Culture and differentiation of PASCs

The pituitary adenoma specimen was resected and immediately divided into two portions during surgery. One portion was snap-frozen and stored in liquid nitrogen for subsequent pathological immunostaining and RNA sequencing analysis. The other portion was transported to the laboratory under sterile conditions for PASCs culture. Tumor specimens were thoroughly washed with 1× PBS and cut into small pieces. Washed in 10 ml DMEM/F-12 and centrifuged at 300×g for 2 min. The resultant cell pellet was lysed in 2 ml of 1× Accutase (Stemcell Technology, USA) and incubated at 37 °C for 5 min. Subsequent centrifugation was performed for 3 min. Erythrocytes were removed from the cell pellet using erythrocyte lysis buffer, with a 5-min treatment. After a washing step, the cells were resuspended and cultured in stem cell medium, which consisted of DMEM/F-12 supplemented with 1% penicillin–streptomycin, 1× B27 (50× concentration, Life Technologies, USA), 20 ng/ml of bFGF (Peprotech, USA), and 20 ng/ml of EGF (Peprotech, USA). The culture was maintained at 37 °C in a 5% CO₂ incubator.