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Selective isolation of nanog-positive human amniotic mesenchymal cells and differentiation into cardiomyocytes.
- Source :
-
Cellular reprogramming [Cell Reprogram] 2013 Feb; Vol. 15 (1), pp. 80-91. Date of Electronic Publication: 2013 Jan 08. - Publication Year :
- 2013
-
Abstract
- Adult cardiomyocytes have little ability to regenerate, thus cardiac regeneration therapy represents a potential method for treating severe heart failure. Human amniotic mesenchymal cells (hAMCs) have the potential to be a useful cell source for cardiac regeneration therapy. We attempted to isolate stem cells from hAMCs and differentiate them into cardiomyocytes. Nanog promoter-Cre plasmid and cytomegalovirus (CMV) promoter-loxP-STOP-loxP-Red-puro(r) plasmid were co-transfected into immortalized hAMCs (iHAMs). Nanog-positive iHAMs were treated with 5-azacytidine (5-aza), trichostatin A (TA), activin A (AA), and bone morphogenetic protein-4 (BMP-4), or co-cultured with murine fetal cardiomyocytes for cardiomyocytes differentiation. Isolated Nanog-positive iHAMs were analyzed by quantitative RT-PCR and immunofluorescent staining before and after differentiation. Expression of Nanog, Oct3/4, Sox2, and Klf4 was significantly higher in Nanog-positive than in Nanog-negative iHAMs. Nanog-positive iHAMs were stained for Nanog and Oct3/4 in the nucleus. Nanog-positive iHAMs treated with 5-aza expressed Nkx2.5, GATA-4, human atrial natriuretic peptide (hANP), cardiac troponin T (cTnT), myocin light chain (Mlc)-2a, Mlc-2v, β-myosin heavy chain (β-MHC), hyperpolarization-activated cyclic nucleotide gated channels (HCN)-4, and inwardly rectifying potassium channels (Kir)-2.1. Although Nanog-positive iHAMs treated with TA, AA, or BMP-4 expressed several cardiac markers, no contraction was observed. Co-cultured Nanog-positive iHAMs with murine fetal cardiomyocytes spontaneously contracted in a synchronized manner and expressed the cardiac markers. In conclusion, Nanog-positive hAMCs with characteristics of stem cells were isolated and differentiated into cardiomyocyte-like cells, suggesting that these isolated hAMCs could be a useful cell source for cardiac regeneration therapy.
- Subjects :
- Animals
Antigens, Differentiation biosynthesis
Antigens, Differentiation genetics
Coculture Techniques
Humans
Kruppel-Like Factor 4
Mice
Plasmids chemistry
Plasmids genetics
Transcription Factors biosynthesis
Transcription Factors genetics
Transfection
Amnion cytology
Amnion metabolism
Cell Differentiation
Mesenchymal Stem Cells cytology
Mesenchymal Stem Cells metabolism
Myocytes, Cardiac cytology
Myocytes, Cardiac metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 2152-4998
- Volume :
- 15
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Cellular reprogramming
- Publication Type :
- Academic Journal
- Accession number :
- 23298400
- Full Text :
- https://doi.org/10.1089/cell.2012.0028