Journal: Journal of Translational Medicine

Article Title: CircPRDM5-mediated regulation of miR-433-3p and HDAC6 in Parkinson's disease: a novel neuroprotective axis

doi: 10.1186/s12967-025-06602-3

Figure Lengend Snippet: HDAC6 is a direct target of miR-433-3p. A miRDB analysis of miR-433-3p binding sites in HDAC6. B qRT-PCR and western blot analysis of HDAC6 expression in SH-SY5Y cells transfected with either miR-NC or miR-433-3p mimics. C Dual-luciferase reporter activity for WT/Mut groups in SH-SY5Y cells transfected with either miR-NC or miR-433-3p mimics. D RNA pull-down assay was performed to assess the binding ability between miR-433-3p and HDAC6. E qRT-PCR analysis of HDAC6 expression in the peripheral blood samples of PD patients (n = 20) and healthy control (n = 20). F , G qRT-PCR and western blot analysis of HDAC6 expression in SH-SY5Y cells either treated with different concentrations of MPP + (0, 0.5, 1 or 2 mM) or with 1 mM of MPP + for different time duration (0, 12, 24 or 48 h). H qRT-PCR analysis of HDAC6 expression in SH-SY5Y cells left untreated or treated with 1 mM of MPP + for 48 h of sh-NC group, sh-circPRDM5 group, sh-circPRDM5 + anti-NC group, and sh-circPRDM5 + anti-miR-433-3p group. Data are shown as the mean ± SD of three independent experiments. P values were calculated using either unpaired Student's t-test or one-way ANOVA followed by Bonferroni post hoc test. **p < 0.01, ***p < 0.001, ## p < 0.01

Article Snippet: After protein transfer, the membranes were blocked with 5% non-fat dry milk in TBST (Tris-buffered saline with 0.1% Tween-20) and then incubated with the following primary antibodies overnight at 4 °C: Bax (5023S, 1:1000, Cell Signaling Technology, USA), Bcl-2 (2876S, 1:1000, Cell Signaling Technology, USA), LC3 I/II (12741S, 1:1000, Cell Signaling Technology, USA), p62/SQSTM1 (5114S, 1:1000, Cell Signaling Technology, USA), HDAC6 (7558S, 1:1000, Cell Signaling Technology, USA), and β-actin (3700, 1:1000, Cell Signaling Technology, USA) as a loading control.

Techniques: Binding Assay, Quantitative RT-PCR, Western Blot, Expressing, Transfection, Luciferase, Activity Assay, Pull Down Assay, Control

Journal: Journal of Translational Medicine

Article Title: CircPRDM5-mediated regulation of miR-433-3p and HDAC6 in Parkinson's disease: a novel neuroprotective axis

doi: 10.1186/s12967-025-06602-3

Figure Lengend Snippet: CircPRDM5 promotes MPP + -induced neuronal damage via HDAC6. SH-SY5Y cells were first transfected with either sh-circPRDM5 alone or together with the HDAC6 overexpression vector (OE-HDAC6) or their corresponding negative control vectors. After transfection, the cells were left untreated or treated with 1 mM MPP + for 48 h. A qRT-PCR analysis was performed to assess HDAC6 expression in indicated groups of SY5Y cells without any treatment. B CCK8 cell viability assay in SH-SY5Y cells of the sh-NC group, sh-circPRDM5 group, sh-circPRDM5 + OE-NC group and sh-circPRDM5 + OE-HDAC6 group. C LDH levels in SH-SY5Y cells of the indicated groups. D Flow cytometry analysis was performed to assess apoptosis rate in SH-SY5Y cells of the indicated groups. E Western blot assay for apoptosis-related proteins in SH-SY5Y cells of the indicated groups. F Western blot assay for autophagy-related proteins in SH-SY5Y cells of the indicated groups. G Immunofluorescence staining for LC3 protein in SH-SY5Y cells of the indicated groups (scale bar = 50 μm). Data are shown as the mean ± SD of three independent experiments. P values were calculated using either unpaired Student's t-test or one-way ANOVA followed by Bonferroni post hoc test. **p < 0.01, ***p < 0.001, # p < 0.05, ## p < 0.01, ### p < 0.001

Article Snippet: After protein transfer, the membranes were blocked with 5% non-fat dry milk in TBST (Tris-buffered saline with 0.1% Tween-20) and then incubated with the following primary antibodies overnight at 4 °C: Bax (5023S, 1:1000, Cell Signaling Technology, USA), Bcl-2 (2876S, 1:1000, Cell Signaling Technology, USA), LC3 I/II (12741S, 1:1000, Cell Signaling Technology, USA), p62/SQSTM1 (5114S, 1:1000, Cell Signaling Technology, USA), HDAC6 (7558S, 1:1000, Cell Signaling Technology, USA), and β-actin (3700, 1:1000, Cell Signaling Technology, USA) as a loading control.

