wga dye solution  (Servicebio Inc)

Journal: Pharmacology Research & Perspectives

Article Title: β2‐Adrenergic Receptor Agonist Clenbuterol Protects Against Acute Ischemia/Reperfusion‐Induced Arrhythmia by Regulation of Akt/ eNOS / NO /Cx43 Signaling Pathway

doi: 10.1002/prp2.70070

Figure Lengend Snippet: Effect of eNOS on Cx43 dephosphorylation and effect of clenbuterol on anti‐arrhythmic effect of clenbuterol in anesthetized rats with acute ischemia/reperfusion arrhythmias. (A, B) Ventricular arrhythmias were induced by ligating the left anterior descending (LAD) artery for 10 min, followed by a 10 min reperfusion. Clenbuterol (Clenb, 0.5 mg/kg, ip ) was administrated 1 h prior to ischemia; ICI 118551 (ICI, 5 mg/kg, ip ) was administrated 1 h before injection of clenbuterol. Representative immunoblots (A), and densitometric quantitative analysis (B) of eNOS and phosphorylation of eNOS (p‐eNOS) in rat ventricular tissue ( n = 3, ** p < 0.01, *** p < 0.001). (C–I) Isolated mice hearts were subjected to 10 min global ischemia and 10 min reperfusion on a Langendorff apparatus. Clenbuterol (10 μmol/L) was administrated 10 min before global ischemia and through reperfusion (10 min); ICI 118551(1 μmol/L) or L‐NAME (100 μmol/L) pretreated the hearts for 10 min before clenbuterol treatment. (C) Arrhythmia score was recorded in indicated mice hearts. ( n = 8–10, *** p < 0.001). (D‐E) Representative immunoblots (D), and densitometric quantitative analysis (E) of eNOS and phosphorylation of eNOS (p‐eNOS) in mice ventricular tissue ( n = 3, *** p < 0.001). (F) NO concentrations were measured in indicated mice hearts. ( n = 3, * p < 0.05, *** p < 0.001). (G–I) Representative immunoblots (G), and densitometric quantitative analysis of p‐Cx43/t‐Cx43 (H) and expression of np‐Cx43 (I) in mice ventricular tissue ( n = 3, * p < 0.05, ** p < 0.01, *** p < 0.001). (J) The distribution of Cx43 and co‐staining of wheat germ agglutinin (WGA) labeling were recorded by immunofluorescence microscopy, Cx43 located at intercalated disks (white arrows) and Cx43 located at lateral edges of the myocytes (yellow arrows) ( n = 4, Scale bar = 10 μm).

Article Snippet: The sections were sequentially performed following incubation with anti‐Cx43 (1:500, Invitrogen, #71–0700) overnight at 4°C and Fluorescein 5‐isothiocyanate (FITC)‐conjugated Affinipure donkey anti‐rabbit IgG(H + L) (Proteintech, #SA00003‐8) for 2 h. Then incubation with wheat germ agglutinin (WGA) dye solution (10 μg/mL, Invitrogen, #W11262) at 37°C in dark for 1 h. The stained sections were viewed under an inverted fluorescence microscope (NIKON ECLIPSE TI‐SR; NIKON, Japan) and imaged with the NIKON DS‐U3 imaging system (NIKON, Japan).

Techniques: De-Phosphorylation Assay, Injection, Western Blot, Isolation, Expressing, Staining, Labeling, Immunofluorescence, Microscopy

Journal: Cell Communication and Signaling : CCS

Article Title: TMEM100 acts as a TAK1 receptor that prevents pathological cardiac hypertrophy progression

doi: 10.1186/s12964-024-01816-2

Figure Lengend Snippet: TMEM100 overexpression attenuated cardiomyocytes enlargement and cardiac fibrosis in vivo . A Representative images of hematoxylin & eosin (HE) staining and wheat germ agglutinin (WGA) staining (left, n = 6) and LV cardiomyocytes’ cross-sectional-area (right, n = 120 cells) in every group. B The mRNA level of cardiac hypertrophic markers was analyzed by RT-PCR ( n = 4). C Representative images of picrosirius red (PSR) staining ( n = 6) and LV collagen volume (right) in every group. D The mRNA level of cardiac fibrosis markers was analyzed by RT-PCR ( n = 4). * p < 0.05 vs. AAV9-GFP sham, ** p < 0.01 vs. AAV9-GFP sham, # p < 0.05 vs. AAV9-GFP TAC, ## p < 0.01 vs. AAV9-GFP TAC

Article Snippet: For WGA staining, a diluted WGA dye solution was added, and the heart sectionslices were incubated at 37 °C in a light-free incubator for 1 h. Then, self-fluorescence quenching agent B was added (G1221, Servicebio), and the mixture was incubated for 5 min and washed with PBS.

Techniques: Over Expression, In Vivo, Staining, Reverse Transcription Polymerase Chain Reaction