EP4 and PKA mediate the in vivo angiogenesis. (A) Images of polyvinyl acetal sponges harvested from mice. The sponges were implanted in the flank of C57BL/6 mice and injected with PGE2 (8.8 ng/g body weight), AH23848 (25 or 125 ng/g body weight), or H89 (11 or 55 ng/g body weight) every 2 days for 2 weeks. (B) Hemoglobin content in the sponges as determined with the Drabkin reagent. Each point represents the mean ± SEM of values obtained from 4 to 5 sponges. *P < .05, **P < .01, compared with corresponding vehicle group. Low, AH23848 (25 ng/g body weight) and H89 (11 ng/g body weight) and high, AH23848 (125 ng/g body weight) and H89 (55 ng/g body weight), groups. (C) Expression of CD31⁺ cells by immunohistochemistry. Harvested sponges were sectioned and stained with the endothelial cell marker CD31 Ab. Sections were counterstained with hematoxylin to visualize nuclei. (D) Blood vessel density in harvested sponges as quantitated by counting red-staining–positive cells or cell clusters. Three 200× magnification fields containing the most prominent hotspots were counted for each section. Each point represents the mean ± SEM of values obtained from 4 to 5 sponges. *P < .05, **P < .01, compared with corresponding vehicle group. Low, AH23848 (25 ng/g body weight) and H89 (11 ng/g body weight) and high, AH23848 (125 ng/g body weight) and H89 (55 ng/g body weight), groups. (E) Schematic presentation of PGE2-induced angiogenic signal in HMVECs (see text for details).