Expansion, increased cell cycle activity, decreased engraftability and inflammatory signature in Abi-1 deficient hematopoietic stem/progenitor cells. (A) Frequencies of Lin- cells in 5 × 10⁵ bone marrow. (B) Frequencies of LSK or LK cells in lineage-negative fractions of Abi-1^WT or Abi-1^KO bone marrow. (C) Frequencies of long-term (LT)- and short-term (ST)-HSCs and multipotent progenitors in LSK fractions were determined by sorting. Sorting was performed using bone marrow obtained from 6 sex-matched 14-week-old Abi-1^WT or Abi-1^KO mice. Sorting strategy is presented in supplemental Figure 5A. Average percentages of (D) EdU-positive LSK cells in G0/G1, S, and G2/M phase and (E) EdU-positive LT-HSCs in G0/G1 and S/G2/M phase of the cell cycle. Data from 4 experiments each using bone marrow from 14-week-old sex-matched Abi-1^WT or Abi-1^KO animals are shown as mean. Noncompetitive bone marrow primary (F) and secondary (G) transplant. Bone marrow cells isolated from Abi-1^WT or Abi-1^KO mice (before poly[I:C] injection) were transplanted into lethally irradiated recipient C57BL/6 wild-type mice in the absence of competitor cells. Four weeks after the transplantation, Abi-1 loss was induced by poly(I:C) administration. Twenty-four weeks posttransplant, bone marrow cells were harvested from primary recipients and transplanted into the secondary recipients. Average donor chimerism in primary and secondary recipients was monitored every 4 weeks for 24 weeks posttransplantation; n = 4 Abi-1^WT or n = 4 Abi-1^KO donor mice (CD45.1) were used for primary or secondary transplants, and 10 recipient lethally irradiated (CD45.2) mice per Abi-1^WT orAbi-1^KO transplant group were used. (H) Western blot analysis of Abi-1 protein levels and (I) average percentage of G0/G1, S or G2/M human CD34⁺ cells (n = 3) exposed to FANA ABI1 silencing antisense oligonucleotide (KD) or scrambled control (Ctrl) for 48 hours. (J) Heat map showing differently expressed transcripts in LSK-enriched cells isolated from the bone marrow of n = 4 Abi-1^KO and n = 5 Abi-1^WT 14-week-old animals. False discovery rate was < 0.05. (K) Cytokine levels detected in the plasma of 14-weeks old sex-matched Abi-1^WT (n = 12) and Abi-1^KO (n = 12) animals. *P < .05, **P < .01.