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Another Step towards the Fully Synthetic Culture of Human Pluripotent Stem Cells

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Review of “Chemically defined and growth-factor-free culture system for the expansion and derivation of human pluripotent stem cells” from Nature Biomedical Engineering by Stuart P. Atkinson

The widespread clinical application of human pluripotent stem cells (hPSCs) and their derivatives will require a means to maintain and expand cells employing cost-effective, chemically defined, and xeno-free conditions. Towards this goal, a new study from the laboratory of Kouichi Hasegawa (Kyoto University, Japan) builds on a previous STEM CELLS Translational Medicine study [1] and now reports a novel defined culture system that lowers the reliance on recombinant human proteins [2]. 

Will this new study help in the construction of a cost-effective, chemically defined, and xeno-free hPSC culture system? 

To help you find out, here are the critical points from the exciting new study by Yasuda et al.:

  • A previous study detailed a novel and simple chemically-defined xeno‐free culture system that allowed for the long‐term expansion of hPSCs without fibroblast growth factor (FGF), transforming growth factor (TGF)-β activation, xenobiotic supplements, serum, serum replacement, or albumin
    • However, the medium still included insulin, transferrin, and the expensive recombinant Wnt3a protein
  • To replace Wnt3a, the authors employed a combination of 1-azakenpaullone, ID-8, and tacrolimus to promote efficient hPSC maintenance and expansion 
    • 1-azakenpaullone inhibits Glycogen synthase kinase 3 beta (GSK3β) [3] and activates the canonical Wnt signaling pathway to support self-renewal
    • ID-8 inhibits Dual specificity tyrosine-phosphorylation-regulated kinase 1A (DYRK1) [4] and prevents the Wnt-induced differentiation caused by 1-azakenpaullone treatment 
    • Tacrolimus inhibits the activity of the Nuclear factor of activated T-cells, cytoplasmic (NFATc) proteins and recovers the growth defect induced by ID-8 treatment
  • Laminin E8 coating (confers integrin-binding activity) supported hPSC maintenance/expansion on tissue culture plastic and the authors employed phosphate buffered saline (PBS) with Ca2+ for cell passaging
  • Both short- and long-term culture employing this novel culture system supported universal hPSC growth and retention of pluripotency and multilineage differentiation potential
    • However, hPSCs grown under these conditions displayed a unique metabolic profile distinct from those described for naive or primed states of pluripotency
  • Additionally, this new culture system also supported the derivation of human induced pluripotent stem cells (hiPSCs) with an efficiency similar to growth-factor-dependent hPSC culture systems

The authors hope that their new study will provide an “optimal starting point” for the further development of a cost-effective, chemically defined, and xeno-free hPSC culture system amenable to industrial applications. However, given the metabolic differences observed, they also see the elucidation of the mechanisms underlying pluripotency, proliferation, survival, and metabolism of hPSCs cultured in these conditions as a potentially informative next step.

For more on the next steps towards the synthetic culture of human pluripotent stem cells, stay tuned to the Stem Cells Portal.

References

  1. Hasegawa K, Yasuda S-y, Teo J-L, et al., Wnt Signaling Orchestration with a Small Molecule DYRK Inhibitor Provides Long‐Term Xeno‐Free Human Pluripotent Cell Expansion. STEM CELLS Translational Medicine 2012;1:18-28.
  2. Yasuda S-y, Ikeda T, Shahsavarani H, et al., Chemically defined and growth-factor-free culture system for the expansion and derivation of human pluripotent stem cells. Nature Biomedical Engineering 2018;2:173-182.
  3. Suemori H, Yasuchika K, Hasegawa K, et al., Efficient establishment of human embryonic stem cell lines and long-term maintenance with stable karyotype by enzymatic bulk passage. Biochemical and Biophysical Research Communications 2006;345:926-932.
  4. Bellmaine SF, Ovchinnikov DA, Manallack DT, et al., Inhibition of DYRK1A disrupts neural lineage specification in human pluripotent stem cells. Elife 2017;6.