Journal: Reviews in Cardiovascular Medicine

Article Title: Enhancer of Zeste Homolog 2 Inhibition Attenuates Atrial Fibrillation Post-Myocardial Infarction Through Reduced Atrial Fibrosis

doi: 10.31083/RCM51343

Figure Lengend Snippet: MI induces atrial fibrosis and upregulates the EZH2/H3K27me3 signaling in mice . (A,B) Representative Masson staining images (A) and quantitative analysis (B) of atrial fibrosis in the sham and MI groups ( n = 5). Scale bar = 0.04 mm under 20× field of view. (C–E) Western blot analysis and quantitative analysis of the expression levels of collagen I, α-SMA, and TGF-β1 in the atria of mice from the Sham and MI groups ( n = 6). (F) qPCR analysis of the expression levels of Ezh2 mRNA in the atria of mice from the Sham and MI groups ( n = 6). (G,H) Western blot analysis and quantitative analysis of the expression levels of EZH2 ( n = 6) and H3K27me3 ( n = 9) in the atria of mice from the Sham and MI groups. Sham, Sham operation group; MI, myocardial infarction group; α-SMA, alpha-smooth muscle actin; TGF-β1, transforming growth factor-beta1; EZH2, Enhancer of Zeste Homolog 2; H3K27me3, trimethylated Histone H3 at Lysine 27. Values are displayed as mean ± standard error of the mean.

Article Snippet: The antibodies utilized in this study comprised the following: Enhancer of Zeste Homolog 2 (EZH2, 5246S, CST, USA), trimethylated Histone H3 at Lysine 27 (H3K27me3, 9733S, CST, USA), Histone 3 (H3, 9715S, CST, USA), collagen I (14695-1-AP, Proteintech, USA), Transforming Growth Factor-beta1 (TGF-β1, 81746-2-RR, Proteintech, USA), Alpha-Smooth Muscle Actin (α-SMA, 14395-1-AP, Proteintech, USA), and Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH, 60004-1-lg, Proteintech, USA).

Techniques: Staining, Analysis, Western Blot, Expressing

Journal: Reviews in Cardiovascular Medicine

Article Title: Enhancer of Zeste Homolog 2 Inhibition Attenuates Atrial Fibrillation Post-Myocardial Infarction Through Reduced Atrial Fibrosis

doi: 10.31083/RCM51343

Figure Lengend Snippet: EZH2 inhibitor GSK126 ameliorates post-MI AF . (A–C) Western blot analysis and quantitative analysis of the expression levels of EZH2 and H3K27me3 in the atria of mice from the MI + Vehicle and MI + GSK126 groups ( n = 6). (D) Representative ECG of mice in the MI + Vehicle and MI + GSK126 groups under resting state and AF-induced state. (E–H) Quantitative analysis of P wave duration (E) and PR interval (F) under resting state, and comparison of AF induction rate (G) and duration time (H) under AF-induced state in the two groups ( n = 7). MI + Vehicle, myocardial infarction with vehicle control group; MI + GSK126, myocardial infarction group treated with GSK126; EZH2, Enhancer of Zeste Homolog 2; H3K27me3, trimethylated Histone H3 at Lysine 27. Values are displayed as mean ± standard error of the mean, except for (G) where values are n .

Article Snippet: The antibodies utilized in this study comprised the following: Enhancer of Zeste Homolog 2 (EZH2, 5246S, CST, USA), trimethylated Histone H3 at Lysine 27 (H3K27me3, 9733S, CST, USA), Histone 3 (H3, 9715S, CST, USA), collagen I (14695-1-AP, Proteintech, USA), Transforming Growth Factor-beta1 (TGF-β1, 81746-2-RR, Proteintech, USA), Alpha-Smooth Muscle Actin (α-SMA, 14395-1-AP, Proteintech, USA), and Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH, 60004-1-lg, Proteintech, USA).

Techniques: Western Blot, Analysis, Expressing, Comparison, Control

Journal: Reviews in Cardiovascular Medicine

Article Title: Enhancer of Zeste Homolog 2 Inhibition Attenuates Atrial Fibrillation Post-Myocardial Infarction Through Reduced Atrial Fibrosis

doi: 10.31083/RCM51343

Figure Lengend Snippet: Inhibition of EZH2 attenuates atrial fibrosis post-MI . (A,B) Representative Masson staining images (A) and quantitative analysis (B) of atrial fibrosis in the MI + Vehicle and MI + GSK126 groups ( n = 5). Scale bar = 0.04 mm under 20× field of view. (C–E) Western blot analysis and quantitative analysis of the expression levels of collagen I, α-SMA, and TGF-β1 in the atria of mice from the MI + Vehicle and MI + GSK126 groups ( n = 6). MI + Vehicle, myocardial infarction with vehicle control group; MI + GSK126, myocardial infarction group treated with GSK126; EZH2, Enhancer of Zeste Homolog 2; α-SMA, alpha-smooth muscle actin; TGF-β1, transforming growth factor-beta1. Values are displayed as mean ± standard error of the mean.

