scholarly journals Distinct iPS Cells Show Different Cardiac Differentiation Efficiency

2013 ◽  
Vol 2013 ◽  
pp. 1-11 ◽  
Author(s):  
Yohei Ohno ◽  
Shinsuke Yuasa ◽  
Toru Egashira ◽  
Tomohisa Seki ◽  
Hisayuki Hashimoto ◽  
...  
Keyword(s):  
Regenerative Medicine ◽  
Cell Lines ◽  
Time Course ◽  
Es Cells ◽  
Ips Cells ◽  
Cardiac Differentiation ◽  
Patient Specific ◽  
Differentiation Potential ◽  
Ips Cell ◽  
Differentiation Efficiency

Patient-specific induced pluripotent stem (iPS) cells can be generated by introducing transcription factors that are highly expressed in embryonic stem (ES) cells into somatic cells. This opens up new possibilities for cell transplantation-based regenerative medicine by overcoming the ethical issues and immunological problems associated with ES cells. Despite the development of various methods for the generation of iPS cells that have resulted in increased efficiency, safety, and general versatility, it remains unknown which types of iPS cells are suitable for clinical use. Therefore, the aims of the present study were to assess (1) the differentiation potential, time course, and efficiency of different types of iPS cell lines to differentiate into cardiomyocytes in vitro and (2) the properties of the iPS cell-derived cardiomyocytes. We found that high-quality iPS cells exhibited better cardiomyocyte differentiation in terms of the time course and efficiency of differentiation than low-quality iPS cells, which hardly ever differentiated into cardiomyocytes. Because of the different properties of the various iPS cell lines such as cardiac differentiation efficiency and potential safety hazards, newly established iPS cell lines must be characterized prior to their use in cardiac regenerative medicine.

scholarly journals Human-induced pluripotent stem cells from blood cells of healthy donors and patients with acquired blood disorders

Blood ◽  
2009 ◽  
Vol 114 (27) ◽  
pp. 5473-5480 ◽  
Author(s):  
Zhaohui Ye ◽  
Huichun Zhan ◽  
Prashant Mali ◽  
Sarah Dowey ◽  
Donna M. Williams ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Lines ◽  
Blood Cells ◽  
Ips Cells ◽  
Patient Specific ◽  
Hematopoietic Differentiation ◽  
Ips Cell ◽  
Healthy Donors ◽  
Cd34 Cells ◽  
Induced Pluripotent

Abstract Human induced pluripotent stem (iPS) cells derived from somatic cells hold promise to develop novel patient-specific cell therapies and research models for inherited and acquired diseases. We and others previously reprogrammed human adherent cells, such as postnatal fibroblasts to iPS cells, which resemble adherent embryonic stem cells. Here we report derivation of iPS cells from postnatal human blood cells and the potential of these pluripotent cells for disease modeling. Multiple human iPS cell lines were generated from previously frozen cord blood or adult CD34+ cells of healthy donors, and could be redirected to hematopoietic differentiation. Multiple iPS cell lines were also generated from peripheral blood CD34+ cells of 2 patients with myeloproliferative disorders (MPDs) who acquired the JAK2-V617F somatic mutation in their blood cells. The MPD-derived iPS cells containing the mutation appeared normal in phenotypes, karyotype, and pluripotency. After directed hematopoietic differentiation, the MPD-iPS cell-derived hematopoietic progenitor (CD34+CD45+) cells showed the increased erythropoiesis and gene expression of specific genes, recapitulating features of the primary CD34+ cells of the corresponding patient from whom the iPS cells were derived. These iPS cells provide a renewable cell source and a prospective hematopoiesis model for investigating MPD pathogenesis.

