Understanding the mesenchymal stem cell and its application to the study of human pluripotent stem cells

AbstractDifferentiation of human pluripotent stem cells into insulin-producing stem cell-derived beta cells harbors great potential for research and therapy of diabetes. The SOX9 gene plays a crucial role during development of the pancreas and particularly in the development of insulin-producing cells as SOX9+ cells form the source for NEUROG3+ endocrine progenitor cells. For the purpose of easy monitoring of differentiation efficiencies into pancreatic progenitors and insulin-producing cells, we generated new reporter lines by knocking in a P2A-H-2Kk-F2A-GFP2 reporter genes into the SOX9 locus and a P2A-mCherry reporter gene into the INS locus mediated by CRISPR/CAS9-technology. The knock-ins enable co-expression of the endogenous genes and reporter genes, report the endogenous gene expression and enable the purification of pancreatic progenitors and insulin-producing cells using FACS or MACS. Using these cell lines we established a new differentiation protocol geared towards SOX9+ cells to efficiently drive human pluripotent stem cells into glucose-responsive beta cells.

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Probing the mechanism for hydrogel-based stasis induction in human pluripotent stem cells: is the chemical functionality of the hydrogel important?

Chemical Science ◽

10.1039/c9sc04734d ◽

2020 ◽

Vol 11 (1) ◽

pp. 232-240 ◽

Cited By ~ 4

Author(s):

M. Sponchioni ◽

C. T. O'Brien ◽

C. Borchers ◽

E. Wang ◽

M. N. Rivolta ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Block Copolymer ◽

Embryonic Stem Cell ◽

Pluripotent Stem Cells ◽

Human Embryonic Stem Cell ◽

Embryonic Stem ◽

Human Pluripotent Stem Cells ◽

Essential Parameter ◽

Chemical Functionality

It is shown that hydroxyl functionality is required to induce stasis in human embryonic stem cell colonies immersed within wholly synthetic block copolymer worm gels with comparable storage moduli. Thus gel softness does not appear to be an essential parameter for stasis induction.

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Towards consistent generation of pancreatic lineage progenitors from human pluripotent stem cells

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2014.0365 ◽

2015 ◽

Vol 370 (1680) ◽

pp. 20140365 ◽

Cited By ~ 19

Author(s):

Maria Rostovskaya ◽

Nicholas Bredenkamp ◽

Austin Smith

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cell Lines ◽

Pluripotent Stem Cells ◽

Pluripotent Stem Cell ◽

Human Pluripotent Stem Cells ◽

Type I ◽

Pancreatic Progenitors ◽

Stem Cell Lines

Human pluripotent stem cells can in principle be used as a source of any differentiated cell type for disease modelling, drug screening, toxicology testing or cell replacement therapy. Type I diabetes is considered a major target for stem cell applications due to the shortage of primary human beta cells. Several protocols have been reported for generating pancreatic progenitors by in vitro differentiation of human pluripotent stem cells. Here we first assessed one of these protocols on a panel of pluripotent stem cell lines for capacity to engender glucose sensitive insulin-producing cells after engraftment in immunocompromised mice. We observed variable outcomes with only one cell line showing a low level of glucose response. We, therefore, undertook a systematic comparison of different methods for inducing definitive endoderm and subsequently pancreatic differentiation. Of several protocols tested, we identified a combined approach that robustly generated pancreatic progenitors in vitro from both embryo-derived and induced pluripotent stem cells. These findings suggest that, although there are intrinsic differences in lineage specification propensity between pluripotent stem cell lines, optimal differentiation procedures may consistently direct a substantial fraction of cells into pancreatic specification.

