Introduction: Mesenchymal stem cells (MSCs) are being developed as proangiogenic agents. However, clinical experience with MSCs is equivocal, and it is unknown if (and why) MSCs from different tissues promote angiogenesis to differing degrees. In an effort to resolve this issue, we examined the angiogenic potential of bone marrow-derived vs thymus-derived MSCs.
Methods: Human neonatal thymus MSCs were compared to subject-matched and unrelated adult bone MSCs. Expression of pericyte markers/genes were evaluated, and angiogenic potency determined in vitro by assessing endothelial cell (EC) tubule formation, spheroid sprouting, angiogenic growth factor expression as well as in vivo using a subcutaneous implant model. Transcriptome differences in subject-matched thymus and bone MSCs were determined by microarray and Western blotting. Downstream gain/loss of function experiments were then performed.
Results: MSCs from both tissue types displayed similar expression of pericyte surface markers, but thymus MSCs had a higher expression of Tbx18, a pericyte transcription factor. Thymus MSCs were also more effective than bone-derived MSCs in promoting angiogenesis in vitro and in promoting the formation of functional new blood vessels in vivo. Microarray analysis identified SLIT3, an axon patterning and stimulatory EC factor, as differentially expressed between thymus and bone derived MSCs, which was confirmed at the protein level. In MSCs, SLIT3 localized to the nucleus, mitochondria, stress actin fibers and cell membrane, with polarized cells displaying higher expression. We found, unexpectedly, that SLIT3 stimulated MSC chemotaxis and motility in 2D and 3D assays, while knockdown of the SLIT receptor, ROBO1, but not ROBO4, blunted the stimulatory effects of SLIT3.
Conclusions: Human neonatal thymus MSCs display superior angiogenic potential relative to bone marrow-derived MSCs. This ability is in part mediated by SLIT3, which stimulates both ECs and MSCs. SLIT3 effects on MSCs are mediated by ROBO1 with SLIT3 regulating angiogenesis in both an autocrine and paracrine fashion. We propose that MSCs exhibiting higher expression levels of endogenous SLIT3 have a greater therapeutic potency.