Fig. 7
From: Salidroside reduces neuropathology in Alzheimer’s disease models by targeting NRF2/SIRT3 pathway
SAL directly inhibits NRF2-KEAP1 complex and facilitates NRF2-induced SIRT3 transcription. A IF of NRF2 (green) and nuclei (blue) in SH-SY5Y cells. Nuclei counterstained with DAPI. Scale bar, 50 μm. B Quantification shows the nucleus-to-cytoplasm ratio of NRF2 levels. C qPCR shows the NRF2 mRNA levels in SH-SY5Y cells treated with indicated dose of SAL. D Immunoblotting images (left) and quantification of band intensity (right) show the protein levels of NRF2 and SIRT3 in SH-SY5Y cells with or without SAL treatment. E As in D, except cytosolic (Cyto) and nuclear (Nuc) protein were isolated and detected. F Immunoprecipitation (IP) shows the protein levels of NRF2 and KEAP1-Flag in SH-SY5Y cells with indicated treatment. An anti-Flag antibody was used for IP. 5% total protein used as input. G Surface plasmon resonance (SPR) analyses show the SAL-NRF2 interaction behavior. Quantitation shows the sensogram of each indicated titration of SAL, as expressed in RU (response units-Y axis) along time (X-axis). H Cellular thermal shift assays (CETSA) show the binding capacity of SAL-NRF2 interaction. Immunoblotting shows NRF2 levels and below graph shows the CETSA melting curve. I Immunoblotting shows the NRF2 levels in protein lysates at 55 °C. The below graph shows the ITDR-CETSA curve. Error bars indicate mean ± SD from at least 3 independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001, n.s., not significant; one-way ANOVA