selective sirt2 inhibitor ak7 n 3 bromophenyl  (Tocris)

Journal: Journal of Neuroinflammation

Article Title: SIRT2 Inhibition promotes microglia LC3-associated phagocytosis via NRF2/CD36 after the experimental subarachnoid hemorrhage

doi: 10.1186/s12974-025-03623-z

Figure Lengend Snippet: Expression of SIRT2 and its prognostic impact after SAH in rats. A-B, Western blot and quantifications of SIRT2 level at different time courses. C-E, Representative images show in the temporal cortex stained with antibodies to SIRT2 and Iba1 (C), NeuN (D), or GFAP (E) at 24 h post-SAH. F, The mean fluorescence intensity (MFI) of SIRT2 immunoreactivity. G, Percentages of Iba1 + SIRT2 + , NeuN + SIRT2 + and GFAP + SIRT2 + in the temporal cortex of each group. H , Neurological score, I , brain water content ( J ) and Evan’s blue in different (Sham, SAH, Vehicle and AK7) groups at 24 h after SAH. K-L , FJC staining in the cortex with quantification. Scale bar: 50 μm. ** P < 0.01, *** P < 0.001, **** P < 0.0001

Article Snippet: SIRT2 inhibitor AK7 (MCE, USA) was dissolved in dimethyl sulfoxide (DMSO).

Techniques: Expressing, Western Blot, Staining, Fluorescence

Journal: Journal of Neuroinflammation

Article Title: SIRT2 Inhibition promotes microglia LC3-associated phagocytosis via NRF2/CD36 after the experimental subarachnoid hemorrhage

doi: 10.1186/s12974-025-03623-z

Figure Lengend Snippet: SIRT2 inhibition promotes microglia phagocytosis. A-B , Western blot and quantitative analysis of SIRT2 expressions in microglia (Con, OxyHb, DMSO and AK7) groups. C-D ,phagocytic index level and analysis by pHrodo. E-F ,Immunofluorescence staining and quantitative analysis of LC3 and Phrodo in microglia. Scale bar: 10 μm. G-K , Western blot and quantitative analysis of Rubicon, NOX2, Beclin1 and LC3 A/B expressions. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001

Article Snippet: SIRT2 inhibitor AK7 (MCE, USA) was dissolved in dimethyl sulfoxide (DMSO).

Techniques: Inhibition, Western Blot, Immunofluorescence, Staining

Journal: Journal of Neuroinflammation

Article Title: SIRT2 Inhibition promotes microglia LC3-associated phagocytosis via NRF2/CD36 after the experimental subarachnoid hemorrhage

doi: 10.1186/s12974-025-03623-z

Figure Lengend Snippet: SIRT2 deacetylates NRF2 and promotes its nuclear translocation in microglia. A-C , Heatmap analyses and q-PCR detects different NRF2 target genes in microglia (SIRT2 and SIRT2 KD) groups. D , Molecular docking shown the obvious interaction force between SIRT2 and NRF2. E-G , Co-immunoprecipitation and quantitative analysis of acetylated NRF2 in SIRT2 WT and KD microglia. H , Western blot and quantitative analysis of SIRT2 in microglia (Con, OxyHb, DMSO and AK7) group with Cyto and nucleus. I-K , The co-localization of SIRT2 and nucleus with laser confocal and analysis in Con group and OxyHb group of microglia. Scale bar: 10 μm. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, n.s., no significant

Article Snippet: SIRT2 inhibitor AK7 (MCE, USA) was dissolved in dimethyl sulfoxide (DMSO).

Techniques: Translocation Assay, Immunoprecipitation, Western Blot

Journal: Journal of Neuroinflammation

Article Title: SIRT2 Inhibition promotes microglia LC3-associated phagocytosis via NRF2/CD36 after the experimental subarachnoid hemorrhage

doi: 10.1186/s12974-025-03623-z

Figure Lengend Snippet: SIRT2 through NRF2/CD36 regulates microglial phagocytosis. A , JASPAR predict NRF2 bind to the promoter region of CD36. B-C , phagocytic index level and analysis by pHrodo in microglia (Con, OxyHb, DMSO, AK7 and AK7+ML385) groups. D-E , Western blot and quantitative analysis of CD36 expression. F-H , ELISA quantitative analysis of pro-inflammatory cytokines (TNF-α, IL-1β and IL-6) level in different groups. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001

Article Snippet: SIRT2 inhibitor AK7 (MCE, USA) was dissolved in dimethyl sulfoxide (DMSO).