Techniques: Transfection, Over Expression, Plasmid Preparation, Negative Control, Quantitative RT-PCR, Expressing, Viability Assay, Flow Cytometry, Western Blot, Immunofluorescence, Staining

Journal: Journal of Translational Medicine

Article Title: CircPRDM5-mediated regulation of miR-433-3p and HDAC6 in Parkinson's disease: a novel neuroprotective axis

doi: 10.1186/s12967-025-06602-3

Figure Lengend Snippet: CircPRDM5 knockdown alleviates PD symptoms in MPTP-induced mice. A – C Pole test, rotarod test and open field test were implemented to evaluate the motor function of mice in the MPTP + sh-NC group (n = 6) and MPTP + sh-circPRDM5 group (n = 6). D qRT-PCR analysis of circPRDM5 expression in substantia nigra (SN) tissue samples from mice in MPTP + sh-NC group (n = 6) and MPTP + sh-circPRDM5 group (n = 6). E HE staining of SN samples from indicated groups of mice (n = 6) (scale bar = 20 μm). F TUNEL staining of SN samples from indicated groups of mice (n = 6) (scale bar = 50 μm). G Immunohistochemical detection of TH and HDAC6 expression in SN samples from mice in the indicated groups (n = 6) (scale bar = 20 μm). H Western blot detection of HDAC6 protein in SN samples from indicated groups of mice (n = 6). Data are shown as the mean ± SD from 6 mice per group, and statistical analyses were performed using unpaired Student’s t-test. *p < 0.05, **p < 0.01, ***p < 0.001

Article Snippet: After protein transfer, the membranes were blocked with 5% non-fat dry milk in TBST (Tris-buffered saline with 0.1% Tween-20) and then incubated with the following primary antibodies overnight at 4 °C: Bax (5023S, 1:1000, Cell Signaling Technology, USA), Bcl-2 (2876S, 1:1000, Cell Signaling Technology, USA), LC3 I/II (12741S, 1:1000, Cell Signaling Technology, USA), p62/SQSTM1 (5114S, 1:1000, Cell Signaling Technology, USA), HDAC6 (7558S, 1:1000, Cell Signaling Technology, USA), and β-actin (3700, 1:1000, Cell Signaling Technology, USA) as a loading control.

Techniques: Knockdown, Quantitative RT-PCR, Expressing, Staining, TUNEL Assay, Immunohistochemical staining, Western Blot

Journal: Oncogene

Article Title: PTEN activation through K163 acetylation by inhibiting HDAC6 contributes to tumour inhibition.

doi: 10.1038/onc.2015.293

Figure Lengend Snippet: Figure 5. Inhibition of HDAC6 induced PTEN acetylation and membrane translocation. (a) Non-selective HDAC inhibitor SAHA (inhibitor for HDAC1–11) and HDAC6-specific inhibitor tubastatin A, but not NaBu (inhibitor for HDAC1–9, except HDAC6), induced PTEN acetylation. PTEN- null U-87 MG cells were transfected with EGFP-PTEN for 24 h and then treated with different inhibitors for 24 h. (b) Confirmation of knockdown of HDAC10 and HDAC11. 293Tcells were transfected with scrambled siRNA, HDAC10 or HDAC11 siRNA for 48 h. Protein expression was detected by western blot analysis, and mRNA expression was quantified through real-time PCR. (c) Knockdown of HDAC10 or HDAC11 did not affect PTEN acetylation. Specific siRNA for HDAC10 or HDAC11 was transfected into 293T cells for 48 h. Whole-cell lysates were immunoprecipitated with PTEN antibody and subjected to western blot with anti-acetyl lysine antibody. (d) Overexpression and knockdown of HDAC6 decreased and increased PTEN acetylation, respectively. 293T cells were transfected with HDAC6 or HDAC6 siRNA for 48 h; the cells and whole-cell lysates were immunoprecipitated with PTEN antibody and detected with acetyl lysine antibody. (e) Overexpression and knockdown of HDAC6 decreased and increased PTEN membrane translocation, respectively. 293T cells were transfected with HDAC6 or HDAC6 siRNA for 48 h. Total protein and membrane protein were subjected to western blot and detected with PTEN antibody. (f) HDAC6 interacted with PTEN. Cell lysates of 293T cells were subjected to co-immunoprecipitation assay with either anti-IgG, anti-PTEN or anti-HDAC6 and subsequently allowed to bind to anti-PTEN or anti-HDAC6. *Po0.01. IP, immunoprecipitation; WB, western blot.

