Figure 2.

Effect of IKKi Treatment on Mesenchymal Lineage Specification of hESCs

(A) Western blot of p65 and phosphorylated p65 at 0, 2, and 4 days of hESC differentiation.

(B) Western blot of IκBα and phosphorylated IκBα at 0, 2, and 4 days of hESC differentiation.

(C) Alkaline phosphatase staining and quantitative alkaline phosphatase activity assay at 2 and 4 days. Scale bar indicates 200 μm.

(D) qRT-PCR results of NANOG, OCT4, and POU5F1 at 2 and 4 days.

(E) qRT-PCR results of PAX6, PDGFR-α, BRACHYURY, and FOXA2 at 2 and 4 days.

(F) qRT-PCR results of CD73 and CD146 at 2, 4, and 7 days.

(G) Flow cytometry analysis of cells with or without IKKi treatment examining MSC marker expression.

(H) Four-color flow cytometry analysis for CD45, CD90, CD73, and CD146 expression to isolate CD73⁺CD90⁺CD146⁺CD45⁻ MSCs following 7 days of hESC differentiation with or without IKKi. Proportions of CD73⁺CD90⁺CD146⁺CD45⁻ MSCs generated. Three independent experiments were performed.

^∗p < 0.05, ^∗∗p < 0.001.