Figure 1. Overexpressing Oct-4 in CAR⁺/mPSCs.

A. Procedure to overexpress Oct-4 in CAR⁺/mPSCs. (i), Representative phase contrast image of primary culture. The magnified image shows the epithelial colony of mPSCs. (ii), Isolation of mPSCs according to CAR-positive expression of primary cultures using flow cytometry and subsequent transfection with retroviral vectors encoding Oct-4 cDNA. (iii), Transfected CAR⁺/mPSCs were co-cultivated with feeder cells at day 2. Cobblestone-like colonies were observed at day 21. The magnified image shows the morphology of one colony. (iv), Isolation and expansion of individual cobblestone-like colonies at day 28. (v), Colonies were established as cell clones, comprising C1, E9, and C7 clones. Representative morphology images of the C1 clone. (Scale bar, 100 μm.) B. Frequency of cobblestone-like colony formation. Data are presented as mean ± SD. CAR⁺/mPSCs with sham control transfection or in CAR⁺/mPSCs-derived type-I pneumocytes transfected with Oct-4 were no detectable (N.D.) colony formation.