Orthotopic ACC PDX recapitulates human ACC and derives sympathetic innervation. (A and B) H&E sections of patient specimens used to generate AH43a and AH45b and corresponding PDX. Scale bar: low power, 200 μm; and high power, 50 μm. (C) AH43a and AH45b cells grown in 2D and 3D cultures. Scale bar: 2D, 100 μm; and 3D, 50 μm. (D and E) Gross, ultrasound, and bioluminescence images and tumor volumes quantification of ACC PDX grown in the SMG, PG, or flank (n = 8). (F) TH⁺ innervation in ACC PDX (T) and adjacent normal salivary gland (N) of 20-μm sections. TH⁺ nerve fibers (arrowheads) sprout from large-caliber (dashed circles) TH⁺ nerves. Scale bar: low power, 100 μm; and high power, 50 μm. (G and H) Immunostaining of TH⁺ nerve sprouts (arrowheads) and large-caliber nerves (dashed circles) from ACC PDX in the mouse SMG and PG. Large-caliber nerves without nerve fiber sprouts are in the flank and lung. Quantification of intratumoral TH⁺ sprouts (AH43a, n = 5; AH45b, n = 8). Data are representative of two independent experiments (E and H). Scale bar: 50 μm. P values were determined by one-way ANOVA and Fisher’s LSD test. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, ns: not significant.