Fig 2: A. TEM observations after upregulating Parkin at 24 h post TNI. Red arrows indicate mitochondria, scale bar = 0.5 μm. B–F. Western blot results of upregulating Parkin on MQC associated molecules including PGC1α, Mfn2, Mfn1 and Drp1, n = 5. G-K. Western blot results of upregulating Parkin on autophagy associated molecules including P62, Timm23, Tomm20 and LC3II, n = 5. ∗P < 0.05 vs. Control group, #P < 0.05 vs. LV-Con group.

Fig 3: A-B. The effect of Parkin overexpression on autophagic flux in cultured primary cortical neurons from CAG-RFP-EGFP-LC3 transgenic mice at 24 h post TNI, scale bar = 5 μm, n = 8. C-D. Effect of Parkin overexpression on mitophagy using mito-Keima reporter lentivirus at 24 h post TNI, scale bar = 5 μm, n = 8. E-G. E-G. The effects of siRNA-PGC1α, SR18292, siRNA-Drp1, mdivi-1, 3-MA, and CQ on Parkin-mediated neuroprotection. ∗P < 0.05 vs. Control group, #P < 0.05 vs. LV-Con group, &P < 0.05 vs. LV-Parkin group.

Fig 4: Metformin mitigates chondrocyte damage via the AMPK/Drp1 signaling axis. (A and B) AMPK expression was silenced in chondrocytes using AMPK siRNA, and AMPK protein levels were evaluated by Western blot. Quantification was performed relative to GAPDH. (C) Chondrocytes were transfected with AMPK siRNA or Scramble siRNA, followed by treatment with or without IL-1β (10 ng/mL) for 24 h, and then metformin (4 mM) for 6 h. Western blot analysis was conducted to assess the expression of AMPK, p-AMPK, Drp1, p-Drp1 (Ser637), and Fis-1. (D) Densitometric analysis of AMPK, p-AMPK, and Fis-1 protein levels normalized to GAPDH. The p-Drp1 (Ser637)/Drp1 ratio was also quantified. (E) EdU staining was performed to assess DNA synthesis as an indicator of chondrocyte proliferative capacity under each treatment condition. DAPI was used to stain nuclei. Scale bar: 20 μm. Scramble siRNA served as the non-specific siRNA control for comparison with AMPK-targeting siRNA. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post-hoc multiple comparisons test: ns, P > 0.05; * P < 0.05; ** P < 0.01; *** P < 0.001. The data represent the mean ± SD from at three independent experiments.

Fig 5: A. Mitochondrial fusion/fission dynamics using Mitotracker, scale bar = 50 μm. B. Quantification of mitochondrial morphology of approximately 100 cells per condition: category 1 (elongated tubulin-like structure), category 2 (large round or short tubular), and category 3 (round or punctate). C-E. Western blot analysis of the influence of Parkin overexpression on phosphorylation of Drp1 on Ser622, Ser643, n = 5. F–H. The effects of LV-Mfn2, LV-Mfn1 and MASM7 on neuronal injury deterioration caused by LV-shParkin, n = 5. I. Western blot analysis of Parkin expression in hippocampus using AAV-Parkin prior to the MWM test, n = 3. J. Real-time quantitative PCR analysis of Parkin mRNA levels prior to the MWM test, n = 6. ∗P < 0.05 vs. Control group, #P < 0.05 vs. Control virus groups, &P < 0.05 vs. LV-shParkin group.