Fig. 4

Involvement of MT2/Bip/NRF1 pathway in melatonin-induced ABCA1 expression. a and b Cholesterol (200 μM, 24 h) was treated to UCB-MSCs and the changes in expression levels of MT1 and MT2 were measured by qPCR or western blotting (n = 5, *p < 0.05 vs control). c Melatonin (1 μM) was pretreated to UCB-MSCs before cholesterol treatment (200 μM, 24 h) and then the expression levels of HSPA5, DERLIN, HERP, SEL1L, and VCP were compared by qPCR (n = 5, *p < 0.05 vs cholesterol). d MT2 inhibitor, 4-P-PDOT (10 μM) were pretreated prior to melatonin treatment (1 μM) and the expression level of BiP was measured by western blotting (n = 5, *p < 0.05 vs control, ^#p < 0.05 vs cholesterol + melatonin). e Immunocytochemistry was conducted with NRF1 (green) and BiP (red) specific antibodies and DAPI (blue) and then visualized by SRRF imaging system. Scale bar was set as 8 μm (magnification × 1000) (n = 5, *p < 0.05 vs control, ^#p < 0.05 vs cholesterol). f, g NRF1 or NT siRNA was transfected to UCB-MSCs using transfection reagents prior to cholesterol treatment (200 μM, 24 h). The expression levels of ABCA1 and NRF1 were measured by western blotting and the level of total cholesterol was quantified with cholesterol quantification kit (n = 5, *p < 0.05 vs NT siRNA, ^#p < 0.05 vs NT siRNA + cholesterol). All blot and images are representative and quantitative data are presented as a mean ± standard error of the mean