scholarly journals Assessment of the In Vitro Cytotoxicity Effects of the Leaf Methanol Extract of Crinum zeylanicum on Mouse Induced Pluripotent Stem Cells and Their Cardiomyocytes Derivatives

2021 ◽  
Vol 14 (12) ◽  
pp. 1208
Author(s):  
Magloire Kanyou Ndjenda II ◽  
Elvine Pami Nguelefack-Mbuyo ◽  
Jürgen Hescheler ◽  
Télesphore Benoît Nguelefack ◽  
Filomain Nguemo
Keyword(s):  
Stem Cells ◽  
Cell Proliferation ◽  
Pluripotent Stem Cells ◽  
Methanol Extract ◽  
Embryoid Body ◽  
In Vitro Cytotoxicity ◽  
Cardiac Biomarkers ◽  
Dependent Manner ◽  
High Concentrations

Crinum zeylanicum (C. zeylanicum) is commonly used in African folk medicine to treat cardiovascular ailments. In the present study, we investigated the cytotoxic effect of the leaf methanol extract of C. zeylanicum (CZE) using mouse pluripotent stem cells (mPSCs). mPSCs and their cardiomyocytes (CMs) derivatives were exposed to CZE at different concentrations. Cell proliferation, differentiation capacity, and beating activity were assessed using xCELLigence system and microscopy for embryoid body (EB) morphology. Expression of markers associated with major cardiac cell types was examined by immunofluorescence and quantitative RT-PCR. Intracellular reactive oxygen species (ROS) levels were assessed by dichlorodihydrofluorescein diacetate staining. The results showed that the plant extract significantly reduced cell proliferation and viability in a concentration- and time-dependent manner. This was accompanied by a decrease in EB size and an increase in intracellular ROS. High concentrations of CZE decreased the expression of some important cardiac biomarkers. In addition, CZE treatment was associated with poor sarcomere structural organization of CMs and significantly decreased the amplitude and beating rate of CMs, without affecting CMs viability. These results indicate that CZE might be toxic at high concentrations in the embryonic stages of stem cells and could modulate the contracting activity of CMs.

scholarly journals Generation and miRNA Characterization of Equine Induced Pluripotent Stem Cells Derived from Fetal and Adult Multipotent Tissues

2019 ◽  
Vol 2019 ◽  
pp. 1-15 ◽  
Author(s):  
Laís Vicari de Figueiredo Pessôa ◽  
Pedro Ratto Lisboa Pires ◽  
Maite del Collado ◽  
Naira Caroline Godoy Pieri ◽  
Kaiana Recchia ◽  
...  
Keyword(s):  
Stem Cells ◽  
Alkaline Phosphatase ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Embryoid Body ◽  
Embryoid Bodies ◽  
Mirna Profile ◽  
Patient Specific ◽  
Induced Pluripotent

Introduction. Pluripotent stem cells are believed to have greater clinical potential than mesenchymal stem cells due to their ability to differentiate into almost any cell type of an organism, and since 2006, the generation of patient-specific induced pluripotent stem cells (iPSCs) has become possible in multiple species. Objectives. We hypothesize that different cell types respond differently to the reprogramming process; thus, the goals of this study were to isolate and characterize equine adult and fetal cells and induce these cells to pluripotency for future regenerative and translational purposes. Methods. Adult equine fibroblasts (eFibros) and mesenchymal cells derived from the bone marrow (eBMmsc), adipose tissue (eADmsc), and umbilical cord tissue (eUCmsc) were isolated, their multipotency was characterized, and the cells were induced in vitro into pluripotency (eiPSCs). eiPSCs were generated through a lentiviral system using the factors OCT4, SOX2, c-MYC, and KLF4. The morphology and in vitro pluripotency maintenance potential (alkaline phosphatase detection, embryoid body formation, in vitro spontaneous differentiation, and expression of pluripotency markers) of the eiPSCs were characterized. Additionally, a miRNA profile analysis of the mesenchymal and eiPSCs was performed. Results. Multipotent cells were successfully isolated, but the eBMmsc failed to generate eiPSCs. The eADmsc-, eUCmsc-, and eFibros-derived iPSCs were positive for alkaline phosphatase, OCT4 and NANOG, were exclusively dependent on bFGF, and formed embryoid bodies. The miRNA profile revealed a segregated pattern between the eiPSCs and multipotent controls: the levels of miR-302/367 and the miR-92 family were increased in the eiPSCs, while the levels of miR-23, miR-27, and miR-30, as well as the let-7 family were increased in the nonpluripotent cells. Conclusions. We were able to generate bFGF-dependent iPSCs from eADmsc, eUCmsc, and eFibros with human OSKM, and the miRNA profile revealed that clonal lines may respond differently to the reprogramming process.

