An in vitro model for the study of human bone marrow angiogenesis: Role ofhematopoietic cytokines

This study describes a human bone marrow endothelial cell culture in which endothelial cells are organized into capillary tubes. These endothelial cel ls were positive for von Willebrand Factor, expressed CD34, CD31, and L-fuc ose residues, took up acetylated low-density lipoproteins, contained Weibel -Palade bodies, and were ensheathed in a basal lamina (which included lamin in beta 1, EDa+ and EDb+ fibronectin, and collagen type iv). Pericytes expr essing alpha-smooth muscle (alpha-SM) actin were spatially associated with the capillary tubes and there was a highly significant correlation between the number of capillary tubes and pericytes. In this model, basal angiogene sis was found to be vascular endothelial growth factor (VEGF)-dependent, be cause neutralization of endogenous VEGF induced a dramatic regression in th e number of tubes. However, the presence of alpha-SM actin-expressing peric ytes in the linings of endothelial tubes partially prevented the VEGF-neutr alized tube regression. We also observed that nitric oxide production contr ibuted to basal angiogenesis and that upregulation of nitric oxide increase d the number of tubes. Tube numbers also decreased when antibodies neutrali zing the integrin alpha v beta 5 were applied to the cultures. Moreover, ad dition of any of the hematopoietic cytokines, erythropoietin, stem cell fac tor, granulocytic colony stimulating factor, or granulomonocytic colony sti mulating factor induced a highly significant increase in tube formation. Wh en erythropoietin and granulocytic colony stimulating factor were added, th is increase was larger than the maximum increase observed with VEGF. Thus, we have described an in vitro model for human bone marrow angiogenesis in w hich pericytes and basal lamina matrix were associated with endothelial cel ls and formed fully organized capillary tubes. In this model, cytokines kno wn to regulate hematopoiesis also seemed to be mediators of angiogenesis. T his culture system may therefore prove to be a valuable tool for the study of hematopoietic cytokines on angiogenesis.