Oxygen tension plays a critical role in the hematopoietic microenvironment in vitro

Abstract

BackgroundIn the bone marrow mesenchymal stromal cells and osteoblasts form functional niches for hematopoietic stem and progenitor cells. This microenvironment can be partially mimicked using in vitro co-culture systems. In this study, we examined the oxygen tension in three distinct compartments in a co-culture system of purified CD34 + cells and mesenchymal stromal cells with regard to different spatial localizations. Design and MethodsHypoxic cells in the co-culture were visualized by pimonidazole staining. Hematopoietic cell distribution, and functional and phenotypic characteristics were analyzed by flow cytometry. The secretion of vascular endothelial growth factor and stromal-derived factor-1 by mesenchymal stromal cells in low oxygen co-cultures was determined by an enzyme-linked immunosorbent assay. The effect of co-culture medium on the hematopoietic cell migration potential was tested in a transwell assay. ResultsIn co-cultures under atmospheric oxygen tension, regions of low oxygen tension could be detected beneath the feeder layer in which a reservoir of phenotypically more primitive hematopoietic cells is located in vitro. In low oxygen co-culture, the adhesion of hematopoietic cells to the feeder layer was decreased, whereas hematopoietic cell transmigration beneath mesenchymal stromal cells was favored. Increased vascular endothelial growth factor-A secretion by mesenchymal stromal cells under low oxygen conditions, which increased the permeability of the monolayer, was responsible for this effect. Furthermore, vascular endothelial growth factor-A expression in low oxygen mesenchymal stromal cells was induced via hypoxia-inducible factor signaling. However, stromal cell-derived factor-1 secretion by mesenchymal stromal cells was down-regulated under low oxygen conditions in a hypoxia-inducible factorindependent manner. ConclusionsWe demonstrate for the first time that differences in oxygen tension cause selective modification of hematopoietic cell and mesenchymal stromal cell interactions in a co-culture system, thus confirming that oxygen tension plays a critical role in the interaction between hematopoietic cells and the niche environment.

Hypoxic regulation of neutrophil function and consequences for Staphylococcus aureus infection

“…Indeed, within the bone marrow structure, haematopoietic stem cells are found sequestered in regions staining most strongly for the hypoxia probe pimonidazole, a 2-nitroimidazole compound, which forms covalent bonds with cellular macromolecules at oxygen levels below 1.3 kPa. Taken together with the evidence that low oxygen tensions favour the maintenance of haematopoietic stem cells in culture [20], hypoxia appears to play a critical role in neutrophil development.…”

Modeling Chemotherapy Resistant Leukemia <em>In Vitro</em>

“…Many groups have shown that co-culture of leukemic cells with BMSC or OB provides a survival advantage when challenged with chemotherapy agents 9,10,[12][13][14][19][20][21] . Work modeling normal immature CD34+ hematopoietic cells in coculture with mesenchymal stem cells (MSC) revealed that hematopoietic cells will interact with the adherent monolayer of MSCs to form three distinct spatial populations of hematopoietic cells 22,23 . Proliferation and differentiation of the CD34+ cells was effected relative to their location within the co-culture 22 .…”

Differential hypoxic regulation of the microRNA-146a/CXCR4 pathway in normal and leukemic monocytic cells: impact on response to chemotherapy

“…[1][2][3][4][5][6] Leukemic cells can infiltrate the niches leading to their enhanced self-renewal and proliferation, quiescence and resistance to chemotherapeutic agents. 7,8 Hypoxia is, therefore, associated with poor prognosis in leukemia, especially in acute myeloid leukemia (AML), [9][10][11][12][13] as also described in cancer and inflammatory diseases.…”