The role of glycolysis and glucose oxidation in hematopoiesis

PROJECT SUMMARY:
The metabolism of nutrients has been studied using unfractionated tissues, or in vitro. An unresolved question
is how nutrients are metabolized by stem cells in vivo. Our understanding of stem cell metabolism has been
limited by the fact that metabolomics typically requires millions of cells, while stem cells are rare. We
developed methods to profile the metabolome and to trace stable isotope labeled nutrients in hematopoietic
stem cells (HSCs) and other rare cell types purified from tissues. We found that T cell progenitors in the
thymus are glucose avid as compared to HSCs, myeloid and B cell restricted progenitors, in contrast to the
prevailing view that HSCs are more glycolytic than hematopoietic progenitors. Stable isotope tracing
experiments showed that in the bone marrow but not the thymus, glycolysis and the TCA cycle are
disconnected. Hematopoietic loss of pyruvate dehydrogenase (PDH), the gatekeeper enzyme that connects
glycolysis to the TCA cycle, reduced the number of double positive (DP) T cell progenitors but did not affect
HSCs or other hematopoietic cell types. Loss of PDH paradoxically did not impair the TCA cycle in the thymus,
but caused accumulation of pyruvate and aberrant redox balance. Cells which do not oxidize glucose in the
TCA cycle are classically thought to ferment glucose through glycolysis to lactate via lactate dehydrogenase
(LDH). Hematopoietic loss of LDHA, one of the two LDH isoforms, impaired development of erythroid
progenitors but not HSCs, T cell progenitors or other restricted hematopoietic progenitors. The cell type
specificity in the requirement of LDH and PDH in the hematopoietic system raises the question of why different
stem or progenitor cell types choose to use LDH-mediated fermentation or PDH-mediated oxidation in vivo.
This application’s objective is to systematically dissect the role of glycolytic as compared to oxidative
metabolism in HSCs and restricted progenitors. Our hypothesis is that T cell progenitors require oxidation of
glucose via PDH to regulate pyruvate levels and redox homeostasis, in contrast to HSCs, myeloid and B cell
progenitors which are metabolically flexible. In Aim 1 we will test the metabolic mechanisms which mediate the
effects of PDH on DP cells. In Aim 2 we will determine the cellular and metabolic effects of blocking LDHA/B or
PDH alone or in combination in HSCs and restricted progenitors. In Aim 3 we will investigate the role of
LDHA/B and PDH in hematopoietic and thymopoietic regeneration. These experiments will identify the
contribution of glucose to metabolite pools in HSCs and progenitors in vivo, systematically test the idea that
HSCs are glycolytic, and identify mechanisms by which central carbon metabolism regulates hematopoietic
differentiation and regeneration. More generally our experiments will address a fundamental metabolic
question by testing if stem or progenitor cells in vivo switch between glucose fermentation or oxidation, as is
the textbook view, or if some cell types in vivo tolerate the loss of both major glucose catabolic pathways.

Grant Number: 5R01HL161387-04
NIH Institute/Center: NIH
Principal Investigator: Michalis Agathocleous