Arpe 19 human rpe cells

ARPE-19 human RPE cells (CRL-2302) obtained from ATCC (Manassas, USA), were cultured in Dulbecco’s Modified Eagle’s Medium/F-12 (Gibco, USA) by the addition of 100 U/ml penicillin (Beyotime, China), 100 μg/ml streptomycin (Beyotime, China), and 10% fetal bovine serum (Gibco, USA). Passage culture of cells was performed every 2–3 days. Isolation of human RPE cells was from the eye tissue of human cadavers in the eye bank of The First Affiliated Hospital of Chongqing Medical University. As per Helsinki Declaration, written consent of the experimental materials for medical research was gained from donors or their families. The method of isolation and culture of RPE cells have been previously described (29). Cells were grown at a constant temperature of 37°C inside a humidified 5% CO₂ condition. The expression of RPE65 was detected by cell immunofluorescence and the cells were identified [25 (link)]. RPE65 antibody (A9615, Abclonal, China) and fluorescent secondary antibody (AS011, Abclonal, China) were purchased from Abclonal, China.

Primary human fetal RPE (H-RPE) cells were purchased at passage one from LONZA (Walkersville, MD, USA), and all experiments were performed with cells at passage two to six. The ARPE-19 human RPE cells were purchased from the ATCC (Manassas, VA, USA). Tamoxifen and 5-ASA were purchased from Enzo Life Sciences (Farmingdale, NY, USA). The rapamycin was purchased from EMD Millipore Corporation (USA). Sulfasalazine, sulfapyridine, bafilomycin A1, and other reagents were purchased from Sigma-Aldrich (St. Louis, MO, USA). NAC (Sigma-Aldrich) and Z-YVAD-FMK (BioVision Inc. Milpitas, CA) were used as inhibitor reagents.