Although correlations between the differentiation capacity of hPSCs and these genetic variations have not been systematically examined, it is feasible that cellular pluripotency and differentiation propensity may differ in different hPSC lines partially due to intrinsic genetic variation that alters cell signaling mediated by protein phosphorylation.
We have successfully used these protocols in four different hPSC lines, including human embryonic stem cells and induced pluripotent stem cells.
The variation in cardiac differentiation potential among different hPSC lines cultured under similar conditions has been well documented [15], [20], [21], [35].
Variations in protein expression and the phosphorylation state of different hPSC lines may affect their responses to environmental stimuli.
To this end, a singularized-cell seeding protocol was established to control more precisely the starting hPSC density, leading to the identification of an optimum day 0 seeding density of 30,000 cells/cm that served to maximize yield and barrier tightness for three different hPSC lines.
Considering the increasing awareness of the broad influence of genetic background (Montgomery et al, 2010) and the biological differences between hPSC lines (Boulting et al, 2011), our experimental system may overcome some of the shortcomings of conventional hiPSC approaches in identifying disease-related phenotypes.
These trisomies appear to occur stochastically between different hNPC lines.
We found variability in the efficiency of differentiation using these protocols between various hPSC lines, indicating the need for line-by-line optimization.
Experiments in additional hPSC lines verified that our monolayer differentiation protocol, consisting of BMP4/GSK-3βI administration in phase 1 and VEGF-A/DAPT administration in phase 2, resulted in robust endothelial differentiation with minimal differences in efficiency between different cell lines that did not reach statistical significance (Supplementary information, Figure S2F).
Between different therapy lines, no difference was observed (DCR second-line 58.6%, third-line 69.7%, fourth-line and more-line 75%, P = 0.542).
