Abstract
Human amniotic fluid (hAF) cells share characteristics of both embryonic and adult stem cells. They proliferate rapidly and can differentiate into cells of all embryonic germ layers but do not form teratomas. Embryoid-bodies obtained from hAF have cardiac differentiation potential, but terminal differentiation to cardiomyocytes (CMs) has not yet been described. Our purpose was to promote cardiac differentiation in hAFcells. Cells were exposed to inducing factors for up to 15 days. Only the subset of hAF cells expressing the multipotency markers SSEA4, OCT4 and CD90 (CardiopoieticAF cells) responded to the differentiation process by increasing the expression of the cardiac transcription factors Nkx2.5 and GATA4, sarcomeric proteins (cTnT, α-MHC, α-SA), Connexin43 and atrial and ventricular markers. Furthermore, differentiated cells were positive for the calcium pumps CACNA1C and SERCA2a, with approximately 30% of CardiopoieticAF-derived CM-like cells responding to caffeine or adrenergic stimulation. Some spontaneous rare beating foci were also observed. In conclusion, we demonstrated that CardiopoieticAF cells might differentiate toward the cardiac lineage giving rise to CM-like cells characterized by several cardiac-specific molecular, structural, and functional properties.
Highlights
Cardiovascular (CV) diseases are the main cause of mortality in the industrialized world, with an estimated 17.7 million deaths by CV in 2015, representing 31% of all global deaths[1]
As in previous studies[15,18,20], we demonstrated that they expressed the mesenchymal stem cell markers CD29, CD73 and CD44, whereas they were negative for the hematopoietic markers CD34 and CD45 and for the endothelial marker PECAM-1/CD31
It has already been described that human amniotic fluid (hAF) cells, such as ESCs, can give rise to EBs16, three-dimensional aggregates representing one of the hallmarks of pluripotent stem cells and an optimal starting point for in vitro lineage-specific differentiation
Summary
Cardiovascular (CV) diseases are the main cause of mortality in the industrialized world, with an estimated 17.7 million deaths by CV in 2015, representing 31% of all global deaths[1]. Different types of stem cells have already been shown to have cardiomyogenic potential[8,9]: For example, embryonic stem (ES) cells and induced pluripotent stem (iPS) cells can be differentiated into beating cells with a cardiac-like phenotype in vitro. The aim of this study was to dissect and promote the cardiomyogenic potential of hAF cells to obtain differentiated progenies with morphological and functional features of CM. For this purpose, 15 hAF samples were analyzed for the expression of embryonic markers and induced to cardiac differentiation through two differentiation protocols: one based on Embryoid Body (EB) formation and the other in which cell monolayers were sequentially exposed to known cardiac differentiation inducers
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