Generation and characterization of cardiac valve endothelial-like cells from human pluripotent stem cells

Linxi Cheng,Yu Song,Weihua Qiao,Yingchao Geng,Nianguo Dong,Yuhua Sun,Kai Huang,Yanyong Zhang,Minghui Xie,Lin Wang,Zheng Wang,Weilin Xu

doi:10.1038/s42003-021-02571-7

Abstract

The cardiac valvular endothelial cells (VECs) are an ideal cell source that could be used for making the valve organoids. However, few studies have been focused on the derivation of this important cell type. Here we describe a two-step chemically defined xeno-free method for generating VEC-like cells from human pluripotent stem cells (hPSCs). HPSCs were specified to KDR+/ISL1+ multipotent cardiac progenitors (CPCs), followed by differentiation into valve endothelial-like cells (VELs) via an intermediate endocardial cushion cell (ECC) type. Mechanistically, administration of TGFb1 and BMP4 may specify VEC fate by activating the NOTCH/WNT signaling pathways and previously unidentified targets such as ATF3 and KLF family of transcription factors. When seeded onto the surface of the de-cellularized porcine aortic valve (DCV) matrix scaffolds, hPSC-derived VELs exhibit superior proliferative and clonogenic potential than the primary VECs and human aortic endothelial cells (HAEC). Our results show that hPSC-derived valvular cells could be efficiently generated from hPSCs, which might be used as seed cells for construction of valve organoids or next generation tissue engineered heart valves.

Highlights

The cardiac valvular endothelial cells (VECs) are an ideal cell source that could be used for making the valve organoids
Western blot analysis confirmed that ISL1 and KDR were abundantly expressed in day 3 human pluripotent stem cells (hPSCs)-derived cardiac progenitor cells (CPCs) (Fig. 1e)
The results demonstrated that endothelialization occurred and cell proliferated after co-culture of de-cellularized porcine aortic valve (DCV) and hPSC-derived valve endothelial-like cells (VELs), and that hPSC-derived VELs exhibited superior proliferative and clonogenic potential than the primary VECs and human aortic endothelial cells (HAEC)

Summary

Introduction

The cardiac valvular endothelial cells (VECs) are an ideal cell source that could be used for making the valve organoids. HPSCs were specified to KDR+/ISL1+ multipotent cardiac progenitors (CPCs), followed by differentiation into valve endothelial-like cells (VELs) via an intermediate endocardial cushion cell (ECC) type. When seeded onto the surface of the de-cellularized porcine aortic valve (DCV) matrix scaffolds, hPSC-derived VELs exhibit superior proliferative and clonogenic potential than the primary VECs and human aortic endothelial cells (HAEC). Previous studies have shown that components of multiple signaling pathways (such as BMP, FGF, WNT, NOTCH, and TGF-β) are expressed in a spatial-temporal manner during endocardial cushion formation and EndoMT12–18, suggesting that these signaling pathways may play key roles in ECC induction and valvular cell formation. When hPSC-derived VELs were seeded onto the surface of the de-cellularized porcine aortic valve (DCV) matrix scaffolds, they exhibit superior proliferative and clonogenic potential than the primary VECs and HAEC

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Conclusion