Article Snippet: The antibodies utilized in this study comprised the following: Enhancer of Zeste Homolog 2 (EZH2, 5246S, CST, USA), trimethylated Histone H3 at Lysine 27 (H3K27me3, 9733S, CST, USA), Histone 3 (H3, 9715S, CST, USA), collagen I (14695-1-AP, Proteintech, USA), Transforming Growth Factor-beta1 (TGF-β1, 81746-2-RR, Proteintech, USA), Alpha-Smooth Muscle Actin (α-SMA, 14395-1-AP, Proteintech, USA), and Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH, 60004-1-lg, Proteintech, USA).

Techniques: Inhibition, Staining, Analysis, Western Blot, Expressing, Control

Journal: Journal of Cell Science

Article Title: Interplay between nuclear survivin and the PRC2 complex and its impact on H3K27me3-directed transcriptional repression

doi: 10.1242/jcs.264572

Figure Lengend Snippet: Hypoxia increases expression of EZH2, H3K27me3 and survivin. (A) Immunoblots of WCEs from U2OS, HeLa and MRC5 lines cultured under normoxic or hypoxic environments (24 h). Blots were immunoprobed with anti-EZH2, anti-H3K27me3 and anti-survivin antibodies. Anti-Hif1a used to prove the hypoxic state had been induced, and anti-tubulin was used as a loading control. (B–D) Quantification of immunoblots represented in A from three independent experiments demonstrating that EZH2, H3K27me3 and survivin are all more abundant under hypoxia. Data presented are means±s.d. * P <0.05, ** P <0.01, *** P <0.001 (two-way ANOVA with Tukey's multiple comparisons post test).

Article Snippet: The following primary antibodies were used at 1:200: anti-survivin (Cell Signaling Technologies 71G4B7 or 6E4), anti-H3K27Me3 (Abcam, Ab192985 ), anti-EZH2 [Cell Signalling Technologies (CST), D269].

Techniques: Expressing, Western Blot, Cell Culture, Control

Journal: Journal of Cell Science

Article Title: Interplay between nuclear survivin and the PRC2 complex and its impact on H3K27me3-directed transcriptional repression

doi: 10.1242/jcs.264572

Figure Lengend Snippet: Survivin and EZH2 interact. (A) Immunoprecipitation was carried out using whole MRC5 extracts using anti-survivin (C60), anti-EZH2, mouse IgG antibodies (negative control). Co-immunoprecipitation was assessed with the alternative antibodies. Co-immunoprecipitation of EZH2 with survivin was evident when anti-EZH2 was used to immunoprecipitate but not when the anti-survivin (C60) antibody was used. (B) GST pulldown assay was carried out with WCEs prepared from RPE cells expressing GST (negative control), GST–survivin and various GST-tagged survivin truncations, (numbering indicating amino acids), used as bait. (C) Quantification of interactions represented in B. EZH2 binds mainly to the first 90 amino acids of survivin. Data are mean±s.d. from three independent experiments. *** P <0.001; **** P <0.0001; ns, not significant (one-way ANOVA with Dunnett's post hoc test). (D) Immunoprecipitation was carried out as in A but using anti-H3K27me3 specific antibodies, rather than anti-EZH2. Co-immunoprecipitation of survivin and H3K27me3 was evident in reciprocal samples. (E) The GST pulldown experiment as in B was repeated using RPE cell lysates with GST or GST–survivin, and interaction with H3K27me3 determined by immunoblotting. (F) Quantification of data represented in E, normalised to the GST or GST–survivin. Data are mean±s.d., n =3. *** P <0.001 (unpaired two-tailed Student's t -test). Blots in A and D are representative of three repeats. Inputs are 7.5%.

Article Snippet: The following primary antibodies were used at 1:200: anti-survivin (Cell Signaling Technologies 71G4B7 or 6E4), anti-H3K27Me3 (Abcam, Ab192985 ), anti-EZH2 [Cell Signalling Technologies (CST), D269].

Techniques: Immunoprecipitation, Negative Control, GST Pulldown Assay, Expressing, Western Blot, Two Tailed Test