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. 2018 May 7;215(5):1337–1347. doi: 10.1084/jem.20171477

Figure 3.

Figure 3.

The spatial relationship of pSHP-1+ HSCs in BM niche. (A) Representative image of a BM section from a C57BL/6J mouse showing pSHP-1+ HSCs were surrounded by MKs. The yellow arrowheads indicate HSCs. Gray, lineage cells; green, MKs; red, HSCs; blue, pSHP-1. Dashed box indicates area of focus. Bars, 25 µm. (B) Percentage of pSHP-1+ HSCs in BM. (C) Distance between pSHP-1+ HSCs and pSHP-1– HSCs to MKs as indicated (n = 311 HSCs from four mice in four independent experiments). (D) Competitive reconstitution analysis of functional HSCs from Pf4-cre–; TGF-β1f/f or from Pf4-cre+; TGF-β1f/f mice. Plots of peripheral blood analysis for total engrafted donor cells at the indicated number of weeks after primary transplantation (1°) and secondary transplantation (2°; n = 10). (E) TGF-β1 protein levels as determined by ELISA in BM from Pf4-cre; TGF-β1f/f mice (cre−, n = 4 mice; cre+, n = 5 mice). The spatial relationship of MKs and Schwann cells. (F) Representative image of a BM section showing CD41+ MKs (green) and GFAP+ Schwann cells (red). Bar, 100 µm. (G) Distance between MKs and Schwann cells. The numbers on the x axis indicate intervals of 10 µm (−10 indicates the interval 0–10; −20 indicates 10–20, and so on). P = 2.7 × 10−14 by two-sample KS test. (H) Probability distributions of the mean distances between MKs and Schwann cells derived from simulations of randomly positioned MKs and actual Schwann on maps of BM. Mean distances observed in situ (red line) are shown in relation to the grand mean (mean of the means) ± 3 SD (3σ, blue and dotted black lines, respectively). Probability P (μ < 25.89 µm) = 2.2 × 10−16. n = 365 MKs from three mice in three independent experiments. (I) The spatial relationship of HSCs (red), MKs (blue), and Schwann cells (green). dMK and dSW indicate the distance between the HSC to the closest MK and Schwann cell, respectively. The arrowheads indicate HSCs. Bars, 25 µm. (J) 2D probability distribution of the distances between HSCs to MKs (dMK) and Schwann cells (dSW) in the BM. Upper panel, HSC distribution in the BM (n = 95 HSCs, from three mice); lower panel, randomized HSC distribution models generated by computational simulations of random HSC localization (P = 1.57 × 10−20 by 2D KS test with two samples). *, P < 0.05; **, P < 0.01; ***, P < 0.001.