DJ-1 Can Replace FGF-2 for Long-Term Culture of Human Pluripotent Stem Cells in Defined Media and Feeder-Free Condition

Conventional human pluripotent stem cell (hPSC) cultures require high concentrations of expensive human fibroblast growth factor 2 (hFGF-2) for hPSC self-renewal and pluripotency in defined media for long-term culture. The thermal instability of the hFGF-2 mandates media change every day, which makes hPSC culture costly and cumbersome. Human DJ-1 (hDJ-1) can bind to and stimulate FGF receptor-1. In this study, for the first time, we have replaced hFGF-2 with hDJ-1 in the essential eight media and maintained the human embryonic stem cells (hESCs), H9, in the defined media at feeder-free condition. After more than ten passages, H9 in both groups still successfully maintained the typical hESC morphology and high protein levels of pluripotency markers, SSEA4, Tra1-60, Oct4, Nanog, and ALP. DNA microarray revealed that more than 97% of the 21,448 tested genes, including the pluripotency markers, Sox2, Nanog, Klf4, Lin28A, Lin28B, and Myc, have similar mRNA levels between the two groups. Karyotyping revealed no chromosome abnormalities in both groups. They also differentiated sufficiently into three germ layers by forming in vitro EBs and in vivo teratomas. There were some variations in the RT-qPCR assay of several pluripotency markers. The proliferation rates and the mitochondria of both groups were also different. Taken together, we conclude that hDJ-1 can replace hFGF-2 in maintaining the self-renewal and the pluripotency of hESCs in feeder-free conditions.

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Rapid induction and long-term self-renewal of primitive neural precursors from human embryonic stem cells by small molecule inhibitors

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1014041108 ◽

2011 ◽

Vol 108 (20) ◽

pp. 8299-8304 ◽

Cited By ~ 232

Author(s):

W. Li ◽

W. Sun ◽

Y. Zhang ◽

W. Wei ◽

R. Ambasudhan ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Small Molecule ◽

Human Embryonic Stem Cells ◽

Small Molecule Inhibitors ◽

Embryonic Stem ◽

Self Renewal ◽

Neural Precursors ◽

Rapid Induction

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Function of Pumilio Genes in Human Embryonic Stem Cells and Their Effect in Stemness and Cardiomyogenesis

10.1101/751537 ◽

2019 ◽

Author(s):

Isabelle Leticia Zaboroski Silva ◽

Anny Waloski Robert ◽

Guillermo Cabrera Cabo ◽

Lucia Spangenberg ◽

Marco Augusto Stimamiglio ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Rna Binding ◽

Rna Binding Proteins ◽

Embryonic Stem ◽

Mrna Levels ◽

Human Escs ◽

Mrna Targets ◽

Cardiomyogenic Differentiation

AbstractPosttranscriptional regulation plays a fundamental role in the biology of embryonic stem cells (ESCs). Many studies have demonstrated that multiple mRNAs are coregulated by one or more RNA binding proteins (RBPs) that orchestrate the expression of these molecules. A family of RBPs, known as PUF (Pumilio-FBF), is highly conserved among species and has been associated with the undifferentiated and differentiated states of different cell lines. In humans, two homologs of the PUF family have been found: Pumilio 1 (PUM1) and Pumilio 2 (PUM2). To understand the role of these proteins in human ESCs (hESCs), we first demonstrated the influence of the silencing of PUM1 and PUM2 on pluripotency genes. OCT4 and NANOG mRNA levels decreased significantly with the knockdown of Pumilio, suggesting that PUMILIO proteins play a role in the maintenance of pluripotency in hESCs. Furthermore, we observed that the hESCs silenced for PUM1 and 2 exhibited an improvement in efficiency of in vitro cardiomyogenic differentiation. Using in silico analysis, we identified mRNA targets of PUM1 and PUM2 expressed during cardiomyogenesis. With the reduction of PUM1 and 2, these target mRNAs would be active and could be involved in the progression of cardiomyogenesis.

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Embryonic Stem Cells Facilitate the Isolation of Persistent Clonal Cardiovascular Progenitor Cell Lines and Leukemia Inhibitor Factor Maintains Their Self-Renewal and Myocardial Differentiation Potential in vitro

Cells Tissues Organs ◽

10.1159/000345804 ◽

2013 ◽

Vol 197 (4) ◽

pp. 249-268 ◽

Cited By ~ 5

Author(s):

Julia Hoebaus ◽

Philipp Heher ◽

Teresa Gottschamel ◽

Matthias Scheinast ◽

Harmen Auner ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Cell Lines ◽

Progenitor Cell ◽

Embryonic Stem ◽

Differentiation Potential ◽

Self Renewal ◽

Leukemia Inhibitor Factor ◽

Myocardial Differentiation

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Primitive and Definitive Erythropoiesis

Blood ◽

10.1182/blood.v120.21.sci-37.sci-37 ◽

2012 ◽

Vol 120 (21) ◽

pp. SCI-37-SCI-37

Author(s):

James Palis

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Embryonic Stem ◽

Yolk Sac ◽

Erythroid Cells ◽

Transient Wave ◽

Self Renewal ◽

Primitive Erythropoiesis ◽

Definitive Erythropoiesis

Abstract Abstract SCI-37 Studies in mammalian and nonmammalian vertebrate embryos indicate that erythropoiesis comes in two flavors: primitive and definitive. The primitive erythroid lineage in mammalian embryos is characterized by a transient wave of lineage-committed progenitors that emerge from the yolk sac and generate a wave of precursors that synchronously mature in the bloodstream. Primitive erythroid precursors dynamically regulate embryonic globin gene expression and ultimately enucleate to form erythrocytes. Primitive erythropoiesis is superseded by definitive erythroid cells that mature extravascularly in association with macrophage cells. Studies in the mouse embryo indicate that definitive erythropoiesis has two distinct developmental origins. The first is a transient wave of erythro-myeloid progenitors (EMP) that emerge from the yolk sac and seed the early fetal liver. The second is a long-term program of erythropoiesis derived from hematopoietic stem cells. Erythropoietin is the central regulator of definitive erythropoiesis, in part by regulating the survival of committed progenitors. In contrast, the role of erythropoietin in primitive erythropoiesis remains poorly understood. Recent studies indicate that erythropoietin does not regulate the primitive erythroid progenitor compartment, but rather plays a critical role in establishing an antiapoptotic state during the terminal maturation of primitive erythroblasts. EMP-derived proerythroblasts are capable of extensive self-renewal in vitro, while primitive erythroid progenitors are incapable of self-renewal under the same conditions. These studies, taken together, indicate that the primitive and definitive forms of erythropoiesis have fundamental differences in the regulation of red cell output. The overlapping emergence of primitive and definitive erythroid lineages in differentiating embryonic stem cells suggests that the transient yolk-sac-derived primitive and EMP-derived definitive erythroid programs are recapitulated in vitro. These studies offer the hope that human embryonic stem cells can serve as a source of functional definitive erythroid cells for transfusion therapy. Disclosures: No relevant conflicts of interest to declare.

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