scholarly journals OncogenicPIK3CApromotes cellular stemness in an allele dose-dependent manner

2019 ◽  
Vol 116 (17) ◽  
pp. 8380-8389 ◽  
Author(s):  
Ralitsa R. Madsen ◽  
Rachel G. Knox ◽  
Wayne Pearce ◽  
Saioa Lopez ◽  
Betania Mahler-Araujo ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Dependent Manner ◽  
Developmental Context ◽  
Epithelial Morphology ◽  
Pi3k Signaling Pathway ◽  
Induced Pluripotent

ThePIK3CAgene, which encodes the p110α catalytic subunit of PI3 kinase (PI3K), is mutationally activated in cancer and in overgrowth disorders known asPIK3CA-related overgrowth spectrum (PROS). To determine the consequences of geneticPIK3CAactivation in a developmental context of relevance to both PROS and cancer, we engineered isogenic human induced pluripotent stem cells (iPSCs) with heterozygous or homozygous knockin ofPIK3CAH1047R. While heterozygous iPSCs remained largely similar to wild-type cells, homozygosity forPIK3CAH1047Rcaused widespread, cancer-like transcriptional remodeling, partial loss of epithelial morphology, up-regulation of stemness markers, and impaired differentiation to all three germ layers in vitro and in vivo. Genetic analysis ofPIK3CA-associated cancers revealed that 64% had multiple oncogenicPIK3CAcopies (39%) or additional PI3K signaling pathway-activating “hits” (25%). This contrasts with the prevailing view thatPIK3CAmutations occur heterozygously in cancer. Our findings suggest that a PI3K activity threshold determines pathological consequences of oncogenicPIK3CAactivation and provide insight into the specific role of this pathway in human pluripotent stem cells.

scholarly journals Analysis of Embryoid Bodies Derived from Human Induced Pluripotent Stem Cells as a Means to Assess Pluripotency

2012 ◽  
Vol 2012 ◽  
pp. 1-9 ◽  
Author(s):  
Steven D. Sheridan ◽  
Vasudha Surampudi ◽  
Raj R. Rao
Keyword(s):  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Embryoid Body ◽  
Embryoid Bodies ◽  
Functional Assay ◽  
Differentiation Potential ◽  
Disease Etiology ◽  
Induced Pluripotent

Human induced pluripotent stem cells (hiPSCs) have core properties of unlimited self-renewal and differentiation potential and have emerged as exciting cell sources for applications in regenerative medicine, drug discovery, understanding of development, and disease etiology. Key among numerous criteria to assess pluripotency includes thein vivoteratoma assay that has been widely proposed as a standard functional assay to demonstrate the pluripotency of hiPSCs. Yet, the lack of reliability across methodologies, lack of definitive clinical significance, and associated expenses bring into question use of the teratoma assay as the “gold standard” for determining pluripotency. We propose use of thein vitroembryoid body (EB) assay as an important alternative to the teratoma assay. This paper summarizes the methodologies for creating EBs from hiPSCs and the subsequent analyses to assess pluripotency and proposes its use as a cost-effective, controlled, and reproducible approach that can easily be adopted to determine pluripotency of generated hiPSCs.

