RESUMO
Cytokine-induced expansion of hematopoietic stem and progenitor cells (HSPCs) is not fully understood. In the present study, we show that whereas steady-state hematopoiesis is normal in basic fibroblast growth factor (FGF-2)-knockout mice, parathyroid hormone stimulation and myeloablative treatments failed to induce normal HSPC proliferation and recovery. In vivo FGF-2 treatment expanded stromal cells, including perivascular Nestin(+) supportive stromal cells, which may facilitate HSPC expansion by increasing SCF and reducing CXCL12 via mir-31 up-regulation. FGF-2 predominantly expanded a heterogeneous population of undifferentiated HSPCs, preserving and increasing durable short- and long-term repopulation potential. Mechanistically, these effects were mediated by c-Kit receptor activation, STAT5 phosphorylation, and reduction of reactive oxygen species levels. Mice harboring defective c-Kit signaling exhibited abrogated HSPC expansion in response to FGF-2 treatment, which was accompanied by elevated reactive oxygen species levels. The results of the present study reveal a novel mechanism underlying FGF-2-mediated in vivo expansion of both HSPCs and their supportive stromal cells, which may be used to improve stem cell engraftment after clinical transplantation.
Assuntos
Proliferação de Células , Quimiocina CXCL12/metabolismo , Fator 2 de Crescimento de Fibroblastos/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Proteínas Proto-Oncogênicas c-kit/metabolismo , Células Estromais/metabolismo , Animais , Sequência de Bases , Transplante de Medula Óssea , Ciclo Celular/efeitos dos fármacos , Células Cultivadas , Quimiocina CXCL12/genética , Regulação para Baixo/efeitos dos fármacos , Fator 2 de Crescimento de Fibroblastos/genética , Fator 2 de Crescimento de Fibroblastos/farmacologia , Citometria de Fluxo , Expressão Gênica/efeitos dos fármacos , Células-Tronco Hematopoéticas/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Modelos Biológicos , Hormônio Paratireóideo/farmacologia , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-kit/genética , Espécies Reativas de Oxigênio/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Transcrição STAT5/metabolismo , Células Estromais/efeitos dos fármacosRESUMO
Fibroblast growth factor 2 (FGF2) positively modulates osteoblast differentiation and bone formation. However, the mechanism(s) is not fully understood. Because the Wnt canonical pathway is important for bone homeostasis, this study focuses on modulation of Wnt/ß-catenin signaling using Fgf2(-/-) mice (FGF2 all isoforms ablated), both in the absence of endogenous FGF2 and in the presence of exogenous FGF2. This study demonstrates a role of endogenous FGF2 in bone formation through Wnt signaling. Specifically, mRNA expression for the canonical Wnt genes Wnt10b, Lrp6, and ß-catenin was decreased significantly in Fgf2(-/-) bone marrow stromal cells during osteoblast differentiation. In addition, a marked reduction of Wnt10b and ß-catenin protein expression was observed in Fgf2(-/-) mice. Furthermore, Fgf2(-/-) osteoblasts displayed marked reduction of inactive phosphorylated glycogen synthase kinase-3ß, a negative regulator of Wnt/ß-catenin pathway as well as a significant decrease of Dkk2 mRNA, which plays a role in terminal osteoblast differentiation. Addition of exogenous FGF2 promoted ß-catenin nuclear accumulation and further partially rescued decreased mineralization in Fgf2(-/-) bone marrow stromal cell cultures. Collectively, our findings suggest that FGF2 stimulation of osteoblast differentiation and bone formation is mediated in part by modulating the Wnt pathway.