Fig. 2: Macrophages are required for β cell proliferation in βVEGF-A mice.
a To deplete macrophages (MΦs) during VEGF-A induction and normalization, βVEGF-A mice were treated with clodronate or control liposomes (150–200 μl i.v.) every other day, beginning 1 day before Dox treatment and continuing 1 week after Dox withdrawal (1wk WD). b Representative flow cytometry plots showing circulating monocytes (CD11b+ Ly6G–) of control and clodronate-treated βVEGF-A mice 24 h after single injection (No Dox). Approximately 10,000 white blood cells (WBCs) were analyzed (monocyte fraction reported as mean + s.e.m.) per each animal. c Islet architecture displayed by labeling for β cells (Insulin; blue), endothelial cells (Caveolin-1; green), and MΦs (Iba1; red) during VEGF-A induction (1wk Dox) and normalization (1wk WD). Scale bar, 50 μm. d Quantification (mean + s.e.m.) of islet MΦ area by immunohistochemistry (5.1 ± 0.7 × 105 μm2 total islet area analyzed per animal). e, f Quantification (mean + s.e.m.) of endothelial cell (EC) area (e) and β cell area (f) in βVEGF-A mice treated with control or clodronate liposomes during VEGF-A induction and normalization. g Rate of β cell proliferation (mean + s.e.m.; 1138 ± 87 β cells counted per animal) during VEGF-A normalization (1wk WD and 2wk WD) in control βVEGF-A mice was significantly reduced in clodronate-treated mice. In panels b–g, each closed circle represents one animal; asterisks indicate unpaired two-tailed t-tests of control vs. clodronate groups; *p < 0.05; **p < 0.01; ****p < 0.0001. Dashed lines in d–g depict average values in βVEGF-A mice at baseline (No Dox).
