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Cell Biology International (2010) 34, 1021–1031 (Printed in Great Britain)

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Intrinsic properties and external factors determine the differentiation bias of human embryonic stem cell lines

Ashish Mehta, Sheena Mathew, Chandra Viswanathan and Anish Sen Majumdar¹

Regenerative Medicine, Reliance Life Sciences Pvt. Ltd., ThaneBelapur Road, Rabale, Navi Mumbai400 701, India

A major goal of human embryonic stem cell (hESC) research is to regulate differentiation through external means to generate specific cell types with high purity for regenerative medicine applications. Although all hESC lines express pluripotency-associated genes, their differentiation ability to various lineages differs considerably. We have compared spontaneous differentiation propensity of the two hESC lines, RelicellhES1 and BG01. Spontaneous differentiation of hESC lines grown in different media conditions was followed by differentiation using two methods. Kinetic data generated by real-time gene expression studies for differentiated cell types were analyzed, and confirmed at protein levels. Both cell lines showed upregulation of genes associated with the 3 germ layers, although stark contrast was evident in the magnitude of upregulation of lineage specific genes. A distinct difference was also found in the rate at which the pluripoteny factors, Oct-4 and Nanog, were downregulated during differentiation. Once differentiation was initiated, both Oct-4 and Nanog gene expression was drastically reduced in RelicellhES1, whereas a gradual decrease was observed in BG01. A clear trend is seen in RelicellhES1 to differentiate into neuroectodermal and mesenchymal lineages, whereas BG01 cells are more prone to mesoderm and endoderm development. In addition, suspension versus plated methods of cell culture significantly influenced the outcome of differentiation of certain types of cells. Results obtained by spontaneous differentiation of hESCs were also amplified by induced differentiation. Thus, differential rates of downregulation of pluripotency markers along with culture conditions seem to play an important role in determining the developmental bias of human ES cell lines.

Key words: differentiation, endoderm, human embryonic stem cell, pluripotency, propensity, neuroectoderm

Abbreviations: DMEM, Dulbecco's modified Eagle's medium, EB, embryoid body, hESC, human embryonic stem cell, MEF, mouse embryonic fibroblast, qRT-PCR, quantitative real time reverse transcriptase polymerase chain reaction

¹To whom correspondence should be addressed (email anish_senmajumdar@relbio.com).

Received 23 April 2010/16 June 2010; accepted 30 June 2010

Published as Cell Biology International Immediate Publication 30 June 2010, doi:10.1042/CBI20100283

© The Author(s) Journal compilation © 2010 Portland Press Limited