Comparative analysis of cardiomyocyte differentiation from human embryonic stem cells under 3-D and 2-D culture conditions.

Pal , Rajarshi and Mamidi, Murali Krishna and Das, Anjan Kumar and Bhonde, Ramesh (2013) Comparative analysis of cardiomyocyte differentiation from human embryonic stem cells under 3-D and 2-D culture conditions. Journal of Bioscience and Bioengineering, 115 (2). pp. 200-206. ISSN 1389-1723

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Abstract

Post-myocardial infarction cardiomyocytes are the most important target cell types for cardiac repair. Many of the applications envisaged for human embryonic stem cells hESC)-derived cardiomyocytes demand that the differentiation procedure be robust, cost effective and high yielding. Various lines of evidence including our earlier study uggest that hESCs have distinct preferences to become heart cells. However, a direct comparison between different protocols has not yet been reported to date. Here, we performed a logical and systematic comparison of ardiomyocytes obtained from hESCs via embryoid bodies (EBs) in suspension versus adherent static cultures of feeder-free hES colonies representing three-dimensional (3-D) and two-dimensional (2-D) culture systems, respectively. An in-depth characterization of the beating cells revealed appropriate cardiac marker expression both at gene and protein levels. Despite using similar media, 3-D and 2-D cultures showed significant variation in growth and ability to form beating areas. While the expression of pre-cardiac mesoderm markers like GATA-4, HAND1, Myf5, Msx1, and BMP-IIR remained unaltered; levels of functional heart-specific markers such as MLC-2A/2V, cTnT, ANP, Phospholamban, a-MHC and KV4.3 were substantially up-regulated in 3-D compared to 2-D cultures. Concurrently we observed a sharp decline in the expression of ESC, ectoderm and endodermmarkers including Oct-4, Sox-2, NFH, Sox-1, Sox-17 and AFP. Further immunocytochemistry and flow cytometry demonstrated a higher percentage of cells positive for Brachyury, desmin and cardiac troponin in 3-D cultures. Our results underscore the higher efficiency of cardiomyocytes derived via 3-D cultures. This finding enriches our basic understanding of the differentiation pattern in hESC-derived cardiomyocytes.

Item Type: Article
Additional Information: Copy right of this article is belongs to Elsevier Science.
Uncontrolled Keywords: Human embryonic stem cells(hESCs); Differentiation;Cardiomyocytes;Cell therapy;3-D and 2-D culture system.
Subjects: Regenerative Medicine > MIRM Bangalore
Depositing User: MCON Library
Date Deposited: 13 Apr 2013 09:22
Last Modified: 13 Apr 2013 09:22
URI: http://eprints.manipal.edu/id/eprint/79691

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