Human haematopoietic stem cells remember inflammatory stress

Ethics statement Human CB samples were obtained with informed consent from Trillium Health, Credit Valley and William Osler Hospitals according to procedures approved by the University Health Network (UHN) Research Ethics Board (REB# 02-0763). The investigation of data from the Ontario Health Study was approved by the University of Toronto Research Ethics Board (protocol #00033112). Consent for use of data and blood samples was previously collected through the Ontario Health Study 63 and CARTaGENE 64 regional cohorts within the Canadian Partnership for Tomorrow’s Health 65 . All research using human material was performed in accordance with relevant guidelines and regulations. Human CB sample processing Mononuclear cells from pools of male and female CB (approximately 1–8 donors) were obtained by centrifugation over a density barrier of Lymphoprep (Multicell). After red cell lysis with ammonium chloride (STEMCELL technologies), mononuclear cells were enriched for HSPCs by positive selection with a CD34 Microbead kit (Miltenyi) per the manufacturer’s instructions, or by lineage depletion as previously described 66 . Resulting HSPC-enriched CB cells were cryopreserved in 50% PBS, 40% fetal bovine serum (FBS) and 10% DMSO and stored at −80 °C short term or −150 °C long term. CB LT-HSC scRNA-seq Human immunophenotypic LT-HSCs were purified from pooled CB samples based on a Lin – CD34 + CD38 − CD45RA − CD90 + CD49f + immunophenotype (see FACS section below). …