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Mesenchymal Stem Cell-Conditioned Medium Enhances Osteogenic and Chondrogenic Differentiation of Human Embryonic Stem Cells and Human Induced Pluripotent Stem Cells by Mesodermal Lineage Induction

Tissue Engineering Part A ◽

10.1089/ten.tea.2013.0265 ◽

2014 ◽

Vol 20 (7-8) ◽

pp. 1306-1313 ◽

Cited By ~ 22

Author(s):

Tae-Jin Lee ◽

Jiho Jang ◽

Seokyung Kang ◽

Suk Ho Bhang ◽

Gun-Jae Jeong ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Mesenchymal Stem Cell ◽

Human Embryonic Stem Cells ◽

Pluripotent Stem Cells ◽

Chondrogenic Differentiation ◽

Embryonic Stem ◽

Induced Pluripotent ◽

Cell Conditioned Medium ◽

Mesodermal Lineage

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Transcriptome-based molecular staging of human stem cell-derived retinal organoids uncovers accelerated photoreceptor differentiation by 9-cis retinal

10.1101/733071 ◽

2019 ◽

Cited By ~ 1

Author(s):

Koray D. Kaya ◽

Holly Y. Chen ◽

Matthew J. Brooks ◽

Ryan A. Kelley ◽

Hiroko Shimada ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Retinoic Acid ◽

Pluripotent Stem Cells ◽

Human Pluripotent Stem Cells ◽

Mitochondrial Morphology ◽

Rod Photoreceptor ◽

Molecular Staging ◽

Organoid Cultures

ABSTRACTRetinal organoids generated from human pluripotent stem cells exhibit considerable variability in temporal dynamics of differentiation. To assess the maturity of neural retina in vitro, we performed transcriptome analyses of developing organoids from human embryonic and induced pluripotent stem cell lines. We show that the developmental variability in organoids was reflected in gene expression profiles and could be evaluated by molecular staging with the human fetal and adult retinal transcriptome data. We also demonstrated that addition of 9-cis retinal, instead of widely-used all-trans retinoic acid, accelerated rod photoreceptor differentiation in organoid cultures, with higher rhodopsin expression and more mature mitochondrial morphology evident by day 120. Our studies thus provide an objective transcriptome-based modality for determining the differentiation state of retinal organoids, which should facilitate disease modeling and evaluation of therapies in vitro.Summary StatementThree-dimensional organoids derived from human pluripotent stem cells have been extensively applied for investigating organogenesis, modeling diseases and development of therapies. However, substantial variations within organoids pose challenges for comparison among different cultures and studies. We generated transcriptomes of multiple distinct retinal organoids and compared these to human fetal and adult retina gene profiles for molecular staging of differentiation state of the cultures. Our analysis revealed the advantage of using 9-cis retinal, instead of the widely-used all-trans retinoic acid, in facilitating rod photoreceptor differentiation. Thus, a transcriptome-based comparison can provide an objective method to uncover the maturity of organoid cultures across different lines and in various study platforms.

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Valproic acid Promotes the in Vitro Differentiation of Human Pluripotent Stem Cells Into Spermatogonial Stem Cell-like Cells

10.21203/rs.3.rs-769984/v1 ◽

2021 ◽

Author(s):