Techniques: Western Blot, Expressing, Enzyme-linked Immunosorbent Assay

Journal: Journal of Neuroinflammation

Article Title: SIRT2 Inhibition promotes microglia LC3-associated phagocytosis via NRF2/CD36 after the experimental subarachnoid hemorrhage

doi: 10.1186/s12974-025-03623-z

Figure Lengend Snippet: SIRT2 via LAP inhibits phagocytosis in microglia. A-F ,Western blot and quantitative analysis of Rubicon, ULK1, CD36, NOX2 and LC3 A/B expressions. G-H , The co-localization of LC3 and pHrodo with laser confocal and analysis indifferent groups of microglia. Scale bar: 10 μm. I-J , Representative TEM images of LAPosomes in microglia. (The black triangle refers to the single membrane structure characteristic of LAPosomes and star refers to the Rubicon particles). Scale bar: 3 μm (upper panels), 1 μm (lower panels). * P < 0.05 vs. Hb, # P < 0.05 vs. Hb+AK7, & P < 0.05 vs. Hb+shRNA, $ P < 0.05 vs. Hb+AK7+shRNA, ☆ P < 0.05 vs. Hb+AK7+ML385, *** P < 0.001, **** P < 0.0001

Article Snippet: SIRT2 inhibitor AK7 (MCE, USA) was dissolved in dimethyl sulfoxide (DMSO).

Techniques: Western Blot, Membrane, shRNA

Journal: Journal of Neuroinflammation

Article Title: SIRT2 Inhibition promotes microglia LC3-associated phagocytosis via NRF2/CD36 after the experimental subarachnoid hemorrhage

doi: 10.1186/s12974-025-03623-z

Figure Lengend Snippet: Inhibition of SIRT2 improves long-term neurological function after SAH. A-B , The falling latency of rotarod test (5RPM and 10 RPM) at 1, 2, and 3 weeks after SAH in rats. C-E ,Escape latency and swimming distance of Morris water maze test on days 23 to 27 after SAH. F-G , Quantification of probe quadrant duration and swimming velocities. H-I , Representative images and neuronal quantifications of Nissl staining in CA1 hippocampal region. Scale bar: 200 μm (upper panels), 50 μm (lower panels). * P < 0.05, ** P< 0.01, *** P < 0.001, **** P < 0.0001, n.s., no significant

Article Snippet: SIRT2 inhibitor AK7 (MCE, USA) was dissolved in dimethyl sulfoxide (DMSO).

Techniques: Inhibition, Staining

Journal: Journal of Neuroinflammation

Article Title: SIRT2 Inhibition promotes microglia LC3-associated phagocytosis via NRF2/CD36 after the experimental subarachnoid hemorrhage

doi: 10.1186/s12974-025-03623-z

Figure Lengend Snippet: Schematic diagram of our study. SIRT2 modified NRF2 by deacetylation and inhibited its entry into the nucleus, affecting the transcription of downstream CD36, thereby reducing LAP in microglia

Article Snippet: SIRT2 inhibitor AK7 (MCE, USA) was dissolved in dimethyl sulfoxide (DMSO).

