Characterization of stable hypoxia-preconditioned dental pulp stem cells compared with mobilized dental pulp stem cells for application for pulp regenerative therapy

Abstract Background Dental pulp stem cells (DPSCs) have been developed as a potential source of mesenchymal stem cells (MSCs) for regeneration of dental pulp and other tissues. However, further strategies to isolate highly functional DPSCs beyond the colony-forming methods are required. We have demonstrated the safety and efficacy of DPSCs isolated by G-CSF-induced mobilization and cultured under normoxia (mobilized DPSCs, MDPSCs) for pulp regeneration. The device for isolation of MDPSCs, however, is not cost-effective and requires a prolonged cell culture period. It is well known that MSCs cultured under hypoxic-preconditions improved MSC proliferation activity and stemness. Therefore, in this investigation, we attempted to improve the clinical utility of DPSCs by hypoxia-preconditioned DPSCs (hpDPSCs) compared with MDPSCs to improve the potential clinical utility for pulp regeneration in endodontic dentistry. Methods Colony-forming DPSCs were isolated and preconditioned with hypoxia in a stable closed cultured system and compared with MDPSCs isolated from the individual dog teeth. We examined the proliferation rate, migration potential, anti-apoptotic activity, and gene expression of the stem cell markers and angiogenic/neurotrophic factors. Trophic effects of the conditioned medium (CM) were also evaluated. In addition, the expression of immunomodulatory molecules upon stimulation with IFN-γ was investigated. The pulp regenerative potential and transplantation safety of hpDPSCs were further assessed in pulpectomized teeth in dogs by histological and immunohistochemical analyses and by chemistry of the blood and urine tests. Results hpDPSCs demonstrated higher proliferation rate and expression of a major regulator of oxygen homeostasis, HIF-1α, and a stem cell marker, CXCR-4. The direct migratory activity of hpDPSCs in response to G-CSF was significantly higher than MDPSCs. The CM of hpDPSCs stimulated neurite extension. However, there were no changes in angiogenic, migration, and anti-apoptotic activities compared with the CM of MDPSCs. The expression of immunomodulatory gene, PTGE was significantly upregulated by IFN gamma in hpDPSCs compared with MDPSCs. However, no difference in nitric oxide was observed. The regenerated pulp tissue was quantitatively and qualitatively similar in hpDPSC transplants compared with MDPSC transplants in dog teeth. There was no evidence of toxicity or adverse events of the hpDPSC transplantation. Conclusions These results demonstrated that the efficacy of hpDPSCs for pulp regeneration was identical, although hpDPSCs improved stem cell properties compared to MDPSCs, suggesting their potential clinical utility for pulp regeneration.

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Characterization of stable hypoxia-preconditioned dental pulp stem cells compared with mobilized dental pulp stem cells for application for pulp regenerative therapy

10.21203/rs.3.rs-102950/v2 ◽

2021 ◽

Author(s):

Mohammed Zayed ◽

Koichiro Iohara ◽

Hideto Watanabe ◽

Mami Ishikawa ◽

Michiyo Tominaga ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Dental Pulp ◽

Clinical Utility ◽

Dental Pulp Stem Cells ◽

Proliferation Rate ◽

Stem Cell Marker ◽

Stem Cell Markers ◽

Pulp Regeneration ◽

Potential Clinical Utility

Abstract Background: Dental pulp stem cells (DPSCs) have been developed as a potential source of mesenchymal stem cells (MSCs) for regeneration of dental pulp and other tissues. However, further strategies to isolate highly functional DPSCs beyond the colony-forming methods are required. We have demonstrated the safety and efficacy of DPSCs isolated by G-CSF-induced mobilization and cultured under normoxia (mobilized DPSCs, MDPSCs) for pulp regeneration. However, The device for isolation of MDPSCs, however, is not cost effective and requires prolonged cell culture period. It is well known that MSCs cultured under hypoxic-preconditions improved MSCs proliferation activity and stemness. Therefore, in this investigation, we attempted to improve the clinical utility of DPSCs by hypoxia-preconditioned DPSCs (hpDPSCs) compared with MDPSCs to improve the potential clinical utility for pulp regeneration in endodontic dentistry.Methods: Colony-forming DPSCs were isolated and preconditioned with hypoxia in a stable closed cultured system and compared with MDPSCs isolated from the individual dog teeth. We examined the proliferation rate, migration potential, anti-apoptotic activity and gene expression of the stem cell markers and angiogenic/neurotrophic factors. Trophic effects of the conditioned medium (CM) were also evaluated. In addition, the expression of immunomodulatory molecules upon stimulation with IFN-γ were investigated. The pulp regenerative potential and transplantation safety of hpDPSCs were further assessed in pulpectomized teeth in dogs by histological and immunohistochemical analyses and by chemistry of blood and urine tests. Results: hpDPSCs demonstrated higher proliferation rate and expression of a major regulator of oxygen homeostasis, HIF-1α, and a stem cell marker, CXCR-4. The direct migratory activity of hpDPSCs in response to G-CSF was significantly higher than MDPSCs. The CM of hpDPSCs stimulated neurite extension. However, there were no changes in angiogenic, migration and anti-apoptotic activities compared with the CM of MDPSCs. The expression of immunomodulatory gene, PTGE was significantly up-regulated by IFN gamma in hpDPSCs compared with MDPSCs. However, no difference in nitric oxide was observed. The regenerated pulp tissue was quantitatively and qualitatively similar in hpDPSC transplants compared with MDPSC transplants in dog teeth. There was no evidence of toxicity or adverse events of the hpDPSC transplantation. Conclusions: These results demonstrated that the efficacy of hpDPSCs for pulp regeneration was identical, although hpDPSCs improved stem cell properties compared to MDPSCs, suggesting their potential clinical utility for pulp regeneration.

