Fig. 3

Chronic stress activates acetylcholinergic pathway of pulmonary epithelial cells with neuroendocrine functions. a. Gating strategy of flow cytometry analyses of ChAT⁺ epithelial cells and CD4⁺ T cells. b-c. FACS analyses of ChAT expression in EpCAM⁺ epithelial cells (b) and CD4⁺ T cells (c) in lungs of breast cancer model mice with or without chronic restraint stress for 2 weeks and their corresponding ChAT positive percentages. d. Immunofluorescence examinations of EpCAM and VAChT in lung tissues of breast cancer model mice in control group and chronic restraint stress group at 2 weeks. e. Immunofluorescence examinations of EpCAM and VAChT in lung tissues of breast cancer model mice in control group and chronic unpredictable stress group. f-g. Percentage (left) and fluorescence intensity (right) of EpCAM-VAChT colocalization in experiments as described in d and e, respectively. ns: no sense, **p < 0.01, ***p < 0.001, ****p < 0.0001