Fibroblast Growth Factor-2 facilitates the growth and chemo-resistance of leukemia cells in the bone marrow by modulating osteoblast functions.

Keiki Sugimoto,Hiroki Mizuno,Hidefumi Kato,Ritsuro Suzuki,Akiyoshi Takami,Tomoki Naoe,Satoshi Nishiwaki,Yasuhiko Miyata,Kyosuke Takeshita,Fumihiko Hayakawa,Shigeki Saito,Ryuzo Ueda,Takayuki Nakayama

doi:10.1038/srep30779

Abstract

Stromal cells and osteoblasts play major roles in forming and modulating the bone marrow (BM) hematopoietic microenvironment. We have reported that FGF2 compromises stromal cell support of normal hematopoiesis. Here, we examined the effects of FGF2 on the leukemia microenvironment. In vitro, FGF2 significantly decreased the number of stromal-dependent and stromal-independent G0-leukemia cells in the stromal layers. Accordingly, CML cells placed on FGF2-treated stromal layers were more sensitive to imatinib. Conversely, FGF2 increased the proliferation of osteoblasts via FGFR1 IIIc, but its effects on osteoblast support of leukemia cell growth were limited. We next treated a human leukemia mouse model with Ara-C with/without systemic FGF2 administration. BM sections from FGF2-treated mice had thickened bone trabeculae and increased numbers of leukemia cells compared to controls. Leukemia cell density was increased, especially in the endosteal region in FGF2/Ara-C -treated mice compared to mice treated with Ara-C only. Interestingly, FGF2 did not promote leukemia cell survival in Ara-C treated spleen. Microarray analysis showed that FGF2 did not alter expression of many genes linked to hematopoiesis in osteoblasts, but modulated regulatory networks involved in angiogenesis and osteoblastic differentiation. These observations suggest that FGF2 promotes leukemia cell growth in the BM by modulating osteoblast functions.

Highlights

Resuspended in PBS containing 1 μg/ml Hoechst (Sigma) for 45 minutes at 37 °C, 5 μl of 100 μg/ml pyronin Y (Sigma) was directly added to the cells
None of FGF receptors (FGFRs)/Fc chimeras added alone to 7F2 cell cultures affected the growth of 7F2 cells or VEGF-A secretion from 7F2. These results demonstrated that FGFR1 IIIc is the receptor that mediates Fibroblast growth factor 2 (FGF2)-induced proliferation and VEGF-A secretion in osteoblasts
Recent evidence suggests that there is little direct association between osteoblasts and HSCs15,16 in accordance with in vitro results that FGF2-treated 7F2 cells showed limited supportive functions toward leukemia cells (Figs 1b and 2b), even though leukemic stem cells localize within the osteoblast-rich area of the bone marrow (BM) where acute myeloid leukemia cells are protected from chemotherapy-induced apoptosis[17]

Summary

Introduction

Resuspended in PBS containing 1 μg/ml Hoechst (Sigma) for 45 minutes at 37 °C, 5 μl of 100 μg/ml pyronin Y (Sigma) was directly added to the cells. (c) Effects of FGF2 on the chemosensitivity of leukemia cell onto stromal layers were measured by using cells transduced with a fluorescent substance. Members of the FGF superfamily function by binding to heparan sulfate proteoglycans and FGF receptors (FGFRs)[8]. Little is known about the effects of FGF2 on the BM microenvironment, even though both mesenchymal stromal cells and osteoblasts express FGFRs10,11. To address this issue further, we evaluated mesenchymal stromal cell and osteoblast mediated support of leukemia cell growth after FGF2 exposure. We describe a novel role of FGF2 as a modulator of osteoblast and mesenchymal stromal cell function, and provide evidence for involvement of FGF2 in leukemia pathogenesis

Methods

Results

Conclusion