scholarly journals Tissue Engineering of Cartilage Using Collagen Scaffold Enriched with Plant Polysaccharides

Cartilage ◽  
2021 ◽  
pp. 194760352110078
Author(s):  
K. Uday Chandrika ◽  
Sapna Kacha ◽  
Anuja S. Nair ◽  
Vijayishwer S. Jamwal ◽  
Shruti Sandilya ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Gum Arabic ◽  
In Vitro Cytotoxicity ◽  
Health Care Economics ◽  
Self Healing ◽  
Induced Pluripotent

Degenerative diseases associated with articular cartilage pose a huge burden on health care economics. The nature of the tissue involved and the changes therein do not allow self-healing; and most of these problems are progressive. Tissue engineering offers some solutions provided we focus on the right kind of cells and the appropriate surrounding niches created for a particular tissue. The present study deals with the formation of polysaccharide rich stable scaffold of collagen after cross-linking with oxidized gum arabic. The scaffold was tested for its biocompatibility and ability to support cells. The in vitro cytotoxicity of the scaffolds toward induced pluripotent stem cells and chondrocytes was evaluated. Evaluation of expression of lineage specific markers indicates differentiation of induced pluripotent stem cells to chondrogenic lineage and maintenance of chondrocytes per se when grown in the scaffold. Animal studies were carried out to study the efficacy of the scaffold to repair the knee injuries. Cells along with the scaffold appeared to be the best filling, in repair of injured cartilage. These studies show that these scaffolds are potential candidates in applications such as tissue engineering of cartilage.

In vitro Cytotoxicity Analysis of Hybrid Graphene Oxide (hGO) Nano Structures in Caprine Wharton’s Jelly Derived Mesenchymal Stem Cells (WJ-MSCs)

2020 ◽  
Author(s):  
S. A. Dhenge ◽  
N. E. Gade ◽  
O. P. Mishra ◽  
Abinash Kumar ◽  
V. N. Khandait
Keyword(s):  
Stem Cells ◽  
Cell Proliferation ◽  
Mesenchymal Stem Cells ◽  
Graphene Oxide ◽  
Growth Curve ◽  
In Vitro Cytotoxicity ◽  
Cell Models ◽  
Nano Structures ◽  
Dose Dependent

Background: Nanotechnology is used in stem cell culture as well as in vivo delivery and tracking of stem cells. Graphene oxide (GO) is a carbon based nanomaterial and it has large surface area as well as good biocompatibility and heteroatoms doped GO exploit its properties. Hybrid GO (hGO) nano structures biocompatibility is depends on its size, dose and exposure time as well as in vitro cell models and hence, need thorough cytotoxicity studies in different species in vitro cell models. Methods: Caprine Wharton’s jelly derived mesenchymal stem cells (WJ-MSCs) were isolated, characterized and dose dependent (100, 50, 25, 10 and 0µg /ml) in vitro cytotoxicity of three different hGO nano structures (phosphorus doped graphene oxide titanium oxide tubes, rods and sheets) were analysed in caprine WJ-MSCs by studying cell cytotoxicity assays. Result: All three hGO nano structures were damaged cell morphology at 100 and 50 µg /ml doses, however, morphologically more good cells were observed in hGO tubes treated group than hGO rods and hGO sheets at 25 and 10 µg/ml doses as compared to control. Cell viability percentage was significantly (P less than 0.01) decreased at dose 100 µg/ml and it was significantly (P less than 0.01) increased at 25 µg/ml dose as compared to 50, 10 and 0 µg/ml doses. But, hGO tubes significantly (P less than 0.01) increased cell viability % as compared to hGO rods and hGO sheets. Cell population doubling time (PDT) was not altered significantly by all hGO nano structures, but 100 and 50 µg/ml doses significantly (P less than 0.01)increased cell PDT as compared to 25, 10 and 0 µg/ml doses. All hGO nano structures were non significantly altered growth curve, however, all hGO nano structures at 25 µg /ml dose altered (inclined) shape of growth curve, while 100 and 50 µg /ml doses significantly declined growth curve shape as compared to 10 and 0 µg /ml doses. Cell proliferation % was significantly (P less than 0.01) increased at 25 and 10 µg/ml doses, while, it was significantly (P less than 0.01) decreased at 100 µg /ml dose as compared to 50 and 0 µg /ml. However, there was no significance difference was observed in cell proliferation % in groups treated by different hGO nanostructures. In last, it was concluded as, hGO nano structures cytotoxicity was dose dependent and hGO nano tubes were least cytotoxic in caprine WJ-MSCs.