Techniques: Modification

Journal: PLOS Pathogens

Article Title: Bacterial infection promotes tumorigenesis of colorectal cancer via regulating CDC42 acetylation

doi: 10.1371/journal.ppat.1011189

Figure Lengend Snippet: (A) CDC42 K153 acetylation is regulated by SIRT family deacetylases. HEK293T cells transfected with Flag-CDC42 were treated with the deacetylase inhibitors TSA (2 μM) and NAM (10 mM) for 16 h before harvesting. K153 acetylation was determined by IP/WB. (B) SIRT2 decreases K153 acetylation in vivo . Overexpression of SIRT2 significantly decreases K153 acetylation. Flag-CDC42 was co-transfected with or without HA-SIRT2 into HEK293T cells, and K153 acetylation was analyzed by IP with anti-Flag antibody and WB with anti-K153Ac antibody. (C) SIRT2 decreases K153 acetylation in vitro . Purified GST-CDC42 protein was incubated with the cell lysate of HEK293T cells expressing HA-SIRT2 or the control vector. K153 acetylation was determined by the GST pull-down assay, followed by WB with anti-K153Ac antibody. (D) A reduced acetylation of K153 was shown in AK7-treated cells. Flag-CDC42 was co-transfected with or without HA-SIRT2, followed by treatment with the SIRT2 specific inhibitor AK7 (10 μM) or dimethyl sulfoxide (DMSO) (as a negative control) for 16 h. K153 acetylation was then determined by IP/WB. SIRT2 knockdown increases K153 acetylation. Flag-CDC42 was transfected with or without HA-PAK4 into HEK293T cells with SIRT2 knocked down, and the cells were infected with S . Typhimurium. Salmonella infection failed to reduce K153 acetylation and binding between CDC42 with PAK4 in the SIRT2 knockdown cells. Cell lysates were used for IP with anti-Flag antibody. K153 acetylation was then analyzed by WB with anti-K153Ac antibody (E), and the association between CDC42 and PAK4 was determined by WB (F).

Article Snippet: Anti-CDC42 (#10155-1-AP, 1:1000 for WB), anti-PAK4 (#14685-1-AP, 1:1000 for WB), anti-PAK1(#21401-1-AP, 1:1000 for WB), monoclonal anti-GST(#66001-2-Ig, 1:10000 for WB), polyclonal anti-HA (#51064-2-AP, 1:5000 for WB), monoclonal anti-SIRT2 (#66410-1-IG, 1:10000 for WB), anti-beta tubulin (#10094-1-AP, 1:2000 for WB), MMP-2 (#10373-2-AP, 1:1000 for WB), MMP-9 (#10375-2-AP, 1:1000 for WB) were purchased from Proteintech; Monoclonal anti-HA (#AB0025, 1:5000 for WB); E-cadherin (#BS1098, 1:1000 for WB) were from Bioworld; Monoclonal anti-CDC42 (#sc-8401, 2 μg per 500 μg of total protein for Immunoprecipitation), anti-p-PAK4 (sc-135775, 1:200 for WB) were from Santa Cruz; Anti-p-PAK1 (#2601, 1:1000 for WB), anti-p-AKT (#4060, 1:1000 for WB), anti-AKT (#4691, 1:1000 for WB), anti-p-ERK (#9101, 1:1000 for WB), anti-ERK (#4370, 1:1000 for WB), anti-p-p38 (#9211, 1:1000 for WB), anti-p38 (#9212, 1:1000 for WB), anti-p-JNK (#9251, 1:1000 for WB), anti-ERK (#9252, 1:1000 for WB), anti-caspase 3 (#14220, 1:1000 for WB) and anti-PARP (#9542, 1:1000 for WB) were from Cell Signaling Technology; Monoclonal anti-HA (#M180, 1:500 for Immunoprecipitation), monoclonal anti-Flag (#M185, 1:500 for Immunoprecipitation), polyclonal anti-Flag (#PM020, 1:1000 for WB) and deacetylase inhibitors TSA (#9950) were from MBL; Deacetylase inhibitor NAM (#N1651) was from APExBIO; CBP/p300 inhibitor A-485 (#N1651) was from MCE; SIRT2 inhibitor AK7 (#4754–10) was from Tocris Bioscience; Staurosporine (#abs810006) was from Absin; Polybrene (#H9268) and puromycin (#P8833) were from Sigma-Aldrich.

