ABSTRACT
Notch signaling is essential for definitive hematopoiesis, but its role in human embryonic hematopoiesis is largely unknown. We show that in hESCs the expression of the Notch ligand DLL4 is induced during hematopoietic differentiation. We found that DLL4 is only expressed in a sub-population of bipotent hematoendothelial progenitors (HEPs) and segregates their hematopoietic versus endothelial potential. We demonstrate at the clonal level and through transcriptome analyses that DLL4(high) HEPs are enriched in endothelial potential, whereas DLL4(low/-) HEPs are committed to the hematopoietic lineage, albeit both populations still contain bipotent cells. Moreover, DLL4 stimulation enhances hematopoietic differentiation of HEPs and increases the amount of clonogenic hematopoietic progenitors. Confocal microscopy analysis of whole differentiating embryoid bodies revealed that DLL4(high) HEPs are located close to DLL4(low/-) HEPs, and at the base of clusters of CD45+ cells, resembling intra-aortic hematopoietic clusters found in mouse embryos. We propose a model for human embryonic hematopoiesis in which DLL4(low/-) cells within hemogenic endothelium receive Notch-activating signals from DLL4(high) cells, resulting in an endothelial-to-hematopoietic transition and their differentiation into CD45+ hematopoietic cells.
Subject(s)
Biomarkers/metabolism , Cell Differentiation , Embryonic Stem Cells/cytology , Endothelium/cytology , Hematopoiesis/physiology , Hematopoietic Stem Cells/cytology , Intracellular Signaling Peptides and Proteins/metabolism , Membrane Proteins/metabolism , Animals , Cell Proliferation , Cells, Cultured , Embryoid Bodies , Embryonic Stem Cells/metabolism , Endothelium/metabolism , Female , Flow Cytometry , Gene Expression Profiling , Hematopoietic Stem Cells/metabolism , Humans , Immunoenzyme Techniques , Intracellular Signaling Peptides and Proteins/genetics , Membrane Proteins/genetics , Mice , Mice, Inbred NOD , Mice, SCID , RNA, Messenger/genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
Morphogens regulate epithelial cell fate decisions in the adult gastrointestinal tract. The authors hypothesized that influx of inflammatory cells into the lamina propria may disturb the normal expression gradients of morphogens (morphogenetic landscape) in gastrointestinal epithelia. Changes in the activity of the bone morphogenetic protein (BMP) pathway in normal and Helicobacter pylori-infected gastric mucosa were therefore examined. It is shown that BMP receptors, the activated (phosphorylated) form of the intracellular BMP signal transduction protein SMAD1, and BMP target ID2 all localize to gastric epithelial cells that are at the end of the axis of epithelial renewal in normal mucosa. Colonization of human gastric mucosa with H. pylori was associated with an increase in BMP2 expression due to influx of inflammatory cells that produce BMP2. Furthermore, whereas no BMP4 was detected in the normal antrum, focal infiltrates of BMP4-expressing cells were found in the H. pylori-infected stomach. This influx of BMP-expressing cells was associated with an increase in epithelial BMP signalling. Interestingly, a shift in activity of the BMP pathway was observed towards the precursor cell compartment (isthmus) of the gastric units. Thus, H. pylori infection results in an influx of inflammatory cells that disturb the normal activity gradient of a morphogenetic pathway with an established role in epithelial cell fate regulation. The data suggest that morphological changes in epithelial histology may result from alterations in the morphogenetic landscape secondary to changes in the cellular composition of the lamina propria.