scholarly journals Chemical Activation of the Hypoxia-Inducible Factor Reversibly Reduces Tendon Stem Cell Proliferation, Inhibits Their Differentiation, and Maintains Cell Undifferentiation

2018 ◽  
Vol 2018 ◽  
pp. 1-13 ◽  
Author(s):  
Alessandra Menon ◽  
Pasquale Creo ◽  
Marco Piccoli ◽  
Sonia Bergante ◽  
Erika Conforti ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Proliferation ◽  
Stem Cell ◽  
Cell Response ◽  
Adult Stem Cells ◽  
Chemical Activation ◽  
Hypoxia Inducible Factor ◽  
Adult Stem Cell ◽  
Dependent Manner

Adult stem cell-based therapeutic approaches for tissue regeneration have been proposed for several years. However, adult stem cells are usually limited in number and difficult to be expanded in vitro, and they usually tend to quickly lose their potency with passages, as they differentiate and become senescent. Culturing stem cells under reduced oxygen tensions (below 21%) has been proposed as a tool to increase cell proliferation, but many studies reported opposite effects. In particular, cell response to hypoxia seems to be very stem cell type specific. Nonetheless, it is clear that a major role in this process is played by the hypoxia inducible factor (HIF), the master regulator of cell response to oxygen deprivation, which affects cell metabolism and differentiation. Herein, we report that a chemical activation of HIF in human tendon stem cells reduces their proliferation and inhibits their differentiation in a reversible and dose-dependent manner. These results support the notion that hypoxia, by activating HIF, plays a crucial role in preserving stem cells in an undifferentiated state in the “hypoxic niches” present in the tissue in which they reside before migrating in more oxygenated areas to heal a damaged tissue.

Ce1-хGdхOy Nanoparticles Stimulate Proliferation of Dental Pulp Stem Cells In Vitro

2017 ◽  
Vol 13 ◽  
pp. 26-31
Author(s):  
Anton L. Popov ◽  
Olga G. Tatarnikova ◽  
Nelly R. Popova ◽  
Irina I. Selezneva ◽  
Azamat Y. Akkizov ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cerium Oxide ◽  
Dental Pulp ◽  
Low Cost ◽  
Dependent Manner ◽  
Urgent Task ◽  
High Concentrations ◽  
Further Development ◽  
Dose Dependent

One of the main reasons for limiting the widespread clinical use of mesenchymal stem cells (MSCs) is a low speed of their proliferation in vitro. In this regard, the search for new safe and effective growth stimulants is an urgent task. In this study, we investigated the effect of nanocrystalline cerium oxide doped with gadolinium (Ce1-х Gdх Oy), on the morphofunctional characteristics and proliferative activity of MSCs derived from dental pulp. It was shown that the introduction of Ce1-х Gdх Oy nanoparticles into the culture of dental MSCs provides the activation of proliferation of the cells in a dose-dependent manner. High concentrations of Ce1-х Gdх Oy nanoparticles inhibit the proliferation of the cells; however, this does not lead to further development of apoptosis and cell death. The obtained results indicate that the nanocrystalline cerium oxide can be considered as a basis for the development of highly efficient and low-cost supplements for culturing MSCs.

