Fig. 5

The beneficial effects of anthocyanins against glutamate-induced toxicity in SH-SY5Y cells. a–c The cell viability histograms of SH-SY5Y cells, as indicated by the MTT assay. a The cells were incubated with three different (indicated) concentrations of anthocyanins for 24 h. b The cell viability histogram of SH-SY5Y cells that were treated with the indicated concentrations of glutamate for 3 h. c The cell viability histogram after pretreating cells (12 h) with three concentrations of anthocyanins and glutamate (30 mM) for additional 3 h. All details are provided in the “Methods” section. d The immunofluorescence images and respective relative IOD histogram of FJB (green)-positive and DAPI (blue) SH-SY5Y cells. Cells were grown in four-well chamber slides and then treated with the mentioned (“Methods” section) concentrations of glutamate and anthocyanins. The FJB staining was repeated in triplicate. e The immunostained images of p-AMPK (green) protein counterstained with DAPI (blue) in the SH-SY5Y cells. The panels represent different treating groups such as untreated control, glutamate, or glutamate and anthocyanins. f The histogram of relative AMPK activity assay conducted with cell lysate experimental treating groups having siRNA of AMPK. g The Western blot analysis of p-AMPK, Nrf2, and HO-1 protein expressions and their relative density histograms after treating cells with glutamate and glutamate and anthocyanins and in the presence or absence of AMPK siRNA. h The immunostaining images and respective relative IOD histograms of Nrf2 protein in SH-SY5Y cells. i The ROS assay histogram after treating SH-SY5Y cells with glutamate or glutamate and anthocyanins with or without compound C. The groups and other experimental details are provided in the “Methods” section. Significance, *P < 0.05, **P < 0.01, ***P < 0.001 and ^## P < 0.01 and ^### P <0.001, respectively