Transient inhibition of the sphingolipid enzyme nSMase2 with GW4869 safeguards hematopoietic stem cell fitness in ex vivo culture. Mitogenic activation from ex vivo culture conditions promotes release of various extracellular vesicles and a decrease in stemness. On nSMase2 blockade with GW4869 in hematopoietic stem and progenitor cells, mitochondrial stress occurs and an activating transcription factor 4-mediated integrated stress response is activated to decrease release of extracellular vesicles that results in enhanced stemness, as measured in competitive transplantation assays. CHOP, CCAAT/enhancer-binding protein homologous protein; eiF2a, eukaryotic translation initiation factor 2A; mTOR, mammalian target of rapamycin; MVB, multivesicular bodies; REDD1, regulated in development and DNA damage response 1, also known as DNA-damage-inducible transcript 4 (DDIT4); ROS, reactive oxygen species. Professional illustration by Somersault18:24.