scholarly journals Oncogenic PIK3CA promotes cellular stemness in an allele dose-dependent manner

2018 ◽  
Author(s):  
Ralitsa R. Madsen ◽  
Rachel G. Knox ◽  
Wayne Pearce ◽  
Saioa Lopez ◽  
Betania Mahler-Araujo ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Dependent Manner ◽  
Developmental Context ◽  
Epithelial Morphology ◽  
Pi3k Signaling Pathway ◽  
Induced Pluripotent

AbstractThe PIK3CA gene, which encodes the p110α catalytic subunit of PI3-kinase (PI3K), is mutationally activated in cancer and in overgrowth disorders known as PIK3CA-related overgrowth spectrum (PROS). To determine the consequences of genetic PIK3CA activation in a developmental context of relevance to both PROS and cancer, we engineered isogenic human induced pluripotent stem cells (iPSCs) with heterozygous or homozygous knock-in of PIK3CAH1047R. While heterozygous iPSCs remained largely similar to wild-type cells, homozygosity for PIK3CAH1047R caused widespread, cancer-like transcriptional remodeling, partial loss of epithelial morphology, upregulation of stemness markers and impaired differentiation to all three germ layers in vitro and in vivo. Genetic analysis of PIK3CA-associated cancers revealed that 64 % had multiple oncogenic PIK3CA copies (39 %) or additional PI3K signaling pathway-activating “hits” (25 %). This contrasts with the prevailing view that PIK3CA mutations occur heterozygously in cancer. Our findings suggest that a PI3K activity threshold determines pathological consequences of oncogenic PIK3CA activation and provide the first insight into the specific role of this pathway in human pluripotent stem cells.

Effects of N-carbamylglutamate and L-arginine on gonadotrophin-releasing hormone (GnRH) gene expression and secretion in GT1-7 cells

2018 ◽  
Vol 30 (5) ◽  
pp. 759 ◽  
Author(s):  
Y. Liu ◽  
J. H. Bai ◽  
X. L. Xu ◽  
Z. L. Chen ◽  
L. J. Spicer ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Proliferation ◽  
Mrna Abundance ◽  
Metastasis Suppressor ◽  
Dependent Manner ◽  
Releasing Hormone ◽  
Gnrh Secretion ◽  
Gonadotrophin Releasing Hormone ◽  
High Concentrations

Recent studies have shown that N-carbamylglutamate (NCG) and arginine (ARG) supplementation improves reproductive performance in livestock. The objectives of the present study were to evaluate the effects of NCG and ARG on GT1-7 cell gonadotrophin-releasing hormone (GnRH) secretion, gene expression and cell proliferation. GT1-7 cells were treated in vitro with different concentrations of NCG (0–1.0 mM) or ARG (0–4.0 mM) in serum-free medium for 12 or 24 h. For GnRH secretion and cell proliferation, GT1-7 cells were more sensitive to NCG than ARG. NCG treatment after 12 h increased cell numbers and inhibited GnRH secretion in a dose-dependent manner (P < 0.05), although there was no significant effect of NCG on these parameters after 24 h culture. ARG treatment decreased GnRH secretion after 24 h (P < 0.05), whereas it had no effect after 12 h. GT1-7 cells express GnRH, Kiss-1 metastasis-suppressor (Kiss1), G-protein coupled receptor 54 (GPR54), neuronal nitric oxide synthase (nNOS) and estrogen receptor α (ERα) genes. High concentrations of NCG (1.0 mM) and ARG (4.0 mM) inhibited (P < 0.05) GnRH and nNOS mRNA abundance in GT1-7 cells. ARG treatment decreased Kiss1 and increased ERα mRNA abundance. Thus, high concentrations of NCG (1.0 mM) and ARG (4.0 mM) may act both directly and indirectly to regulate GnRH neuron function by downregulating genes related to GnRH synthesis and secretion to slow GnRH production while stimulating GT1-7 cell proliferation.

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