sirtuin 1  (Proteintech)

Journal: Frontiers in Molecular Neuroscience

Article Title: Caffeine improves mitochondrial dysfunction in the white matter of neonatal rats with hypoxia-ischemia through deacetylation: a proteomic analysis of lysine acetylation

doi: 10.3389/fnmol.2024.1394886

Figure Lengend Snippet: Caffeine promoted the effects of deacetylation after hypoxia-ischemia-induced WMD in neonatal rats. (A) Western blots of proteins in the white matter of brains of neonatal rats on day 14 using an anti-acetylated-lysine antibody. (B) Analyses of relative acetylated-lysine levels, with β-tubulin used for normalization. (C) Gene Ontology (GO) of SIRT2. (D) Western blot detection of SIRT2. (E) Relative protein level of SIRT2. (F) Analysis of the protein-protein interaction (PPI) networks of differential acetylation modification and proteome protein. (G) Polymerase chain reaction (PCR) analysis to determine mRNA levels of Mapt. Mapt levels are normalized against those of glyceraldehyde 3-phosphate dehydrogenase (GADPH) and expressed as the fold change. (H) Mapt immunoprecipitation with anti-acetyl-lysine antibody and analysis with anti-tau. (I) Analyses of relative total Mapt levels, with β-actin used for normalization. (J) Analyses of relative acetyl-Mapt levels, with Mapt used for normalization. Data is presented as the mean ± standard error of the mean. Statistical analyses involved one-way ANOVA. The artwork with “ * ” indicate statistically significant differences between groups. * p < 0.05, ** p < 0.01, *** p < 0.001. Sham group ( n = 6); HI group ( n = 6); Caffeine group ( n = 6); Caffeine+AK-7 group ( n = 6).

Article Snippet: Furthermore, in the Caffeine+AK-7 group, rats were subjected to treatment with both caffeine and the sirtuin 2 (SIRT2) inhibitor 3-(1-azepanylsulfonyl)-N-(3-bromphenyl) benzamide (AK-7) (s591401, Selleck Chemicals, USA).

Techniques: Western Blot, Modification, Polymerase Chain Reaction, Immunoprecipitation

Journal: Frontiers in Molecular Neuroscience

Article Title: Caffeine improves mitochondrial dysfunction in the white matter of neonatal rats with hypoxia-ischemia through deacetylation: a proteomic analysis of lysine acetylation

doi: 10.3389/fnmol.2024.1394886

Figure Lengend Snippet: Caffeine deacetylated Mapt through SIRT2 after hypoxia-ischemia-induced WMD in neonatal rats. (A) Western blot detection of Bcl-2, Caspase-3, cytochrome C (Cyt-C) and mitochondrial transcription factor A (TFAM). (B) Analyses of relative Bcl-2, (C) Caspase-3, (D) Cyt-C, and (E) TFAM levels, with β-actin used for normalization. (F) Mapt was immunoprecipitated with anti-acetyl-lysine antibody and analyzed with anti-tau. (G) Analyses of relative total Mapt levels, with β-actin used for normalization. (H) Analyses of relative acetyl-Mapt levels, with Mapt used for normalization. (I) Western blot detection of nuclear Mapt. (J) Analyses of relative nuclear Mapt protein levels, with Lamin B used for normalization. (K) ELISA to determine ATP levels. (L) Polymerase chain reaction (PCR) analysis to determine mitochondrial DNA (mtDNA) levels. Levels are normalized against those of glyceraldehyde 3-phosphate dehydrogenase (GADPH) and are expressed as the fold change. (M) Mitochondrial Membrane Potential (MMP) assay. (N) Change rate of MMP. Data is presented as the mean ± standard error of the mean. Statistical analyses involve one-way ANOVA. The artwork with “*” indicate statistically significant differences between groups. * p < 0.05, ** p < 0.01, *** p < 0.001. Sham group ( n = 6); HI group ( n = 6); Caffeine group ( n = 6); Caffeine+AK-7 group ( n = 6).

Article Snippet: Furthermore, in the Caffeine+AK-7 group, rats were subjected to treatment with both caffeine and the sirtuin 2 (SIRT2) inhibitor 3-(1-azepanylsulfonyl)-N-(3-bromphenyl) benzamide (AK-7) (s591401, Selleck Chemicals, USA).

Techniques: Western Blot, Immunoprecipitation, Enzyme-linked Immunosorbent Assay, Polymerase Chain Reaction, Membrane, Mmp Assay

Journal: Frontiers in Molecular Neuroscience

Article Title: Caffeine improves mitochondrial dysfunction in the white matter of neonatal rats with hypoxia-ischemia through deacetylation: a proteomic analysis of lysine acetylation

doi: 10.3389/fnmol.2024.1394886

Figure Lengend Snippet: Mechanism diagram of caffeine inhibiting Mapt acetylation by upregulating SITR2, promoting its nuclear translocation, and improving mitochondrial damage. (A) In normal brain tissue, Mapt is deacetylated and translocated into the nucleus, thereby promoting mitochondrial transcription in the nucleus and maintaining mitochondrial function. (B) After hypoxia-ischemia-induced WMD, the acetylation level of Mapt increased, nuclear translocation decreased, mitochondrial transcription decreased, and damage increased. Induced to caffeine treatment, SIRT2 was activated, Mapt deacetylation increased, nuclear translocation increased, mitochondrial transcription increased, and damage improved.

Article Snippet: Furthermore, in the Caffeine+AK-7 group, rats were subjected to treatment with both caffeine and the sirtuin 2 (SIRT2) inhibitor 3-(1-azepanylsulfonyl)-N-(3-bromphenyl) benzamide (AK-7) (s591401, Selleck Chemicals, USA).

Techniques: Translocation Assay