Human Amniotic Mesenchymal Stem Cell-Derived Induced Pluripotent Stem Cells May Generate a Universal Source of Cardiac Cells

1 Division of Cardiovascular Medicine, Department of Medicine, Stanford University School of Medicine , Stanford, California.
Stem cells and development (Impact Factor: 3.73). 04/2012; 21(15):2798-808. DOI: 10.1089/scd.2011.0435
Source: PubMed


Human amniotic mesenchymal stem cells (hAMSCs) demonstrated partially pluripotent characteristics with a strong expression of Oct4 and Nanog genes and immunomodulatory properties characterized by the absence of HLA-DR and the presence of HLA-G and CD59. The hAMSCs were reprogrammed into induced pluripotent stem cells (iPSCs) that generate a promising source of universal cardiac cells. The hAMSC-derived iPSCs (MiPSCs) successfully underwent robust cardiac differentiation to generate cardiomyocytes. This study investigated 3 key properties of the hAMSCs and MiPSCs: (1) the reprogramming efficiency of the partially pluripotent hAMSCs to generate MiPSCs; (2) immunomodulatory properties of the hAMSCs and MiPSCs; and (3) the cardiac differentiation potential of the MiPSCs. The characteristic iPSC colony formation was observed within 10 days after the transduction of the hAMSCs with a single integration polycistronic vector containing 4 Yamanaka factors. Immunohistology and reverse transcription-polymerase chain reaction assays revealed that the MiPSCs expressed stem cell surface markers and pluripotency-specific genes. Furthermore, the hAMSCs and MiPSCs demonstrated immunomodulatory properties enabling successful engraftment in the SVJ mice. Finally, the cardiac differentiation of MiPSCs exhibited robust spontaneous contractility, characteristic calcium transience across the membrane, a high expression of cardiac genes and mature cardiac phenotypes, and a contractile force comparable to cardiomyocytes. Our results demonstrated that the hAMSCs are reprogrammed with a high efficiency into MiPSCs, which possess pluripotent, immunomodulatory, and precardiac properties. The MiPSC-derived cardiac cells express a c-kit cell surface marker, which may be employed to purify the cardiac cell population and enable allogeneic cardiac stem cell therapy.

