Spatial relationship between uPA-expressing cells and PAI-expressing cells during capillary sprouting in vitro. Cultured rat aortic ring explants, displaying branching capillary-like sprouts, are shown 3 weeks after initial culture. (A) A phase-contrasted micrograph showing the tips of sprouts and the preformed monolayer through which sprouts extend. (B) Immunostaining with vWF antibodies. vWF-positive cells are organized in cords, indicating that all cells seen as surrounding the cords in (A) and (C through E) are nonendothelial cells. (C) In situ hybridization with a uPA-specific probe. Note that expressing cells are endothelial cells forming the cords. (D and E) In situ hybidization with a PAI-1–specific probe. Note that strongest hybridization signals are in cells closest to cords (highlighted by arrows) and that fibroblasts residing more distally show much weaker hybridization (arrowheads). Magnification: (A, C, and E) ×200 original magnification; (B) ×100 original magnification; (D) ×300 original magnification.