Augmenting regenerative hematopoietic stem cell self-renewal by intervening in peroxisomal β-oxidation

Robust tissue regeneration relies on extensive stem cell self-renewal. However, the molecular mechanisms promoting stem cell self-renewal during regeneration remain unclear. Here we investigate the role of peroxisomes, a key metabolic organelle, in regulating hematopoietic stem cells (HSCs). We found that disrupting peroxisome biogenesis by deleting Pex5 does not significantly impact HSC number or hematopoiesis at steady state. Strikingly, Pex5-deficient HSCs self-renew extensively and give rise to robust multilineage reconstitution upon multiple rounds of serial transplantation into recipients. Pex5-deficient HSCs are more resistant to stress and have enhanced capacity to regenerate the hematopoietic system following chemo drug treatment or irradiation. Deletion of Pex5 promotes the expansion of HSCs and hematopoietic progenitors with extensive serial ex vivo plating capacity. Pharmacological inhibition of peroxisomal β-oxidation also promotes extensive plating by HSCs and hematopoietic progenitors. We propose that intervening in peroxisomal β-oxidation can be exploited to promote HSC and progenitor function for regenerative medicine. Mx1-cre and Pex5fl mice were obtained from the Jackson Laboratory and maintained on C57BL/6 background. pIpC was injected to young adult mice for five doses every other day to Pex5fl/fl mice and Mx1-cre; Pex5fl/fl mice. Samples were collected at least three weeks after pIpC injection. The mice for sequencing were analyzed at three-month-old. DAPI-lineage-cKit+ cells from bone marrow were double-sorted in a yield-purity order of sorting modes on BD SORP FACSAria. Immediately following FACS isolation, 10x Genomics Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 were used to prepare the library according to the manufacturer’s instructions. Individual libraries were pooled for sequencing on the NovaSeq 6000 Sequencing System (Illumina). Control group samples were collected from Pex5fl/fl mice (LY001, LY003); Mutant group samples were collected from Mx1-cre; Pex5fl/fl mice (LY002, LY004).