Fig 2: Aspirin alleviates Aß-induced senescence and mitochondrial dysfunctions via upregulation of SIRT pathway. (a–g) HMC3 cells were preincubated with aspirin for 4 h followed by treatment with 10 µM Aß for 72 h. (a) Protein levels of SIRT1, PAI-1, and p21 were detected by western blotting. (b) The quantification of relative protein levels in (a). (c) The representative SA-ß-gal staining images. (d) Quantification of the percentage of SA-ß-gal cells. Scar bar, 50 µm. (e) The cells were stained with JC-1 dye and the fluorescence intensity was quantified using BioTek reader. (f) HMC3 cells were incubated with DCFH-DA and Hoechst; then, the fluorescence intensity was detected by BioTek reader. (g) The fluorescent latex beads were added to the medium and incubated at 37°C for 3 h. The fluorescence intensity was quantified using BioTek reader. The data are presented as mean ± SEM, n = 3 independent experiments; *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001, analyzed by one-way ANOVA followed by Bonferroni's test.

Fig 3: Aß-induced gene expression of senescence in human microglial cells. (a–h) Human microglial cells were treated with 10 µM Aß for different times. (a–d) Relative genes expression levels of PAI-1, p21, CCND1, and CCNA1 were measured by qPCR. (e) Protein levels of p53, PAI-1, and p21 were detected by western blotting. (f–h) The quantification of relative protein levels in (e). (i–n) Human microglial cells were stimulated with Aß (1-10 µM) for 72 h. (i) Western blotting was used to investigate protein levels of p53, PAI-1, and p21. (j–l) The quantification of relative protein levels in (i). (m) The representative SA-ß-gal staining images. (n) Quantification of the percentage of SA-ß-gal cells. Scar bar, 100 µm. (o–q) Senescence-associated proinflammatory cytokines such as TNF-a, IL-1ß, and IL-6 were detected by qPCR. The data are presented as mean ± SEM, n = 3 independent experiments; *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001, analyzed by one-way ANOVA followed by Bonferroni's test.