Srt1720

A549 cells (human lung epithelial cell line) were cultured in DMEM containing 10% fetal bovine serum, 100 U/mL penicillin, and 100 μg/mL streptomycin. For siRNA transfection, the cells were transfected with an siRNA mixture against human CD9, CD81, or control random RNAs (B-Bridge International) using Lipofectamine RNAiMAX (Invitrogen). The cells were cultured for 2 days, and the gene-silencing effect of the siRNAs was assessed by immunoblotting with anti-CD9 and anti-CD81 monoclonal antibodies (Abs). In some experiments, the cells were trypsinized 2 days after transfection, re-cultured in DMEM overnight, and then stimulated with 50 ng/mL TNF-α. The cells were harvested after a further 24-h incubation. Alternatively, 5 µM SRT1720 (Cayman Chemical) was added to the culture at 4 h after CD9 and CD81 siRNA transfection, and the cells were harvested 2 days after transfection. Concentrations of IL-8 in culture supernatants were measured by enzyme-linked immunosorbent assay using Quantikine (R&D Systems).

CAY10591 and SRT1720 were purchased from Cayman Chemicals (Ann Arbor, MI, USA). Primary antibodies against β-actin, CREB (sc-25785), and NF-κΒ p65 (sc-7151) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Antibodies against phospho-AMPK^Thr172 (#2535), acetyl-p53^Lys379 (#2570), phospho-p53^Ser15, phospho-mTOR^Ser2448, and phospho-CREB^Ser133 (#9198) were purchased from Cell Signaling Technology. The primary antibodies against iNOS (610431) and p53 were purchased from BD Transduction Lab (Lexington, KY, USA). The primary antibody against COX-2 (aa 570-598) was purchased from Cayman Chemicals (Ann Arbor, MI, USA). Antibodies against α-tubulin (T5168), acetylated tubulin^Lys40 (T7451), HDAC6, EX527 (E7034), and TSA (T8552) were purchased from Sigma Aldrich (St. Louis, MO). Antibodies against HO-1 were purchased from Enzo Life Sciences, Inc. (Farmingdale, NY, USA). Antibodies against β-actin (ab6267) and SIRT1 (ab50517) were purchased from Abcam (Cambridge, MA, USA).