Figure 6. Knockout of 14-3-3t attenuates the anti-apoptotic effect of NSC-sEVs in neuronal cells. (A) Western blot analysis of changes in neuronal apoptosis-related proteins. (B, C) Semi-quantitative detection of relative expression levels of apoptosis-related proteins, normalized to GAPDH. (D) TUNEL detection of neuronal apoptosis. TUNEL-positive apoptotic cells (red). Nuclear staining using DAPI (blue). Scale bar = 100um. (E) Quantitative estimation of the proportion of apoptotic cells in each of the three experimental groups. (F) Annexin V/FITC/PI double staining and flow cytometry was used to detect neuronal apoptosis induced by Glu with or without NSC-sEVs. (G) Quantitative results of NSC-sEVs treatment and non-treatment of apoptotic neurons. * p < 0.05, compared to the Glu group, # p < 0.05, compared to shGFP-sEVs. NSC-sEVs, neural stem cell-derived small extracellular vesicles; DAPI, 4’,6-diamidino-2-phenylindole; FITC, fluorescein isothiocyanate; PI, propidium iodide; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; Glu, Glutamate; TUNEL, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay.