bafilomycin a1 bfa  (InvivoGen)

Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology

Article Title: Amphiregulin improves ventricular remodeling after myocardial infarction by modulating autophagy and apoptosis.

doi: 10.1096/fj.202302385R

Figure Lengend Snippet: FIGURE 4 Areg deficiency upregulates autophagosome formation and inhibits autophagosome clearance. (A) LC3-II, p62, and GAPDH protein levels in the infarct border myocardium from WT and Areg−/− mice were quantified by western blotting on day 7 after MI. Left, representative immunoblot. Right, summary data. n = 4 per group. (B) WT and Areg−/− mice treated with 3-MA (30 μg/g/day) or vehicle were administered at the same time each day for 7 days after MI. LC3-II and GAPDH protein levels in the infarct border myocardium from WT and Areg−/− mice treated with 3-MA or vehicles were quantified by western blotting. Top, representative immunoblot. Bottom, summary data. n = 6 per group. (C) Seven days after MI, WT, and Areg−/− mice were injected with BFA (1.5 μg/g) or vehicles and sacrificed 2 h after for the assays. LC3-II and GAPDH protein levels in the infarct border myocardium from WT and Areg−/− mice treated with BFA or vehicles were quantified by western blotting. Top, representative immunoblot. Bottom, summary data. n = 5 per group. (D) Electron micrographs of the infarct border myocardium from WT (left) and Areg−/− (middle) hearts, were analyzed for the autophagic vacuole density (right). n = 4–5 per group. Scale bar, 1 μm. Data are expressed as means ± SEM. *p < .05, **p < .01. Data in (A) were analyzed by a two-tailed unpaired t-test. One-way ANOVA with the Tukey post hoc test was performed in (B) and (C). Data in (D) were analyzed with the Mann–Whitney test. GAPDH, glyceraldehyde 3-phosphate dehydrogenase; i.p., intraperitoneal injection; 3-MA, 3-methyladenine; BFA, bafilomycin-A1.

Article Snippet: Bafilomycin- A1 (BFA) (No. HY- 100558; MedChemExpress) was dissolved in corn oil.

Techniques: Western Blot, Injection, Two Tailed Test, MANN-WHITNEY

Journal: Advanced Science

Article Title: Single‐Cell RNA Sequencing of Retina Reveals Nna1 Upregulation in Myopic Diabetic Retinopathy as a Protective Factor Against Diabetic Damage

doi: 10.1002/advs.202500438

Figure Lengend Snippet: Nna1 overexpression inhibited HG‐induced autophagy and apoptosis pathways in rMC‐1 cells by regulating Tubulin Polyglutamylation. A‐B) Western blots images depicting the protein expression of LC3B in rMC‐1 cells transfected with oe‐NC or oe‐Nna1. BFA (400 n m ) was administered to the groups for 6 h prior to protein extraction. ( n = 3/group). C,D) Western blots images showing the protein level of Nna1, P62, Bcl2, Beclin1, Casp3, Bax, and Bim in rMC‐1 cells treated with HG or untreated, and transfected with oe‐NC or oe‐Nna1. ( n = 3/group). E,F) Immunofluorescence staining and quantifications of rMC‐1 cells transfected with oe‐NC or oe‐Nna1 with or without HG treatment. Cells were stained with LC3 (green), P62 (red), and DAPI (Blue). Scale bars indicate 10 µm. ( n = 6/group). G,H) Immunofluorescence staining and quantifications of rMC‐1 cells transfected with oe‐NC or oe‐Nna1 with or without HG treatment. Cells were stained with Casp3 (green), BAX (red), and DAPI (Blue). Scale bars indicate 10 µm. ( n = 6/group). I) The binding affinity between Nna1 and a‐tubulin and its effect on polyglutamylation tubulin after Nna1 overexpression with or without HG treatment was evaluated using Co‐IP assays. ( n = 3/group). Results expressed as mean ± SEM. * p <0.05, ** p <0.01, *** p <0.001. P values were determined by one‐way ANOVA (A,B,D,F,H).

Article Snippet: To induce autophagy, cells were treated with 100 n m BFA (S1413, Selleck).

Techniques: Over Expression, Western Blot, Expressing, Transfection, Protein Extraction, Immunofluorescence, Staining, Binding Assay, Co-Immunoprecipitation Assay