Xiaotong Wang ◽

Mengyuan Qu ◽

Zili Li ◽

Yuting Long ◽

Kai Hong ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Wnt Signaling ◽

Signaling Pathway ◽

Pluripotent Stem Cells ◽

Wnt Signaling Pathway ◽

Human Pluripotent Stem Cells ◽

Sequencing Analysis ◽

Upregulated Genes

Abstract Background: Studying human germ cell development and male infertility is heavily relied on mouse models. In vitro differentiation of human pluripotent stem cells into spermatogonial stem cell-like cells (SSCLCs) can be used as a model to study human germ cells and infertility. The current study aimed to develop the SSCLC induction protocol and assess the effects of the developed protocol on the SSCLC induction. Methods: We examined the effects of valproic acid (VPA), vitamin C (VC) and the combination of VPA and VC on the SSCLC induction efficiency and determined the expression of spermatogonial genes of differentiated cells. The percentage of haploid cells and cells expressed meiotic and spermatid genes were also detected. RNA-sequencing analysis was performed to compare the transcriptome between cells at 0 and 12 days of differentiation and differently expressed genes were confirmed by RT-qPCR. We further evaluated the alteration in histone marks (H3K9ac and H3K27me3) at 12 days of differentiation. Moreover, the SSCLC induction efficiency of two hiPSC lines of non-obstructive azoospermia (NOA) patients was assessed using different induction protocols.Results: The combination of low concentrations of VPA and VC in the induction medium was most effective to induce SSCLCs expressing several spermatogonial genes from human pluripotent stem cells at 12 days of differentiation. High concentration of VPA was more effective to induce cells expressing meiotic genes and haploid cells. RNA-sequencing analysis revealed that the induction of SSCLC involved the upregulated genes in Wnt signaling pathway, and cells at 12 days of differentiation showed increased H3K9ac and decreased H3K27me3. Additionally, two hiPSC lines of NOA patients showed low SSCLC induction efficiency and the expression of genes in Wnt signaling pathway. Conclusions: VPA robustly promotes the differentiation of human pluripotent stem cell lines into SSCLCs, which involved the upregulated genes in Wnt signaling pathway and epigenetic changes. hiPSCs from NOA patients showed decreased SSCLC induction efficiency and Wnt signaling pathway gene expression, suggesting that inactivation of Wnt signaling pathway might be a cause of SSC depletion in azoospermia testes. Our developed SSCLC induction protocol provides a reliable tool and model to study human germ cell development and male infertility.

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Expression of miRNAs from the Imprinted DLK1/DIO3 Locus Signals the Osteogenic Potential of Human Pluripotent Stem Cells

Cells ◽

10.3390/cells8121523 ◽

2019 ◽

Vol 8 (12) ◽

pp. 1523 ◽

Cited By ~ 1

Author(s):

Laetitia Barrault ◽

Jacqueline Gide ◽

Tingting Qing ◽

Lea Lesueur ◽

Jorg Tost ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Osteogenic Differentiation ◽

Cell Lines ◽

Pluripotent Stem Cells ◽

Pluripotent Stem Cell ◽

Human Pluripotent Stem Cells ◽

Osteogenic Potential ◽

Human Pluripotent Stem Cell ◽

Stem Cell Lines

Substantial variations in differentiation properties have been reported among human pluripotent cell lines (hPSC), which could affect their utility and clinical safety. We characterized the variable osteogenic capacity observed between different human pluripotent stem cell lines. By focusing on the miRNA expression profile, we demonstrated that the osteogenic differentiation propensity of human pluripotent stem cell lines could be associated with the methylation status and the expression of miRNAs from the imprinted DLK1/DIO3 locus. More specifically, quantitative analysis of the expression of six different miRNAs of that locus prospectively identified human embryonic stem cells and human-induced pluripotent stem cells with differential osteogenic differentiation capacities. At the molecular and functional levels, we showed that these miRNAs modulated the expression of the activin receptor type 2B and the downstream signal transduction, which impacted osteogenesis. In conclusion, miRNAs of the imprinted DLK1/DIO3 locus appear to have both a predictive value and a functional impact in determining the osteogenic fate of human pluripotent stem cells.

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Generating Kidney from Stem Cells

Annual Review of Physiology ◽

10.1146/annurev-physiol-020518-114331 ◽

2019 ◽

Vol 81 (1) ◽

pp. 335-357 ◽

Cited By ~ 8

Author(s):

Melissa H. Little ◽

Lorna J. Hale ◽

Sara E. Howden ◽

Santhosh V. Kumar

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Pluripotent Stem Cells ◽

Cellular Therapy ◽

Kidney Diseases ◽

Kidney Tissue ◽

Human Kidney ◽

Human Pluripotent Stem Cells ◽

Directed Differentiation ◽

Cellular Therapies

Human kidney tissue can now be generated via the directed differentiation of human pluripotent stem cells. This advance is anticipated to facilitate the modeling of human kidney diseases, provide platforms for nephrotoxicity screening, enable cellular therapy, and potentially generate tissue for renal replacement. All such applications will rely upon the accuracy and reliability of the model and the capacity for stem cell–derived kidney tissue to recapitulate both normal and diseased states. In this review, we discuss the models available, how well they recapitulate the human kidney, and how far we are from application of these cells for use in cellular therapies.

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