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CXCL14 and MCP1 are potent trophic factors associated with cell migration and angiogenesis leading to higher regenerative potential of dental pulp side population cells.

Hayashi Y, Murakami M, Kawamura R, Ishizaka R, Fukuta O, Nakashima M - Stem Cell Res Ther (2015)

Bottom Line: Furthermore, the trophic factors responsible for the regenerative potential were identified as the upregulated genes present in pulp CD31(-) SP cells when compared with the genes in both bone marrow and adipose CD31(-) SP cells by using microarray analysis, real-time RT-PCR, and Western blot analysis.Pulp CM also demonstrated significantly increased cell migration, anti-apoptosis, and angiogenesis in C2C12 cells.The stimulatory effects on both migration and angiogenesis of CXCL14 and MCP1 were demonstrated in vitro.

View Article: PubMed Central - PubMed

Affiliation: Department of Dental Regenerative Medicine, Center of Advanced Medicine for Dental and Oral Diseases, National Center for Geriatrics and Gerontology, Research Institute, Morioka 7-430, Obu, Aichi, 474-8511, Japan. hys8y9@gmail.com.

ABSTRACT

Introduction: The release of trophic factors from mesenchymal stem cells (MSCs) is critical for tissue regeneration. A systematic investigation of the regenerative potential of trophic factors from different MSCs, however, has not been performed. Thus, in the present study, the regenerative potential of conditioned medium (CM) from dental pulp, bone marrow, and adipose tissue-derived CD31(-) side population (SP) cells from an individual source was compared in an ectopic tooth transplantation model.

Methods: The tooth root transplantation in an ectopic site model was used for investigation of the regenerative potential and trophic effects in vivo. Either pulp CD31(-) SP cell populations (1×10(6) cells) at the third to fourth passage or 5 μg/ml of CM from dental pulp, bone marrow, and adipose stem cells from four different individuals were injected into the root with collagen TE. Each root was transplanted subcutaneously in 5-week-old severe combined immunodeficiency mice. Each root with surrounding tissue was harvested for histology on days 7, 21, and 28 and for Western blot analysis and real-time reverse transcription-polymerase chain reaction (RT-PCR) analysis on day 28. Furthermore, the trophic factors responsible for the regenerative potential were identified as the upregulated genes present in pulp CD31(-) SP cells when compared with the genes in both bone marrow and adipose CD31(-) SP cells by using microarray analysis, real-time RT-PCR, and Western blot analysis.

Results: Transplantation of pulp CM yielded increased volume of pulp regeneration, more bromodeoxyuridine (BrdU)-positive migrated cells, and fewer caspase 3-positive cells in the regenerated pulp compared with the others. Pulp CM also demonstrated significantly increased cell migration, anti-apoptosis, and angiogenesis in C2C12 cells. Higher expression of CXCL14 and MCP1 in pulp SP cells suggested candidate trophic factors. The stimulatory effects on both migration and angiogenesis of CXCL14 and MCP1 were demonstrated in vitro. In the regenerated tissue, BrdU-positive migrated cells expressed CXCR4 and CCR2, receptors for CXCL14 and MCP1, respectively.

Conclusions: The higher regenerative potential of pulp SP cells may be due to potent trophic factors, including CXCL14 and MCP1, which promote migration and angiogenesis.

No MeSH data available.


