The analysis of vascular structures has been reported frequently during the last decades and vascularization quantified in tissues (e.g. retina, tumors) and in vitro assays (e.g. CAM-assay), including angiogenic processes in cultured endothelial cells.
In co-cultures of outgrowth endothelial cells (OEC) and osteogenic cells, angiogenesis was followed for three weeks by imaging the cells after double fluorescent staining for CD31 (PECAM-1) and CD11b, a cellular marker of myeloid cells which have a key role in the regulation of angiogenesis during inflammation and wound healing.
The images were analyzed using ImageJ in a two step procedure:
(1) From images stained for PECAM-1, capillary-like structures were extracted and analyzed, resulting in the vessel area, length and position of branching- and endpoints.
(2) Colocalization of angiogenic structures with myeloid cells was assessed by measuring the relative density of CD11b fluorescence along the area of the angiogenic structures and around the branching- and endpoints.
The relative density of myeloid cells (CD11b positive) is increased along the angiogenic structures, indicating that endogenous myeloid cells are involved in vascularisation processes of outgrowth endothelial cells.
Colocalization, angiogenesis, OEC, myeloid cells
Presenting author: Harald Schmidt a
Organisation: MetaPhysiol, Essenheim, Germany
co-authors: Gerhard Kramer b, Yang Shi c, Charles James Kirkpatrick b, Sabine Fuchs b,c