
Neurodegenerative Diseases ◽  
Dental Pulp ◽  
Therapeutic Potential ◽  
Permanent Teeth ◽  
Deciduous Teeth ◽  
Proliferative Potential ◽  
Pulp Tissue ◽  
Human Exfoliated Deciduous Teeth

: Most conventional treatments for neurodegenerative diseases fail due to their focus on neuroprotection rather than neurorestoration. Stem cell‐based therapies are becoming a potential treatment option for neurodegenerative diseases as they can home in, engraft, differentiate and produce factors for CNS recovery. Stem cells derived from human dental pulp tissue differ from other sources of mesenchymal stem cells due to their embryonic neural crest origin and neurotrophic property. These include both dental pulp stem cells [DPSCs] from dental pulp tissues of human permanent teeth and stem cells from human exfoliated deciduous teeth [SHED]. SHED offer many advantages over other types of MSCs such as good proliferative potential, minimal invasive procurement, neuronal differentiation and neurotrophic capacity, and negligible ethical concerns. The therapeutic potential of SHED is attributed to the paracrine action of extracellularly released secreted factors, specifically the secretome, of which exosomes is a key component. SHED and its conditioned media can be effective in neurodegeneration through multiple mechanisms, including cell replacement, paracrine effects, angiogenesis, synaptogenesis, immunomodulation, and apoptosis inhibition, and SHED exosomes offer an ideal refined bed-to-bench formulation in neurodegenerative disorders. However, in spite of these advantages, there are still some limitations of SHED exosome therapy, such as the effectiveness of long-term storage of SHED and their exosomes, the development of a robust GMP-grade manufacturing protocol, optimization of the route of administration, and evaluation of the efficacy and safety in humans. In this review, we have addressed the isolation, collection and properties of SHED along with its therapeutic potential on in vitro and in vivo neuronal disorder models as evident from the published literature.

Mini Review; Differentiation of Human Pluripotent Stem Cells into Oocytes

2020 ◽  
Vol 15 (4) ◽  
pp. 301-307 ◽  
Author(s):  
Gaifang Wang ◽  
Maryam Farzaneh
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Embryonic Stem ◽  
Human Pluripotent Stem Cells ◽  
Human Oocyte ◽  
Differentiation Potential ◽  
Cell Lineages ◽  
Cell Based Therapy ◽  
Induced Pluripotent

Primary Ovarian Insufficiency (POI) is one of the main diseases causing female infertility that occurs in about 1% of women between 30-40 years of age. There are few effective methods for the treatment of women with POI. In the past few years, stem cell-based therapy as one of the most highly investigated new therapies has emerged as a promising strategy for the treatment of POI. Human pluripotent stem cells (hPSCs) can self-renew indefinitely and differentiate into any type of cell. Human Embryonic Stem Cells (hESCs) as a type of pluripotent stem cells are the most powerful candidate for the treatment of POI. Human-induced Pluripotent Stem Cells (hiPSCs) are derived from adult somatic cells by the treatment with exogenous defined factors to create an embryonic-like pluripotent state. Both hiPSCs and hESCs can proliferate and give rise to ectodermal, mesodermal, endodermal, and germ cell lineages. After ovarian stimulation, the number of available oocytes is limited and the yield of total oocytes with high quality is low. Therefore, a robust and reproducible in-vitro culture system that supports the differentiation of human oocytes from PSCs is necessary. Very few studies have focused on the derivation of oocyte-like cells from hiPSCs and the details of hPSCs differentiation into oocytes have not been fully investigated. Therefore, in this review, we focus on the differentiation potential of hPSCs into human oocyte-like cells.

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