Session: 311. Disorders of Platelet Number or Function: Poster I
Hematology Disease Topics & Pathways:
HSCs, platelets, Cell Lineage
PF4Cre/Tgfb1flox/flox mice had >50% reduction in TGFβ1 levels in BM cells and exudates (TGFβ1 levels in BM exudates were 1.4 ± 0.033 ng in WT and 0.68 ± 0.065 ng in PF4CreTgfb1flox/flox mice, p<0.01; and in BM cells 50 ± 9 ng/ml in WT and 22 ± 4.2 ng/ml in PF4CreTgfb1flox/flox; p<0.001). MK numbers were ~25% higher in PF4Cre/Tgfb1flox/flox mice (n=6) compared to combined littermate controls (n=3) and WT (n=3) (MK was 0.30 ± 0.02% in PF4Cre/Tgfb1flox/flox and 0.23 ± 0.16% in combined controls; p<0.001 (n=6), whereas blood platelet counts were only marginally higher in PF4Cre/Tgfb1flox/flox (1114 ± 300) vs. controls (806 ± 116; p<0.05). There was a ~2-fold higher plasma TPO levels in PF4CreTgfb1flox/flox mice vs. WT (p=0.04, n=4). Increased DMS and nucleus areas in MK have been shown to correlate with proplatelets formation and platelet production. However, DMS and nuclear areas remained unchanged between genotypes [(DMS area was 197 ± 46 in PF4CreTgfb1flox/flox and 228 ± 50 um2 in combined WT and littermate controls (p=0.3), and nucleus size was 154 ± 23 in PF4CreTgfb1flox/flox and 160 ± 33 um2 in controls (p=0.7)], indicating that the role of TGFβ1 is limited to megakaryopoiesis. To test whether the in vivo phenotype was recapitulated, we cultured whole BM isolated from WT and PF4Cre/Tgfb1flox/flox mice, which showed a ~2.5-fold increase in MK numbers vs. WT when cultured for 5 days in TPO-supplemented medium. The addition of recombinant TGFβ1 in culture medium inhibited MK numbers, and a neutralizing antibody against TGFβ1 resulted in increased MK numbers.
We conclude that MK-derived TGFβ1 negatively regulates megakaryopoiesis in mice. Further investigation is needed to determine the mechanism by which TGFβ1 regulates TPO-induced megakaryopoiesis. Our study may be important in megakaryocyte generation in vitro and may have important implications in vivo under normal and stress-inducing conditions where variable megakaryopoiesis is observed, such as essential thrombocythemia and primary myelofibrosis.
Disclosures: No relevant conflicts of interest to declare.
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