Deteriorated integrity of retinal vessels in the absence of MCs. (a) H&E staining of retinal sections of control (top) and APB5-treated (bottom) mice. APB5 treatment induced progressive exacerbation of retinal edema in the nerve fiber layer (NFL) at around P8 to P9, resulting in retinal collapse and massive hemorrhage at P10. (b) Retinal cup of APB5-treated mouse at P9 depicting widespread hemorrhage and funnel-shaped retinal collapse. (c and d) Visualization of vascular leakage in P8 retina by TRITC-dextran perfusion. Multifocal leakage of systemically perfused dextran demonstrated the increased vascular permeability in the absence of MCs (d). No leakage was found in the control retina (c). (e and f) Scanning electron microscopy demonstrating numerous cleft formations (arrowheads in e) in denuded endothelial sheets and extravasation of blood cells (f). (g–j) Ultrastructural analysis depicting vessel walls with MC coverage in the control retina (g and h) and MC-free endothelial walls in the APB5-treated retina (i and j). In the absence of MCs, extraluminal spaces were enlarged with fluid accumulation. Note the swelling of periendothelial basement membranes with cell debris (arrowhead in j). EC, endothelial cell; MC, mural cell. (k–m) Double immunofluorescent staining of PECAM (green) and fibronectin (k), type IV collagen (l), and laminin (m) (red) in retinal vessels of APB5-treated mice. Major extracellular matrix components of basement membranes were deposited around EC surfaces in the absence of MCs. Bars represent 5 μm.