Abstract 3460: Molecular Characterization, Safety and Feasibility of Induced Pluripotent Stem (iPS) Cell Derived Cardiomyocytes for Heart Regenerative Therapy

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Yohei Ohno ◽  
Shinsuke Yuasa ◽  
Takeshi Onizuka ◽  
Toru Egashira ◽  
Kenichiro Shimoji ◽  
...  
Keyword(s):  
Molecular Characterization ◽  
Es Cells ◽  
Sinus Node ◽  
Ips Cells ◽  
Embryoid Bodies ◽  
Marker Genes ◽  
List Type ◽  
Cardiomyocyte Differentiation ◽  
Differentiation Potential ◽  
Ips Cell

[Background] We recently reported that mouse and human iPS cells could be generated from somatic cells by gene transfer of Oct3/4, Sox2, c-Myc and Klf4. Although the morphology, growth characteristics and pluripotency of iPS cells are believed to be similar to those of ES cells, there are several versions such as Fbx-iPS, Nanog-iPS cells, and it remains unknown whether what type of iPS cell could be appropriate for the cardiovascular regeneration therapy. This study investigated the cardiomyocyte differentiation potential of mouse iPS cells, molecular characterization of iPS-induced cardiomyocytes, and the safety and feasibility of their transplantation into the heart. [Methods and Results] We used several lines of Fbx-iPS and Nanog-iPS cells and induced cardiomyocyte differentiation using the hanging drop culture method. Mouse ES cells were used as control. Both Fbx-iPS and Nanog-iPS cells could differentiate into cardiomyocytes, and the incidence of beating embryoid bodies was similar to that of ES cells. RT-PCR analyses and immunohistochemistry revealed that both iPS cells expressed all the cardiomyocyte marker genes including Nkx2.5, GATA4, MEF2C, myosin light chain-2v, α-myosin heavy chain, atrial natriuretic peptide, and α-sarcomeric actinin, and they showed normal structures. Electron microscopic study revealed that iPS cell derived cardiomyocytes had the typical morphological cardiomyocyte structure, which is difficult to distinguish from that of ES cell derived cells. Electrical physiological study showed that iPS cell derived cardiomyocytes had sinus node-like or fetal ventricular type action potentials. Purified iPS cell derived cardiomyocytes could be successfully transplanted into murine hearts and the engrafted cells survived and were integrated into host myocardium. Nanog-iPS cell derived cells showed no severe complication after transplantation, however Fbx-iPS cell derived cells showed teratoma in the host tissue. Fbx-iPS cells expressed extrinsic c-myc after differentiation. [Conclusions] The iPS cells proved to be an attractive cell source for the regeneration of cardiomyocytes, however safety and careful characterization is necessary before clinical application.

Reduced Production of Mature Neutrophils From Induced Pluripotent Stem Cells Derived From a Severe Congenital Neutropenia Patient with HAX1 Gene Deficiency

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2402-2402 ◽  
Author(s):  
Tatsuya Morishima ◽  
Ken-ichiro Watanabe ◽  
Akira Niwa ◽  
Takayuki Tanaka ◽  
Katsutsugu Umeda ◽  
...  
Keyword(s):  
Cell Lines ◽  
Normal Control ◽  
Annexin V ◽  
Ips Cells ◽  
Patient Specific ◽  
Es Cell ◽  
Severe Congenital Neutropenia ◽  
Congenital Neutropenia ◽  
Ips Cell ◽  
Induced Pluripotent

