Podoplanin-positive periarteriolar stromal cells promote megakaryocyte growth and proplatelet formation in mice by CLEC-2

Abstract

Key Points• BM FRC-like cells regulate megakaryocytic clonal expansion via CLEC-2/PDPN interactions.• CLEC-2/PDPN binding stimulates BM FRC-like cells to secrete the proplatelet formation-promoting factor, CCL5.Megakaryopoiesis is the hierarchical differentiation of hematopoietic stem cells into megakaryocytes. Differentiating megakaryocytes undergo maturation characterized by endomitosis and produce numerous platelets through proplatelet formation. C-type lectin-like receptor 2 (CLEC-2) is a podoplanin (PDPN) receptor mainly expressed on platelets and megakaryocytes. Deletion of platelet/megakaryocyte CLEC-2 causes thrombocytopenia in mice; however, its contribution to megakaryopoiesis remains unknown.Here, we show that megakaryopoiesis is promoted through the CLEC-2/PDPN interaction in the vicinity of arterioles in the bone marrow (BM). We have also identified PDPNexpressing BM arteriolar stromal cells, tentatively termed as BM fibroblastic reticular cell (FRC)-like cells. Platelet/megakaryocyte-specific CLEC-2 conditional knockout (cKO) mice showed a decrease in the number of immature megakaryocytes. CLEC-2 wild-type megakaryocyte expansion was augmented in vitro by the addition of recombinant PDPN, but not cKO megakaryocytes. Moreover, megakaryocyte colonies were colocalized with periarteriolar BM FRC-like cells in the BM. Coculture of megakaryocytes with BM FRC-like cells augmented megakaryocyte expansion, which was dependent upon the CLEC-2/PDPN interaction. Furthermore, we found that the CLEC-2/PDPN interaction induces BM FRClike cells to secrete chemokine (C-C motif) ligand 5 (CCL5) to facilitate proplatelet formation. These observations indicate that a reciprocal interaction between CLEC-2 on megakaryocytes and PDPN on BM FRC-like cells contributes to the periarteriolar megakaryopoietic microenvironment in mouse

Hematopoietic stem cell loss and hematopoietic failure in severe aplastic anemia is driven by macrophages and aberrant podoplanin expression

“…However CLEC-2/PDPN ligation also elicits bidirectional signaling, eliciting RANTES production from PDPN-expressing cells. 33,34 We noted increased RANTES in SAA, relative to radiation controls. Thus, our observation that PDPN blockade did not impact SAA-induced RANTES was somewhat surprising.…”

“…PDPN-CLEC-2 interaction was reported to induce RANTES, 33 which can support platelet production, thus our finding that PDPN blockade improved thrombocytopenia was somewhat surprising. Consistent with a role for PDPN in RANTES production we observed increased RANTES during SAA ( Online Supplementary Figure S9C ), where we also observe increased PDPN + Mϕs.…”

“…Because SAA was associated with severe thrombocytopenia we examined expression of podoplanin (PDPN), recently identified in the BM and shown to increase platelet production. 33 We noted a striking loss in PDPN + stromal cells in SAA ( Online Supplementary Figure S7C and D ). At the same time, however, we observed induction of PDPN on hematopoietic cells that appeared to be entirely restricted to Mϕs and a majority were CD11b lo/− Mϕs (Figure 6A and B, and Online Supplementary Figure S8A ).…”

Platelet-activating factor podoplanin: from discovery to drug development

“…Not only through the platelet interaction, but CLEC-2-expressed dendritic cells cause stretching of stroma by affecting podoplanin-expressed FRC, which leads to lymph node expansion in immune response [ 45 ]. And also the interaction between CLEC-2 on megakaryocytes and podoplanin on FRC-like stroma cells is suggested to promote megakaryocyte expansion and proplatelet formation in bone marrow [ 46 ]. Recent accumulated reports suggest physiological functions of podoplanin in the development of lymphatic vessels, lymph nodes, and immune responses.…”

Hematopoietic Stem Cell Regulation by Type I and II Interferons in the Pathogenesis of Acquired Aplastic Anemia

“…Sterile, IFNα/β-driven inflammation relocates HSPCs away from quiescence-enforcing arteriolar niches (118), though it is unclear whether this is cause or consequence of changes in HSC cycling. IFNα/β can also stimulate endothelial chemokine expression, including that of CCL5 or RANTES (177), which can impact platelet production by megakaryocytes (178). The role of megakaryocytes in HSC regulation is dynamic, as homeostatic expression of CXCL4 and TGFβ1 promotes quiescence, while concomitant increases in FGF-1 and decreases in TGFβ1 facilitate regeneration (174, 175).…”