From left: Dr Lee Dong-Yup, Yeo Hock Chuan, Sherwin Ting, Dr Steve Oh
Hock Chuan Yeo1,2, Sherwin Ting1, Romulo Martin Brena3, Geoffrey Koh1, Allen Chen1, Siew Qi Toh2, Yu Ming Lim1, Steve Kah Weng Oh1 & Dong-Yup Lee1,2,4
1 Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), Singapore
2 Department of Chemical and Biomolecular Engineering, National University of Singapore, Singapore
3 USC Epigenome Center, University of Southern California, USA
4 NUS Synthetic Biology for Clinical and Technological Innovation (SynCTI), Life Sciences Institute, National University of Singapore, Singapore
Published in Scientific Reports 2016 6: 31068 (Online Version)
The advent of induced pluripotent stem cells has accelerated the growth of personalized medicine. Researchers can now take a personal sample of skin cells and ‘reset’ them into the pluripotent state. The reprogrammed cells can then be differentiated into any desired cell-type, in order to replace donor tissues that are damaged, test drug efficacies and chemical toxicity, and to study unique disease progression. While there have been remarkable developments in the generation of heart muscle cells, there has been a lack of understanding on the interactions between cell lines, culture platforms and differentiation protocols.
In this study, the authors investigate such interplay by comparing the same pluripotent stem cells grown under three different culture conditions (Figure 1). Besides conventional culture using a layer of mouse feeder cells on Matrigel coating, they also designed an in-between condition whereby cells grown on Matrigel were then replated on feeders. Differentiation efficiencies were then quantified using a protocol that has worked well for feeder-based cells but not those grown on Matrigel. To explain the contrasting outcomes of these cell cultures, the authors analyzed the global silencing of genes via ‘methylation’ signals marked on DNA, the DNA binding sites of gene regulators called transcription factors (TFs), and the resulting gene expressions. Through bioinformatics analysis, crucial changes in cellular signaling and the downstream expression of ‘Forkhead box’ (FOX) TFs were determined to mark the differentiation potential of pluripotent stem cells. The authors further demonstrated the correlative utility of such FOX markers in the differentiation of two distinct cell lines. The study underpinned the importance of computational biology elucidating biological phenomena, and the authors envision even closer collaboration between experimentalists and computational biologists for such studies.
Figure 1. Experimental design involving different cell lines, culture platforms and differentiation protocols.
Factors determining the efficiency of heart muscle generation are identified by profiling and analysing the interplay between differentiation outcomes, global gene expressions (transcriptome), the DNA binding sites of transcription factors, and gene silencing by DNA-methylation. The hypothesis is then tested in a different set of conditions.