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Activation of the FGF signaling pathway and subsequent induction of mesenchymal stem cell differentiation by inorganic polyphosphate.

Kawazoe Y, Katoh S, Onodera Y, Kohgo T, Shindoh M, Shiba T - Int. J. Biol. Sci. (2008)

Bottom Line: The effect of poly(P) on the osteogenic differentiation of HDPCs and human MSCs (hMSCs) were also investigated.Furthermore, induced expression of MMP1, OPN and OC genes in both cells was confirmed by real-time PCR.The results suggest that the activation of the FGF signaling pathway by poly(P) induces both proliferation and mineralization of stem cells.

View Article: PubMed Central - PubMed

Affiliation: Regenetiss Inc., 1-9-4, Asahigaoka, Hino, Tokyo 191-0065, Japan.

ABSTRACT
Inorganic polyphosphate [poly(P)] is a biopolymer existing in almost all cells and tissues, although its biological functions in higher eukaryotes have not been completely elucidated. We previously demonstrated that poly(P) enhances the function of fibroblast growth factors (FGFs) by stabilizing them and strengthening the affinity between FGFs and their cell surface receptors. Since FGFs play crucial roles in bone regeneration, we further investigated the effect of poly(P) on the cell differentiation of human stem cells via FGF signaling systems. Human dental pulp cells (HDPCs) isolated from human dental pulp show the characteristics of multipotent mesenchymal stem cells (MSCs). HDPCs secreted FGFs and the proliferation of HDPCs was shown to be enhanced by treatment with poly(P). Cell surface receptor-bound FGF-2 was stably maintained for more than 40 hours in the presence of poly(P). The phosphorylation of ERK1/2 was also enhanced by poly(P). The effect of poly(P) on the osteogenic differentiation of HDPCs and human MSCs (hMSCs) were also investigated. After 5 days of treatment with poly(P), type-I collagen expression of both cell types was enhanced. The C-terminal peptide of type-I collagen was also released at higher levels in poly(P)-treated HDPCs. Microarray analysis showed that expression of matrix metalloproteinase-1 (MMP1), osteopontin (OPN), osteocalcin (OC) and osteoprotegerin was induced in both cell types by poly(P). Furthermore, induced expression of MMP1, OPN and OC genes in both cells was confirmed by real-time PCR. Calcification of both cell types was clearly observed by alizarin red staining following treatment with poly(P). The results suggest that the activation of the FGF signaling pathway by poly(P) induces both proliferation and mineralization of stem cells.

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Multipotentiality of HDPCs isolated from human dental pulp. Isolated HDPCs were differentiated into chondrocytes (A, B, C) and adipocytes (D, E) under appropriate conditions described in Materials and Methods. (A) Differentiated chondrocytes were stained with alcian blue, and (B) the expression of aggrecan (a chondrocyte marker) was visualized by immunostaining. (C) Cell morphology of the differentiated chondrocyte was visualized by HE staining. Hypertrophic cells were observed at the peripheral area of the cell pellet. (D) Cell morphology of the differentiated adipocyte and (E) its high magnified image. Lipid accumulation was observed in the differentiated cells. Bars: 50 μm scale.
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Figure 1: Multipotentiality of HDPCs isolated from human dental pulp. Isolated HDPCs were differentiated into chondrocytes (A, B, C) and adipocytes (D, E) under appropriate conditions described in Materials and Methods. (A) Differentiated chondrocytes were stained with alcian blue, and (B) the expression of aggrecan (a chondrocyte marker) was visualized by immunostaining. (C) Cell morphology of the differentiated chondrocyte was visualized by HE staining. Hypertrophic cells were observed at the peripheral area of the cell pellet. (D) Cell morphology of the differentiated adipocyte and (E) its high magnified image. Lipid accumulation was observed in the differentiated cells. Bars: 50 μm scale.

Mentions: In order to confirm that HDPCs isolated from human dental pulp contain multipotent MSCs, the cells were treated with both the chondrogenic cocktail and the adipogenic cocktail (see Materials and Methods) to induce cell differentiation into chondrocytes and adipocytes, respectively. After 24 days of treatment with the chondrogenic cocktail, HDPCs were stained with both alcian blue (Fig. 1A) and anti-aggrecan antibody (Fig. 1B), indicating that the cells had differentiated into chondrocytes. Hypertrophic cells, which correspond to differentiated chondrocytes, were also observed at the peripheral area of the cell pellet (Fig. 1C). In addition, some populations of HDPCs produced lipid particles after 30 days of treatment with the adipogenic cocktail described above (Fig. 1D and 1E). These results suggest that HDPCs have multipotent ability to differentiate into both chondrocytes and adipocytes. These findings are consistent with the results of previous studies showing the existence of multipotent mesenchymal stem cells (MSCs) in human dental pulp that have the potential to differentiate into adipocytes 21, 22. Also, we have found that human dental pulp cells have the ability to differentiate into chondrocytes. Based on these results, we decided to use HDPCs in this study as our MSC-containing population of primary culture cells.


