Fig. 3

ROS accumulation blocks autophagosome degradation by activating the RIP1/RIP3 pathway after OGD/R. a Immunofluorescence images showing autophagy flux labeled by mRFP-GFP-LC3 in OGD/R-treated SH-SY5Y cells after NAC treatment (bar = 10 μm, n = 5/group). b Western blotting analysis and Co-IP assay showing RIP3 phosphorylation and RIP1-RIP3 binding ability in OGD/R-treated SH-SY5Y cells after NAC treatment (n = 8/group). c Western Blotting analysis and Co-IP assay showing RIP3 phosphorylation and RIP1-RIP3 binding ability in SH-SY5Y cells after Drp1 S616A mutation (n = 8/group). d Immunofluorescence images showing autophagy flux labeled by mRFP-GFP-LC3 in SH-SY5Y cells after Drp1 S616A mutation and Nec1 treatment (bar = 10 μm, n = 5/group). e Western Blotting analysis showing LC3 II/I ratio in SH-SY5Y cells after Drp1 S616A mutation and Nec1 treatment (n = 8/group). ^*P < 0.05, compared with normal group; ^#P < 0.05, compared with OGD/R or Drp1 S616A group; ^&P < 0.05, compared with OGD/R + NAC (1 mmol/L) or OGD/R + Nec1 (10 μmol/L) group. Co-IP co-immunoprecipitation, OGD/R oxygen–glucose deprivation/reoxygenation treatment, ROS reactive oxygen species