Extended Data Figure 9 |. Xenografted WA25 skin organoid hair follicles have sebaceous glands comparable to second-trimester (18 weeks) foetal and adult facial hair.

a, Brightfield image of two day-115 (in vitro) WA25 skin organoid hair follicles with visible hair shafts and sebaceous glands; represents hair follicles produced in 9 independent experiments. b, LipidTOX staining of day-120 WA25 skin organoid hair follicles reveals lipid-rich cells, such as sebocytes and adipocytes (APs; outlined by dashed lines). (left) Immunostaining for KRT15 labels outer root sheath of hair follicle cells. (right) High magnification image of an adjacent cryosection of the specimen in the left panel. Dashed line highlights a horizontally-cross-sectioned hair shaft and a sebaceous gland. c, TEM image of a day-140 WA25 skin organoid sebaceous gland; TEM was performed once on 2 different skin organoids from separate experiments. Peripheral layer cells (PLCs) and a maturing sebocyte (MS) containing sebum vacuoles (SV) have been pseudo-coloured in yellow and green, respectively. d, d’, SCD1⁺ sebaceous glands in xenografted day-140 WA25 skin organoids. Xenografts were extracted and fixed in 4% PFA, >49 days after transplantation. Dashed boxes and asterisks indicate magnified regions presented in following image panels. Dashed lines distinguish between the grafted human skin organoid tissue and the tissues of host mouse (NU/J). Arrows indicate SCD1⁺ adipocytes. e, SCD1⁺ sebaceous glands in the NU/J mouse skin adjacent to xenografts. Note that the size of mouse sebaceous glands is smaller than those of human that the origin of sebaceous glands is distinguishable within the extracted xenograft samples. f, f’, SCD1⁺ sebaceous glands in 18-week human foetal forehead skin. g, g’, SCD1⁺ sebaceous glands and adipocytes in 18-week human foetal cheek skin. Note that the adipocytes are prominently abundant in foetal cheek tissue compared to forehead tissue (f vs. g). Dashed boxes and symbols highlight the magnified regions in following images. h-i, SCD1⁺ sebaceous glands (SG) in adult human facial skin. Epithelium is visualized by ECAD immunostaining. Dashed line outlines several lobes of sebaceous glands of a hair follicle. Immunostaining images represent one of 5 independent staining on 3–5 different samples per tissue type. Scale bars: 250 μm (a, f, g, h), 100 μm (b; left, d, d’, f’, g’, g’; **, h’, i), 50 μm (b; right, d’; *, **, separate cryosection (SC), e, f’; *, SC, g’; #, SC, ##, *), 5 μm (c). Corresponds with data in Fig. 4.