Activation of the FGF signaling pathway and subsequent induction of mesenchymal stem cell differentiation by inorganic polyphosphate.

Kawazoe Y, Katoh S, Onodera Y, Kohgo T, Shindoh M, Shiba T - Int. J. Biol. Sci. (2008)

Multipotentiality of HDPCs isolated from human dental pulp. Isolated HDPCs were differentiated into chondrocytes (A, B, C) and adipocytes (D, E) under appropriate conditions described in Materials and Methods. (A) Differentiated chondrocytes were stained with alcian blue, and (B) the expression of aggrecan (a chondrocyte marker) was visualized by immunostaining. (C) Cell morphology of the differentiated chondrocyte was visualized by HE staining. Hypertrophic cells were observed at the peripheral area of the cell pellet. (D) Cell morphology of the differentiated adipocyte and (E) its high magnified image. Lipid accumulation was observed in the differentiated cells. Bars: 50 μm scale.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2238184&req=5

Figure 1: Multipotentiality of HDPCs isolated from human dental pulp. Isolated HDPCs were differentiated into chondrocytes (A, B, C) and adipocytes (D, E) under appropriate conditions described in Materials and Methods. (A) Differentiated chondrocytes were stained with alcian blue, and (B) the expression of aggrecan (a chondrocyte marker) was visualized by immunostaining. (C) Cell morphology of the differentiated chondrocyte was visualized by HE staining. Hypertrophic cells were observed at the peripheral area of the cell pellet. (D) Cell morphology of the differentiated adipocyte and (E) its high magnified image. Lipid accumulation was observed in the differentiated cells. Bars: 50 μm scale.
Mentions: In order to confirm that HDPCs isolated from human dental pulp contain multipotent MSCs, the cells were treated with both the chondrogenic cocktail and the adipogenic cocktail (see Materials and Methods) to induce cell differentiation into chondrocytes and adipocytes, respectively. After 24 days of treatment with the chondrogenic cocktail, HDPCs were stained with both alcian blue (Fig. 1A) and anti-aggrecan antibody (Fig. 1B), indicating that the cells had differentiated into chondrocytes. Hypertrophic cells, which correspond to differentiated chondrocytes, were also observed at the peripheral area of the cell pellet (Fig. 1C). In addition, some populations of HDPCs produced lipid particles after 30 days of treatment with the adipogenic cocktail described above (Fig. 1D and 1E). These results suggest that HDPCs have multipotent ability to differentiate into both chondrocytes and adipocytes. These findings are consistent with the results of previous studies showing the existence of multipotent mesenchymal stem cells (MSCs) in human dental pulp that have the potential to differentiate into adipocytes 21, 22. Also, we have found that human dental pulp cells have the ability to differentiate into chondrocytes. Based on these results, we decided to use HDPCs in this study as our MSC-containing population of primary culture cells.

Bottom Line: The effect of poly(P) on the osteogenic differentiation of HDPCs and human MSCs (hMSCs) were also investigated.Furthermore, induced expression of MMP1, OPN and OC genes in both cells was confirmed by real-time PCR.The results suggest that the activation of the FGF signaling pathway by poly(P) induces both proliferation and mineralization of stem cells.

View Article: PubMed Central - PubMed

Affiliation: Regenetiss Inc., 1-9-4, Asahigaoka, Hino, Tokyo 191-0065, Japan.

ABSTRACT
Inorganic polyphosphate [poly(P)] is a biopolymer existing in almost all cells and tissues, although its biological functions in higher eukaryotes have not been completely elucidated. We previously demonstrated that poly(P) enhances the function of fibroblast growth factors (FGFs) by stabilizing them and strengthening the affinity between FGFs and their cell surface receptors. Since FGFs play crucial roles in bone regeneration, we further investigated the effect of poly(P) on the cell differentiation of human stem cells via FGF signaling systems. Human dental pulp cells (HDPCs) isolated from human dental pulp show the characteristics of multipotent mesenchymal stem cells (MSCs). HDPCs secreted FGFs and the proliferation of HDPCs was shown to be enhanced by treatment with poly(P). Cell surface receptor-bound FGF-2 was stably maintained for more than 40 hours in the presence of poly(P). The phosphorylation of ERK1/2 was also enhanced by poly(P). The effect of poly(P) on the osteogenic differentiation of HDPCs and human MSCs (hMSCs) were also investigated. After 5 days of treatment with poly(P), type-I collagen expression of both cell types was enhanced. The C-terminal peptide of type-I collagen was also released at higher levels in poly(P)-treated HDPCs. Microarray analysis showed that expression of matrix metalloproteinase-1 (MMP1), osteopontin (OPN), osteocalcin (OC) and osteoprotegerin was induced in both cell types by poly(P). Furthermore, induced expression of MMP1, OPN and OC genes in both cells was confirmed by real-time PCR. Calcification of both cell types was clearly observed by alizarin red staining following treatment with poly(P). The results suggest that the activation of the FGF signaling pathway by poly(P) induces both proliferation and mineralization of stem cells.

Show MeSH
Related in: MedlinePlus