Abstract Abstract 2402 Induced pluripotent stem (iPS) cells are reprogrammed somatic cells with embryonic stem (ES) cell-like characteristics. As iPS cells can be generated from somatic cells of patients with a certain disease, they are expected to be a novel model to study pathogenesis of various diseases. Recently, we established a neutrophil differentiation system from human iPS cells (Morishima T, et al. J Cell Physiol. 2011). In an attempt to apply the system to investigate pathophysiology of neutrophil-affected disorders, we generated iPS cells from a severe congenital neutropenia (SCN) patient with HAX1 gene deficiency. The patient was an 11-year-old boy with severe congenital neutropenia as well as developmental delay and epilepsy. DNA sequence analysis revealed HAX1 gene mutation in exon 2 (Matsubara K, et al. Haematologica. 2007). Four iPS cell lines were generated from skin fibroblasts of the patient by retroviral overexpression of the three or four transcription factors Oct3/4, Sox2, and Klf4, with or without c-Myc. These patient-derived iPS cell lines showed human ES cell like morphology and could be maintained under human ES cell culture condition. They also expressed typical human ES cell markers and were capable of differentiating into the cell lineages and tissues representing three germ layers by teratoma formation in vivo. These cells had normal karyotype and short tandem repeat analysis indicated that they were derived from parental skin fibroblast. DNA sequencing analysis of the iPS cell lines identified the same mutation carried in the parental skin fibroblasts, thus confirmed that we had established the HAX1 deficiency patient-specific iPS cells (HAX1-iPSCs). Next these HAX1-iPSCs and the healthy-person derived iPS cells were differentiated into neutrophils in vitro using feeder-free culture protocols established in our laboratory. In this culture system, small human iPS cell clumps were cultured on the matrigel-coated dish with recombinant cytokines and without any feeder cells and fetal calf serum. Around day 25 of culture, mature neutrophils were obtained as floating cells. Morphologically, the majority of HAX1-iPSCs-derived cells were classified into myeloblast or promyelocyte stage and there were only a few mature neutrophils. The proportion of mature neutrophils was only less than 10% in HAX1-iPSCs-derived cells whereas more than 40% in normal control. Flow cytometric analysis revealed that the percentage of immature CD34 positive cells was significantly higher and that of myeloid-committed CD11b positive cells was lower in the HAX1-iPSCs-derived cells than normal control. Immunocytochemical analysis for neutrophil specific granules showed that lactoferrin- and gelatinase-positive cells decreased in the HAX1-iPSCs-derived cells compared with normal control, confirming that HAX1-iPSCs-derived cells contained less mature neutrophils than normal control. Apoptosis assay by Annexin V staining revealed that HAX1-iPSCs-derived cells showed higher percentage of Annexin V-positive cells compared with normal control. Overall, these HAX1 deficiency patient-specific iPS cell lines recapitulate the hematological phenotype in the patient. These results indicate that patient-derived iPS cells provide us a novel disease model and make a contribution to the understanding of the pathophysiology of the diseases that affect neutrophils. Disclosures: No relevant conflicts of interest to declare.

Comprehensive Comparison Of Gene Expression, Genomic DNA Methylation, and In Vitro Hematopoietic Differentiation Among Many Human iPS and ES Cell Lines

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1187-1187
Author(s):  
Masatoshi Nishizawa ◽  
Kazuhisa Chonabayashi ◽  
Akiko Oishi ◽  
Ikue Takei ◽  
Misato Nishikawa ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Cell Lines ◽  
Genomic Dna ◽  
Ips Cells ◽  
Hematopoietic Differentiation ◽  
Es Cell ◽  
Differentiation Potential ◽  
Genome Wide ◽  
Differentiation Efficiency

