Journal: Advanced Science
Article Title: Pharmaceutical Manipulation of Mitochondrial F0F1‐ATP Synthase Enables Imaging and Protection of Myocardial Ischemia/Reperfusion Injury Through Stress‐induced Selective Enrichment
doi: 10.1002/advs.202307880
Figure Lengend Snippet: The uptake mechanism of IR‐780 by cardiomyocytes following I/R. A) Representative NIR imaging and confocal images of the heart tissues 2 h following reperfusion in rats treated with IR‐780 or HSA‐IR‐780. Scale bar: 75 µm. B) The absorbance of the IR‐780 dispersed in PBS and HSA. C,D) The albumin‐bond site of IR‐780 was determined with specific inhibitors (warfarin, ibuprofen, digoxin, and quinidine, used as competitive inhibitors of albumin‐binding sites I, II, III, and α1‐glycolipoprotein, respectively) (n = 3 per group). E) The absorbance changes of IR‐780 dispersed in HSA within different pH conditions. F,G) Representative images and quantitative data of the NIR fluorescent signals in H9C2 cells exposed to IR‐780 pre‐treatment with ice condition, sulfobromophthalein (BSP), amiloride (an actin inhibitor), chlorpromazine (a clathrin inhibitor) and MβCD (a caveolae inhibitor) (n = 3 per group). Scale bar: 75 µm. H) Schematic of the in vivo protocols to verify the roles of OATPs on the IR‐780 uptake. I) Representative NIR photographs of the whole heart (left panel), transverse sections (middle panel) and micro‐photographs of the heart tissue received IR‐780 injection with or without pre‐BSP local administration in rat I/R models. Scale bar: 75 µm. J,K) Representative images and quantitative flow cytometric data of the NIR fluorescent signals in H9C2 cells exposed to IR‐780 within different pH conditions (n = 3 per group). Scale bar: 50 µm. L) Quantitative flow cytometric data of the mitochondrial membrane potential of the H9C2 cells exposed to hypoxia and I/R (n = 3 per group). M,N) Representative images and quantitative flow cytometric data of the NIR fluorescent signals in H9C2 cells exposed to IR‐780 pre‐treatment with FCCP, Rotenone, Oligomycin A (n = 3 per group). Scale bar: 50 µm. All the p values are present in the graphs.
Article Snippet: The cells were treated with DMEM with amiloride (75 µg mL −1 , HY‐B0285, MCE), BSP (250 µ m , S0252, Sigma), chlorpromazine (15 µg mL −1 , 31 679, Sigma), methyl‐β‐cyclodextrin (MβCD, 7.5 m m , 779 776, Sigma), FCCP (10 µ m , C2920, Sigma), rotenone (10 µ m , 45 656, Sigma), oligomycin A (10 µ m , 495 455, Sigma), HSA (1 µ m , A1887, Sigma), HSA (1 µ m , A1887, Sigma) + ibuprofen (100 µ m , I0415, TCI), or ice or hypoxic conditions (5% O 2 ) or different pH conditions (5.2, 6.2, 7.4, and 8.9) for 30 min; subsequently, IR‐780 (0.1 µ m , 425 311, Sigma) was added into the medium for another 20 min. Next, the cells were washed thrice with PBS and were for further NIR imaging using the Leica NIR fluorescent microscope and flow cytometric analysis using a BD Accuri C6 Flow Cytometer.
Techniques: Imaging, Binding Assay, In Vivo, Injection, Membrane