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Available from: Renee A Reijo Pera, Jul 07, 2015
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    • "Recently, several ASCs have been successfully isolated and cultured in vitro, including hematopoietic stem cells (HSCs)11, mesenchymal stem cells (MSCs)1213, epidermis stem cells14, neural stem cells (NSCs)15, adipose-derived stem cells (ADSCs)161718, islet stem cells1920, and germ line stem cells212223. Human mesenchymal stem cells originate mainly from bone marrow2425, cord blood262728, placenta293031, and endometrium32, but epidermis-derived MSCs have not yet been isolated. In the present study, we isolated small spindle-shaped cells with strong proliferative potential from human epidermis. "
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    ABSTRACT: We isolated human epidermis-derived mesenchymal stem cell-like pluripotent cells (hEMSCPCs) and demonstrate efficient harvesting, maintenance in vitro for at least 30 passages, reprogramming into multiple phenotypes in vivo, and integration into adult host tissues after injection into the mouse blastocyst to create chimeras. Cell phenotype was examined by karyotyping, immunostaining, immunofluorescence, and flow cytometry. A nested PCR protocol using primers specific for human SRY genes was designed to detect hEMSCPC-derived cells in female chimeric mice. FISH was used to validate the results of nested PCR. Results indicated that hEMSCPCs were derived from epidermis but were distinct from epidermal cells; they resembled mesenchymal stem cells (MSCs) morphologically and expressed the main markers of MSCs. About half of all female offspring of mice implanted with embryos injected with hEMSCPCs at the blastocyst stage harbored the human Y chromosome and tissue-specific human protein, thereby demonstrating the transdifferentiation of hEMSCPCs.
    Full-text · Article · Jun 2013 · Scientific Reports
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    • "Human amniocytes are considered an embryonic or fetal multipotent stem cell due to expression of transcriptional regulators [11]–[14] and cell surface antigens [15]–[18] characteristic of stem cells. Interestingly, amniocytes can be efficiently reprogrammed into a primitive pluripotent state by DNA-integrating [19]–[25] and non-integrating methods [18], and subsequently differentiated along multiple lineages [17], [18], [22], [26]–[32]. Alternatively, they can be reprogrammed through direct methods, which are thought to bypass pluripotency altogether [33], or as our data suggests, use some of the innate pluripotency of amniocytes. "
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    ABSTRACT: Human amniotic fluid contains cells that potentially have important stem cell characteristics, yet the programs controlling their developmental potency are unclear. Here, we provide evidence that amniocytes derived from multiple patients are marked by heterogeneity and variability in expression levels of pluripotency markers. Clonal analysis from multiple patients indicates that amniocytes have large pools of self-renewing cells that have an inherent property to give rise to a distinct amniocyte phenotype with a heterogeneity of pluripotent markers. Significant to their therapeutic potential, genome-wide profiles are distinct at different gestational ages and times in culture, but do not differ between genders. Based on hierarchical clustering and differential expression analyses of the entire transcriptome, amniocytes express canonical regulators associated with pluripotency and stem cell repression. Their profiles are distinct from human embryonic stem cells (ESCs), induced-pluripotent stem cells (iPSCs), and newborn foreskin fibroblasts. Amniocytes have a complex molecular signature, coexpressing trophoblastic, ectodermal, mesodermal, and endodermal cell-type-specific regulators. In contrast to the current view of the ground state of stem cells, ESCs and iPSCs also express high levels of a wide range of cell-type-specific regulators. The coexpression of multilineage differentiation markers combined with the strong expression of a subset of ES cell repressors in amniocytes suggests that these cells have a distinct phenotype that is unlike any other known cell-type or lineage.
    Full-text · Article · Jan 2013 · PLoS ONE
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    ABSTRACT: Mesenchymal stromal/stem cells (MSCs) are widely distributed in different tissues such as bone marrow, adipose tissues, peripheral blood, umbilical cord and amnionic fluid. Recently, MSC-like cells were also found to exist in rat olfactory bulb and are capable of inducing differentiation into mesenchymal lineages - osteocytes, chondrocytes and adipocytes. However, whether these cells can differentiate into myocardial cells is not known. In this study, we examined whether olfactory bulb-derived MSCs could differentiate into myocardial cells in vitro. Fibroblast-like cells isolated from the olfactory bulb of neonatal rats were grown under four conditions: no treatment; in the presence of growth factors (neuregulin-1, bFGF and forskolin); co-cultured with cardiomyocytes; and co-cultured with cardiomyocytes plus neuregulin-1, bFGF and forskolin. Cell differentiation into myocardial cells was monitored by RT-PCR, light microscopy immunofluorescence, western blot analysis and contractile response to pharmacological treatments. The isolated olfactory bulb-derived fibroblast-like cells expressed CD29, CD44, CD90, CD105, CD166 but not CD34 and CD45, consistent with the characteristics of MSCs. Long cylindical cells that spontaneously contracted were only observed following 7 days of co-culture of MSCs with rat cardiomyocytes plus neuregulin-1, bFGF and forskolin. RT-PCR and western blot analysis indicated that the cylindrical cells expressed myocardial markers, such as Nkx2.5, GATA4, sarcomeric α-actinin, cardiac troponin I, cardiac myosin heavy chain, atrial natriuretic peptide and connexin 43. They also contained sarcomeres and gap junction and were sensitive to pharmacological treatments (adrenal and cholinergic agonists and antagonists). These findings indicate that rat olfactory bulb-derived fibroblast-like cells with MSC characteristics can differentiate into myocardial-like cells. Copyright © 2013 John Wiley & Sons, Ltd.
    No preview · Article · Mar 2013 · Journal of Tissue Engineering and Regenerative Medicine
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