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Sample records for pulp stem cells

  1. Dental pulp stem cells

    PubMed Central

    Ashri, Nahid Y.; Ajlan, Sumaiah A.; Aldahmash, Abdullah M.

    2015-01-01

    Inflammatory periodontal disease is a major cause of loss of tooth-supporting structures. Novel approaches for regeneration of periodontal apparatus is an area of intensive research. Periodontal tissue engineering implies the use of appropriate regenerative cells, delivered through a suitable scaffold, and guided through signaling molecules. Dental pulp stem cells have been used in an increasing number of studies in dental tissue engineering. Those cells show mesenchymal (stromal) stem cell-like properties including self-renewal and multilineage differentiation potentials, aside from their relative accessibility and pleasant handling properties. The purpose of this article is to review the biological principles of periodontal tissue engineering, along with the challenges facing the development of a consistent and clinically relevant tissue regeneration platform. This article includes an updated review on dental pulp stem cells and their applications in periodontal regeneration, in combination with different scaffolds and growth factors. PMID:26620980

  2. Dental pulp stem cells in regenerative dentistry.

    PubMed

    Casagrande, Luciano; Cordeiro, Mabel M; Nör, Silvia A; Nör, Jacques E

    2011-01-01

    Stem cells constitute the source of differentiated cells for the generation of tissues during development, and for regeneration of tissues that are diseased or injured postnatally. In recent years, stem cell research has grown exponentially owing to the recognition that stem cell-based therapies have the potential to improve the life of patients with conditions that span from Alzheimer's disease to cardiac ischemia to bone or tooth loss. Growing evidence demonstrates that stem cells are primarily found in niches and that certain tissues contain more stem cells than others. Among these tissues, the dental pulp is considered a rich source of mesenchymal stem cells that are suitable for tissue engineering applications. It is known that dental pulp stem cells have the potential to differentiate into several cell types, including odontoblasts, neural progenitors, osteoblasts, chondrocytes, and adipocytes. The dental pulp stem cells are highly proliferative. This characteristic facilitates ex vivo expansion and enhances the translational potential of these cells. Notably, the dental pulp is arguably the most accessible source of postnatal stem cells. Collectively, the multipotency, high proliferation rates, and accessibility make the dental pulp an attractive source of mesenchymal stem cells for tissue regeneration. This review discusses fundamental concepts of stem cell biology and tissue engineering within the context of regenerative dentistry.

  3. Pulp stem cells: implication in reparative dentin formation.

    PubMed

    Dimitrova-Nakov, Sasha; Baudry, Anne; Harichane, Yassine; Kellermann, Odile; Goldberg, Michel

    2014-04-01

    Many dental pulp stem cells are neural crest derivatives essential for lifelong maintenance of tooth functions and homeostasis as well as tooth repair. These cells may be directly implicated in the healing process or indirectly involved in cell-to-cell diffusion of paracrine messages to resident (pulpoblasts) or nonresident cells (migrating mesenchymal cells). The identity of the pulp progenitors and the mechanisms sustaining their regenerative capacity remain largely unknown. Taking advantage of the A4 cell line, a multipotent stem cell derived from the molar pulp of mouse embryo, we investigated the capacity of these pulp-derived precursors to induce in vivo the formation of a reparative dentin-like structure upon implantation within the pulp of a rodent incisor or a first maxillary molar after surgical exposure. One month after the pulp injury alone, a nonmineralized fibrous matrix filled the mesial part of the coronal pulp chamber. Upon A4 cell implantation, a mineralized osteodentin was formed in the implantation site without affecting the structure and vitality of the residual pulp in the central and distal parts of the pulp chamber. These results show that dental pulp stem cells can induce the formation of reparative dentin and therefore constitute a useful tool for pulp therapies. Finally, reparative dentin was also built up when A4 progenitors were performed by alginate beads, suggesting that alginate is a suitable carrier for cell implantation in teeth.

  4. Functionalized scaffolds to control dental pulp stem cell fate

    PubMed Central

    Piva, Evandro; Silva, Adriana F.; Nör, Jacques E.

    2014-01-01

    Emerging understanding about interactions between stem cells, scaffolds and morphogenic factors has accelerated translational research in the field of dental pulp tissue engineering. Dental pulp stem cells constitute a sub-population of cells endowed with self-renewal and multipotency. Dental pulp stem cells seeded in biodegradable scaffolds and exposed to dentin-derived morphogenic signals give rise to a pulp-like tissue capable of generating new dentin. Notably, dentin-derived proteins are sufficient to induce dental pulp stem cell differentiation into odontoblasts. Ongoing work is focused on developing ways of mobilizing dentin-derived proteins and disinfecting the root canal of necrotic teeth without compromising the morphogenic potential of these signaling molecules. On the other hand, dentin by itself does not appear to be capable of inducing endothelial differentiation of dental pulp stem cells, despite the well known presence of angiogenic factors in dentin. This is particularly relevant in the context of dental pulp tissue engineering in full root canals, where access to blood supply is limited to the apical foramina. To address this challenge, scientists are looking at ways to use the scaffold as a controlled release device for angiogenic factors. The aim of this manuscript is to present and discuss current strategies to functionalize injectable scaffolds and customize them for dental pulp tissue engineering. The long-term goal of this work is to develop stem cell-based therapies that enable the engineering of functional dental pulps capable of generating new tubular dentin in humans. PMID:24698691

