Hematopoietic cytokine production by human brain endothelial cells (Hubec) Is associated with the ex vivo expansion of CD34+CD38− cells

We have recently demonstrated that primary human brain endothelial cells (HUBEC) supplemented with GMCSF+IL-3+ IL-6+SCF+Flt-3 ligand support a 400-fold increase in CD34+CD38− cells in 7 day cultures. In order to identify soluble factors which might contribute to this hematopoietic effect, we performed ELISAs on supernatant collected from HUBEC monolayers and compared this with supernatant from human umbilical vein endothelial cells (HUVEC) which do not support the expansion of CD34+CD38− cells. In unconcentrated HUBEC supernatant, we detected high levels of VEGF (1951 pg/mL±561), IL-11 (1832 pg/mL±142), IL-6 (2625 pg/mL±39), and SCF (172 pg/mL±2), and surprisingly low or nonmeasurable levels of GMCSF (nm), GCSF (22 pg/mL±1), Epo (nm), IL-3 (nm), and Flt-3 ligand (18 pg/mL±0.1). In contrast, supernatant collected from HUVEC contained no measurable VEGF or IL-11, but had very high levels of GCSF (2278 pg/mL±76) and PDGF (2369 pg/mL±235). In addition, HUVEC supernatant contained IL-6 (767 pg/mL±21), GMCSF (67 pg/mL±3), and SCF (340 pg/ml±6), but Flt-3 ligand and Epo were not detectable. In summary, HUBEC secrete high levels of IL-11 and VEGF which have both been identified to induce the proliferation of hematopoietic stem cells (HSC) in vitro. In contrast, HUVEC produce neither IL-11 nor VEGF but do secrete high levels of GCSF which likely contributes to the differentiation of progenitor cells in vitro. We are currently studying the capacity of HUBEC supernatant to induce HSC proliferation in the absence of cell-to-cell contact and the contribution of known or novel cytokines toward this effect.