Comparison of day 40 cardiomyocyte spheroids and adult heart gene expression profiles.

(A) Human Affymetrix exon microarray data for Neor undifferentiated hESCs and Puror CSs (isolated as in Figure 6a and analyzed at day 40 of culture) were combined with a previously described neural differentiation dataset (Cythera, HUES6 and fetal hCNS stem cells) in addition to adult tissue samples using the RMA algorithm (ExpressionConsole 1.0). A non-clustered heatmap of well-characterized cardiac and embryonic stem cell pluripotency genes are displayed for all combined differentiation and adult tissues, relative to their respective undifferentiated hESC controls (adult tissue compared to Neor hESCs from this study). (B) Individual array fold changes for day 40 CS and adult heart, relative to Neor undifferentiated hESCs were clustered using the HOPACH algorithm (Bioconductor), for 3030 genes up or down-regulated in the day 40 CS comparison. Genes clustered into one of nine top-level clusters (indicated), where red and green indicate gene up- and down-regulation, respectively, relative to undifferentiated hESCs. (C–E) Pathway over-representation analysis with the tool GO-Elite of the 1466 and 2502 upregulated genes in day 40 CS and adult heart samples, respectively, relative to undifferentiated hESCs as a function of the percentage of the genes changed per GO term. Cellular cardiac differentiation and complex tissue developmental pathways were enriched in both samples (C). Genes disproportionately enriched in adult heart aligned with in vivo tissue processes typical of whole heart and endothelial function (D). Genes disproportionately enriched in day 40 CS samples (E) included stem cell and early developmental processes.