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. 2022 Jun 7;25(7):104542. doi: 10.1016/j.isci.2022.104542

Figure 2.

Figure 2

CYP3A4 induction by PXR ligands in monolayer organoid-derived IECs

(A and B) hiPSO-derived monolayer IECs in collagen I-coated 12-well plates were treated with 0, 0.2, 2, and 20 μM rifampicin (A) or 0, 0.1, 1, 10 μM SR-12813 (B) for 48 h. Cells were harvested, and CYP3A4 and VIL1 mRNA levels were determined by qRT-PCR and normalized to 18S rRNA levels. Assays were performed in n = 3 independent biological replicates (mean ± S.D.). Statistical significance was determined by one-way analysis of variance with the Bonferroni test. ∗p < 0.05 (versus no treatment group).

(C) Differentiated Caco-2 cells cultured in collagen I-coated 12-well plates were treated with 0, 0.2, 2, and 20 μM rifampicin for 48 h. After cells were harvested, the mRNA levels of each gene were determined by qRT-PCR and normalized to 18S rRNA levels. Assays were performed in n = 3 independent biological replicates (mean ± S.D.).

(D) Representative bright-field images of hiPSO- or PIO-derived IECs observed under a phase-constant microscope. Scale bar, 200 μm.

(E) TER values of PIO-derived IECs cultured in collagen-coated Transwells were measured on the indicated days. Assays were performed in n = 4 independent biological replicates (mean ± S.D.).

(F) Immunohistochemistry analysis of frozen sections of PIO-derived monolayer IECs stained with 4′,6-Diamidino-2-phenylindole (DAPI) (blue), anti-E-cadherin (green), and anti-Villin1 (red) antibodies. Scale bar, 20 μm. Right, magnified image of the boxed area.

(G) hiPSO- or PIO-derived monolayer IECs cultured in collagen I-coated 6-well plates were harvested, and the mRNA levels of each gene were determined by qRT-PCR and normalized to 18S rRNA levels. Assays were performed in n = 3 biological replicates (mean ± S.D.).

(H) PIO-derived monolayer IECs or cultured in collagen I-coated 12-well plates were treated with 0, 0.2, 2, and 20 μM rifampicin for 48 h. After cells were harvested, the mRNA levels of each gene were determined by qRT-PCR and normalized to 18S rRNA levels. Assays were performed in n = 3 independent biological replicates (mean ± S.D.). Statistical significance was determined by one-way analysis of variance with the Bonferroni test. ∗p < 0.05 (versus no treatment group).

See also Figure S3.