Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Hofmann, NA.
OXYGEN SENSING ADULT STEM AND PROGENITOR CELLS
[ Dissertation ] Medical University of Graz; 2012. pp. 138
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- Autor*innen der Med Uni Graz:
- Hofmann Nicole
- Betreuer*innen:
- Graier Wolfgang
- Strunk Dirk
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- Abstract:
- Hypoxia is a major stimulus of new vessel formation (neo-vasculogenesis). In a hypoxic environment endothelial colony-forming progenitor cells (ECFCs) arrange into tubular structures which can connect to the pre-existing vasculature and form functional perfused vessels. The current view is that mesenchymal stem and progenitor cells (MSPCs) are recruited subsequently to stabilize vessels. Stem cell therapy to re-vascularize ischemic tissue has been a promising tool for various therapeutic targets including stroke, myocardial infarction and peripheral artery disease. So far, clinical applications of endothelial progenitors have largely failed to meet medical needs. Previous work showing that ECFCs require MSPCs in vivo to form patent vessels has provoked the hypothesis that MSPCs have a decisive role in the vasculogenic response to hypoxia. This work demonstrates that ECFCs in hypoxic conditions in vivo need the presence of functional MSPCs not only to stabilize but primarily to initiate neo-vasculogenesis by a hypoxia-inducible transcription factor (HIF)-dependent mechanism. Adult human ECFCs were isolated from peripheral blood and MSPCs from bone marrow aspirates and expanded under humanized conditions. Progenitor cell phenotype, long-term proliferation, HIF stabilization, wound repair, migration and vasculogenic functions were monitored under severe hypoxia (1% O2), venous oxygen levels (5% O2) and standard ambient air culture conditions (20% O2). ECFC and MSPC crosstalk in vivo was studied in immune-deficient NSG mice after subcutaneous implantation using various extracellular matrices. Chemical and genetic inhibition of HIF (YC-1, shRNA) was used to define the cell type-specific role of hypoxia sensing in MSPCs and ECFCs during vasculogenesis in vivo. To determine the capacity of downstream target proteins of HIF-1¿ to substitute for MSPC presence during vasculogenesis, selected growth factors and cytokines were tested. Progenitor proliferation and function in vitro were reduced with declining oxygen levels. HIF-1¿ was stabilized by ECFCs only at 1% O2, while MSPCs stabilized HIF-1¿ already at 5% O2. In an NSG mouse model, ECFCs transplanted into a hypoxic environment did not stabilize HIF-1¿, while transplanted MSPCs alone or MSPCs in co-transplants showed strong nuclear HIF-1¿ stabilization as early as 1 day after transplantation. In the absence of MSPCs, ECFCs injected alone largely underwent apoptosis within 24h in vivo. Chemical as well as genetic inhibition of HIF-1¿ stabilization in MSPCs but not in ECFCs significantly abrogated vessel formation in vivo. Blocking the HIF-1¿ down-stream target VEGF resulted in inhibition of neo-vasculogenesis. Interestingly, substitution of VEGF alone could not restore vessel formation in vivo; neither when injected together with ECFCs alone nor in a model where ECFCs were co-transplanted with HIF-depleted MSPCs. Supplementation of a complex mixture of platelet-derived factors in vivo could only in part substitute the vasculogenic function of HIF-competent MSPCs. MSPCs react to a low oxygen environment by stabilizing HIF-1¿ faster and more sensitively than ECFCs. In the initial phase of vasculogenesis MSPCs promote vessel formation at least in part by rescuing ECFCs from hypoxia-induced apoptosis by a HIF-dependent trophic mechanism. Surprisingly, neo-vasculogenesis can occur independently of endothelial HIF stabilization. These results argue in favor of MSPC/ECFC co-transplantation as a promising cellular therapy for vascular regeneration. The finding that VEGF alone could not compensate for the vasculogenic function of MSPCs in vivo highlights the complexity of the hypoxia-induced cytokine network. The fact that HIF stabilization in MSPCs but not in ECFCs is crucial to initiating vascular regeneration supports a shift of focus from endothelial cells to perivascular mesenchymal cells as a therapeutic target in regenerative medicine and anti-angiogenic therapy.