Related in: MedlinePlus

Localization of chemokine (C-X-C motif) ligand 14 (CXCL14) and its receptor, CXCR4, related to the transplanted cells and the endogenous migrated cells in the regenerated pulp on day 7. a, b Immunostaining of CXCL14 (red) merged with glutamic-oxaloacetic transaminase 2 (GOT2) or bromodeoxyuridine (BrdU) (green) and Hoechst 33342 (blue) after transplantation of pulp CD31− side population (SP) cells. c-e In situ hybridization and immunohistochemical analysis of expression of C-X-C chemokine receptor type 4 (CXCR4) (red) merged with BrdU (green) after transplantation of conditioned media (CM) from pulp (c), bone marrow (d), and adipose CD31− SP cells (e). f Ratio of CXCR4-positive cell numbers to BrdU-positive cell numbers in regenerated pulp tissues. Data are expressed as mean ± standard deviation of four determinations. *P < 0.05
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Fig6: Localization of chemokine (C-X-C motif) ligand 14 (CXCL14) and its receptor, CXCR4, related to the transplanted cells and the endogenous migrated cells in the regenerated pulp on day 7. a, b Immunostaining of CXCL14 (red) merged with glutamic-oxaloacetic transaminase 2 (GOT2) or bromodeoxyuridine (BrdU) (green) and Hoechst 33342 (blue) after transplantation of pulp CD31− side population (SP) cells. c-e In situ hybridization and immunohistochemical analysis of expression of C-X-C chemokine receptor type 4 (CXCR4) (red) merged with BrdU (green) after transplantation of conditioned media (CM) from pulp (c), bone marrow (d), and adipose CD31− SP cells (e). f Ratio of CXCR4-positive cell numbers to BrdU-positive cell numbers in regenerated pulp tissues. Data are expressed as mean ± standard deviation of four determinations. *P < 0.05

Mentions: Recent studies have demonstrated that CXCL14 and MCP1 promoted cell migration and angiogenesis/vasculogenesis, respectively [34, 35]. Thus, localization of CXCL14 and MCP1 related to the migrated cells and to the newly formed vessels in the regenerated pulp tissue 7 days after transplantation of pulp CD31− SP cells was examined. CXCL14- or MCP1-positive reactivity colocalized with GOT-positive transplanted cells (Figs. 6a and 7a) but did not colocalize with BrdU-positive migrated cells in the regenerated tissue after transplantation of pulp CD31− SP cells (Figs. 6b and 7b), indicating that CXCL14 and MCP1 are secreted by the transplanted cells and not by the endogenous migrated cells. Confocal laser microscopic analysis demonstrated that MCP1-positive cells were not co-localized to vessels but were in proximity to two different sized vessels of approximately 40 μm and 70-130 μm in diameter (Fig. 7g-k), suggesting the trophic effect of MCP1 on angiogenesis, especially for arterioles and venules (small veins).Fig. 6


CXCL14 and MCP1 are potent trophic factors associated with cell migration and angiogenesis leading to higher regenerative potential of dental pulp side population cells.

Hayashi Y, Murakami M, Kawamura R, Ishizaka R, Fukuta O, Nakashima M - Stem Cell Res Ther (2015)

Localization of chemokine (C-X-C motif) ligand 14 (CXCL14) and its receptor, CXCR4, related to the transplanted cells and the endogenous migrated cells in the regenerated pulp on day 7. a, b Immunostaining of CXCL14 (red) merged with glutamic-oxaloacetic transaminase 2 (GOT2) or bromodeoxyuridine (BrdU) (green) and Hoechst 33342 (blue) after transplantation of pulp CD31− side population (SP) cells. c-e In situ hybridization and immunohistochemical analysis of expression of C-X-C chemokine receptor type 4 (CXCR4) (red) merged with BrdU (green) after transplantation of conditioned media (CM) from pulp (c), bone marrow (d), and adipose CD31− SP cells (e). f Ratio of CXCR4-positive cell numbers to BrdU-positive cell numbers in regenerated pulp tissues. Data are expressed as mean ± standard deviation of four determinations. *P < 0.05
© Copyright Policy - open-access
Related In: Results  -  Collection

