Disorganized vascular remodeling in the absence of MCs. (a and b) Double immunostaining with PECAM (brown) and SMA (blue) of P8 retina. In the control (a), advancing vessels had reached the edges of the retina, while in the APB5-treated mice, the peripheral retina remained unvascularized (b). Note the strong SMA expression around arteries in the control retina. (c and d) PECAM staining of P8 retina of control (c) and APB5-treated mice (d). Note the remaining capillary pruning and intense PECAM staining in arteries even in the absence of MCs. (e and f) Morphometric analysis of retinal vasculature at P3, P5, and P8. Black bars, control mice; gray bars, APB5-treated mice. Data (mean ± SD) are expressed as percentages relative to the control retina on P3. *P < 0.05 by Student t test. (g and h) High-magnification view of PECAM staining in P5 retina. The shape of each EC is illustrated by white lines. Increases in the number and size of ECs contributed to vessel enlargement in the absence of MCs. Bars represent 20 μm. (i and j) Detection of dying ECs by TUNEL assay (i) and in vivo propidium iodide labeling (j) in P5 retina. Endothelial apoptosis (arrowheads) was not accelerated even in the absence of MCs. (k–n) Double labeling of BrdU-incorporating cells (green) and PECAM⁺ ECs (red) in P5 retina. ECs were mitotically active, particularly in the regions of advancing retinal vessels in both APB5-treated mice (m and n) and controls (k and l). A, artery; V, vein.