High-efficiency derivation of human embryonic stem cell lines using a culture system with minimized trophoblast cell proliferation

Abstract

BackgroundDue to their extensive self-renewal and multilineage differentiation capacity, human embryonic stem cells (hESCs) have great potential for studying developmental biology, disease modeling, and developing cell replacement therapy. The first hESC line was generated in 1998 by culturing inner cell mass (ICM) cells isolated from human blastocysts using an immunosurgery technique. Since then, many techniques including mechanical ICM isolation, laser dissection, and whole embryo culture have been used to derive hESC lines. However, the hESC derivation efficiency remains low, usually less than 50%, and it requires a large number of human embryos to derive a significant number of hESC lines. Due to a shortage of and restricted access to human embryos, a novel approach with better hESC derivation efficiency is badly needed to decrease the number of embryos used.MethodsWe hypothesized that the low hESC derivation efficiency might be due to extensive proliferation of trophoblast (TE) cells which could interfere with ICM proliferation. We therefore developed a methodology to minimize TE cell proliferation by culturing ICM in a feeder-free system for 3 days before transferring them onto feeder cells.ResultsThis minimized trophoblast cell proliferation (MTP) technique could be successfully used to derive hESCs from normal, abnormal, and frozen–thawed embryos with better derivation efficiency of more than 50% (range 50–100%; median 70%).ConclusionsWe successfully developed a better hESC derivation methodology using the “MTP” culture system. This methodology can be effectively used to derive hESCs from both normal and abnormal embryos under feeder-free conditions with higher efficiency when compared with other methodologies. With this methodology, large-scale production of clinical-grade hESCs is feasible.Electronic supplementary materialThe online version of this article (10.1186/s13287-018-0866-5) contains supplementary material, which is available to authorized users.

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“…Stem cells can undergo immunomodulation, provide anti-inflammatory responses, and repair and regenerate injured tissues [24,25]. Stem cells are generally of two major types based on their origin: embryonic stem cells (ESCs) [26] and adult stem cells. Human embryonic stem cells are pluripotent in origin, while adult stem cells are multipotent cells that have the potential to differentiate into particular cell lineages.…”

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“…The efficiency of derivation, in other words, the number of times inner cell mass is plated onto feeder cells and the number of times an ES cell line is generated is relatively low. The reported derivation efficiencies of ES cell lines vary from 50 to 100 per cent [1]. As passaging of the ES cell line is the most critical activity during ongoing derivations, a manual passaging method has given rise to the high percentage of undifferentiated colonies and continuity of the cell line during critical early passages [2].…”

Role of YAP as a Mechanosensing Molecule in Stem Cells and Stem Cell-Derived Hematopoietic Cells

“…Unlike adult and perinatal stem cells with limited multipotent differentiation capacity, PSCs (ESCs and iPSCs) can self-renew and differentiate into all cell types in the body, including blood cells [ 62 ]. ESCs are derived from the inner cell mass of an embryo [ 63 ].…”