Fig. 1
Mitigation of motor decline by preserving nigral DA neurons during aging. a Immunofluorescence staining for TH and SIRT3 in the SNpc of young and aged mice (scale bar, 50 μm; left upper panels) and Western blotting of TH, p-TH, and SIRT3 in the SN (right upper panels). A.U., arbitrary unit. Immunofluorescence staining and quantification for cellular colocalization of AAV-GFP and AAV-SIRT3-induced GFP in the SN 4 weeks post-injection (scale bars, 200 μm and 50 μm; lower panels). b GO enrichment analysis for BP, CC, and GSEA of ATP production and DA signaling in the SN 4 weeks post-injection of AAV-GFP or AAV-SIRT3 (upper panels). Western blot analysis for SIRT3, p16INK4a, and mitochondrial proteins in the SN 4 weeks and 18 months post-injection of AAV-GFP or AAV-SIRT3 (lower panels). c Immunohistochemical staining for TH in the SN of 3- and 20-month-old intact controls, as well as 20-month-old experimental mice 18 months post-injection with AAV-GFP or AAV-SIRT3 (scale bar, 500 μm), along with quantification of TH-positive neurons in the SNpc (upper panels). VTA, ventral tegmental area. Western blot analysis for cleaved Caspase-3 (c-Caspase-3), cleaved poly (ADP-ribose) polymerase 1 (c-PARP-1), phosphorylated-mixed lineage kinase domain-like protein (p-MLKL), and phosphorylated-Receptor-interacting serine/threonine-protein kinase 3 (p-RIP3), markers associated with cell death in the SN (middle panels). Immunohistochemical staining for TH in the striatum (STR; scale bar, 500 μm; left lower panels), and HPLC analysis for DA and HVA in the STR of mice (right lower panels). d Behavioral tests (rotarod and grip strength) were performed every 2 months until 20 months of age (upper panels). DXA scans for body composition and ROI analysis (middle panels). Western blotting for p16INK4a and synaptophysin in muscle tissues (lower panels). Statistical analyses were performed using Student’s t-test, one-way or two-way ANOVA with Tukey’s post hoc test, or the Kruskal–Wallis test (for HVA and LBM). Data as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001 vs. 3-month-old intact or between the indicated groups (n = 14 for behavioral tests; n = 6 for other analyses)
