VEGF-A promotes intussusceptive angiogenesis in the developing chicken chorioallantoic membrane

Abstract

Objective: To assess the impact of vascular endothelial growth factor (VEGF) on intussusceptive angiogenesis.

Methods and results: Polyurethane casts of the microvasculature of chicken chorioallantoic membrane (CAM) were prepared on embryonic days (E) 8, 10, 12, and 14. At light microscopy level, minute holes (<2 microm in diameter) and hollows (>2 microm) were observed in the casts. Transmission electron microscopy disclosed the minute holes to mainly represent transluminal pillars characteristic for intussusceptive angiogenesis. The numerical density of the holes/pillars was highest at an early (E8) and a late (E12-E14) stage. Only mRNA of VEGF-A-122 and VEGF-A-166 isoforms was detected in the CAM. The transcription rate of VEGF-A mRNA peaked on E8/9 and E12, while VEGF-A protein expression increased on E8/9 and E11/12 to rapidly decrease thereafter as determined by immunoblotting. At all time points investigated, VEGF-A immunohistochemical reactivity was restricted to cells of the chorionic epithelium in direct contact to the capillary plexus. When the VEGF-R-inhibitor PTK787/ZK222584 (0.1 mg/mL) was applied on E9 CAM, the microvasculature topology on E12 was similar to that on E10.

Conclusions: The temporal course of intussusception corresponded to the expression of VEGF-A in CAM microvasculature. Inhibition of VEGF-signaling retarded intussusceptive-dependent capillary maturation. These data suggest that VEGF promotes intussusception.