Leibovitz s l 15 medium

Human PC cells SW1990 were purchased from Procell (China) and cultured in Leibovitz's L-15 medium (Solarbio, China) with 10% FBS. PANC-1, AsPC-1, and BxPC-3 cell lines were purchased from iCell Bioscience (China). PANC-1 cells were treated with 10% FBS in DMEM medium. In addition, RPMI-1640 medium with 10% FBS was used for AsPC-1 and BxPC-3 cells. The above cells were cultured in an incubator at 37°C and 5% CO₂.
PANC-1 and AsPC-1 cells were used to silence or overexpress ITGBL1. For ITGBL1 downregulation, PANC-1 and AsPC-1 cells were transfected with short hairpin RNAs (shRNAs) specifically targeting ITGBL1 or the negative control (NC) for 48 h. For ITGBL1 overexpression, PANC-1 and AsPC-1 cells were transfected with ITGBL1 overexpression vector or empty vector for 48 h. For co-transfection, PANC-1 cells were co-transfected with ITGBL1 overexpression vector and JDP2 overexpression vector for 24 h. The transfections were mediated by Lipofectamine 3000 (Invitrogen, USA), according to the manufacturer's instruction.

The ovary was quickly removed and placed into a special oocyte medium (OCM). The OCM consisted of 90% Leibovitz’s L-15 medium (Solarbio, Beijing, China), 0.5 mg/mL bovine serum albumin (BSA, Amresco, Solon, OH, USA) and 3% Penicillin-Streptomycin (PS, Gibco, Waltham, MA, USA). Intact follicles about 0.625 mm in diameter were isolated from ovarian fragments and then placed into the oocyte maturation medium (OMM, 90% Leibovitz’s L-15 medium, 0.5 mg/mL BSA, 1% PS and 1 ug/mL 17α-20β-dihydroxy-4 pregnen-3-one) (17α-20β-dihydroxy-4 pregnen-3-one, DHP, Medbio, Shanghai, China). After incubation at 26 °C for 24 h, the oocytes’ maturation and survival rates were examined. To determine the oocytes’ maturation, they were stained away from light with 20 umol/L brilliant cresyl blue (Yuanye, Shanghai, China) for 150 min, after which, mature cells were stained blue. Moreover, to calculate the survival rate, the oocytes were stained with 0.4% Trypan blue (Sigma-Aldrich, St. Louis, MO, USA) at room temperature for 5 min, and any dead cells were stained blue.

SW982 cell line was purchased from the Cell Bank of Type Culture Collection of Chinese Academy of Sciences and was cultured in Leibovitz's L-15 Medium (Solarbio, Beijing, China) containing 10% fetal bovine serum (Thermo Fisher, Waltham, MA, USA), 100 U/mL, penicillin and 100 mg/mL streptomycin at 37°C in 5% CO2, respectively. For autophagy inhibition or initiation, cells were treated with 10 mmol/L of 3-MA or rapamycin (10 nmol/L; Sigma Aldrich, St. Louis, MO, USA) for 24 hours, after transfection with recombinant plasmid.