Figure 3

NMN partially rescues cuproptosis via upregulating the expression of sirt2. HeLa cells were cultured in DMEM with 10% copper free serum replacement and pulsed Es/Cu with or without NMN or only NMN, the sirt2 protein level were determined by western-blot (A) and the band intensity were analyzed (B), the mRNA levels were determined by qPCR (C). HeLa cells were transfected with pEX-1-sirt2 (sirt2) or pEX-1 empty vector (vector) as control, the protein expression of sirt2 was determined by western-blot (D) and the bands density were analyzed (E) and the mRNA levels were determined by qPCR (F). HeLa cells transfected with sirt2 or empty vector for 24 h, and then treated with ES/Cu, the cell viabilities were determined by CCK-8 assay (G). HeLa cells were transfected with siRNA against sirt2 (si-Sirt2) or nonspecific siRNA (si–C), the expression of protein and mRNA were determined (H–J). 24 h after siRNA transfected, cells were treated with ES/Cu, and the cell viabilities were determined by CCK-8 assay (K). Results were the mean from 3 to 5 independent experiments. Data were represented as mean ± SEM. **** P < 0.0001.