Microbioreactor Arrays for Full Factorial Screening of Exogenous and Paracrine Factors in Human Embryonic Stem Cell Differentiation
PLoS ONE, ISSN: 1932-6203, Vol: 7, Issue: 12, Page: e52405
2012
- 44Citations
- 75Captures
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Example: if you select the 1-year option for an article published in 2019 and a metric category shows 90%, that means that the article or review is performing better than 90% of the other articles/reviews published in that journal in 2019. If you select the 3-year option for the same article published in 2019 and the metric category shows 90%, that means that the article or review is performing better than 90% of the other articles/reviews published in that journal in 2019, 2018 and 2017.
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Metrics Details
- Citations44
- Citation Indexes44
- 44
- CrossRef38
- Captures75
- Readers75
- 75
Article Description
Timed exposure of pluripotent stem cell cultures to exogenous molecules is widely used to drive differentiation towards desired cell lineages. However, screening differentiation conditions in conventional static cultures can become impractical in large parameter spaces, and is intrinsically limited by poor spatiotemporal control of the microenvironment that also makes it impossible to determine whether exogenous factors act directly or through paracrine-dependent mechanisms. We detail here the development of a continuous flow microbioreactor array platform that combines full-factorial multiplexing of input factors with progressive accumulation of paracrine factors through serially-connected culture chambers, and further, the use of this system to explore the combinatorial parameter space of both exogenous and paracrine factors involved in human embryonic stem cell (hESC) differentiation to a MIXL1-GFP primitive streak-like population. We show that well known inducers of primitive streak (BMP, Activin and Wnt signals) do not simply act directly on hESC to induce MIXL1 expression, but that this requires accumulation of surplus, endogenous factors; and, that conditioned medium or FGF-2 supplementation is able to offset this. Our approach further reveals the presence of a paracrine, negative feedback loop to the MIXL1-GFP population, which can be overcome with GSK-3β inhibitors (BIO or CHIR99021), implicating secreted Wnt inhibitory signals such as DKKs and sFRPs as candidate effectors. Importantly, modulating paracrine effects identified in microbioreactor arrays by supplementing FGF-2 and CHIR in conventional static culture vessels resulted in improved differentiation outcomes. We therefore demonstrate that this microbioreactor array platform uniquely enables the identification and decoding of complex soluble factor signalling hierarchies, and that this not only challenges prevailing strategies for extrinsic control of hESC differentiation, but also is translatable to conventional culture systems. © 2012 Titmarsh et al.
Bibliographic Details
10.1371/journal.pone.0052405; 10.1371/journal.pone.0052405.g004; 10.1371/journal.pone.0052405.g002; 10.1371/journal.pone.0052405.g001; 10.1371/journal.pone.0052405.g003
http://www.scopus.com/inward/record.url?partnerID=HzOxMe3b&scp=84871593878&origin=inward; http://dx.doi.org/10.1371/journal.pone.0052405; http://www.ncbi.nlm.nih.gov/pubmed/23300662; https://dx.plos.org/10.1371/journal.pone.0052405.g004; http://dx.doi.org/10.1371/journal.pone.0052405.g004; https://dx.plos.org/10.1371/journal.pone.0052405; https://dx.plos.org/10.1371/journal.pone.0052405.g002; http://dx.doi.org/10.1371/journal.pone.0052405.g002; https://dx.plos.org/10.1371/journal.pone.0052405.g001; http://dx.doi.org/10.1371/journal.pone.0052405.g001; https://dx.plos.org/10.1371/journal.pone.0052405.g003; http://dx.doi.org/10.1371/journal.pone.0052405.g003; https://dx.doi.org/10.1371/journal.pone.0052405.g003; https://journals.plos.org/plosone/article/figure?id=10.1371/journal.pone.0052405.g003; https://dx.doi.org/10.1371/journal.pone.0052405.g004; https://journals.plos.org/plosone/article/figure?id=10.1371/journal.pone.0052405.g004; https://dx.doi.org/10.1371/journal.pone.0052405.g001; https://journals.plos.org/plosone/article/figure?id=10.1371/journal.pone.0052405.g001; https://dx.doi.org/10.1371/journal.pone.0052405; https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0052405; https://dx.doi.org/10.1371/journal.pone.0052405.g002; https://journals.plos.org/plosone/article/figure?id=10.1371/journal.pone.0052405.g002; http://dx.plos.org/10.1371/journal.pone.0052405.g001; http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0052405; https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0052405&type=printable; http://dx.plos.org/10.1371/journal.pone.0052405.g003; http://dx.plos.org/10.1371/journal.pone.0052405.g002; http://dx.plos.org/10.1371/journal.pone.0052405; http://www.plosone.org/article/metrics/info:doi/10.1371/journal.pone.0052405; http://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0052405&type=printable; http://dx.plos.org/10.1371/journal.pone.0052405.g004; http://europepmc.org/abstract/med/23300662; http://europepmc.org/articles/PMC3530582; http://journals.plos.org/plosone/article?id=10.1371%2Fjournal.pone.0052405
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