Abstract
BackgroundComparative analysis of the cellular biology of the microvasculature in different tissues requires the availability of viable primary endothelial cells (ECs). This study describes a novel method to isolate primary ECs from healthy organs, repair blastemas and tumors as examples of non-proliferating and proliferating benign and malignant tissues and their functional characterization.Methodology/Principal FindingsSingle cell suspensions from hearts, lungs, repair blastemas and tumors were incubated consecutively with an anti-CD31 antibody and magnetic micro-beads, coupled to a derivative of biotin and streptavidin, respectively. Following magnetic bead separation, CD31-positive ECs were released by biotin-streptavidin competition. In the absence of micro-beads, ECs became adherent to plastic surfaces. ECs from proliferating repair blastemas and tumors were larger and exhibited higher expression densities of CD31, CD105 and CD102 compared to those from non-proliferating normal tissues such as heart and lung. The expression density of CD34 was particularly high in tumor-derived ECs, and that of CD54 and CD144 in ECs of repair blastemas. Functionally, ECs of non-proliferating and proliferating tissues differed in their capacity to form tubes in matrigel and to align under flow conditions.Conclusions/SignificanceThis method provides a powerful tool to generate high yields of viable, primary ECs of different origins. The results suggest that an altered expression of adhesion molecules on ECs in proliferating tissues contribute to loss of EC function that might cause a chaotic tumor vasculature.
Highlights
The diffusion limit of oxygen from the capillary to non-vascular tissue in the body ranges from 100 to 200 mm
It is assumed that the recovery rate of primary endothelial cells (ECs) are higher because the mean cell surface density of CD31 on heart, repair blastema and tumor ECs (8568, 479614, 211667) are higher compared to that of H5V cells (3164)
ECs could only be isolated from growing tissues of very young mice that have the capacity to proliferate and have the chance to get rid of the attached beads with increasing cell divisions
Summary
The diffusion limit of oxygen from the capillary to non-vascular tissue in the body ranges from 100 to 200 mm. A dense network of blood vessels is necessary to provide an adequate supply of oxygen and nutrients [1,2]. Microvascular ECs fulfill important functions in wound healing and blood flow regulation e.g. by preventing thrombosis. It is known that the microvascular architecture of tumors differs from that of normal tissues. Comparative analysis of the cellular biology of the microvasculature in different tissues requires the availability of viable primary endothelial cells (ECs). This study describes a novel method to isolate primary ECs from healthy organs, repair blastemas and tumors as examples of non-proliferating and proliferating benign and malignant tissues and their functional characterization
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