Techniques: Transfection, Histone Deacetylase Assay, In Vivo, Over Expression, In Vitro, Purification, Incubation, Expressing, Control, Plasmid Preparation, Pull Down Assay, Negative Control, Knockdown, Infection, Binding Assay

Journal: PLOS Pathogens

Article Title: Bacterial infection promotes tumorigenesis of colorectal cancer via regulating CDC42 acetylation

doi: 10.1371/journal.ppat.1011189

Figure Lengend Snippet: (A) Human CRC specimens were confirmed by HE staining, and the expression levels of CDC42 and CDC42 K153 acetylation and the localization of bacteria in tissues of 17 human CRC specimens were detected by IHC. CDC42 K153 acetylation level was significantly lower in the colorectal adenocarcinoma tissues than in the adjacent normal colorectal tissues as determined by IHC. The average of IHC intensity ± SD were quantitated by modified H-score from two groups of 17 patient samples (B) and 69 patients with different CRC stages (C) is shown. (D) Kaplan–Meier analysis of overall survival of 69 patients with CRC according to CDC42 K153 acetylation level. (E) Model of CDC42 K153 acetylation effect on colorectal tumorigenesis. Under physiological conditions, acetylated CDC42 K153 can maintain the normal physiological function of cells through MAPK phosphorylation (including ERK, JNK and p38) mediated by the CDC42-PAK4 signaling pathway. When Salmonella infects a host cell, SIRT2 is activated to deacetylate CDC42 K153, which causes an impaired binding of its downstream effector PAK4 and an attenuated phosphorylation of p38 and JNK, consequently reduces cell apoptosis. Moreover, low acetylation level of CDC42 K153 may contribute to the migration and invasion abilities of CRC cells, and promoting tumorigenesis, which may activate tumors mainly through the CDC42-PAK signaling axis.

Article Snippet: Anti-CDC42 (#10155-1-AP, 1:1000 for WB), anti-PAK4 (#14685-1-AP, 1:1000 for WB), anti-PAK1(#21401-1-AP, 1:1000 for WB), monoclonal anti-GST(#66001-2-Ig, 1:10000 for WB), polyclonal anti-HA (#51064-2-AP, 1:5000 for WB), monoclonal anti-SIRT2 (#66410-1-IG, 1:10000 for WB), anti-beta tubulin (#10094-1-AP, 1:2000 for WB), MMP-2 (#10373-2-AP, 1:1000 for WB), MMP-9 (#10375-2-AP, 1:1000 for WB) were purchased from Proteintech; Monoclonal anti-HA (#AB0025, 1:5000 for WB); E-cadherin (#BS1098, 1:1000 for WB) were from Bioworld; Monoclonal anti-CDC42 (#sc-8401, 2 μg per 500 μg of total protein for Immunoprecipitation), anti-p-PAK4 (sc-135775, 1:200 for WB) were from Santa Cruz; Anti-p-PAK1 (#2601, 1:1000 for WB), anti-p-AKT (#4060, 1:1000 for WB), anti-AKT (#4691, 1:1000 for WB), anti-p-ERK (#9101, 1:1000 for WB), anti-ERK (#4370, 1:1000 for WB), anti-p-p38 (#9211, 1:1000 for WB), anti-p38 (#9212, 1:1000 for WB), anti-p-JNK (#9251, 1:1000 for WB), anti-ERK (#9252, 1:1000 for WB), anti-caspase 3 (#14220, 1:1000 for WB) and anti-PARP (#9542, 1:1000 for WB) were from Cell Signaling Technology; Monoclonal anti-HA (#M180, 1:500 for Immunoprecipitation), monoclonal anti-Flag (#M185, 1:500 for Immunoprecipitation), polyclonal anti-Flag (#PM020, 1:1000 for WB) and deacetylase inhibitors TSA (#9950) were from MBL; Deacetylase inhibitor NAM (#N1651) was from APExBIO; CBP/p300 inhibitor A-485 (#N1651) was from MCE; SIRT2 inhibitor AK7 (#4754–10) was from Tocris Bioscience; Staurosporine (#abs810006) was from Absin; Polybrene (#H9268) and puromycin (#P8833) were from Sigma-Aldrich.

Techniques: Staining, Expressing, Bacteria, Modification, Binding Assay, Migration