Establishing and Characterizing Pancreatic Cell Lines

The HapT1 cancer cell line was commercially procured from Sigma, and human PSCs (hu-PSCs) were procured from Sciencell Research Laboratories. Both the cell lines were propagated and frozen immediately after receipt. The frozen stocks were revived and used within three months. HapT1 cells were maintained in 10% FBS that contained MEM medium (Pan-Biotech) supplemented with 1x NEAA (Life Technologies). Human PSCs were maintained in 2% FBS containing Stellate Cell Medium (SteCM; Sciencell) supplemented with growth factors. With the approval of Institutional Animal Ethics Committee (Institute of Life Sciences, Bhubaneswar, India), rat and hamster PSCs were isolated from normal pancreas. The PSCs were confirmed to have the typical stellate cell morphology and were positive for α-SMA (a marker for activated PSCs). All of the established cell lines were used between 3-6 passages. Rat PSCs were maintained in 10% FBS containing Iscove's Modified Dulbecco's Medium (IMDM; Pan-Biotech), and hamster PSCs were maintained in 5% hamster serum containing IMDM medium (Pan-Biotech). To check cell viability in vitro for HapT1 and PSCs, the MTT assay, crystal violet staining, and colony formation assay were performed as reported earlier [44 (link), 45 (link)]. To determine the role of ROS-induced cell death, cells were pre-treated with 1 mM NAC for 12 hours before adding DSF or DSF+Cu.