Purine Metabolism and Hematopoietic Stem and Progenitor Stem Cells Under Stress

Hematopoietic stem cells (HSCs) play a key role in the lifelong maintenance of hematopoiesis through self-renewal and multi-lineage differentiation. Adult HSCs reside within a specialized microenvironment of the bone marrow (BM), called "niche", in which they are maintained in a quiescent state in cell cycle. Most of HSCs within BM show quiescence under the hypoxic niche. Since the loss of HSC quiescence leads to the exhaustion or aging of stem cells through excess cell division, the regulation of quiescence in HSCs is essential for hematopoietic homeostasis. On the other hand, cellular metabolism has been suggested to play a critical role in many biological processes including the regulation of stem cell properties and functions. However, the metabolic condition and adaptation of stem cells remain largely unaddressed.

First, we have analyzed HSC metabolism using metabolomics approaches. With step-wise differentiation of stem cells, the cell metabolism associated with each differentiation stage may be different. A feature of quiescent HSCs is their low baseline energy production; quiescent HSCs rely on glycolysis and exhibit low mitochondrial membrane potential (ΔΨ_m). Likewise, HSCs with a low ΔΨ_m show higher reconstitution activity in BM hematopoiesis, compared to cells with high ΔΨ_m. By contrast, upon stress hematopoiesis, HSCs actively divide and proliferate. However, the underlying mechanism for the initiation of HSC division still remains unclear. In order to elucidate the mechanism underlying the transition of cell cycle state in HSCs, we analyzed the change of mitochondria activity in HSCs after BM suppression induced by 5-fluoruracil (5-FU). Upon 5-FU treatment, cycling progenitors are depleted and then quiescent HSCs start to divide. We found that HSCs initiate cell division after exhibiting enhanced ΔΨ_m, as a result of increased intracellular Ca²⁺ level. We hypothesize that extracellular adenosine, derived from hematopoietic progenitors, inhibits the calcium influx and mitochondrial metabolism. While further activation of Ca²⁺-mitochondria pathway led to loss of the stem cell function after cell division, the appropriate suppression of intracellular Ca²⁺ level by nifedipine, a blocker of L-type voltage-gated Ca²⁺ channels, prolonged cell division interval in HSCs, and simultaneously achieved both cell division and HSC maintenance (self-renewal division). Thus, our results indicate that the adenosine-Ca²⁺-mitochondria pathway induces HSC division critically to determine HSC cell fate. Next, to examine the mitochondria oxidative metabolism and purinergic pathways, we introduced the study on a tumor suppressor, Folliculin (FLCN). Conditional deletion of FLCN in HSC compartment using the Mx1-Cre or Vav-iCre system disrupted HSC quiescence and BM homeostasis dependently on the lysosomal stress response induced by TFE3. Together all, we propose that the change in cellular metabolism involving mitochondria is crucial for HSC homeostasis in the stress settings.