Article Snippet: Anti-PTEN antibodies (#9552), anti-phospho-AKT (Thr308) antibodies (#9275), anti-GFP antibodies (#2555), anti-acetylated-lysine rabbit monoclonal antibodies (Ac-K-103) (#9068), anti-acetylated-lysine mouse mAb (Ac-K-103) (#9681) and anti-HDAC6 antibodies (#7612) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Inhibition, Membrane, Translocation Assay, Transfection, Knockdown, Expressing, Western Blot, Real-time Polymerase Chain Reaction, Immunoprecipitation, Over Expression, Co-Immunoprecipitation Assay

Journal: Oncogene

Article Title: PTEN activation through K163 acetylation by inhibiting HDAC6 contributes to tumour inhibition.

doi: 10.1038/onc.2015.293

Figure Lengend Snippet: Figure 6. Inhibition of HDAC6 induced PTEN acetylation at K163 and membrane translocation. (a–d) Inhibition of HDAC6 by tubastatin A or tubacin induced the acetylation and membrane translocation of wild-type PTEN, but not K163R mutant. U-87 MG or NCl-H1650 cells were transfected with EGFP-PTEN or EGFP-PTEN-K163R for 24 h and then treated with tubastatin A or tubacin for 24 h. Experiments were performed as in Figures 2d and 3a. *Po0.01. (e) Fluorescent microphotographs of 293T cells transfected with EGFP-PTEN or EGFP-PTEN-K163R and treated with tubastatin A for 24 h. (f, g) Knockdown of HDAC6 increased acetylation and membrane translocation of wild-type PTEN, but not K163R mutant. HDAC6 siRNA was co-transfected with EGFP-PTEN or EGFP-PTEN-K163R into U-87 MG or NCl-H1650 cells for 48 h. Experiments were performed as described above. *Po0.01. IP, immunoprecipitation; WB, western blot.

Article Snippet: Anti-PTEN antibodies (#9552), anti-phospho-AKT (Thr308) antibodies (#9275), anti-GFP antibodies (#2555), anti-acetylated-lysine rabbit monoclonal antibodies (Ac-K-103) (#9068), anti-acetylated-lysine mouse mAb (Ac-K-103) (#9681) and anti-HDAC6 antibodies (#7612) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Inhibition, Membrane, Translocation Assay, Mutagenesis, Transfection, Knockdown, Immunoprecipitation, Western Blot

Journal: Scientific Reports

Article Title: Selective HDAC6 inhibitor WT161 modulates the VLA-4/FAK pathway by inhibiting PKA activity in acute lymphoblastic leukemia

doi: 10.1038/s41598-025-23887-y

Figure Lengend Snippet: HDAC6 expression in ALL cell lines and the effect of WT161 on HDAC6, acetyl-α-tubulin, and cell proliferation. ( a ) HDAC6 protein expression in four ALL cell lines (BALL-1, NALM-6, Jurkat, and MOLT-4) as detected by Western blot. ( b ) Relative HDAC6 mRNA levels measured by quantitative PCR (qPCR), showing expression trends consistent with ( a ). ( c ) Western blot analysis of HDAC6 and acetyl-α-tubulin protein levels in cell lines treated with or without 5 µM WT161 for 72 h. ( d ) Concentration- and time-dependent inhibition of cell proliferation by WT161 across all four ALL cell lines. IC 50 values were calculated to quantify compound sensitivity.

Article Snippet: The following antibodies were used for Western blot and other applications: HDAC6, β1 integrin, and α-tubulin (ABclonal, China); acetylated α-tubulin, α4 integrin, phospho-FAK, and phospho-EGFR (Affinity Biosciences, China); AP-conjugated α4 integrin (Sigma-Aldrich, China); FAK, EGFR, goat anti-rat IgG-HRP, and α-tubulin (Wanlei Biotechnology, China).

Techniques: Expressing, Western Blot, Real-time Polymerase Chain Reaction, Concentration Assay, Inhibition