Human embryonic and induced pluripotent stem cells maintain phenotype but alter their metabolism after exposure to ROCK inhibitor

• Authors: Spyros I. Vernardis, Imperial College London; Konstantinos Terzoudis, Imperial College London; Nicki Panoskaltsis, Emory University; Athanasios Mantalaris, Imperial College London
• Journal Title: Scientific Reports
• Volume: Volume 7, Number 1
• Publisher: Nature Research (part of Springer Nature): Fully open access journals | 2017-02-06, Pages 42138-42138
• Type of Work: Article | Final Publisher PDF
• Publisher DOI: http://doi.org/10.1038/srep42138
• Subjects: Engineering, Biomedical
• Keywords: Science & Technology; Multidisciplinary Sciences; Science & Technology - Other Topics; FOCAL ADHESION KINASE; IN-VITRO; GLUCOSE-METABOLISM; CANCER-CELLS; IPS CELLS; HUMAN ES; EFFICIENT; SURVIVAL; CRYOPRESERVATION; APOPTOSIS

Abstract

Human pluripotent stem cells (hPSCs) are adhesion-dependent cells that require cultivation in colonies to maintain growth and pluripotency. Robust differentiation protocols necessitate single cell cultures that are achieved by use of ROCK (Rho kinase) inhibitors. ROCK inhibition enables maintenance of stem cell phenotype; its effects on metabolism are unknown. hPSCs were exposed to 10 μM ROCK inhibitor for varying exposure times. Pluripotency (TRA-1-81, SSEA3, OCT4, NANOG, SOX2) remained unaffected, until after prolonged exposure (96 hrs). Gas chromatography-mass spectrometry metabolomics analysis identified differences between ROCK-Treated and untreated cells as early as 12 hrs. Exposure for 48 hours resulted in reduction in glycolysis, glutaminolysis, the citric acid (TCA) cycle as well as the amino acids pools, suggesting the adaptation of the cells to the new culture conditions, which was also reflected by the expression of the metabolic regulators, mTORC1 and tp53 and correlated with cellular proliferation status. While gene expression and protein levels did not reveal any changes in the physiology of the cells, metabolomics revealed the fluctuating state of the metabolism. The above highlight the usefulness of metabolomics in providing accurate and sensitive information on cellular physiological status, which could lead to the development of robust and optimal stem cell bioprocesses.