Abstract Objective Hematopoietic differentiation from human induced pluripotent stem (iPS)/embryonic stem (ES) cell attracts much attention due to its huge potential for regenerative medicine. As indicated by some earlier papers, there is large variation in differentiation potential among pluripotent stem cell (PSC) lines, and this is one of major concerns in clinical application of PSCs. If it becomes possible to predict which PSC line has high differentiation potential without real differentiation experiment, it would greatly contribute to clinical application of PSCs. Although some papers reported about presence of epigenetic memories of parental somatic cells in iPS cells, the amount of the influence on differentiation potential remains to be known. Furthermore, especially in studies using human PSCs, genetic difference among individual donors of iPS/ES cells seems to be large, thus the study using many PSC lines from many donors is warranted. To address these issues, we planned to collect data of many iPS/ES cell lines on genome-wide gene expression and genomic DNA methylation, and differentiation potentials of individual lines, and identify the factors which affected difference in differentiation potential among PSC lines. The final goal of this study is to create data base about gene expression and DNA methylation profile and differentiation potentials of many PSC lines. We believe that this dataset will allow us to predict differentiation potentials of individual PSC lines, and accelerate clinical application of PSC lines in hematology field. Method We utilized 39 iPS/ES lines (iPS 35 lines, ES 4 lines) in this study. The iPS cell lines were derived from dermal fibroblast (n = 16), cord blood (n = 3), peripheral blood (n = 10), keratinocyte (n = 3), and dental pulp cell (n = 3), and were generated by retrovirus vector (n = 9), episomal vector (n = 25), and sendai virus vector (n = 1). The iPS cells were derived from 15 donors, and the ES cells were derived from 4 donors. We assessed hematopoietic differentiation potential by investigating hematopoietic differentiation efficiency for the first 15 days from start of differentiation, and colony forming potential of hematopoietic precursor cells (CD34+CD38-CD43+lineage marker- population) generated from PSC lines using semi-solid methylcellulose based-media. In addition, we collected genome-wide mRNA expression and DNA methylation profile of PSC lines, parental lines of iPS cells, hematopoietic precursor cells generated from PSCs by using mRNA microarray, genomic methylation beads array, and next generation sequencers, and analyzed correlation of these data with differentiation potentials of individual PSC lines. Result We have found that there is large variation in hematopoietic differentiation efficiency and colony forming ability as reported previously. Genome-wide investigation of gene expression and genomic DNA methylation revealed that expression of some genes or some factors were significantly correlated with hematopoietic differentiation efficiency or colony forming ability of hematopoietic precursor cells. Importantly, the factors affecting differentiation efficiency for first 15 days and those affecting colony-forming ability were absolutely different. More importantly, by combining several factors discovered in this analysis, we can predict hematopoietic differentiation potential of individual iPS/ES cell lines regardless of what parental cell lines iPS cells are derived or whether it is an iPS cell or ES cell. Conclusion From genome-wide analysis of gene expression and genomic DNA methylation, and hematopoietic differentiation experiments, we discovered the factors that were associated with difference in differentiation potential among PSC lines. Now, we are focusing on investigating molecular mechanisms by which the discovered factors are responsible for the difference in hematopoietic differentiation potentials among PSC lines. We believe that our findings will contribute not only to clinical application of hematopoietic cells generated from human PSCs, but also to further understanding of human developmental hematopoiesis. Disclosures: No relevant conflicts of interest to declare.

292 GENERATION OF MOUSE INDUCED PLURIPOTENT STEM CELLS FROM VARIOUS GENETIC BACKGROUND BY SLEEPING BEAUTY TRANSPOSON-MEDIATED GENE TRANSFER

2011 ◽  
Vol 23 (1) ◽  
pp. 243 ◽  
Author(s):  
S. Muenthaisong ◽  
O. Ujhelly ◽  
E. Varga ◽  
A. C. Carstea ◽  
Z. Ivics ◽  
...  
Keyword(s):  
Cell Lines ◽  
Pluripotent Stem Cells ◽  
Ips Cells ◽  
Sleeping Beauty ◽  
Embryoid Bodies ◽  
Es Cell ◽  
Differentiation Potential ◽  
Ips Cell ◽  
Sleeping Beauty Transposon ◽  
Induced Pluripotent

