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. 2022 Jan 30;13(5):1449–1467. doi: 10.1016/j.jcmgh.2022.01.018

Figure 5.

Figure 5

CD73+CD104+cells represent a less-differentiated and slow-cycling cell population within the basal zone. (A and B) EPC2-hTERT organoids were harvested on day 11 and dissociated for FACS. (A) The cells in EPC2-hTERT organoids were sorted based on CD73 and CD104 expression. (B) CD73, CD104, IVL, and FLG relative gene expression in CD73+CD104+ and CD73+CD104- populations was validated by qRT-PCR. (C) EPC2-hTERT cells in 2-dimensional culture (organoid day 0) and EPC2-hTERT organoid (day 7 and day 11) were harvested for flow cytometry and the representative flow cytometry plots for CD73 and CD104 expression are shown. (D) Representative IHC images for CD73 staining in EPC2-hTERT organoids harvested on day 5, day 7, day 9, and day 11. Scale bars: 100 μm. (E–G) EPC2-hTERT organoids were harvested on day 11 after incubation in culture medium with EdU for 2 hours, and dissociated for flow cytometry. (E) A representative FACS dot plot of cells from EPC2-hTERT organoids subjected to EdU, with the frequencies of each population according to CD73 and CD104 expression, is shown. (F) Representative histograms of EdU staining in each cell population. The percentage of EdU+ cells in each population is shown. (G) Percentages of EdU+ cells in each population. All data are shown as means ± SD (n = 3). A 2-tailed Student t test or analysis of variance were performed for comparing 2 or multiple comparisons, respectively. ∗P < .05, ∗∗P < .01, ∗∗∗P < .001, and ∗∗∗∗P < .0001.