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JQ1 induces proliferation of HSC
to downregulation of c-Myb, which is known to be a potent negative modulator of megakaryopoiesis.17 As Brd4 is also highly expressed in MK (Figure 1A), we examined their prevalence in the BM and found numerically increased absolute numbers of CD41+CD61+ MK with enhanced colony forming unit-megakaryocyte (CFU-MK) upon JQ1 treatment (Figure 1H and I). Wright’s-Giemsa staining of BM and splenocytes confirmed the presence of more MK upon JQ1 treatment (Figure 1J). Thus, JQ1 treat-
ment induces quantitative and phenotypic changes within cell populations showing the highest level of Brd4 expres- sion.
JQ1 blocks B-cell maturation and induces T-cell apoptosis
Since BET inhibition by JQ1 reduced the numbers of mature B cells, we next analyzed the different stages of B- cell development in the BM and spleen. The earliest pro-
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Figure 1. JQ1 treatment affects normal hematopoiesis in cell context-dependent manner. (A) Sorted megakaryocytes (MK), hematopoietic stem cells (HSC), granu- locyte-monocyte progenitors (GMP), Gr1+ cells (GR1+) and monocytes (Mo) from bone marrow (BM) of mice were analyzed for baseline RNA expression of Brd4 via qRT-PCR (n=2-3; *P<0.05). (B-J) Animals received daily intraperitoneal (i.p.) injections of 50 mg/kg JQ1 for 21 days after which the different parameters were ana- lyzed. (B) Mononucleated cells in peripheral blood (n=8; *P<0.05). (C) Mononucleated cells in BM (n=8; *P<0.05). (D and E) Spleen (D) and thymus (E) weight (n=7- 8; *P<0.05). (F and G) B-cell (F) and T-cell (G) counts in BM (n=8; *P<0.05). (H) Flowcytometric quantification of MK in BM (n=4; *P<0.05). (I) Colony formation assay from BM to quantify megakaryocytic progenitors (n=5-7; *P<0.05). (J) Representative images showing increased numbers of MK (arrows) upon JQ1 treatment in BM. Scale bars represent 100 mm.
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