MSCs have generally been shown to express Sca-1, PDGR- (CD140a), NST, and CD133 (9, 13, 15, 30), whereas GFP-negative cells do not express CD133 and express variable levels of PDGR-. immunofluorescence analysis for Cytochalasin B clean muscle-specific caldesmon, nearly all cells were immunoreactive ( em SI Appendix /em , Fig. S12 em D /em , em 1C4 /em ). These results demonstrate that fibroblast-like cells from embryos have the capacity to mature into cell types much like those of pericytes. Conversation This study establishes that embryonic cells of the fibroblast class constitute a rich human population of subtypes. From a transgenic reporter mouse collection in which a VEGF promoter fragment drives the manifestation of GFP, ethnicities containing Cytochalasin B subpopulations of GFP-positive and GFP-negative cells with the typical characteristics of fibroblasts could be initiated. Within the GFP-negative cells, further subdivisions could be made based on the large quantity of various cell surface proteins. Our analysis identified a large number of unique subpopulations in ethnicities initiated from embryos. Cells with differing cell surface phenotypes were not very easily distinguished from one another by morphology; for example, despite obvious variations in functionality, GFP-negative and GFP-positive fibroblasts were not distinguishable ultrastructurally. GFP-negative fibroblasts displayed a well-spread morphology with prominent stress fibers and could become induced to differentiate into cell types of extra fat, muscle, and bone lineages. These features will also be exhibited from the fibroblast-like cells identified as MSCs (29); however, the patterns of phenotypic manifestation recognized here diverge from previously recognized compilations of MSC phenotypes. MSCs from bone marrow have been purified by bad selection for antibodies against CD34, CD45, and CD14. In contrast, we found that GFP-negative cells indicated detectable levels of CD34 and CD14. GFP-negative cells show low levels of CD105 and CD73, in contrast to MSCs. MSCs have generally been shown to express Sca-1, PDGR- (CD140a), NST, and CD133 (9, 13, 15, 30), whereas GFP-negative cells do not express CD133 and express variable levels Cytochalasin B of PDGR-. GFP-negative cells communicate CD24, unlike preparations of MSCs. GFP-negative cells communicate PDPN, a marker suggested to be absent from MSCs (10, 11). The pattern of surface protein markers of the GFP-negative cells does not coincide with that of additional known MSCs or the more primitive multipotent adult progenitor cells (14). Although GFP-negative cells communicate some proteins found in ES cells, such as NST, nestin, and Fra-1, they do not communicate additional stem cell markers, such as Sox-2, Klf-4, Oct-4, cMyc, CD31, SSEA-1, SSEA-3, and Tra-1-81. Both GFP-positive and GFP-negative cells express proteins found in DcR2 contractile and intermediate filaments, such as SMA, sM22, and vimentin, but lack others, such as desmin and smMHC. GFP-positive cells express ER-TR7 and FAP, whereas GFP-negative cells express only FAP. Following the identification of a small number of surface markers that could be used to subdivide the population by expression pattern, the fibroblast-like cells were found to be extremely heterogeneous, belying what appear to be relatively common assumptions about MEF uniformity. Multicolor circulation cytometry revealed a complex populace structure of subtypes with varying degrees of stability of the cell surface phenotype. Gating cells by CD73 and CD146 expression gave three subtypes that were further analyzed for CD54 and CD71 expression, generating 12 unique patterns. Additional gating on CD24, CD80, and CD90.1 allowed further differentiation of subtypes. On growth of sorted populations in culture, the most frequently observed behavior was retention of the expression pattern used to select the population. Consistent with the observed retention of characteristics following culture of sorted subpopulations, cells from individual colonies showed unimodal large quantity distributions and distinguishable characteristics, a obtaining inconsistent with the interpretation that this heterogeneous expression may be attributed to gene or gene network noise (31). Cells with differing surface phenotypes could be coerced into bone, muscle, and excess fat lineages, indicating that at least.