Endothelial cell apoptosis in the context of quantification of angiogenesis in solid human adenocarcinomas: a novel double immunolabelling technique to identify endothelial cell apoptosis.

Standardised methods of microvessel density quantification have been published. However, a reliable and reproducible method to visualise endothelial cell apoptosis is lacking, which is a shortcoming in assessing vascular remodelling during angiogenesis. The aim of this study was to validate a newly developed technique to demonstrate endothelial cell apoptosis by double immunolabelling with anti-CD34 and anti-activated caspase-3 in human adenocarcinomas. Double immunolabelling was used to identify apoptotic endothelial cells in six tumours of eight different human adenocarcinomas. Microvessel density and rate of apoptotic endothelial cells were quantified. The technique revealed endothelial cell apoptosis simultaneously with the identification of microvessel density on one slide. These characteristics were reproducible in adenocarcinomas of various sites. In conclusion, apoptotic endothelial cells and microvessel density can now be evaluated simultaneously within one and the same area, allowing a more reliable histological quantification of angiogenesis.