Fig. 3

The inhibition of I/R-induced neuronal apoptosis by NRP-1 overexpression. A Representative immunofluorescence image exhibiting the overexpression of NRP-1 using GFP-containing lentivirus in cultured neurons. Scale bar: 200 μm. B Representative immunoblots and C Western Blot analysis of NRP-1 in cultured neurons of different groups; n = 4. Data are expressed as mean ± SEM. and analyzed by one-way analysis of ANOVA with Bonferroni’s post hoc test. **p < 0.01, as compared with the Cont group; ^##p < 0.01, as compared with the OGD/R + LV-vector group. D qPCR analysis of NRP-1 in cultured neurons of different groups; n = 4. Data are expressed as mean ± SEM. and analyzed by one-way analysis of ANOVA with Bonferroni’s post hoc test. *p < 0.05, **p < 0.01, as compared with the Cont group; ^###p < 0.001, as compared with the OGD/R + LV-vector group. E Cell viability was detected by CCK-8 assay; n = 3. Data are expressed as mean ± SEM. and analyzed by one-way analysis of ANOVA with Bonferroni’s post hoc test. ***p < 0.001, as compared with the Cont group; ^#p < 0.05, as compared with the OGD/R + LV-vector group. F Cell cytotoxicity was determined by LDH release assay; n = 3. Data are expressed as mean ± SEM. and analyzed by one-way analysis of ANOVA with Bonferroni’s post hoc test. ***p < 0.001, as compared with the Cont group; ^###p < 0.001, as compared with the OGD/R + LV-vector group. G Flow cytometry of JC-1 expression and H quantified results of red/green in cultured neurons of different groups; ***p < 0.001, as compared with the Cont group; ^#p < 0.05, as compared with the OGD/R + LV-vector group. (Cont = Control). I Representative terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) images of different groups. Black arrows indicate TUNEL-positive cells. J the percentage of TUNEL-positive cells in the penumbra region of different groups; n = 3. Data are expressed as mean ± SEM. and analyzed by one-way analysis of ANOVA with Bonferroni’s post hoc test. ***p < 0.001, as compared with the Sham group; ^#p < 0.05, as compared with the tMCAO + AAV-empty group. Scale bar: 100 μm. K Representative immunoblots and L Western Blot analysis of Bcl-2, M Cleaved caspase-3, N Caspase-3 and O Bax in the penumbra region of different groups; n = 4. Data are expressed as mean ± SEM. and analyzed by one-way analysis of ANOVA with Bonferroni’s post hoc test. **p < 0.01, ***p < 0.001, as compared with the Sham group; ^#p < 0.05, ^##p < 0.01, as compared with the tMCAO + AAV-empty group