Late outgrowth endothelial cells resemble mature endothelial cells and are not derived from bone marrow.

A decade of research has sought to identify circulating endothelial progenitor cells (EPC) in order to harness their potential for cardiovascular regeneration. Endothelial outgrowth cells (EOC) most closely fulfil the criteria for an EPC, but their origin remains obscure. Our aim was to identify the source and precursor of EOC and to assess their regenerative potential compared to mature endothelial cells. EOC are readily isolated from umbilical cord blood (6/6 donors) and peripheral blood mononuclear cells (4/6 donors) but not from bone marrow (0/6) or peripheral blood following mobilization with granulocyte-colony stimulating factor (0/6 donors). Enrichment and depletion of blood mononuclear cells demonstrated that EOC are confined to the CD34(+)CD133(-)CD146(+) cell fraction. EOC derived from blood mononuclear cells are indistinguishable from mature human umbilical vein endothelial cells (HUVEC) by morphology, surface antigen expression, immunohistochemistry, real-time polymerase chain reaction, proliferation, and functional assessments. In a subcutaneous sponge model of angiogenesis, both EOC and HUVEC contribute to de novo blood vessel formation giving rise to a similar number of vessels (7.0 ± 2.7 vs. 6.6 ± 3.7 vessels, respectively, n = 9). Bone marrow-derived outgrowth cells isolated under the same conditions expressed mesenchymal markers rather than endothelial cell markers and did not contribute to blood vessels in vivo. In this article, we confirm that EOC arise from CD34(+)CD133(-)CD146(+) mononuclear cells and are similar, if not identical, to mature endothelial cells. Our findings suggest that EOC do not arise from bone marrow and challenge the concept of a bone marrow-derived circulating precursor for endothelial cells.

Dendritic cells previously exposed to mannan-binding lectin enhance cytokine production in allogeneic mononuclear cell cultures.

Mannan (or mannose)-binding lectin (MBL) can bind to monocytes and dendritic cells, but the significance of such interactions is unknown. We hypothesized that the presence of MBL might prevent the differentiation of monocytes into monocyte-derived dendritic cells or interfere with the development of dendritic cells in some way. We therefore investigated the influence of recombinant human MBL on surface antigen expression and on secretion of selected cytokines. By these means, no direct influence of rhMBL on dendritic cell differentiation or maturation was detected. However, mature dendritic cells prepared in the presence of rhMBL and subsequently co-cultured with allogeneic mononuclear cells, markedly promoted production of interleukin-1ß, interleukin-6, and tumor necrosis factor-α in vitro. In most dendritic cell-mononuclear cell combinations, IFN-γ production was also enhanced. This influence required the presence of rhMBL during dendritic cell maturation and was critically dependent on the presence of monocytes. This observation provides evidence that MBL can influence cellular immunity in addition to its established role as an opsonin.

Highly efficient differentiation of hESCs to functional hepatic endoderm requires ActivinA and Wnt3a signaling.

Human embryonic stem cells (hESCs) are a valuable source of pluripotential primary cells. To date, however, their homogeneous cellular differentiation to specific cell types in vitro has proven difficult. Wnt signaling has been shown to play important roles in coordinating development, and we demonstrate that Wnt3a is differentially expressed at critical stages of human liver development in vivo. The essential role of Wnt3a in hepatocyte differentiation from hESCs is paralleled by our in vitro model, demonstrating the importance of a physiologic approach to cellular differentiation. Our studies provide compelling evidence that Wnt3a signaling is important for coordinated hepatocellular function in vitro and in vivo. In addition, we demonstrate that Wnt3a facilitates clonal plating of hESCs exhibiting functional hepatic differentiation. These studies represent an important step toward the use of hESC-derived hepatocytes in high-throughput metabolic analysis of human liver function.

Inactivation of Apc perturbs mammary development, but only directly results in acanthoma in the context of Tcf-1 deficiency.

Apc (adenomatous polyposis coli) encodes a tumour suppressor gene that is mutated in the majority of colorectal cancers. Recent evidence has also implicated Apc mutations in the aetiology of breast tumours. Apc is a component of the canonical Wnt signal transduction pathway, of which one target is Tcf-1. In the mouse, mutations of both Apc and Tcf-1 have been implicated in mammary tumorigenesis. We have conditionally inactivated Apc in both the presence and absence of Tcf-1 to examine the function of these genes in both normal and neoplastic development. Mice harbouring mammary-specific mutations in Apc show markedly delayed development of the mammary ductal network. During lactation, the mice develop multiple metaplastic growths which, surprisingly, do not spontaneously progress to neoplasia up to a year following their induction. However, additional deficiency of Tcf-1 completely blocks normal mammary development and results in acanthoma.