Abstract
The fate of hematopoietic stem cells (HSCs) can be directed by microenvironmental factors including extracellular calcium ion concentration ([Ca2+]e), but the local [Ca2+]e around individual HSCs in vivo remains unknown. Here we develop intravital ratiometric analyses to quantify the absolute pH and [Ca2+]e in the mouse calvarial bone marrow, taking into account the pH sensitivity of the calcium probe and the wavelength-dependent optical loss through bone. Unexpectedly, the mean [Ca2+]e in the bone marrow (1.0 ± 0.54 mM) is not significantly different from the blood serum, but the HSCs are found in locations with elevated local [Ca2+]e (1.5 ± 0.57 mM). With aging, a significant increase in [Ca2+]e is found in M-type cavities that exclusively support clonal expansion of activated HSCs. This work thus establishes a tool to investigate [Ca2+]e and pH in the HSC niche with high spatial resolution and can be broadly applied to other tissue types.
Highlights
The fate of hematopoietic stem cells (HSCs) can be directed by microenvironmental factors including extracellular calcium ion concentration ([Ca2+]e), but the local [Ca2+]e around individual HSCs in vivo remains unknown
In contrast to the tightly regulated calcium ion concentration ([Ca2+]) in the blood serum with a setpoint near 1 mM3–5, the local extracellular calcium concentration ([Ca2+]e) at the endosteal surface can reach as high as 40 mM at sites of bone resorption, and the difference may help to specify the bone marrow (BM) as the destination for HSC homing during development and after systemic transplantation[1]
An average [Ca2+]e of 0.5 mM was found in BM interstitial fluid extracted from tibia, much lower than serum calcium, suggesting that HSCs are maintained in a BM microenvironment with low [Ca2+]e3
Summary
The fate of hematopoietic stem cells (HSCs) can be directed by microenvironmental factors including extracellular calcium ion concentration ([Ca2+]e), but the local [Ca2+]e around individual HSCs in vivo remains unknown. In contrast to the tightly regulated calcium ion concentration ([Ca2+]) in the blood serum with a setpoint near 1 mM3–5, the local extracellular calcium concentration ([Ca2+]e) at the endosteal surface can reach as high as 40 mM at sites of bone resorption, and the difference may help to specify the BM as the destination for HSC homing during development and after systemic transplantation[1]. An average [Ca2+]e of 0.5 mM was found in BM interstitial fluid extracted from tibia, much lower than serum calcium, suggesting that HSCs are maintained in a BM microenvironment with low [Ca2+]e3 These conflicting observations highlight the need to further examine the role of extracellular calcium in regulating the fate and function of HSCs in vivo. We show that long-term (LT)-HSCs reside in BM locations with elevated [Ca2+]e ranging from ~0.8 to 2.6 mM
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