Extended Data Figure 3: Depletion of nestin⁺ cells or their Cxcl12 production accelerates MPN.

a, Experimental design for in vivo nestin⁺ cell depletion. Nes-cre^ERT2;iDTA and control iDTA mice were transplanted with BM cells from pIpC-treated Mx1-cre;JAK2(V617F) and control mice and treated with tamoxifen (n = 3). b–e, Unlike Schwann cells, MSCs are reduced in Nes-cre^ERT2;iDTA BM. b, c, Frequency of BM CD45⁻CD31⁻Ter119⁻CD90⁺ cells (b) and fibroblastic colony-forming units (CFU-F, c) in BM CD45⁻Ter119⁻ cells from Nes-cre^ERT2;iDTA and control mice after 4-week tamoxifen treatment. d, Expression of glial fibrillary acidic protein (Gfap) mRNA (normalized to Gapdh) in the BM of mice in a. e, Quantification of Gfap immunostaining of femoral BM from mice in b, c. f–k, Early mutant HSC expansion and mobilisation correlates with reduced Cxcl12 expression in BM nestin⁺ cells. f, g, BM lin⁻sca-1⁺c-kit⁺ (f) CD34⁻Flt3⁻ long-term (LT) and CD34⁺Flt3⁻ short-term (ST) HSCs, and CD34⁻Flt3⁻ multipotent progenitors (MPP) and (g) frequency of haematopoietic colony-forming units in culture (CFU-C) from BM nucleated cells, spleen or peripheral blood from Nes-Gfp;Mx1-cre;JAK2(V617F) (n = 6) and control (n = 11) mice 6 weeks after pIpC treatment. h, Appearance (inset; scale bar, 1 cm) and haematoxylin and eosin stainings of spleens (magnification, ×100). i–k, Cxcl12 protein (i) and mRNA levels (j, k) in BM extracellular fluid (i), nucleated cells (j) and stromal Nes-GFP⁺ cells (k) of Mx1-cre;JAK2(V617F) and control mice 6 weeks after pIpC treatment (i, j) or Nes-gfp mice 10 weeks after transplantation with mutant BM (n = 4) or control cells (n = 7) (k). l, Circulating platelets in Nes-cre^ERT2;Cxcl12^fl/fl (n = 11) and control Cxcl12^fl/fl littermates (n = 14) 18 weeks after transplantation with BM cells from MPN mice and 12 weeks after tamoxifen treatment. Mean ± s.e.m. *P < 0.05, **P < 0.01 (unpaired two-tailed t test).