Extended Data Fig. 4: Gram-negative bacteria contribute to the expansion of pre-leukaemic HSCs.
From: Microbial metabolite drives ageing-related clonal haematopoiesis via ALPK1
a, Experimental design to examine the contribution of bacteria (gram-positive vs gram-negative) following gut injury. BM cells from Dnmt3af/f;Mx1-Cre mice (n = 4) treated with poly(IC) to induce recombination were transplanted into WT mice (n = 32) to generate chimeric mice. Chimeric mice established using low-dose irradiation were pre-treated with antibiotics targeting gram-negative bacteria (metronidazole, gentamicin, and neomycin) or antibiotics targeting gram-positive bacteria (vancomycin) for 4 weeks and then treated with DSS for 1 week. b, Number of donor HSCs (Lin-cKit+Sca1+ CD150+CD48-) in the BM 2 weeks after treatment (n = 8 mice per group). c, Experimental design to examine the contribution of bacteria (gram-positive vs gram-negative) in young and old mice. BM cells from Dnmt3af/f;Mx1-Cre mice (n = 6) treated with poly(IC) to induce recombination were transplanted into WT mice (n = 48) to generate chimeric mice. Chimeric mice established using low-dose irradiation were pre-treated with antibiotics targeting gram-negative bacteria (metronidazole, gentamicin, and neomycin) or antibiotics targeting gram-positive bacteria (vancomycin) for 4 weeks. d, Number of donor HSCs (Lin-cKit+Sca1+ CD150+ CD48−) in the BM 2 weeks after treatment (n = 12 mice per group). e, Experimental design to examine the contribution of the microbiome using faecal microbial transplant (related to Fig. 1f,g). BM cells from Dnmt3af/f;Mx1-Cre mice or Dnmt3a+/+;Mx1-Cre mice (n = 2 per group) treated with poly(IC) to induce recombination were transplanted into WT mice (n = 12 mice per group) to generate chimeric mice. Chimeric mice established using low-dose irradiation were pre-treated with broad-spectrum antibiotics (ABX) for 4 weeks and then colonized with faecal material from either H2O- or DSS-treated WT mice. f, ADP-heptose levels in plasma from Dnmt3aWT or Dnmt3aKO mice colonized by FMT from H2O- or DSS-treated WT mice (H2O: n = 6, DSS: n = 6 biological replicates). Error bars represent the SEM. *, P < 0.05, **; P < 0.01, ***; P < 0.001; ****, P < 0.0001. P values were calculated for the indicated comparisons using a two-tailed unpaired Student’s t-test (b, d, and f).