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Characterization of Stable Hypoxia-Preconditioned Dental Pulp Stem Cells: Implication for Pulp Regeneration

10.21203/rs.3.rs-102950/v1 ◽

2020 ◽

Author(s):

Mohammed Zayed ◽

Koichiro Iohara ◽

Hideto Watanabe ◽

Mami Ishikawa ◽

Michiyo Tominaga ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Dental Pulp ◽

Clinical Utility ◽

Dental Pulp Stem Cells ◽

Proliferation Rate ◽

Stem Cell Marker ◽

Stem Cell Markers ◽

Pulp Tissue ◽

Pulp Regeneration

Abstract Background: Dental pulp stem cells (DPSCs) have been developed as a potential source of mesenchymal stem cells (MSCs) for regeneration of dental pulp and other tissues. However, further strategies to isolate highly functional DPSCs beyond the colony-forming methods are required. Our clinical study has demonstrated safety and efficacy of DPSCs isolated by G-CSF-induced mobilization and cultured under normoxia (mobilized DPSCs, MDPSCs) for pulp regeneration. It is well known that the oxygen concentration is closely linked to the maintenance of stemness. Thus, in this investigation, hypoxia-preconditioned DPSCs (hpDPSCs) was characterized to develop and improve the clinical utility for regeneration of dental pulp in endodontics.Methods: Colony-forming DPSCs were isolated and preconditioned with hypoxia in a stable closed cultured system and compared with MDPSCs isolated from the individual dog teeth. We examined the proliferation rate, migration potential, anti-apoptotic activity and gene expression of the stem cell markers and angiogenic/neurotrophic factors. Trophic effects of the conditioned medium (CM) were also evaluated. In addition, the expression of immunomodulatory molecules upon stimulation with IFN-γ were investigated. The pulp regenerative potential and transplantation safety of hpDPSCs were further assessed in pulpectomized teeth in dogs by histological and immunohistochemical analyses and by chemistry of blood and urine. tests Results: hpDPSCs demonstrated higher proliferation rate and expression of a major regulator of oxygen homeostasis, HIF-1α, and a stem cell marker, CXCR-4. The direct migratory activity of hpDPSCs in response to G-CSF was significantly higher than MDPSCs. The CM of hpDPSCs stimulated neurite extension. However, there were no changes in angiogenic, migration and anti-apoptotic activities compared with the CM of MDPSCs. The expression of immunomodulatory gene, PTGE was significantly up-regulated by IFN gamma in hpDPSCs compared with MDPSCs. However, no difference in nitric oxide was observed. The regenerated pulp tissue was quantitatively and qualitatively similar in hpDPSC transplants compared with MDPSC transplants in dog teeth. There was no evidence of toxicity or adverse events of the hpDPSC transplantation Conclusions: These results demonstrated that hpDPSCs improved stem cell properties compared to MDPSCs, suggesting their potential clinical utility for pulp regeneration.

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C5L2 Regulates DMP1 Expression during Odontoblastic Differentiation

Journal of Dental Research ◽

10.1177/0022034518820461 ◽

2019 ◽

Vol 98 (5) ◽

pp. 597-604 ◽

Cited By ~ 1

Author(s):

F. Chmilewsky ◽

R. Liang ◽

M. Kanazawa ◽

I. About ◽

L.F. Cooper ◽

...

