(A) Viability of SH-SY5Y cells incubated with 25–400 mM D-galactose for 24 and 48h. (B) Viability of SH-SY5Y cells incubated with 200 mM D-galactose (DG) with or without 24h pretreated crocin (25–500 μM). (C) Viability of SH-SY5Y cells incubated with crocin (25–500 μM) alone for 72h. Values are expressed as means ± SD (n=6). #P<0.05, ##p < 0.01 and ###p < 0.001 vs. control, *p < 0.05 and **p < 0.01 vs, D-gal at the same incubation time.

Effect of crocin on D-gal induced intracellular ROS formation in SH-SY5Y cells after 30 min incubation with 20μM, 70–dichlorofluorescin (DCF). Data are expressed as means ± SD (n=6). ###P < 0.05 vs. control, *P < 0.05 vs. D-gal treated cells at the same incubation time.

(A) Effect of crocin on D-gal induced cellular senescence in SH-SY5Y cells. Cells were treated with SA-b-gal for at least 12h. Arrows show staining senescent cells. (B) The percentage of SA-B-gal positive cells. Data are expressed as means ± SD. ##p < 0.01 vs. control, **p < 0.01 vs. D-gal treated cells at the same incubation time. A, control group B, D-gal 200mM group C, D-gal 200 mM + crocin 500μMgroup D, crocin 500μM group.

Expressions of N-ɛ-(carboxylmethyl)lysine (CML) protein in SH-SY5Ycells treated of 200 mM D-gal with or without crocin pretreatment (500 μM) for 24 h. (A) Western blots of CML protein (B) The quantitative protein levels of CML. Data are expressed as the mean ± SD of four separate experiments. ##P < 0.01 vs. control, *P < 0.05, **p < 0.01 vs. D-gal treated cells.