Hemangioblastic characteristics of fetal bone marrow-derived Flk1⁺CD31^-CD34^- cells

Objective. To investigate whether Flk1⁺CD31^-CD34^- cells isolated from fetal bone marrow (BM) have characteristics of hemangioblasts, i.e., progenitors of endothelial and hematopoietic cells. Materials and Methods. Mononuclear cells from fetal BM were negatively sorted by CD45, GlyA, and CD34 micromagnetic beads, then cultured to form cell colonies. A single colony was harvested. Culture-expanded cells were seeded on ECM gel or semisolid media supplemented with endothelial and hematopoietic growth factors, respectively. Immunochemistry staining and RT-PCR were performed for cell characterization. Results. 99% of cells from the single colony maintained Flk1⁺ and CD31/CD34^- during passaging. On ECM gel, Flk1⁺CD31^-CD34^- cells could grow into vascular structure that was positive for CD31 and vWF. There were round CD34⁺ cells around the vascular structure. When angiogenesis inhibitor suramin was added before tube formation, formation of vascular structure was blocked. Additionally, Flk1⁺CD31^-CD34^- cells cultured on hematopoietic condition could differentiate into hematopoietic cells which expressed GATA-1, 2, and γ, β globin gene. After being replated in methylcellulose medium, they formed typical erythroid colonies. Conclusions. Flk1⁺CD31^-CD34^- cells derived from fetal BM could differentiate into endothelial and hematopoietic cells. The results suggested that these Flk1⁺CD31^-CD34^- cells after embryo stage bear characteristics of hemangioblast and may have potential application for the hematopoietic and vascular diseases.