Fig. 6.

Engineered tubules as a tool for studying human UB developmental processes and defects. (A) Experimental design. Fragments of iPSC-derived macrotubules are cultured with mouse embryonic kidneys or with growth factors in the 3D collagen culture system. (B,C) Healthy donor-derived tubule co-cultured with mouse embryonic kidneys. (B) At 1 day, long primary buds emerge (left panel). At 2 days, a lateral ramification (arrow) arises from the primary bud. (C) A ramified bud consists of aligned E-cadherin-positive cells (white). (D) Percentage of the ramified buds in the total buds emerged from healthy donor-derived tubules. Data are expressed as means ± SEM from three independent experiments. Number of fields analyzed: n = 48 for H + G, n = 30 for H + G + F1 and H + G + F7, n = 22 for H + G + F1 + F7, n = 21 for mouse embryonic kidneys. *P < 0.05 versus H + G, °P < 0.05 versus H + G + F1, #P < 0.01 versus H + G by one-way ANOVA with Holm-Sidak's multiple comparisons test. (E,F) Healthy donor-derived tubule cultured with all growth factors displays primary and secondary buds (arrows) some of which showing terminal bifid branching (asterisks). (F) A primary bud arising from healthy donor-derived tubule cultured as that in (E). (G) Percentage of the ramified buds in the total buds emerged in healthy donor- and patient-derived tubules cultured with all growth factors. Data are expressed as means ± SEM from three independent experiments. Number of fields analyzed: n = 22 for healthy donor- and n = 27 for patient-derived tubules. **P < 0.0005 by two-tailed Student's t-test. (H,I) Patient-derived tubule cultured with all growth factors shows mainly primary buds. (I) A primary bud arising from patient-derived tubule cultured as that in (H). DAPI, blue-stained nuclei; H, HGF; G, GDNF; F1, FGF1; F7, FGF7. Scale bars: 100 μm (B, left panel; E, left panel), 50 μm (E, right panel; H), 20 μm (B, right panel; C,F,I).