Joris van Dongen

166 Chapter 7 Enzymatic isolation and culture of those precursor cells orADSC results in a large series of cells that can be used in regenerative medicine. After several days of culture the in vivo phenotype of precursor cells changes into an in vitro specific phenotype. Most of the cells will lose their CD34 expression and almost all of the cells gain expression of CD105 61,62 . The CD105 marker is also known as endoglin and is a TGF- β type III receptor, which is expressed on virtually all cells of mesenchymal origin, but also on e.g. endothelial cells. Ten to twenty percent of the subpopulations remain CD34 pos , but their proliferation rate and adipogenic differentiation ability is significantly lower as compared to the CD34 min subpopulation 61,63 . This suggests that 80%-90% of the so-called ADSC, characterized by their phenotype in vitro (CD34 min /CD105 pos ), are not present in vivo: in other words: the majority of ADSC acquire their phenotype through culturing. Culturing of ADSC also causes dramatic shifts in secretome, as will be discussed within a few sentences below. The different components and cell types of all fractions of adipose tissue are summarized in Figure 2. Some studies have described that regenerative potencies of ADSC is caused by secretion of trophic factors or differentiation into other cells 65 . In vivo , little is known about the secretion of trophic factors by ADSC. In vitro , secretion of trophic factors by ADSC in medium (called ADSC conditioned medium) is affected by many aspects: differences in culture conditions, donors, methods and medium and cell counts results in different expression of growth factors. For instance, hypoxia culture upregulates VEGF, platelet derived growth factor, placental growth factor and insulin-like growth factor II 66 . A3D culture structure results in thousands of genes with a significant higher mRNA expression related to extracellular matrix (ECM), cell adhesion, wound healing and growth factors as compared to a 2D structure 67 . Concentrations of proteins related to angiogenesis, ECM remodeling and regeneration increase as well 67 . The regenerative potency of SVF might be caused by the interaction between cells and growth factors. For example, angiogenesis is significant greater when pericytes and endothelial cells are combined rather than the use of pericytes or endothelial cells alone 68 . Growth factors like VEGF, hepatocyte growth factor and TGF- β and extracellular matrix (ECM) stimulate angiogenesis 69 .