Single-cell RNA sequencing of adipocyte-specific hM4Di-DREADD expressing mice 24 h after cardiac ischemia.
Single-cell RNA sequencing of adipocyte-specific hM4Di-DREADD expressing mice 24 h after cardiac ischemia.
After mouse sacrifice hearts were excised and briefly washed in ice-cold PBS. Aorta was cannulated and coronaries retrograde flushed with 5 ml PBS/Heparin followed by 5 ml Collagenase/DNAse solution. Heart tissue was minced with scissors, transferred into 5ml Collagenase/DNAse solution and digested for 20 minutes at 37°C on a rotator. Cells were filtered over a 100µm and 40µm filter and pelleted at 800g, 4°C for 10 minutes. Cells were stained with a-TER antibody to identify erythrocytes and propidium iodide to identify dead cells. Cells were sorted and single, TER negative, PI negative cells were further used for sequencing. All used samples showed a cell vitality >80%. Chromium Single Cell 3' NextGEM Reagent Kit v3.1 according to the manufacturer's instructions Chromium Single Cell 3' NextGEM Reagent Kit v3.1 according to the manufacturer's instructions