Abstract
Cell-cell adhesion ensures tight contacts between neighbouring cells, which is necessary for cell segregation, as well as for the morphological and functional differentiation of different tissues. Evidently there are cell-cell recognition systems that make cells of the same type preferentially adherent to one another. Homotypic cell adhesion is particularly important in mediating a range of physiological processes such as cell survival, migration and remodeling of vessels. Thus in the present study we selected two populations of endothelial progenitor cells which are from the same donor to investigate the possible effect of a small molecule compound Icariin on homotypic cell adhesion. Many angiogenic factors can destabilize the organization of intercellular junctions, causing endothelial barrier opening. In the present study, we observed that Icariin treatment reduced the level of VE-cadherin expression in EPCs indicating a decrease in cell-cell adhesion-a proof of the pro-angiogenic effect of Icariin. In summary, the observed loss of homotypic adhesion of EPCs may contribute to the enhanced angiogenic effect exerted by Icariin.
Keywords: Endothelial progenitor cell, Intercellular adhesion, Cell differentiation
Materials and Reagents
Equipment
Procedure
Representative data
Figure 1. Images of cells collected from the interface between the Ficoll-Paque PLUS and plasma. A. Bone marrow sample was diluted with equal volume of DPBS, then the diluted sample was carefully layered onto the top of Ficoll-Paque PLUS. B. After centrifuging at 400 x g for 30 min, the mononuclear cells were harvested from the interface between the Ficoll-Paque PLUS and plasma. Figure 2. Light microscope image of mononuclear cells. The mononuclear cells were carefully harvested from the interface between the Ficoll-Paque PLUS and plasma using a sterile Pasteur pipette, and transfered to sterile centrifuge tubes. After washing, the isolated mononuclear cells were cultivated in flasks coated with human fibronectin (25 μg/ml) and induced by ECGM2 medium at 37 °C with 5% CO2 in humidified air at a density of 3-5 x 106/cm2. Figure 3. Fluorescence microscope images of cells after intercellular adhesion assay. A. Cells were plated overnight to a confluent monolayer in ECGM2, and labeled the cells with Hoechst 33258. B. Representative fluorescence micrograph demonstrating cells after intercellular adhesion assay.
Notes
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Acknowledgments
The concept of this protocol was adapted from our previous study, in which the intercellular adhesion assay was used to measure cell-cell adhesion ability of endothelial progenitor cells (Tang et al., 2015).This study was supported by the German Academic Exchange Service and Federal Ministry of Education and Research (D/09/04774).
References
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