Keyword(s):

Stem Cells ◽

Dental Pulp ◽

Matrix Protein ◽

Dental Pulp Stem Cells ◽

Negative Regulator ◽

Immunofluorescent Staining ◽

Stem Cell Markers ◽

C5a Receptor ◽

Pulp Regeneration ◽

Odontoblastic Differentiation

The presence of stem cells within the dental-pulp tissue as well as their differentiation into a new generation of functional odontoblast-like cells constitutes an important step of the dentin-pulp regeneration. Recent investigations demonstrated that the complement system activation participates in 2 critical steps of dentin-pulp regeneration: pulp progenitor’s recruitment and pulp nerve sprouting. Surprisingly, its implication in odontoblastic differentiation has not been addressed yet. Since the complement receptor C5a receptor-like 2 (C5L2) is expressed by different stem cells, the aim of this study is to investigate if the dental pulp stem cells express C5L2 and if this receptor participates in odontoblastic differentiation. Immunohistochemistry performed on human third molar pulp sections showed a perivascular co-localization of the mesenchymal stem cell markers STRO1 and C5L2. In vitro immunofluorescent staining confirmed that hDPSCs express C5L2. Furthermore, we determined by real-time polymerase chain reaction that the expression of C5L2 is highly modulated in human dental pulp stem cells (hDPSCs) undergoing odontoblastic differentiation. Moreover, we showed that this odontogenesis-regulated expression of C5L2 is specifically potentiated by the proinflammatory cytokine TNFα. Using a C5L2-siRNA silencing strategy, we provide direct evidence that C5L2 constitutes a negative regulator of the dentinogenic marker DMP1 (dentin matrix protein 1) expression by hDPSCs. Our findings suggest a direct correlation between the odontoblastic differentiation and the level of C5L2 expression in hDPSCs and identify C5L2 as a negative regulator of DMP1 expression by hDPSCs during the odontoblastic differentiation and inflammation processes. This work is the first to demonstrate the involvement of C5L2 in the biological function of stem cells, provides an important knowledge in understanding odontoblastic differentiation of dental pulp stem cells, and may be useful in future dentin-pulp engineering strategies.

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Effect of laser phototherapy on gene expression of stem cell markers of human dental pulp stem cells

Medicina Oral Patología Oral y Cirugia Bucal ◽

10.4317/medoral.17643599 ◽

2012 ◽

pp. S100-. ◽

Cited By ~ 1

Author(s):

LS Ferreira ◽

MFSS Destro ◽

CMC Maranduba ◽

SPH Miyagi ◽

MM. Marques

Keyword(s):

Gene Expression ◽

Stem Cells ◽

Stem Cell ◽

Dental Pulp ◽

Dental Pulp Stem Cells ◽

Stem Cell Markers ◽

Cell Markers ◽

Laser Phototherapy ◽

Human Dental Pulp

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Cell-derived micro-environment helps dental pulp stem cells promote dental pulp regeneration

Cell Proliferation ◽

10.1111/cpr.12361 ◽

2017 ◽

Vol 50 (5) ◽

pp. e12361 ◽

Cited By ~ 7

Author(s):

Xuexin Zhang ◽

Hui Li ◽

Jingjing Sun ◽

Xiangyou Luo ◽

Hefeng Yang ◽

...

Keyword(s):

Stem Cells ◽

Dental Pulp ◽

Dental Pulp Stem Cells ◽

Pulp Regeneration

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A Cell-Based Approach to Dental Pulp Regeneration Using Mesenchymal Stem Cells: A Scoping Review

International Journal of Molecular Sciences ◽

10.3390/ijms22094357 ◽

2021 ◽

Vol 22 (9) ◽

pp. 4357

Author(s):

Sahng G. Kim

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Stem Cell Transplantation ◽

Cell Transplantation ◽

Scoping Review ◽

Clinical Studies ◽

Dental Pulp ◽

Animal Studies ◽

Risk Of Bias ◽

Pulp Regeneration

Despite the recent explosion of investigations on dental pulp regeneration using various tissue engineering strategies, the translation of the findings from such studies into therapeutic applications has not been properly achieved. The purpose of this scoping review was to systematically review the efficacy of mesenchymal stem cell transplantation for dental pulp regeneration. A literature search was conducted using five electronic databases from their inception to January 2021 and supplemented by hand searches. A total of 17 studies, including two clinical trials and 15 animal studies using orthotopic pulp regeneration models, were included for the review. The risk of bias for the individual studies was assessed. This scoping review demonstrated that the regeneration of vascularized pulp-like tissue was achieved using the stem cell transplantation strategy in animal models. Autologous cell transplantation in two clinical studies also successfully regenerated vascularized vital tissue. Dental pulp stem cell subpopulations, such as mobilized dental pulp stem cells, injectable scaffolds such as atelocollagen, and a granulocyte-colony forming factor, were the most commonly used for pulp regeneration. The overall risk of bias was unclear for animal studies and was moderate or judged to raise some concerns for clinical studies. More high-quality clinical studies are needed to further determine the safety and efficacy of the stem cell transplantation strategy for dental pulp regeneration.

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