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. 2021 Sep 23;16(10):2432–2441. doi: 10.1016/j.stemcr.2021.08.014

Figure 2.

Figure 2

Generation of iN-VOs and iMB-VOs

(A) General strategy for the generation of vascularized organ tissues via introduction of parenchymal cell types in VOs.

(B) Schematic of iN-VO and iMB-VO culture protocol.

(C) Immunofluorescence 100-μm z stack, maximum projection, confocal micrographs of MAP2- and CDH5-labeled uninduced (iN-VO, −Dox) and induced (iN-VO, +Dox) day 15 iN-VO organoids (scale bars, 50 μm).

(D) qRT-PCR analysis of signature endothelial genes CDH5 and VEPTP; signature smooth muscle gene SMA; and signature neuronal genes MAP2, VGLUT2, BRN2, and FOXG1 at day 15 of culture for iN-VO organoids. Data represent the mean ± SD (n = 7 organoids, from three independent experiments).

(E) Immunofluorescence 100-μm z stack, maximum projection, confocal micrographs of MYH- and CDH5-labeled uninduced (iMB-VO, −Dox) and induced (iMB-VO, +Dox) day 15 iMB-VO organoids (scale bars, 50 μm).

(F) qRT-PCR analysis of signature endothelial genes CDH5 and VEPTP; signature smooth muscle gene SMA; and signature skeletal muscle genes MYOG, MYH8, TNNC1, and RYR at day 15 of culture for iMB-VO organoids. Data represent the mean ± SD (n = 7 organoids, from three independent experiments).

(G) Pan-organoid tile-scan immunofluorescence confocal micrograph of a CDH5- and MAP2-labeled day 15 neuro-vascular organoid (scale bars, 500 μm). Image is a 200-μm z stack maximum intensity projection.

(H) Pan-organoid tile-scan immunofluorescence confocal micrograph of CDH5- and MYH-labeled day 15 myo-vascular organoid (scale bars, 500 μm). Image is a 200-μm z stack maximum intensity projection. (D and F) ∗∗p ≤ 0.01, ∗∗∗p ≤ 0.001, and ∗∗∗∗p ≤ 0.0001; ns, not significant.