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Fig6: Localization of chemokine (C-X-C motif) ligand 14 (CXCL14) and its receptor, CXCR4, related to the transplanted cells and the endogenous migrated cells in the regenerated pulp on day 7. a, b Immunostaining of CXCL14 (red) merged with glutamic-oxaloacetic transaminase 2 (GOT2) or bromodeoxyuridine (BrdU) (green) and Hoechst 33342 (blue) after transplantation of pulp CD31− side population (SP) cells. c-e In situ hybridization and immunohistochemical analysis of expression of C-X-C chemokine receptor type 4 (CXCR4) (red) merged with BrdU (green) after transplantation of conditioned media (CM) from pulp (c), bone marrow (d), and adipose CD31− SP cells (e). f Ratio of CXCR4-positive cell numbers to BrdU-positive cell numbers in regenerated pulp tissues. Data are expressed as mean ± standard deviation of four determinations. *P < 0.05
Mentions: Recent studies have demonstrated that CXCL14 and MCP1 promoted cell migration and angiogenesis/vasculogenesis, respectively [34, 35]. Thus, localization of CXCL14 and MCP1 related to the migrated cells and to the newly formed vessels in the regenerated pulp tissue 7 days after transplantation of pulp CD31− SP cells was examined. CXCL14- or MCP1-positive reactivity colocalized with GOT-positive transplanted cells (Figs. 6a and 7a) but did not colocalize with BrdU-positive migrated cells in the regenerated tissue after transplantation of pulp CD31− SP cells (Figs. 6b and 7b), indicating that CXCL14 and MCP1 are secreted by the transplanted cells and not by the endogenous migrated cells. Confocal laser microscopic analysis demonstrated that MCP1-positive cells were not co-localized to vessels but were in proximity to two different sized vessels of approximately 40 μm and 70-130 μm in diameter (Fig. 7g-k), suggesting the trophic effect of MCP1 on angiogenesis, especially for arterioles and venules (small veins).Fig. 6

Bottom Line: Furthermore, the trophic factors responsible for the regenerative potential were identified as the upregulated genes present in pulp CD31(-) SP cells when compared with the genes in both bone marrow and adipose CD31(-) SP cells by using microarray analysis, real-time RT-PCR, and Western blot analysis.Pulp CM also demonstrated significantly increased cell migration, anti-apoptosis, and angiogenesis in C2C12 cells.The stimulatory effects on both migration and angiogenesis of CXCL14 and MCP1 were demonstrated in vitro.

View Article: PubMed Central - PubMed

Affiliation: Department of Dental Regenerative Medicine, Center of Advanced Medicine for Dental and Oral Diseases, National Center for Geriatrics and Gerontology, Research Institute, Morioka 7-430, Obu, Aichi, 474-8511, Japan. hys8y9@gmail.com.

ABSTRACT

Introduction: The release of trophic factors from mesenchymal stem cells (MSCs) is critical for tissue regeneration. A systematic investigation of the regenerative potential of trophic factors from different MSCs, however, has not been performed. Thus, in the present study, the regenerative potential of conditioned medium (CM) from dental pulp, bone marrow, and adipose tissue-derived CD31(-) side population (SP) cells from an individual source was compared in an ectopic tooth transplantation model.

Methods: The tooth root transplantation in an ectopic site model was used for investigation of the regenerative potential and trophic effects in vivo. Either pulp CD31(-) SP cell populations (1×10(6) cells) at the third to fourth passage or 5 μg/ml of CM from dental pulp, bone marrow, and adipose stem cells from four different individuals were injected into the root with collagen TE. Each root was transplanted subcutaneously in 5-week-old severe combined immunodeficiency mice. Each root with surrounding tissue was harvested for histology on days 7, 21, and 28 and for Western blot analysis and real-time reverse transcription-polymerase chain reaction (RT-PCR) analysis on day 28. Furthermore, the trophic factors responsible for the regenerative potential were identified as the upregulated genes present in pulp CD31(-) SP cells when compared with the genes in both bone marrow and adipose CD31(-) SP cells by using microarray analysis, real-time RT-PCR, and Western blot analysis.

Results: Transplantation of pulp CM yielded increased volume of pulp regeneration, more bromodeoxyuridine (BrdU)-positive migrated cells, and fewer caspase 3-positive cells in the regenerated pulp compared with the others. Pulp CM also demonstrated significantly increased cell migration, anti-apoptosis, and angiogenesis in C2C12 cells. Higher expression of CXCL14 and MCP1 in pulp SP cells suggested candidate trophic factors. The stimulatory effects on both migration and angiogenesis of CXCL14 and MCP1 were demonstrated in vitro. In the regenerated tissue, BrdU-positive migrated cells expressed CXCR4 and CCR2, receptors for CXCL14 and MCP1, respectively.

Conclusions: The higher regenerative potential of pulp SP cells may be due to potent trophic factors, including CXCL14 and MCP1, which promote migration and angiogenesis.

No MeSH data available.


Related in: MedlinePlus