Human subcutaneous fat-derived stem cells were recently shown to have the potential to differentiate in vitro into a variety of cell types. Herein it is shown that upon prolonged cell culture fat-derived stem cells spontaneously changed their phenotype from fibroblast-like- to cobble-stone appearance. Undifferentiated fat-derived stem cells expressed the stem cell markers CD13, Stro-1, CD105, Oct-4 and Flk-1. Upon differentiation CD13, Stro-1 and CD105 were downregulated whereas an upregulation of von Willebrand factor, VE-cadherin, Tie-2, Flk-1 and Flt-1 was observed, thus indicating endothelial cell differentiation. When grown on matrigel differentiated stem cells formed cord-like structures comparably to HUVEC cells. Upon cultivation on tumor tissues fat-derived stem cells invaded into the depth of the tissue and formed capillary-like cell structures which were positive for von Willebrand factor. In summary, our data indicate that fat-derived stem cells spontaneously acquire an endothelial cell phenotype, and may be used for cell replacement studies.