Induced pluripotent stem (iPS) cell technology allows the reprogramming of somatic cells to a pluripotent state; however, it requires viral gene transduction and permanent existence of the exogenous genes in the genome, which is a potential risk for abnormalities in the derived iPS cells. Recently, there was report that iPS cells have been made with piggyBack transposon. Here, we first reported that nonviral transfection of a Sleeping Beauty transposon, which comprises c-Myc, Klf-4, Oct3/4 (Pou5f1), and Sox-2, can reprogram mouse fibroblasts from 3 different genetic backgrounds: ICR (outbred), C57BL/6 (inbred), and F1 hybrid (C57BL/6 × DBA/2J), with parallel robust expression of all exogenous (c-Myc, Klf-4, Oct3/4, and Sox-2) and endogenous (e.g. Nanog) pluripotency genes. The iPS cells were cultured under standard conditions with promotion of differentiate by withdrawal of leukemia inhibitory factor. We chose 6 cloned of each line that exhibited characteristics typical for undifferentiated embryonic stem (ES) cell: ES-cell-like morphology, alkaline phosphatase positivity, and gene expression pattern [quantitative real-time PCR and immunofluorescence of ES cell markers (e.g. Oct-4, SSEA1, Nanog]. Furthermore, cells were able to form embryoid bodies and beat rhythmically and expressed cardiac markers assayed by immunofluorescence (e.g. cardiac Troponin T, desmin). In vivo testing of iPS cell lines for their developmental potential (diploid and tetraploid embryo complementation assay) is currently underway. The iPS cell lines generated from the ICR strain appeared the earliest in time (ICR-d11, F1 day-2 and Bl6-d12), with higher efficiency than colonies from the other 2 backgrounds. The differentiation potential of the iPS lines derived from the 3 genetic backgrounds was similar. Interestingly, the ICR-iPS lines had higher differentiation potential than did the ICR-ES cell lines: the rate of embryoid bodies forming rhythmically beating cardiomyocytes was 4% in ICR-ES and 79% in ICR-iPS cells, respectively. Our results suggest that the iPS technology provide a new tool to generate pluripotent stem cells from genetic backgrounds where good-quality ES cell generation is difficult. These studies provide new insights into virus-free iPS technology and contribute to defining future cell-based therapies, drug screening methods, and production of transgenic animals with genetically modified iPS cells. This study was financed by EU FP6 (CLONET, MRTN-CT-2006-035468), EU FP7 (PartnErS, PIAP-GA-2008-218205; InduHeart, PEOPLE-IRG-2008-234390; InduVir, PEOPLE-IRG-2009-245808; InduStem, PIAP-GA-2008-230675; PluriSys, HEALTH-2007-B-223485); NKTH-OTKA-EU FP7-HUMAN-2009-MB08-C 80205, and NKTH/KPI (Jedlik NKFP_07_1-ES2HEART-HU OM-00202-2007).

Cell Culture, iPS Cells and Neurodegenerative Diseases

2016 ◽  
Author(s):  
Roxana Nat ◽  
Andreas Eigentler
Keyword(s):  
Cell Culture ◽  
Muscular Atrophy ◽  
Ips Cells ◽  
Patient Specific ◽  
Neural Cells ◽  
Differentiation Potential ◽  
Ips Cell ◽  
Human Neurons ◽  
Culture Models ◽  
Induced Pluripotent

Somatic reprogramming technology, which enables the conversion of adult human non-neural cells into neurons, has progressed rapidly in recent years. The derivation of patient-specific induced pluripotent stem (iPS) cells has become routine. The inherent broad differentiation potential of iPS cells makes possible the generation of diverse types of human neurons. This constitutes a remarkable step in facilitating the development of more appropriate and comprehensive preclinical human disease models, as well as for high throughput drug screenings and cell therapy. This chapter reviews recent progress in the human iPS cell culture models related to common and rare NDDs, such as Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, amyotrophic lateral sclerosis, spinal muscular atrophy, and degenerative ataxias. It focuses on the pathophysiological features revealed in cell cultures, and the neuronal subtypes most affected in NDDs. The chapter discusses the validity, limitation, and improvements of this system in faithfully and reproducibly recapitulating disease pathology.

scholarly journals Nonhuman Primate Induced Pluripotent Stem Cells in Regenerative Medicine

