Adult Multipotent Cardiac Progenitor-Derived Spheroids: A Reproducible Model of In Vitro Cardiomyocyte Commitment and Specification
Author:
Scalise Mariangela1, Marino Fabiola1ORCID, Salerno Luca1, Amato Nunzia2, Quercia Claudia2, Siracusa Chiara2, Filardo Andrea2, Chiefalo Antonio1, Pagano Loredana2, Misdea Giuseppe1, Salerno Nadia1, De Angelis Antonella3, Urbanek Konrad4, Viglietto Giuseppe1, Torella Daniele1ORCID, Cianflone Eleonora2ORCID
Affiliation:
1. Department of Experimental and Clinical Medicine, Magna Graecia University, 88100 Catanzaro, Italy 2. Department of Medical and Surgical Sciences, Magna Graecia University, 88100 Catanzaro, Italy 3. Department of Experimental Medicine, University of Campania “L. Vanvitelli”, 80138 Naples, Italy 4. Department of Molecular Medicine and Medical Biotechnology, Federico II University, 88121 Naples, Italy
Abstract
Background: Three-dimensional cell culture systems hold great promise for bridging the gap between in vitro cell-based model systems and small animal models to study tissue biology and disease. Among 3D cell culture systems, stem-cell-derived spheroids have attracted significant interest as a strategy to better mimic in vivo conditions. Cardiac stem cell/progenitor (CSC)-derived spheroids (CSs) provide a relevant platform for cardiac regeneration. Methods: We compared three different cell culture scaffold-free systems, (i) ultra-low attachment plates, (ii) hanging drops (both requiring a 2D/3D switch), and (iii) agarose micro-molds (entirely 3D), for CSC-derived CS formation and their cardiomyocyte commitment in vitro. Results: The switch from a 2D to a 3D culture microenvironment per se guides cell plasticity and myogenic differentiation within CS and is necessary for robust cardiomyocyte differentiation. On the contrary, 2D monolayer CSC cultures show a significant reduced cardiomyocyte differentiation potential compared to 3D CS culture. Forced aggregation into spheroids using hanging drop improves CS myogenic differentiation when compared to ultra-low attachment plates. Performing CS formation and myogenic differentiation exclusively in 3D culture using agarose micro-molds maximizes the cardiomyocyte yield. Conclusions: A 3D culture system instructs CS myogenic differentiation, thus representing a valid model that can be used to study adult cardiac regenerative biology.
Funder
Ministry of Education, Universities and Research Ministero della Salute
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