Human umbilical vein endothelial cells (HUVECs) were purchased from Lonza (C2519A-lonza; Walkersville, MD, USA) and used in the experiments. The cells were grown in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 5% heat-inactivated fetal calf serum, 1% penicillin streptomycin, 4 mM L-glutamine, 4.5 g/L glucose, and 1.5 g/L sodium bicarbonate. Cells were kept in a humidified incubator with 5% CO2-95% air at 37°C, and the media were refreshed two to three times weekly. The cells were maintained in six-well culture plates, and the passages were performed by trypsinization. The experiments were performed when the plates became at least 80% confluent.
LPS, interleukin (IL) 1α, IL-1β, and media were purchased from Sigma-Aldrich (St. Louis, MO, USA). Anti-fractalkine (CX3CL1) antibody was purchased from Torrey Pines Biolabs (Secaucus, NJ, USA). Calpain inhibitor (BML-PI116-0005) and caspase-1 inhibitor (ZYVAD[OMe]-FMK) were purchased from Enzo Life Sciences (Farmingdale, NY, USA). Caspase-3 inhibitor (Ac-DEVD-CHO) and caspase-9 inhibitor (Z-LEHD-FMK) were purchased from BD Biosciences (San Jose, CA, USA).
LPS, interleukin (IL) 1α, IL-1β, and media were purchased from Sigma-Aldrich (St. Louis, MO, USA). Anti-fractalkine (CX3CL1) antibody was purchased from Torrey Pines Biolabs (Secaucus, NJ, USA). Calpain inhibitor (BML-PI116-0005) and caspase-1 inhibitor (ZYVAD[OMe]-FMK) were purchased from Enzo Life Sciences (Farmingdale, NY, USA). Caspase-3 inhibitor (Ac-DEVD-CHO) and caspase-9 inhibitor (Z-LEHD-FMK) were purchased from BD Biosciences (San Jose, CA, USA).