Fig. 7.

Characterization of human type II cells cultured in ALI system compare to adding WNT inhibitors at day 7 of differentiation towards type I cells (A–D) Immunofluorescence analysis of ATI markers; (A, B) show T1α and Caveolin-1 positive cells cultured in ALI system; (C, D) show T1α and Caveolin-1 positive cells cultured in media supplemented with IWR-1. Scale bar, 63 µm (E) Flow cytometric analysis of T1α and caveolin-1 during ALI–mediated and IWR-1–mediated induction of type I phenotype in human ATII cells at day 7 (compare to hATI cells stained with corresponding isotype). (F) Comparison of mRNA expression of T1α, AQ5, caveolin-1 and SPC in human ATII cells cultured in ALI system compared to cells submerged in media and submerged in media+IWR-1. Data expressed as quantification of mRNA normalized to GAPDH and compared to hATII cells that were derived from fresh human lung. Fold change of GOI transcript levels between iPS derived-ATII and human type II cells equals 2^−ΔΔCt, where ΔCt = Ct_(GOI) Ct_(GAPDH), and ΔΔCt = ΔCt_(AETII) - ΔCt_(ATII). Bar indicate ± SEM and n= 3 independent experiments for qRT-PCR and flow cytometry. ‘a’ marks statistically significant difference (p < 0.05) when comparing gene expression in cells cultured in ALI vs. cells submerged in media. ‘b’ marks statistically significant difference (p < 0.05) in gene expression in cells exposed to IWR-1 vs. cells submerged in media.