The biochemical and electrophysiological profiles of amniotic fluid-derived stem cells following Wnt signaling modulation cardiac differentiation

Yen-Wen Liu,Ping-Yen Liu,Chi-Ting Su,Sheng-Nan Wu,Shiaw-Min Hwang,Yi-Hsein Fang

doi:10.1038/s41420-019-0143-0

Abstract

Owing to the beneficial properties of amniotic fluid-derived stem cells (AFSCs), including pluripotency and the lack of ethical issues associated with embryonic stem cells (ESCs), they should be a promising cell source for regenerative medicine. However, how to differentiate AFSCs into contracting cardiomyocytes has not been established. In this study, a well-established, direct cardiac differentiation protocol involving the modulation of Wnt signaling was used to differentiate Oct 3/4+ AFSCs into cardiomyocytes. By day 14 of cardiomyocyte differentiation, these AFSCs expressed cardiac-specific genes (i.e., cardiac troponin T and myosin light chain 2v) and proteins but could not spontaneously contract. Using the patch-clamp technique, we further characterized the electrophysiological properties of human ESC-derived cardiomyocytes (hESC-CMs) and differentiated AFSCs. We used different configurations to investigate membrane potentials and ion currents in differentiated AFSCs and hESC-CMs. Under cell-attached voltage- or whole-cell current-clamp modes, we recorded spontaneous action currents (ACs) or action potentials (APs) in hESC-CMs but not in differentiated AFSCs. Compared to hESC-CMs, differentiated AFSCs showed significantly diminished activity of both BKCa and IKCa channels, which might lead to a lack of spontaneous ACs and APs in differentiated AFSCs. These results indicated that this well-established Wnt signaling modulating cardiac differentiation protocol was insufficient to induce the differentiation of functional cardiomyocytes from Oct 3/4+ AFSCs. Therefore, AFSC may not be an ideal candidate for cardiomyocyte differentiation.

Highlights

After severe myocardial injury, such as myocardial infarction, the regenerative ability of mammalian hearts is very limited,[1] which may lead to impaired cardiac systolic function, heart failure or even death
Flow cytometry indicated that undifferentiated amniotic fluid-derived stem cells (AFSCs) and Human embryonic stem cells (hESCs) expressed the pluripotent stem cell markers, i.e., Nanog, Oct3/4, and SSEA4 (Table 1; Fig. 1b)
At cardiac differentiation day 14, the expression of these 3 pluripotent stem cell markers significantly reduced in both differentiated AFSCs and hESC-CMs (Table 1; Fig. 1b)

Summary

Introduction

After severe myocardial injury, such as myocardial infarction, the regenerative ability of mammalian hearts is very limited,[1] which may lead to impaired cardiac systolic function, heart failure or even death. Post-infarct cardiac contractility could be restored by replacing scar. It was reported that exogenous bone-marrow-derived ckit+ hematopoietic stem cells[3] and endogenous c-kit+ cardiac progenitor cells[4] restored the infarcted myocardium, supporting the concept that stem cells may be effective for cardiac regeneration. Several studies have shown that c-kit+ stem cells, including hematopoietic stem cells and cardiac progenitor cells, do not efficiently differentiate into cardiomyocytes.[5,6,7] over the last decade, hundreds of patients have received c-kit+ stem cell therapy, with conflicting results regarding the improvement in cardiac function.[8,9,10,11,12,13].

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