2 nbdg glucose uptake assay

Glucose uptake was determined by the 2-NBDG Glucose Uptake Assay (BioVision, Milpitas, CA, USA). Briefly, the Ishikawa and ECC-1 cells on 96-well plates were treated with PA for 16–18 h. The cells were washed with HBSS and incubated in a glucose-free medium with 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-D-glucose (2-NBDG, 100 μg/mL) for 15 min. After washing the plates with Hanks’ Balanced Salt Solution (HBSS) twice, the fluorescence intensity (Ex/Em = 465/540) of cellular 2-NBDG in each well was measured using a Tecan plate reader. The experiments were performed in triplicate and repeated three times.

After lentiviral transduction as indicated, 5 x 10⁵ AsPC-1 and BxPC-3 cells were seeded into six‐well plates and incubated for 48 hours. Glucose uptake and lactate levels in culture supernatants were quantified using the 2-NBDG Glucose Uptake Assay (BioVision) and the Lactate Assay kit (BioVision), respectively. Intracellular ATP content was determined using an ATP Assay Kit (Roche) according to the manufacturer's instructions.