NE increases ACC cell proliferation in vitro. (A) Expression of β-adrenergic receptors from ACC RNA-seq datasets (n = 74). (B) Immunohistochemistry of β2R using a human salivary ACC tissue microarray (n = 28). Scale bar: low power, 100 μm; and high power, 20 μm. (C) RT-PCR of β-adrenergic receptors (n = 3 per group) and IF staining of β2R in AH43a and AH45b. Scale bar: 50 μm. (D) MTT [3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide] assay of AH43a and AH45b treated with NE, propranolol, or both (n = 6 per group). (E) Western blot of signaling pathways in AH43a and AH45b treated with NE at varying concentrations. Western blot quantification of pERK/GAPDH and pCREB/GAPDH compared with controls. (F) Tiled and stitched images of AH43a and AH45b spheroids cultured in Matrigel without (control) or with SG. Scale bar: 200 μm. Quantification of spheroid numbers for control and SG (n = 8 per group, 4 AH43a control). (G) Confocal images of P0⁺ TdTomato-lined SG neurites infiltrating into AH43a and AH45b spheroids (arrowheads show infiltrating neurites). Scale bar: low power, 100 μm; and high power, 20 μm. (H and I) Quantification of AH43a and AH45b spheroid numbers grown in Matrigel alone (control) or co-cultured with SG with propranolol or grown with CM from SG (n = 6 per group, 5 AH45b SG). (J) Left: Bright-field microscopy images of SG grown in regular media (control) or CM from AH43a after 7 days. Middle: Processed images showing the largest contiguous neurite area (red) emanating from the center (ganglion body). Right: Quantification of the neurite area (n = 5 per group). Data are representative of two independent experiments (D, F, and H–J). P values were determined by one-way ANOVA, Fisher’s LSD test (F and H), and Student’s t test. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, ns: not significant (D and I). Source data are available for this figure: SourceData F2.