Figure 4. The vhCOs possess functional vascular system.

(a) Depiction of subcutaneous implantation of control hCOs and vhCOs in the right and left leg of immune-deficient mice. (b) Left, in vivo T₂ map of the implanted control hCOs and vhCOs. Right, anatomical image of hCOs after 10- and 30-day post-implantation (dpi). Both images detected the vhCO region, but area of implanted control hCO was not apparent. Data are representative of three independent experiments. (c) Top, tissue concentration of gadolinium contrast agent as a function of time in the left leg muscle (gray trace) and implanted vhCOs (black trace). Muscle tissue indicates a rapid uptake and backflow into the vasculature, but vhCOs showed a slower and irreversible uptake. Bottom, map of the area under the curve (AUC) of the concentration curve, with ROIs outlined for the vhCOs (green) and muscle (blue). Data are representative of three independent experiments. (d) Schematic of the method for FITC-dextran perfusion. Host blood vessels are filled with FITC-dextran, shown green, and endogenous vessels in vhCOs were shown as magenta. (e) Explanted organoids from FITC-perfused mice were stained for human-specific CD31 and hNuclei at day 30 dpi. The scale bar represents 50 μm, n=3 animals and 3 organoids from three independent batches for MRI and n=7 animals and 7 organoids from four independent batches for FITC-perfusion (*p=0.00412, **p=0.000183, the unpaired two-tail t-test was used for all comparisons). Mean ± SEM are shown.