Direct measurement of local oxygen concentration in the bone marrow of live animals

Abstract

Characterizing how the microenvironment, or niche, regulates stem cell activity is central to understanding stem cell biology and to developing strategies for therapeutic manipulation of stem cells1. Low oxygen tension (hypoxia) is commonly thought to be a shared niche characteristic in maintaining quiescence in multiple stem cell types2–4. However, support for the existence of a hypoxic niche has largely come from indirect evidence such as proteomic analysis5, expression of HIF-1 and related genes6, and staining with surrogate hypoxic markers (e.g. pimonidazole)6–8. Here we perform direct in vivo measurements of local oxygen tension (pO2) in the bone marrow (BM) of live mice. Using two-photon phosphorescence lifetime microscopy (2PLM), we determined the absolute pO2 of the BM to be quite low (<32 mmHg) despite very high vascular density. We further uncovered heterogeneities in local pO2, with the lowest pO2 (~9.9 mmHg, or 1.3%) found in deeper peri-sinusoidal regions. The endosteal region, by contrast, is less hypoxic as it is perfused with small arteries that are often positive for the marker nestin. These pO2 values change dramatically after radiation and chemotherapy, pointing to the role of stress in altering the stem cell metabolic microenvironment.

Hypoxic regulation of neutrophil function and consequences for Staphylococcus aureus infection

“…Neutrophils are generated within the bone marrow, a significantly hypoxic microenvironment even under healthy physiological conditions, with murine in vivo measurements of local oxygen tension recorded as low as 1.3 kPa [19]. 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.…”

Quantification and three-dimensional microanatomical organization of the bone marrow

“…Many of the niches are constructed around diverse microvessel types, which distinctly regulate access to nutrients, oxygen, and metabolites derived from circulating blood. [64][65][66] Nevertheless, for the most part, available data on BM subcompartments remain fragmentary, and a broader picture integrating comprehensive spatial patterns and number of niches in an organ-wide context is lacking. In fact, only a limited number of studies have analyzed cell subset localization at a global scale and thus explored the hypothesis that hematopoietic cells distribute in a nonarbitrary fashion in accordance to a defined zonation of the BM landscape.…”

Assessment of bone vascularization and its role in bone remodeling

“…However, the peri-endosteal region (zone located at 0-20 mm from the bone surface) in which the smallest vessels were observed exhibited intermediate values (21.9 mm Hg in the vessels and 13.5 mm Hg outside the vessels). 40 A growing number of reports suggested a critical role of local hypoxia in the hematopoietic niche related to the stemness maintenance of the hematopoietic stem cells. These data emphasize the fact that vessel count cannot be a surrogate marker of blood supply or perfusion in bone and that the combination of morphological and functional analyses are necessary to better understand the bone/vessel relationship.…”