RESUMO
Stem and progenitor cells are currently being investigated for their applicability in cell-based therapy for periodontal tissue regeneration. We recently demonstrated that the transplantation of adipose tissue-derived multi-lineage progenitor cells (ADMPCs) enhances periodontal tissue regeneration in beagle dogs. However, the molecular mechanisms by which transplanted ADMPCs induce periodontal tissue regeneration remain to be elucidated. In this study, trophic factors released by ADMPCs were examined for their paracrine effects on human periodontal ligament cell (HPDL) function. ADMPC conditioned medium (ADMPC-CM) up-regulated osteoblastic gene expression, alkaline phosphatase activity and calcified nodule formation in HPDLs, but did not significantly affect their proliferative response. ADMPCs secreted a number of growth factors, including insulin-like growth factor binding protein 6 (IGFBP6), hepatocyte growth factor and vascular endothelial growth factor. Among these, IGFBP6 was most highly expressed. Interestingly, the positive effects of ADMPC-CM on HPDL differentiation were significantly suppressed by transfecting ADMPCs with IGFBP6 siRNA. Our results suggest that ADMPCs transplanted into a defect in periodontal tissue release trophic factors that can stimulate the differentiation of HPDLs to mineralized tissue-forming cells, such as osteoblasts and cementoblasts. IGFBP6 may play crucial roles in ADMPC-induced periodontal regeneration.
Assuntos
Tecido Adiposo/citologia , Meios de Cultivo Condicionados/farmacologia , Cemento Dentário/efeitos dos fármacos , Osteoblastos/efeitos dos fármacos , Ligamento Periodontal/efeitos dos fármacos , Células-Tronco/citologia , Tecido Adiposo/metabolismo , Fosfatase Alcalina/genética , Fosfatase Alcalina/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem da Célula/efeitos dos fármacos , Células Cultivadas , Cemento Dentário/citologia , Cemento Dentário/metabolismo , Regulação da Expressão Gênica , Fator de Crescimento de Hepatócito/genética , Fator de Crescimento de Hepatócito/metabolismo , Humanos , Proteína 6 de Ligação a Fator de Crescimento Semelhante à Insulina/antagonistas & inibidores , Proteína 6 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Proteína 6 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Osteoblastos/citologia , Osteoblastos/metabolismo , Osteogênese/efeitos dos fármacos , Ligamento Periodontal/citologia , Ligamento Periodontal/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Células-Tronco/metabolismo , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Osteopontin is a protein found in the bone-related matrix and plays multiple regulatory roles in mineralizing and non-mineralizing tissue. In osteogenic cell-lines, the expression of osteopontin increases with the progression of differentiation, but both the expression and function of osteopontin vary with the cell type and its activation state. In this study, we examined the expression of osteopontin by clones established from mouse periodontal ligament, in response to inorganic phosphate and fibroblast growth factor (FGF)-2, which can induce periodontal tissue regeneration. The involvement of inorganic phosphate in the expression of osteopontin during the course of cell differentiation of a clone MPDL22 was confirmed by addition of foscarnet, an inorganic phosphate transport inhibitor. Although FGF-2 decreased the mRNA expression of almost every bone-related protein in MPDL22, FGF-2 upregulated the expression of osteopontin in MPDL22 at both mRNA and protein levels. Interestingly, FGF-2 enhanced the concentration of osteopontin in the culture supernatant of MPDL22, whereas inorganic phosphate did not. The FGF-2-induced osteopontin in the culture supernatant seems to be involved in cell survival activity. An immunohistochemical study showed that the FGF-2-induced osteopontin was mainly present in perinuclear matrices while the inorganic phosphate-induced osteopontin was associated with extracellular matrices in addition to perinuclear matrices. The present results indicated that FGF-2 induces unique expression of osteopontin, which may play a role different from the other bone-related proteins during the process of periodontal tissue regeneration by FGF-2.
Assuntos
Fator 2 de Crescimento de Fibroblastos/metabolismo , Regulação da Expressão Gênica , Osteopontina/metabolismo , Ligamento Periodontal/citologia , Animais , Apoptose , Diferenciação Celular/fisiologia , Membrana Celular/metabolismo , Sobrevivência Celular , Células Cultivadas , Matriz Extracelular/química , Matriz Extracelular/metabolismo , Fator 2 de Crescimento de Fibroblastos/genética , Camundongos , Osteopontina/genética , Ligamento Periodontal/metabolismo , Fosfatos/química , Fosfatos/metabolismoRESUMO
INTRODUCTION: Homeostasis and tissue repair of dentin-pulp complex are attributed to dental pulp tissue and several growth factors. Dental pulp cells play a pivotal role in homeostasis of dentin-pulp complex and tissue responses after tooth injury. Among these cytokines, fibroblast growth factor (FGF)-2 has multifunctional biologic activity and is known as a signaling molecule that induces tissue regeneration. In this study, we examined the effects of FGF-2 on growth, migration, and differentiation of human dental pulp cells (HDPC). METHODS: HDPC were isolated from healthy dental pulp. Cellular response was investigated by [(3)H]-thymidine incorporation into DNA. Cytodifferentiation was examined by alkaline phosphatase (ALPase) assay and cytochemical staining of calcium by using alizarin red. Migratory activity was determined by counting the cells migrating into cleared area that had introduced with silicon block. RESULTS: FGF-2 activated HDPC growth and migration but suppressed ALPase activity and calcified nodule formation. Interestingly, HDPC, which had been pretreated with FGF-2, showed increased ALPase activity and calcified nodule formation when subsequently cultured without FGF-2. These results suggest that FGF-2 potentiates cell growth and accumulation of HDPC that notably did not disturb cytodifferentiation of the cells later. Thus, FGF-2 is a favorable candidate for pulp capping agent. CONCLUSIONS: These results provide new evidence for the possible involvement of FGF-2 not only in homeostasis but also in regeneration of dentin-pulp complex.