1 Introduction Formation of the embryonic vascular system happens before nutrition by diffusion runs into its limitation, and represents the first organ system in the early embryo (Carmeliet, 2003; Noden, 1989). Vasculogenesis, the process of de novo formation of vascular structures, is characterized by the differentiation of mesodermal derived angioblast precursor cells into endothelial cells forming the primary capillary plexus (Risau, 1997; Risau and Flamme, 1995). Subsequently, during angiogenesis this early vascular system undergoes a period of pruning, remodeling and maturation. Additionally, it takes two cell types to generate functional blood vessels, endothelial cells and perivascular cells (Gerhardt and Betsholtz, 2003). Perivascular cells (PVC) can be classified as pericytes in capillaries or vascular smooth muscle cells (vSMC) in larger vessels, depending on their morphology and location. Both may represent subtypes of a continuum of related cell types. PVCs are essential for the development of functional vessel walls and contribute to the structural integrity and contractility of vessels. However, developmental origins of PVCs remain unclear. Different speculations were proposed, such as derivation from mesenchymal and epicardial cells or neural crest (Gerhardt and Betsholtz, 2003), but also trans- differentiation of endothelial cells is discussed (DeRuiter et al., 1997). Recent data support the idea of a common progenitor of perivascular and endothelial cells – the FLK1-positive angioblast (Ema et al., 2003; Yamashita et al., 2000). Pericytes were originally defined by their morphology and close contact to endothelial cells located within a shared basement membrane (Sims, 1986; Tilton, 1991). They represent a heterogeneous population of cells involved in normal and pathological angiogenesis, such as diabetic microangiopathy (Cogan et al., 1961; Hirschi and D’Amore, 1996), atherosclerosis (Bostrom et al., 1993) or cancer (Wesseling et al., 1995). Recruitment and coverage of vessels with pericytes is essential for the development of vasculature, as has been shown by genetic ablation of PDGFB signaling. In mice with disrupted signaling, the expansion and spreading of pericytes is impaired and leads to perinatal lethality due to leakage and hemorrhage (Leveen et al., 1994; Lindahl et al., 1997; Soriano, 1994). Presently, no ‘pan-specific’ marker is available that defines the pericyte phenotype unambiguously (Gerhardt and Betsholtz, 2003). Common markers such as smooth muscle actin (Nehls and Drenckhahn, 1991), NG2- The annexin A5 gene (Anxa5) was recently found to be expressed in the developing and adult vascular system as well as the skeletal system. In this paper, the expression of an Anxa5-lacZ fusion gene was used to define the onset of expression in the vasculature and to characterize these Anxa5-lacZ-expressing vasculature-associated cells. After blastocyst implantation, Anxa5-lacZ-positive cells were first detected in extra-embryonic tissues and in angioblast progenitors forming the primary vascular plexus. Later, expression is highly restricted to perivascular cells in most blood vessels resembling pericytes or vascular smooth muscle cells. Viable Anxa5-lacZ + perivascular cells were isolated from embryos as well as adult brain meninges by specific staining with fluorescent X-gal substrates and cell-sorting. These purified lacZ + cells specifically express known markers of pericytes, but also markers characteristic for stem cell populations. In vitro and in vivo differentiation experiments show that this cell pool expresses early markers of chondrogenesis, is capable of forming a calcified matrix and differentiates into adipocytes. Hence, Anxa5 expression in perivascular cells from mouse defines a novel population of cells with a distinct developmental potential. Key words: Annexin A5, Perivascular cells, Pericytes, Adult stem cells Summary Perivascular cells expressing annexin A5 define a novel mesenchymal stem cell-like population with the capacity to differentiate into multiple mesenchymal lineages Bent Brachvogel 1, *, Helga Moch 2 , Friederike Pausch 2 , Ursula Schlötzer-Schrehardt 3 , Clementine Hofmann 4 , Rupert Hallmann 5 , Klaus von der Mark 2 , Thomas Winkler 6 and Ernst Pöschl 2 1 Department of Cell and Matrix Biology, MCRI, 3052 Parkville Victoria, Australia 2 Department of Experimental Medicine I, University Erlangen-Nürnberg, 91054 Erlangen, Germany 3 Department of Ophthalmology, University Erlangen-Nürnberg, 91054 Erlangen, Germany 4 Max-Planck-Institute of Psychiatry, 80804 München, Germany 5 Department of Experimental Pathology, Lund University, 22363 Lund, Sweden 6 Department of Genetics, University Erlangen-Nürnberg, 91054 Erlangen, Germany *Author for correspondence (e-mail: bent.brachvogel@mcri.edu.au) Accepted 22 March 2005 Development 132, 000-000 Published by The Company of Biologists 2005 doi:10.1242/dev.01846 Research article Development and disease Development Development ePress online publication date 27 April 2005