Figure 7.
Chimeric transplantation demonstrates a dose-dependent effect of HDAC11 depletion in attenuating MPLW515L-induced disease. (A-C) WT recipients (n = 5 per group) were transplanted with MPLWT- or MPLW515L-expressing BM, or with a 1:1 mixture of MPLW515L-expressing Hdac11+/+ and Hdac11−/− BM. Hdac11−/− recipients also received MPLW515L-transduced Hdac11−/− BM to assess potential effects of HDAC11 deficiency on the BM niche. (A) WBC, (B) platelet, and (C) RBC counts were determined in the indicated transplant recipients (Rec) on day 11 (supplemental Figure 6) and day 18 after transplantation. Statistical analysis was performed by using ANOVA, followed by Dunnett’s multiple comparison test with P < .05. (D) Western blot analyses of pSTAT3 and pSTAT5 were performed with BM cells isolated from the indicated transplant recipient mice. (E-F) Spleen and liver (supplemental Figure 6) weights were measured in the indicated transplant recipient mice on day 20. (G) Colony formation assays were performed with BM cells from indicated groups in methylcellulose media without cytokine addition except in the WT control group, which was supplemented with cytokines. ANOVA was performed followed by multiple comparison test using MPLW515L-transplanted WT donors as control. Two-tailed unpaired Student t test was used to compare WT donors receiving MPLW515L-transduced BM from each group. (H) Histological analysis of spleen and BM harvested at day 20. Hematoxylin and eosin (H&E) staining of the spleen and BM shown at an original magnification of ×10 or ×60 (insert) or reticulin stain shown at an original magnification of ×20 using an Olympus BVC51 microscope fitted with a UPlanFL N lens. (I) Hdac11+/+MPLWT, Hdac11+/+MPLW515L, and Hdac11−/−MPLW515L BM cells were transplanted into recipients (n = 3 to 5 per group). Survival was monitored over 70 days. (J) Lin– cells were enriched from BM of recipient mice that received either Hdac11+/+MPLW515L (n = 9) or Hdac11−/−MPLW515L (n = 10) cells after disease was established to similar levels on the basis of GFP expression (supplemental Figure 7B). Enriched Lin– cells were transplanted into lethally irradiated secondary recipients (n = 6 to 8 per group) and survival was monitored. KO, knockout. Survival statistics were performed by using log-rank (Mantel-Cox) test with P value indicated.37 ****P < .0001; ***P < .001; *P < .05.

Chimeric transplantation demonstrates a dose-dependent effect of HDAC11 depletion in attenuating MPLW515L-induced disease. (A-C) WT recipients (n = 5 per group) were transplanted with MPLWT- or MPLW515L-expressing BM, or with a 1:1 mixture of MPLW515L-expressing Hdac11+/+ and Hdac11−/− BM. Hdac11−/− recipients also received MPLW515L-transduced Hdac11−/− BM to assess potential effects of HDAC11 deficiency on the BM niche. (A) WBC, (B) platelet, and (C) RBC counts were determined in the indicated transplant recipients (Rec) on day 11 (supplemental Figure 6) and day 18 after transplantation. Statistical analysis was performed by using ANOVA, followed by Dunnett’s multiple comparison test with P < .05. (D) Western blot analyses of pSTAT3 and pSTAT5 were performed with BM cells isolated from the indicated transplant recipient mice. (E-F) Spleen and liver (supplemental Figure 6) weights were measured in the indicated transplant recipient mice on day 20. (G) Colony formation assays were performed with BM cells from indicated groups in methylcellulose media without cytokine addition except in the WT control group, which was supplemented with cytokines. ANOVA was performed followed by multiple comparison test using MPLW515L-transplanted WT donors as control. Two-tailed unpaired Student t test was used to compare WT donors receiving MPLW515L-transduced BM from each group. (H) Histological analysis of spleen and BM harvested at day 20. Hematoxylin and eosin (H&E) staining of the spleen and BM shown at an original magnification of ×10 or ×60 (insert) or reticulin stain shown at an original magnification of ×20 using an Olympus BVC51 microscope fitted with a UPlanFL N lens. (I) Hdac11+/+MPLWT, Hdac11+/+MPLW515L, and Hdac11−/−MPLW515L BM cells were transplanted into recipients (n = 3 to 5 per group). Survival was monitored over 70 days. (J) Lin– cells were enriched from BM of recipient mice that received either Hdac11+/+MPLW515L (n = 9) or Hdac11−/−MPLW515L (n = 10) cells after disease was established to similar levels on the basis of GFP expression (supplemental Figure 7B). Enriched Lin– cells were transplanted into lethally irradiated secondary recipients (n = 6 to 8 per group) and survival was monitored. KO, knockout. Survival statistics were performed by using log-rank (Mantel-Cox) test with P value indicated.37  ****P < .0001; ***P < .001; *P < .05.

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