LT-HSC engraftment after BMT is severely impaired by blockade of host megakaryocytes. (A) GFP⁺ donor engraftment in competitive secondary (2°) transplant (BMT) assays in which GFP⁺ donor BM was transplanted (5 × 10⁶ BM cells) into post-TBI WT, anti-CD41–treated WT, and mpl^−/− primary (1°) recipients. (B) In the initial assay, BM was harvested 24 hours after 1° BMT and transplanted with 2 × 10⁵ WT competitor BM cells into post-TBI WT 2° recipients. GFP⁺ peripheral RBC, platelet, total WBC, myeloid (GR-1⁺), and B-cell (B220⁺) engraftment at 3 weeks after 2° BMT is shown. *P < .05 vs 2° recipients receiving WT 1° recipient BM. (C) Separate competitive transplant assay showing GFP⁺ peripheral RBC, platelet, total WBC, myeloid, B-cell, and T-cell (CD3⁺) engraftment at 12 weeks after competitive 2° BMT with 10⁵ WT competitor BM cells, and either WT, WT + αCD41, or mpl^−/− 1° recipient BM, harvested 3 weeks after 1° recipient BMT with 2 × 10⁵ GFP⁺ donor BM cells. *P < .05 vs 2° recipients receiving WT 1° recipient cells. (D) Percentage of competitively transplanted 2° recipients receiving WT, WT + αCD41, or mpl^−/− 1° recipients cells 3 weeks after 1° BMT demonstrating >3% GFP⁺ cells in specified lineage from 3 to 24 weeks after 2° BMT. *P < .05 vs percentage of 2° recipients of anti-CD41–treated or mpl^−/− 1° recipient BM.