2012 ◽  
Vol 2012 ◽  
pp. 1-7 ◽  
Author(s):  
Yuehong Wu ◽  
Anuja Mishra ◽  
Zhifang Qiu ◽  
Steven Farnsworth ◽  
Suzette D. Tardif ◽  
...  
Keyword(s):  
Stem Cells ◽  
Regenerative Medicine ◽  
Induced Pluripotent Stem Cells ◽  
Cell Lines ◽  
Pluripotent Stem Cells ◽  
Patient Specific ◽  
Ips Cell ◽  
Disease States ◽  
Autologous Cell Therapy ◽  
Induced Pluripotent

Among the various species from which induced pluripotent stem cells have been derived, nonhuman primates (NHPs) have a unique role as preclinical models. Their relatedness to humans and similar physiology, including central nervous system, make them ideal for translational studies. We review here the progress made in deriving and characterizing iPS cell lines from different NHP species. We focus on iPS cell lines from the marmoset, a small NHP in which several human disease states can be modeled. The marmoset can serve as a model for the implementation of patient-specific autologous cell therapy in regenerative medicine.

Abstract 340: From Stem Cells to Cardiomyocytes: HDAC1 Induces Cardiovascular Differentiation by Regulating SOX17-BMP2 Expression in Early Development.

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Eneda Hoxha ◽  
Erin Lambers ◽  
Veronica Ramirez ◽  
Prasanna Krishnamurthy ◽  
Suresh Verma ◽  
...  
Keyword(s):  
Stem Cells ◽  
Molecular Mechanisms ◽  
Es Cells ◽  
Ips Cells ◽  
Full Potential ◽  
Pluripotent Cells ◽  
Differentiation Potential ◽  
Histone Deacetylase 1 ◽  
Specific Differentiation ◽  
Induced Pluripotent

Cardiomyocytes derived from embryonic and induced pluripotent stem cells (ES/iPS) provide an excellent source for cell replacement therapies following myocardial ischemia. However, some of the obstacles in the realization of the full potential of iPS/ES cells arise from incomplete and poorly understood molecular mechanisms and epigenetic modifications that govern their cardiovascular specific differentiation. We identified Histone Deacetylase 1 (HDAC1) as a crucial regulator in early differentiation of mES and iPS cells. We propose a novel pathway in which HDAC1 regulates cardiovascular differentiation by regulating SOX17 which in turn regulates BMP2 signaling in differentiating pluripotent cells. Utilizing stable HDAC1 knock-down (HDAC1-KD) cell lines, we report an essential role for HDAC1 in deacetylating regulatory regions of pluripotency-associated genes during early cardiovascular differentiation. HDAC1-KD cells show severely repressed cardiomyocyte differentiation potential. We propose a novel HDAC1-BMP2-SOX17 dependent pathway through which deacetylation of pluripotency associated genes leads to their suppression and allows for early cardiovascular-associated genes to be expressed and differentiation to occur. Furthermore, we show that HDAC1 affects DNA methylation both during pluripotency and differentiation and plays a crucial, non-redundant role in cardiovascular specific differentiation and cardiomyocyte maturation. Our data elucidates important differences between ES and iPS HDAC1-KD cells that affect their ability to differentiate into cardiovascular lineages. As varying levels of chromatin modifying enzymes are likely to exist in patient derived iPS cells, understanding the molecular circuitry of these enzymes in ES and iPS cells is critical for their potential therapeutic applications in regenerative medicine. Further research in the molecular mechanisms involved in this process will greatly aid our understanding of the epigenetic circuitry of pluripotency and differentiation in pluripotent cells.

scholarly journals Isolation, Characterization and Differentiation Potential of Chicken Spermatogonial Stem Cell Derived Embryoid Bodies

