Tim23 Antibody - #DF12052

Fig. 1 Morphine induced NLRX1-mediated mitophagy in microglia. A The mitochondrial DNA (mtDNA) levels were decreased significantly in primary microglia with 1.0 μM morphine as measured by mtDNA/nDNA analysis (n = 4). B Co-staining of HSP60 or Tim23 and LC3B in primary microglia. Bar = 10 μm. C The quantified results of AD-mCherry-GFP-LC3B transfection in BV2 cells with morphine treatment (n > 50 cells). The Kruskal–Wallis test was employed to indicate statistical significance. The results of mCherry-dots are shown in red and the mCherry-GFP-dots are shown in yellow. D Representative Western blots of BV2 cells in control or morphine treatment group (n = 3–6). E The expression of NLRX1 mRNA was peaked in primary microglia with 1.0 μM morphine as measured by qPCR (n = 3). One-way ANOVA were employed in A, E. F Quantitative graphs of NLRX1 mRNA in BV2 cells (n = 6). G, H Confocal microscopy analysis of NLRX1 (red), LC3B (green) and DAPI (light blue). Bar = 2 μm. The Pearson’s correlation coefficients of NLRX1 and LC3B were elevated in morphine-treated BV2 cells (H, n = 6–9 fields). I Co-immunoprecipitation analysis of NLRX1 and LC3 in BV2 cells (n = 3). J, K Western blots analysis of NLRX1-mediated mitophagy in BV2 cells by siRNA (J, n = 5) or shRNA (K, n = 3). L The quantified results of AD-mCherry-GFP-LC3B transfection in BV2 cells with NC-siRNA or NLRX1-siRNA pre-treatment (n > 50 cells). Data represent the mean ± SEM. Student's t-test or Mann–Whitney U test were used to measure significance between two groups. (* p < 0.05, ** p < 0.01, *** p < 0.001 and ns p > 0.05)

Fig. 2 NLRX1-mediated mitophagy occurred in microglia independent of PINK1–Parkin pathway, but not astrocyte (A, B) Quantitative graphs of NLRX1 mRNA (A, n = 6) and representative Western blots of mitophagy (B, n = 3) in primary astrocyte. Quantitative graphs of NLRX1 mRNA (C, n = 6) and representative Western blots of mitophagy (D, n = 3) in MA cells. E Co-staining of HSP60 or Tim23 and LC3B in primary astrocyte. Bar = 10 μm. F The quantitative graphs of JC-1 assays as measured by flow cytometry (n = 4). G The NLRX1 mRNA expression of BV2 cells with or without CCCP treatment (n = 6). Data represent the mean ± SEM. Two-sided Student’s t tests were used to measure significance between two groups (* p < 0.05, ** p < 0.01, *** p < 0.001 and ns p > 0.05)

Fig. 8. The expression of mitochondrial-related genes in L6 myoblasts after PINK1 knockdown by lentivirus. (A) Tom 20 mRNA expression in different groups. (B) Tim 23 mRNA expression in different groups. (C) VDAC1 mRNA expression in different groups. (D) Tom 20 protein expression in different groups. (E) Tim 23 protein expression in different groups. (F) VDAC1 protein expression in different groups. #P