  5. Stem cells in dental practice: perspectives in conservative pulp therapies.

    PubMed

    Casagrande, Luciano; Mattuella, Letícia Grando; de Araujo, Fernando Borba; Eduardo, Jacques

    2006-01-01

    Stem cells are undifferentiated cells that have the capacity to self-renew. They have been discovered in many adult tissues, including teeth. The Dental Pulp Stem Cells are involved in dentinal repair by activation of growth factors, released after caries process and have the ability to regenerate the dentin-pulp-like complex. The molecular/cellular research raises the possibilities to grow new tissues and biological structures for clinical application, providing cells for therapies including cell transplantation and tissue engineering.

  6. A modified efficient method for dental pulp stem cell isolation

    PubMed Central

    Raoof, Maryam; Yaghoobi, Mohammad Mehdi; Derakhshani, Ali; Kamal-abadi, Ali Mohammadi; Ebrahimi, Behnam; Abbasnejad, Mehdi; Shokouhinejad, Noushin

    2014-01-01

    Background: Dental pulp stem cells can be used in regenerative endodontic therapy. The aim of this study was to introduce an efficient method for dental pulp stem cells isolation. Materials and Methods: In this in-vitro study, 60 extracted human third molars were split and pulp tissue was extracted. Dental pulp stem cells were isolated by the following three different methods: (1) digestion of pulp by collagenase/dispase enzyme and culture of the released cells; (2) outgrowth of the cells by culture of undigested pulp pieces; (3) digestion of pulp tissue pieces and fixing them. The cells were cultured in minimum essential medium alpha modification (αMEM) medium supplemented with 20% fetal bovine serum(FBS) in humid 37°C incubator with 5% CO 2. The markers of stem cells were studied by reverse transcriptase polymerase chain reaction (PCR). The student t-test was used for comparing the means of independent groups. P <0.05 was considered as significant. Results: The results indicated that by the first method a few cell colonies with homogenous morphology were detectable after 4 days, while in the outgrowth method more time was needed (10-12 days) to allow sufficient numbers of heterogeneous phenotype stem cells to migrate out of tissue. Interestingly, with the improved third method, we obtained stem cells successfully with about 60% efficiency after 2 days. The results of RT-PCR suggested the expression of Nanog, Oct-4, and Nucleostemin markers in the isolated cells from dental pulps. Conclusion: This study proposes a new method with high efficacy to obtain dental pulp stem cells in a short time. PMID:24932197

  7. Allogenic banking of dental pulp stem cells for innovative therapeutics.

    PubMed

    Collart-Dutilleul, Pierre-Yves; Chaubron, Franck; De Vos, John; Cuisinier, Frédéric J

    2015-08-26

    Medical research in regenerative medicine and cell-based therapy has brought encouraging perspectives for the use of stem cells in clinical trials. Multiple types of stem cells, from progenitors to pluripotent stem cells, have been investigated. Among these, dental pulp stem cells (DPSCs) are mesenchymal multipotent cells coming from the dental pulp, which is the soft tissue within teeth. They represent an interesting adult stem cell source because they are recovered in large amount in dental pulps with non-invasive techniques compared to other adult stem cell sources. DPSCs can be obtained from discarded teeth, especially wisdom teeth extracted for orthodontic reasons. To shift from promising preclinical results to therapeutic applications to human, DPSCs must be prepared in clinical grade lots and transformed into advanced therapy medicinal products (ATMP). As the production of patient-specific stem cells is costly and time-consuming, allogenic biobanking of clinical grade human leukocyte antigen (HLA)-typed DPSC lines provides efficient innovative therapeutic products. DPSC biobanks represent industrial and therapeutic innovations by using discarded biological tissues (dental pulps) as a source of mesenchymal stem cells to produce and store, in good manufacturing practice (GMP) conditions, DPSC therapeutic batches. In this review, we discuss about the challenges to transfer biological samples from a donor to HLA-typed DPSC therapeutic lots, following regulations, GMP guidelines and ethical principles. We also present some clinical applications, for which there is no efficient therapeutics so far, but that DPSCs-based ATMP could potentially treat.

