Abstract
Proper control of mitochondrial function is a key factor in the maintenance of hematopoietic stem cells (HSCs). Mitochondrial content is commonly measured by staining with fluorescent cationic dyes. However, dye staining can be affected, not only by xenobiotic efflux pumps, but also by dye intake, which is dependent on the negative charge of mitochondria. Therefore, mitochondrial membrane potential (ΔΨmt) must be considered in these measurements because a high ΔΨmt due to respiratory chain activity can enhance dye intake, leading to the overestimation of mitochondrial volume. Here, we show that HSCs exhibit the highest ΔΨmt of the hematopoietic lineages and, as a result, ΔΨmt-independent methods most accurately assess the relatively low mitochondrial volumes and DNA amounts of HSC mitochondria. Multipotent progenitor stage or active HSCs display expanded mitochondrial volumes, which decline again with further maturation. Further characterization of the controlled remodeling of the mitochondrial landscape at each hematopoietic stage will contribute to a deeper understanding of the mitochondrial role in HSC homeostasis.