Inhibition of autophagy enhanced the blood perfusion into the brain b (A) Example of images (each representative of 6 zebrafishes) depicting autophagosome (green) of endothelial cells and autolysosome (red) of endothelial cells in 0, 10 Gy group, 10 Gy + chloroque group, 10 Gy + wortamanin group and 10 Gy + Ly294002 group at 4-day post radiation. Scale bars, 100 µm. (B) Live example of images (each representative of 6 zebrafishes) depicting endothelial cells (green) of cerebral capillaries (top) and the tracer Dextran blue (MW 10 kDa) pericardial injected (bottom) in 0, 10 Gy group, 10 Gy + chloroque group, 10 Gy + wortamanin group and 10 Gy + Ly294002 group at 4-day post radiation. Scale bars, 100 µm. (C) Morphometric analyses of autophagical puncta in 0, 10 Gy group, 10 Gy + chloroque group, 10 Gy + wortamanin group and 10 Gy + Ly294002 group at 4-day post radiation (n = 6 zebrafishes per group). (D) Morphometric analyses of blood-perfused vessel branches in 0, 10 Gy group, 10 Gy + chloroque group, 10 Gy + wortamanin group and 10 Gy + Ly294002 group at 2-day and 4-day post radiation (n = 6 zebrafishes per group). (E) The survival analysis of zebrafishes in 0, 10 Gy group, 10 Gy + Cloroquine (n = 10, p = 0.0126), 10 Gy + ly294002 (n = 10, p < 0.01) and 10 Gy + Wortamanin (n = 10, p < 0.01). Statistical analysis in C and D was performed using t-test: **P < 0.05, ***P < 0.01. Data represent the mean ± s.e.m.
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