2016 ◽  
Vol 16 (1) ◽  
pp. 115-128 ◽  
Author(s):  
Thanh Luan Nguyen ◽  
Jae Gyu Yoo ◽  
Neelesh Sharma ◽  
Sung Woo Kim ◽  
Yong Jun Kang ◽  
...  
Keyword(s):  
Developmental Biology ◽  
Regenerative Medicine ◽  
Connexin 43 ◽  
Es Cells ◽  
Periodic Acid ◽  
Embryonic Stem ◽  
Embryoid Bodies ◽  
Significant Finding ◽  
Differentiation Potential

Abstract Human, murine and monkey spermatogonial stem cells (SSCs) have the capability to undergo self-renewal and differentiation into different body cell types in vitro, which are expected to serve as a powerful tool and resource for the developmental biology and regenerative medicine. We have successfully isolated and characterized the chicken SSCs from 3-day-old chicken testicular cells. The pluripotency was using Periodic Acid-Schiff (PAS ) staining or alkaline phosphatase staining, and antibodies to stage-specific embryonic antigens. In suspension culture conditions SSCs formed embryoid bodies (EBs) like embryonic stem (ES) cells. Subsequently EB differentiated into osteoblasts, adipocytes and most importantly into cardiomyocytes under induced differentiation conditions. The differentiation potential of EBs into cardiomyocyte-like cells was confirmed by using antibodies against sarcomeric α-actinin, cardiac troponin T and connexin 43. Cardiomyocytes-like cells were also confirmed by RT-PCR analysis for several cardiac cell genes like GATA-4, Nkx2-5, α-MHC, and ANF. We have successfully established an in vitro differentiation system for chicken SSCs into different body cells such as osteoblasts, adipocytes and cardiomyocytes. The most significant finding of this study is the differentiation potential of chicken SSCs into cardiomyocytes. Our findings may have implication in developmental biology and regenerative medicine by using chicken as the most potential animal model.

scholarly journals Generation of multipotent cell lines from a distinct population of male germ line stem cells

Reproduction ◽  
2008 ◽  
Vol 135 (6) ◽  
pp. 771-784 ◽  
Author(s):  
Fariborz Izadyar ◽  
Francis Pau ◽  
Joel Marh ◽  
Natalia Slepko ◽  
Tracy Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Germ Cell ◽  
Cell Lines ◽  
Germ Line ◽  
Es Cells ◽  
Embryonic Stem ◽  
Complex Mixture ◽  
Neural Cells ◽  
Differentiation Potential

Spermatogonial stem cells (SSCs) maintain spermatogenesis by self-renewal and generation of spermatogonia committed to differentiation. Under certain in vitro conditions, SSCs from both neonatal and adult mouse testis can reportedly generate multipotent germ cell (mGC) lines that have characteristics and differentiation potential similar to embryonic stem (ES) cells. However, mGCs generated in different laboratories showed different germ cell characteristics, i.e., some retain their SSC properties and some have lost them completely. This raises an important question: whether mGC lines have been generated from different subpopulations in the mouse testes. To unambiguously identify and track germ line stem cells, we utilized a transgenic mouse model expressing green fluorescence protein under the control of a germ cell-specific Pou5f1 (Oct4) promoter. We found two distinct populations among the germ line stem cells with regard to their expression of transcription factor Pou5f1 and c-Kit receptor. Only the POU5F1+/c-Kit+ subset of mouse germ line stem cells, when isolated from either neonatal or adult testes and cultured in a complex mixture of growth factors, generates cell lines that express pluripotent ES markers, i.e., Pou5f1, Nanog, Sox2, Rex1, Dppa5, SSEA-1, and alkaline phosphatase, exhibit high telomerase activity, and differentiate into multiple lineages, including beating cardiomyocytes, neural cells, and chondrocytes. These data clearly show the existence of two distinct populations within germ line stem cells: one destined to become SSC and the other with the ability to generate multipotent cell lines with some pluripotent characteristics. These findings raise interesting questions about the relativity of pluripotency and the plasticity of germ line stem cells.

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