TGF-β Signaling Contributes to Myelofibrosis and Clonal Dominance of Myeloproliferative Neoplasms

TGF-β expression is elevated in most cases of myeloproliferative neoplasms (MPNs). However, the contribution of TGF-β to disease pathogenesis is not well understood. Prior studies have shown that TGF-β regulates hematopoietic stem cell (HSC) quiescence. There also is published evidence that increased TGF-β may contribute to myelofibrosis. However, this is controversial, as recent studies have implicated other inflammatory cytokines in the development of myelofibrosis. Here, we test two specific hypotheses. First, we hypothesize that increased TGF-β signaling in mesenchymal stromal cells (MSCs) is required for the development of myelofibrosis. Second, we hypothesize that Jak2 mutated HSCs are resistant to the growth suppressive effect of TGF-β, resulting in a competitive advantage that contributes to their clonal expansion in MPN and clonal hematopoiesis. To test the first hypothesis, we abrogated TGF-β signaling in mesenchymal stem/progenitor cells by deleting Tgfbr2 using a doxycycline-repressible Osterix-Cre transgene (Osx-Cre), which targets all mesenchymal stromal cells in the bone marrow. Osx-Cre was induced at birth (by removal of doxycycline), since we recently reported that the post-natal loss of TGF-β signaling in mesenchymal stromal cells has no discernible effect on basal hematopoiesis or the stem cell niche. We transplanted bone marrow cells from UBC-CreERT2; Jak2V617F mice or c-kit+ cells transduced with MPLW515L retrovirus into irradiated wildtype or Osx-Cre; Tgfbr2f/fmice. For the Jak2V617F model, mice were treated with tamoxifen 6 weeks post transplantation to induce mutant Jak2 expression. Of note, elevated Tgfb1 was present in both MPN models. MPLW515L induced a rapidly fatal MPN with reticulin fibrosis in the bone marrow. A similar hematopoietic phenotype was observed in Osx-Cre; Tgfbr2f/f recipients. However, myelofibrosis, as measured by reticulin staining and Collagen III (Col3a1) mRNA expression, was reduced, but not completely abrogated in Osx-Cre; Tgfbr2f/fmice. Likewise, in the Jak2V617F MPN model, the hematopoietic phenotype was similar in wildtype and Osx-Cre; Tgfbr2f/fmice. Although overt myelofibrosis was not observed in this MPN model, increased RNA and protein expression of Collagen III were detected in the bone marrow. Although still above baseline, Col3a1 expression was significantly reduced in Osx-Cre; Tgfbr2f/frecipient mice. To examine the role of canonical TGF-β signaling in the induction of myelofibrosis, we cultured