In vitro model for the evaluation of angiogenic potential of MSCs derived from dental pulp

Mesenchymal stem cells (MSCs) are adult multipotent cells distributed in different anatomic sites where they are thought to play a key role in tissues homeostasis. Several data demonstrate that MSCs could be both efficiently propagated in vitro and induced to differentiate into cells of mesenchymal lineage in the appropriate culture conditions. However, several aspects remain to be elucidated, including the degree of potency belonging to MSCs from different tissues, and the best culture conditions suitable to reach the translational objective. In our study, we aimed to explore the phenotypic and functional characteristics of human MSCs isolated from dental pulp (DPSCs), and in particular, we investigated the ability of DPSCs to modulate in vitro angiogenesis. For this purpose, we performed co-culture tubule-formation assays using human endothelial cells (HUVECs) in combination with DPSCs. DPSCs analysed by FACS within two weeks since cell isolation expressed high levels of stem cell markers, including CD90, CD105 and CD44. When used in the co-culture model, DPSCs were able to interact with HUVECs and, significantly, they stabilize HUVEC tubules generating stratified cell structures, in which DPSCs wrap HUVEC tubules. The resulting angiogenic net remained stable for several days; otherwise, in absence of DPSCs, HUVECs underwent apoptotic cell death after 24h. Expression of N-cadherin on both cell types was responsible for heterotypic interaction. However, the pro-angiogenic effect was evident only when DPSCs have previously been cultured in complete endothelial growth medium (EGM-2). Accordingly, FACS and western blot analyses revealed that EGM-2 stimulated a significant phenotypic switch in DPSCs, with an increased expression of pericyte markers, including NG-2, respect to DPSCs cultured in standard conditions. The low expression of endothelial markers in EGM-2-conditioned DPSCs further supported their role as perivascular cells in the angiogenic model. In physiological condition, pericytes envelop the surface of the vascular tube contributing in stabilization of the vessel, and supporting wound repair. Thus, the further characterization of factors needed for pericyte differentiation, could render DPSCs a new useful therapeutic tool in angiogenic defective clinical situations.