FIGURE 4.

Autophagy flux analysis in quiescent satellite cells by immunofluorescence. (A) Representative images of GFP⁺-autophagosomes (green) and nuclei (blue) in freshly isolated QSCs from GFP-LC3 reporter mice (left panel) with the corresponding quantification of GFP-LC3⁺ puncta per cell (right panel). Treatment with vehicle (DMSO) or BafA1 was performed for 4 h prior QSC isolation by FACS (n = 4). Scale bar, 2 μm. (B) Representative images of LC3-stained autophagosomes and nuclei (blue) in freshly isolated QSCs from WT mice (left panel) with its corresponding quantification of LC3⁺ puncta per cell (right panel). Treatment with vehicle (DMSO) or BafA1 was performed for 4 h prior QSC isolation by FACS (n = 3). Scale bar, 2 μm. (C) Comparison of autophagy flux in GFP-LC3 or WT QSCs. Autophagy flux was determined as the ratio of the number of autophagosome puncta in BafA1 treated QSCs divided by the number of autophagosome puncta in vehicle (DMSO) treated QSCs (for GFP-LC3: n = 4; for WT: n = 3). Means ± SD; two-tailed unpaired t-test. ****p < 0.0001.