Fig. 4: Comprehensive Characterization of ISCs3D-hIO Cultured on XF-DISC. | Nature Communications

Fig. 4: Comprehensive Characterization of ISCs3D-hIO Cultured on XF-DISC.

From: Xenogeneic-free culture of human intestinal stem cells on functional polymer-coated substrates for scalable, clinical-grade stem cell therapy

Fig. 4

a Heatmap of gene expression in hPSCs and ISCs3D-hIO cultured on Matrigel-coated surfaces and XF-DISC for 4, 6, and 8 days at passage 3, as quantified by qRT‒PCR (n = 3). b Gene expression of intestinal markers (LGR5, EPHB3, CD44 and SOX9) and a proliferation marker (MKI67) of ISCs3D-hIO cultured on a Matrigel-coated surface and XF-DISC for 7 days at passages 3, 8 and 18 quantified by qRT‒PCR. Data represent the mean ± SD (n = 3 biological samples), and a two-tailed t-test was applied to measure p values. c Cell population of markers (CD44, SOX9 and CD34) of ISCs3D-hIO cultured on a Matrigel-coated surface and XF-DISC for 7 days, as determined by FACS analysis (Red: negative control cells in each sample, black: specific antibody stained-ISCs3D-hIO cultured on Matrigel-coated surface and blue: specific antibody stained-ISCs3D-hIO cultured on XF-DISC). d Representative images showing the immunostaining of SOX9, Ki67, EIF3E, FABP1, LDHB, CHGA and MUC2 expression in ISCs cultured on a Matrigel-coated surface and XF-DISC for 7 days at passage 2 (n = 3 samples/group). Scale bar: 100 µm. e Volcano plot depicting gene abundance changes in ISCs cultured on a Matrigel-coated surface and XF-DISC for 7 days at passage 17. The vertical red lines indicate ± 4-fold changes, and the horizontal red line indicates a minimum significance of 20 (p value = 0.01). Multidimensional scaling analysis of the pairwise distances of the sample was conducted. f MDS plot of the pairwise distances between samples. Six homogeneous sample groups were observed; Matrigel (black, n = 3), XF-DISC (red, n = 3), ALI (blue, n = 3), hSI (green, n = 2), iPSC (CRL, pink, n = 1), and ESC (H9, yellow, n = 1). g Volcano plot depicting protein abundance changes in ISCs cultured on a Matrigel-coated surface and XF-DISC for 3 days at passage 12. The vertical red lines indicate ± 4-fold changes, and the horizontal red line indicates a minimum significance of 20 (p value = 0.01). Label-free quantification-based proteomic analysis was performed via ANOVA.

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