Side population (SP) cells are highly able to exclude the Hoechst 33342 dye through membrane transporters, a feature associated with cell immaturity and therefore proposed as a marker of stem cells. Herein we demonstrate that the adipose tissue derived stromal vascular fraction (SVF) contains a novel population of non-haematopoietic "side population" (SPCD45(-)) cells. Simultaneous qRT-PCR of 64 genes revealed that the freshly isolated SPCD45(-) was highly enriched for cells expressing genes related to stem cells, the Notch pathway, and early vascular precursors. Notably, the expression of smooth muscle actin, C-met and Cd34 together with Angpt2, Flk1, VE-cadherin, and Cd31 suggested a phenotypic resemblance to pericytes and aorta-derived mesoangioblasts. Recent evidence suggests that cells residing within the vascular niche may participate in regeneration of skeletal muscle and although skeletal muscle repair mainly relies on the satellite cell, several reports have shown that vessel-associated cells may adopt a myogenic phenotype when exposed to a muscle environment. In accordance with these findings, we also observed in vitro myogenic specification of SPCD45(-) cells when cocultured with myoblasts. Furthermore, immediate intramuscular engraftment of non-cultured SPCD45(-) cells gave rise to myofibres and cells lining blood vessels, whereas the SVF only provided donor derived mononuclear cells. We therefore conclude that the SPCD45(-) fraction of adipose-derived SVF is enriched for cells expressing vascular associated markers and that the myogenic differentiation potential of these cells does not depend on prior in vitro expansion.