The role of oxygen availability in embryonic development and stem cell function

Abstract

Low levels of oxygen (O2) occur naturally in developing embryos. Cells respond to their hypoxic microenvironment by stimulating several hypoxia-inducible factors (and other molecules that mediate O2 homeostasis), which then coordinate the development of the blood, vasculature, placenta, nervous system and other organs. Furthermore, embryonic stem and progenitor cells frequently occupy hypoxic 'niches' and low O2 regulates their differentiation. Recent work has revealed an important link between factors that are involved in regulating stem and progenitor cell behaviour and hypoxia-inducible factors, which provides a molecular framework for the hypoxic control of differentiation and cell fate. These findings have important implications for the development of therapies for tissue regeneration and disease.

Figure 1. Branch morphogenesis during D. melanogaster tracheal development and mammalian blood vessel formation are regulated by O₂ levels

(A) Models for O₂ sensing and patterning in D. melanogaster. Cells within a target tissue experiencing low O₂ (blue), due to their distance from an existing tracheal branch (red), begin expressing Branchless (Bnl, the orthologue of mammalian FGF). Branchless expression increases in these O₂-starved cells and the tracheal cells respond by sprouting terminal branches that grow toward each Bnl signalling centre. When the branch approaches the source, it starts to arborize (adapted from 12). (B) Model for vascular morphogenesis. Haemangioblasts are putative mesodermal progenitor cells giving rise to both haematopoietic stem cells (HSCs) and angioblasts, the forerunner of endothelial cells which line the vasculature. Vascular endothelial growth factor (VEGF) is required to generate haemangioblasts in the developing embryo. Vasculogenesis, the formation of a primary endothelial cell plexus, also depends on VEGF. Angiogenic remodelling into a mature vascular system (including arteries and veins), involves other important endothelial cell receptors and their ligands, such as Tie2, Tie1, angiopoietin-1 (Ang-1), angiopoietin-2 (Ang-2), and transforming growth factor-β (TGFβ)/TGF-receptor (TGFR) interactions. Of note, virtually all of these vasculogenic and angiogenic regulatory factors (VEGF, Tie2, angiopoietins, TGFβ, platelet-derived growth factor-β (PDGFβ), etc.) are regulated by both decreased O₂ levels and the HIFs.

Figure 1. Branch morphogenesis during D. melanogaster tracheal development and mammalian blood vessel formation are regulated by O₂ levels

(A) Models for O₂ sensing and patterning in D. melanogaster. Cells within a target tissue experiencing low O₂ (blue), due to their distance from an existing tracheal branch (red), begin expressing Branchless (Bnl, the orthologue of mammalian FGF). Branchless expression increases in these O₂-starved cells and the tracheal cells respond by sprouting terminal branches that grow toward each Bnl signalling centre. When the branch approaches the source, it starts to arborize (adapted from 12). (B) Model for vascular morphogenesis. Haemangioblasts are putative mesodermal progenitor cells giving rise to both haematopoietic stem cells (HSCs) and angioblasts, the forerunner of endothelial cells which line the vasculature. Vascular endothelial growth factor (VEGF) is required to generate haemangioblasts in the developing embryo. Vasculogenesis, the formation of a primary endothelial cell plexus, also depends on VEGF. Angiogenic remodelling into a mature vascular system (including arteries and veins), involves other important endothelial cell receptors and their ligands, such as Tie2, Tie1, angiopoietin-1 (Ang-1), angiopoietin-2 (Ang-2), and transforming growth factor-β (TGFβ)/TGF-receptor (TGFR) interactions. Of note, virtually all of these vasculogenic and angiogenic regulatory factors (VEGF, Tie2, angiopoietins, TGFβ, platelet-derived growth factor-β (PDGFβ), etc.) are regulated by both decreased O₂ levels and the HIFs.

