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. 2016 Jun 2;6(6):940–956. doi: 10.1016/j.stemcr.2016.05.002

Figure 4.

Figure 4

Cell-Autonomous and Non-cell-autonomous Roles of TRIF and MYD88 in LPS-Induced HSC Expansion and Myelosuppression

(A) Chimeras: WT recipients of TRIF−/− donor cells, TRIF−/− recipients of WT donor cells, and WT recipients of WT cells were challenged with LPS or PBS and analyzed after 24 hr (n = 4). WT and TRIF−/− mice (parental) were used as a reference (n = 11). Bar graph shows the mean percentage of LT-HSC and MPPs in the Lin− population.

(B) WT recipients (CD45.1) were transplanted with WTGFP+ + MYD88−/− BM cells or WTGFP+ + TRIF−/− BM cells, mixed 1:1 ratio. After 8 weeks mice were challenged with LPS or PBS for 24 hr. Bar graphs show the mean percentage of LT-HSC and MPPs in gated WTGFP+ donor cells (light and dark gray bars, n = 4) and in gated GFP− (CD45.2) KO donor cells (red and black bars, n = 4) within the same recipient.

(C) Bar graph shows the percentage of CMP and GMP in LPS- versus PBS-treated mice: in chimeras WT recipients of TRIF−/− or MYD88−/− cells, TRIF−/− and MYD88−/− recipients of WT cells, and in parental WT, MYD88−/−, and TRIF−/− mice. Parental, n = 8–19; chimeras, n = 4–6.

(D) WT recipients of MYD88−/− donor cells, MYD88−/− recipients of WT donor cells, and parental WT and MyD88−/− mice were challenged with LPS or PBS and analyzed after 24 hr. Bar graph shows the mean percentage of GR1+MAC1+ cells in total BM cells (n = 8–17).

(E) WT recipients (CD45.1) were transplanted with WTGFP+ + MYD88−/− BM cells or WTGFP+ + TRIF−/− BM cells, mixed 1:1 ratio. After 8 weeks mice were challenged with LPS or PBS for 24 hr. Bar graphs show the mean percentage of GR1+MAC1+ cells in in gated WTGFP+ donor cells (WT; n = 4) and in gated GFP− (CD45.2) KO donor cells (KO, n = 4) within the same recipient. PBS controls are WT-GFP+ of all chimeras challenged with PBS (n = 8).

(F) LPS activation of TLR4-MYD88 or -TRIF signaling exerts its effects on hematopoietic cells (HC) through direct and indirect mechanisms: (1) microenvironment-dependent: activation of MYD88 in the BM stroma cells results in the release of factors that act on HC; (2) hematopoietic-restricted: activation of MYD88 in HC stimulates the release of factors affecting neighboring HC in a paracrine fashion; (3) cell-intrinsic (or cell-autonomous): activation of TRIF in the HC causes a direct effect on the cell.

Data represent mean ± SEM. LPS versus PBS: ∗p < 0.05, ∗∗p < 0.01; WT versus MYD88−/− or TRIF−/−: ˆp < 0.05, ˆˆp < 0.01.