MiR-146a is required for emergence of macrophages during early zebrafish embryogenesis. (A) Conservation of pre–miR-146a sequences in the genomes of human, mouse, and zebrafish. Highlighted are the seed regions and the mature miR sequence. The target regions of the 2 independent ASOs to induce miR KD are indicated. Scale bar indicates 100 mm. Comparison of L-plastin⁺ macrophages (indicated by arrowheads) in WT, miR-146a KD (drosha cleavage), miR-146a KD (stem loop), and miR-430 KD in 24-28 hpf zebrafish embryos. Each control/KD embryo is shown with a lateral magnified view of the head. (C) Quantification of the L-plastin⁺ macrophage numbers in z-stack image files of whole-mount in situ hybridizations from controls or KD zebrafish as shown in Figure 5B (n = 5). **P < .001 by Student t test. (D-E) Generation of erythrocytes and macrophages in WT and ASO-treated zebrafish determined by FACS analysis of scatter plots of whole fish single-cell suspensions. Representative FACS plots (D) and statistical analysis (E) are displayed (n = 8). **P < .001 by Student t test. Error bars indicate SD.