Figure 2. Oxygen gradients are generated in developing human and mouse placentae

(A) Diagrammatic representation of the differentiation pathway that cytotrophoblast stem cells undertake in vivo. These cells detach from the underlying uterine basement membrane and either fuse to form multinucleated syncytiotrophoblasts or columns of mononuclear cells that attach the conceptus to the uterine wall. A subset of these cells stops proliferating and differentiates into invasive cytotrophoblasts that breach and enlarge maternal blood vessels to generate an utero–placental circulation. The differentiation of proliferating cytotrophoblasts into invasive cytotrophoblasts is an O₂-dependent process, with O₂ levels increasing as cells migrate towards the maternal spiral arteries. (B) Placentation is regulated by changes in O₂ availability. An E8.0 mouse embryo is shown to illustrate the O₂ gradient generated during murine placentation. Similar to human placentae, the early murine placenta generates an O₂ gradient where cells that migrate dorsally experience increasing O₂ levels. Trophoblast stem cells adopt specific cell fates in the placenta when they encounter discrete O₂ levels: low O₂ enforces a spongiotrophoblast cell fate, whereas higher O₂ levels enforce a giant cell fate.

Figure 3. Distinct populations of stem cells occupy microenvironments that contain different O₂ levels

As described in the main text, some stem cells (such as those in the endosteal bone marrow compartment) occupy extremely low O₂ microenvironments (less than 0.5% O₂) as shown in (A). Other stem cells (as those described as perivascular SLAM⁺ (for Signalling Lymphocyte Activation Molecule) stem cells can occupy relatively well-oxygenated environments as they are in close proximity to blood vessel endothelial cells (B). However, it should be noted that although stem cells can be perivascular, the vessels might be associated with venous structures and therefore be relatively hypoxic.

Figure 4. Models depicting O₂ availability and transcriptional activity

(A) Under hypoxic conditions, hypoxia-inducible factor-1α (HIF-1α) typically interacts with HIF-1β (also known as aryl hydrocarbon receptor nuclear translocator (ARNT)) to stimulate target genes such as vascular endothelial growth factor (VEGF), fibroblast growth factor 2 (FGF-2), and platelet-derived growth factor-β (PDGF- β). HIF-1α can also interact with the intracellular domain (ICD) of Notch in the nucleus at Notch-responsive promoters. In the nucleus, Notch interacts with the CSL (C-promoter-binding factor/Suppressor-of-Hairless/Lag1) DNA-binding protein and coactivators such as CBP/p300 and Mastermind (Mam1) to activate target genes such as Hes and Hey. It is currently not known if the initial HIF-1α–Notch interaction occurs outside or within the nucleus. Furthermore, the actual relationship between components of the Notch complex at promoters is unclear. HIF-1α could directly interact with ICD, an unidentified ‘bridging’ protein, or with Maml . (B) In cells where the Oct-4 locus is accessible as a result of open chromatin, its transcription is induced directly by HIF-2α–ARNT dimers in response to hypoxia.

Figure 5. Multiple pathways responding to changes in O₂ availability affect developmental processes as well as social behaviour

As described in the text, HIFs regulate many aspects of cardiovascular morphogenesis and stem and/or progenitor cell maintenance. Mutagenesis of mammalian target of rapamycin (mTOR) and its associated proteins, such as raptor, rictor, and mLST8, has revealed an important role for mTORC1 and mTORC2 during embryonic development. However, whether mTOR is responding to hypoxia in embryonic microenvironments to regulate development remains to be determined. The “unfolded protein response” (UPR)-regulated kinase PERK (and its substrate eIF2α) is necessary for pancreatic β cell production during development or shortly after birth. Inositol-requiring-1 (IRE1) is another ER-associated UPR effector that activates X box-binding protein 1 (XBP-1), promoting the transcription of ER chaperone genes such as BiP and c/EBP-homologous protein (CHOP) . Finally, cyclic guanosine monophosphate (cGMP) regulation promotes neuronal activity and social feeding behaviour in C. elegans, allowing them to avoid O₂ levels outside the range of 5–12% O₂. This leads to specific appearances of nematode colonies, causing “bordering” or “aggregation”.