Fig. 4

CASP8 is a pivotal participant in RIR injury. a Volcano plot showing a total of 980 genes, of which 385 were upregulated and 595 were downregulated in retinal tissue in the RIR group compared with the sham group. A fold change > 2 or < 0.5 was considered statistically significant (n = 5 mice/group). b KEGG pathway enrichment analysis of differentially expressed genes between the ischemia/reperfusion and sham groups (n = 5 mice/group). The enriched pathways of hypoxia and inflammatory signaling are shown. The positive regulation of I-kappaB kinase/NF-kappaB signaling was chosen for further analysis. c Heat map showing the contents of the positive regulation of I-kappaB kinase/NF-kappaB signaling in the retinas of mice after the RIR process (n = 5 mice/group). RIR mice were sacrificed 7 days after reperfusion, and retinal tissue was collected for RNA sequence analysis. d HE staining of retinas from mice under RIR with or without CASP8 knockdown (20 μΜ). The retinal tissue was harvested on the seventh day after reperfusion (n = 6). Scale bar: 50 μm. The total retinal thicknesses were quantified for the three groups (n = 6). e Representative immunofluorescence images of RGCs in the retina from mice treated with or without CASP8 knockdown. Primary antibody against RBPMS was used to label RGCs (n = 6). Scale bar: 100 μm. f Retrograde FG staining of RGCs from RIR injury in the absence or presence of CASP8 interference (20 μΜ, n = 6). Scale bar: 200 μm. RGCs survival were analyzed comparing to the controls (n = 6). RIR: retinal ischemia-reperfusion; GCL: ganglion cell layer; IPL: inner plexiform layer; INL: inner nuclear layer; OPL: outer plexiform layer; ONL: outer nuclear layer; CASP8 si: caspase-8 siRNA. All of the data are representative of at least three independent experiments. Data are represented as the mean ± SD. *P < 0.05, **P < 0.01, one-way ANOVA