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

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 cells(1). Low oxygen tension (hypoxia) is commonly thought to be a shared niche characteristic in maintaining quiescence in multiple stem cell types(2–4). However, support for the existence of a hypoxic niche has largely come from indirect evidence such as proteomic analysis(5), expression of HIF-1 and related genes(6), and staining with surrogate hypoxic markers (e.g. pimonidazole)(6–8). Here we perform direct in vivo measurements of local oxygen tension (pO(2)) in the bone marrow (BM) of live mice. Using two-photon phosphorescence lifetime microscopy (2PLM), we determined the absolute pO(2) of the BM to be quite low (<32 mmHg) despite very high vascular density. We further uncovered heterogeneities in local pO(2), with the lowest pO(2) (~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 pO(2) values change dramatically after radiation and chemotherapy, pointing to the role of stress in altering the stem cell metabolic microenvironment.