Isolation and characterization of mouse bone marrow-derived Lin⁻/VEGF-R2⁺ progenitor cells

Circulating endothelial progenitor cells (EPCs) in the peripheral blood (PB) have physiological roles in the maintenance of the existing vascular beds and rescue of vascular injury. In this study, we have evaluated the properties of Lin⁻/VEGF-R2⁺ progenitor cells isolated from the mouse bone marrow (BM) and further studied their distribution and integration in an animal model of laser-induced retinal vascular injury. Lin⁻/VEGF-R2⁺ cells were enriched from C57BL/6 mice BM using magnetic cell sorting with hematopoietic lineage (Lin) depletion followed by VEGF-R2 positive selection. Lin⁻/VEGF-R2⁺ BM cells were characterized using flow cytometry and immunocytochemistry and further tested for colony formation during culture and tube formation on Matrigel®. Lin⁻/VEGF-R2⁺ BM cells possessed typical EPC properties such as forming cobble-stone shaped colonies after 3 to 4 weeks of culture, CD34⁺ expression, take up of Dil-acLDL and binding to Ulex europaeus agglutinin. However, they did not form tube-like structures on Matrigel®. The progenitor cells retained their phenotype over extended period of culture. After intravitreal transplantation in eyes subjected to the laser-induced retinal vascular injury, some Lin⁻/VEGF-R2⁺ cells were able to integrate into the damaged retinal vasculature but the level of cell integration seemed less efficient when compared with previous reports in which EPCs from the human PB were employed. Our results indicate that Lin⁻/VEGF-R2⁺ cells isolated from the mouse BM share some similarities to EPCs from the human PB but most of them are at a very early stage of maturation and remain quiescent during culture and after intravitreal transplantation.