Introduction: There is growing evidence that the regenerative potential of stem cells relies primarily
on their secretion of soluble factors. In the present study we investigated whether
paracrine factors derived from cultured endothelial progenitor cells (EPC) may support
brain endothelial cell angiogenesis. Moreover we addressed the signalling pathways
that may be involved.
Methods: Cultures from rat brain endothelial cells were incubated with EPC-derived conditioned
medium (EPC-CM). Angiogenic response of the microvascular endothelial cells was assessed
by measuring tubulogenesis and cell number after incubation with EPC-CM. Cell number
in culture was measured by the MTT assay after 24 hours of incubation. The morphogenic
response of endothelial cells to EPC-CM was investigated by means of tube formation
assay on growth factor reduced Matrigel™ for 8 hours. The specific PI3K/AKT inhibitor
LY294002 and the MAPK/ERK inhibitor PD98059 were used to analyze the involvement of
these two signalling pathways in the transduction of the effects of EPC-CM.
Results: By means of an antibody array we found that EPC-CM contains a variety of growth factors
and cytokines. Incubation of brain microvascular endothelial cells with EPC-CM resulted
in a two- and ninefold increase of tubule and network complexity. The cell number
was likewise augmented after incubation with EPC-CM. Inhibition of the PI3K/AKT signaling
pathway resulted in a significant reduction of the length, the complexity of the tubule
network formation as well as the cell number. Conversely the inhibition of the MAPK/ERK
pathway did not affect the angiogenic response of the brain microvascular cells to
EPC-CM.
Conclusion: The present study shows that EPC-derived paracrine factors substantially promote
the angiogenic response of brain microvascular endothelial cells. The big variety
of factors contained in the EPC-CM likely activates multiple signaling cascades; however,
our findings identified the PI3K/AKT pathway to play a central role.
