Gastrin 1

Briefly, UCB tumor tissues were minced at about 0.5 cm diameter and disassociated into individual cells via digestion with 5 mg/mL collagenase type II (Life Technologies) with 10 μM Y-27632 dihydrochloride and incubated at 37 °C for 1 h. Then, we filtered the suspension of dissociated cells through a 70 μm mesh filter. The UCB cells were then embedded in Matrigel and cultured with Advanced Dulbecco's modified Eagle's medium/F12 mixed with 500 ng/mL Rspo1, 100 ng/mL Noggin, 50 ng/mL EGF, 10 mM nicotinamide, 500 nM A830-1 (Tocris), 3 mM SB202190 (Sigma), 10 nM prostaglandin E2 (Sigma), penicillin/streptomycin, 10 mM HEPES, 2 mM GlutaMAX, 1 × B27 (Life Technologies), 10 nM gastrin I (Sigma), and 1 mM N-acetylcysteine (Sigma) at 37 °C under 5% CO₂. The medium was obtained from OuMel (Hai Shang, China). For the generation of stable lines, organoids were first digested into individual cells by TrypLE (GIBICO) and incubated with purified lentivirus for 6 h, followed by selection with 2 μg/mL puromycin. For organoids formation assay, the individual organoid cells were seeded onto a 24-well plate with a density of 1000 cells in 50 μL/well. The number and size of the organoids were determined after 2 weeks, and the growth area of organoids was measured and quantified by ImageJ.