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Generation of NKX2.5GFP Reporter Human iPSCs and Differentiation Into Functional Cardiac Fibroblasts

Authors :
Leyre López-Muneta
Javier Linares
Oscar Casis
Laura Martínez-Ibáñez
Arantxa González Miqueo
Jaione Bezunartea
Ana Maria Sanchez de la Nava
Mónica Gallego
María Eugenia Fernández-Santos
Juan Roberto Rodriguez-Madoz
Xabier L. Aranguren
Francisco Fernández-Avilés
José Carlos Segovia
Felipe Prósper
Xonia Carvajal-Vergara
Source :
Frontiers in Cell and Developmental Biology, Vol 9 (2022)
Publication Year :
2022
Publisher :
Frontiers Media S.A., 2022.

Abstract

Direct cardiac reprogramming has emerged as an interesting approach for the treatment and regeneration of damaged hearts through the direct conversion of fibroblasts into cardiomyocytes or cardiovascular progenitors. However, in studies with human cells, the lack of reporter fibroblasts has hindered the screening of factors and consequently, the development of robust direct cardiac reprogramming protocols.In this study, we have generated functional human NKX2.5GFP reporter cardiac fibroblasts. We first established a new NKX2.5GFP reporter human induced pluripotent stem cell (hiPSC) line using a CRISPR-Cas9-based knock-in approach in order to preserve function which could alter the biology of the cells. The reporter was found to faithfully track NKX2.5 expressing cells in differentiated NKX2.5GFP hiPSC and the potential of NKX2.5-GFP + cells to give rise to the expected cardiac lineages, including functional ventricular- and atrial-like cardiomyocytes, was demonstrated. Then NKX2.5GFP cardiac fibroblasts were obtained through directed differentiation, and these showed typical fibroblast-like morphology, a specific marker expression profile and, more importantly, functionality similar to patient-derived cardiac fibroblasts. The advantage of using this approach is that it offers an unlimited supply of cellular models for research in cardiac reprogramming, and since NKX2.5 is expressed not only in cardiomyocytes but also in cardiovascular precursors, the detection of both induced cell types would be possible. These reporter lines will be useful tools for human direct cardiac reprogramming research and progress in this field.

Details

Language :
English
ISSN :
2296634X
Volume :
9
Database :
Directory of Open Access Journals
Journal :
Frontiers in Cell and Developmental Biology
Publication Type :
Academic Journal
Accession number :
edsdoj.18096c79bf8a442fbff2dd48d63ad000
Document Type :
article
Full Text :
https://doi.org/10.3389/fcell.2021.797927