  8. Allogenic banking of dental pulp stem cells for innovative therapeutics

    PubMed Central

    Collart-Dutilleul, Pierre-Yves; Chaubron, Franck; De Vos, John; Cuisinier, Frédéric J

    2015-01-01

    Medical research in regenerative medicine and cell-based therapy has brought encouraging perspectives for the use of stem cells in clinical trials. Multiple types of stem cells, from progenitors to pluripotent stem cells, have been investigated. Among these, dental pulp stem cells (DPSCs) are mesenchymal multipotent cells coming from the dental pulp, which is the soft tissue within teeth. They represent an interesting adult stem cell source because they are recovered in large amount in dental pulps with non-invasive techniques compared to other adult stem cell sources. DPSCs can be obtained from discarded teeth, especially wisdom teeth extracted for orthodontic reasons. To shift from promising preclinical results to therapeutic applications to human, DPSCs must be prepared in clinical grade lots and transformed into advanced therapy medicinal products (ATMP). As the production of patient-specific stem cells is costly and time-consuming, allogenic biobanking of clinical grade human leukocyte antigen (HLA)-typed DPSC lines provides efficient innovative therapeutic products. DPSC biobanks represent industrial and therapeutic innovations by using discarded biological tissues (dental pulps) as a source of mesenchymal stem cells to produce and store, in good manufacturing practice (GMP) conditions, DPSC therapeutic batches. In this review, we discuss about the challenges to transfer biological samples from a donor to HLA-typed DPSC therapeutic lots, following regulations, GMP guidelines and ethical principles. We also present some clinical applications, for which there is no efficient therapeutics so far, but that DPSCs-based ATMP could potentially treat. PMID:26328017

  9. Influence of different types of pulp treatment during isolation in the obtention of human dental pulp stem cells

    PubMed Central

    Viña-Almunia, Jose; Borras, Consuelo; Gambini, Juan; El Alamy, Marya; Viña, Jose

    2016-01-01

    Background Different methods have been used in order to isolate dental pulp stem cells. The aim of this study was to study the effect of different types of pulp treatment during isolation, under 3% O2 conditions, in the time needed and the efficacy for obtaining dental pulp stem cells. Material and Methods One hundred and twenty dental pulps were used to isolate dental pulp stem cells treating the pulp tissue during isolation using 9 different methods, using digestive, disgregation, or mechanical agents, or combining them. The cells were positive for CD133, Oct4, Nestin, Stro-1, CD34 markers, and negative for the hematopoietic cell marker CD-45, thus confirming the presence of mesenchymal stem cells. The efficacy of dental pulp stem cells obtention and the minimum time needed to obtain such cells comparing the 9 different methods was analyzed. Results Dental pulp stem cells were obtained from 97 of the 120 pulps used in the study, i.e. 80.8% of the cases. They were obtained with all the methods used except with mechanical fragmentation of the pulp, where no enzymatic digestion was performed. The minimum time needed to isolate dental pulp stem cells was 8 hours, digesting with 2mg/ml EDTA for 10 minutes, 4mg/ml of type I collagenase, 4mg/ml of type II dispase for 40 minutes, 13ng/ml of thermolysine for 40 minutes and sonicating the culture for one minute. Conclusions Dental pulp stem cells were obtained in 97 cases from a series of 120 pulps. The time for obtaining dental pulp stem cells was reduced maximally, without compromising the obtention of the cells, by combining digestive, disgregation, and mechanical agents. Key words:Dental pulp stem cells, mesenchymal stem cells, isolation method. PMID:26946201

  10. Pulp tissue from primary teeth: new source of stem cells

    PubMed Central

    TELLES, Paloma Dias; MACHADO, Maria Aparecida de Andrade Moreira; SAKAI, Vivien Thiemy; NÖR, Jacques Eduardo

    2011-01-01

    SHED (stem cells from human exfoliated deciduous teeth) represent a population of postnatal stem cells capable of extensive proliferation and multipotential differentiation. Primary teeth may be an ideal source of postnatal stem cells to regenerate tooth structures and bone, and possibly to treat neural tissue injury or degenerative diseases. SHED are highly proliferative cells derived from an accessible tissue source, and therefore hold potential for providing enough cells for clinical applications. In this review, we describe the current knowledge about dental pulp stem cells and discuss tissue engineering approaches that use SHED to replace irreversibly inflamed or necrotic pulps with a healthy and functionally competent tissue that is capable of forming new dentin. PMID:21625731

  11. Mobilized dental pulp stem cells for pulp regeneration: initiation of clinical trial.

    PubMed

    Nakashima, Misako; Iohara, Koichiro

    2014-04-01

    Stem cell therapy is a potential strategy to regenerate the dentin-pulp complex, enabling the conservation and restoration of functional teeth. We assessed the efficacy and safety of pulp stem cell transplantation as a prelude before the initiation of clinical trials. Granulocyte-colony stimulating factor (G-CSF) induces subsets of dental pulp stem cells to form mobilized dental pulp stem cells (MDPSCs). Good manufacturing practice is a prerequisite for the isolation and expansion of MDPSCs that are enriched in stem cells, expressing a high level of trophic factors with properties of high proliferation, migration, and antiapoptotic effects and endowed with regenerative potential. The quality of clinical-grade MDPSCs was assured by the absence of abnormalities/aberrations in karyotype and the lack of tumor formation after transplantation in immunodeficient mice. Autologous transplantation of MDPSCs with G-CSF in pulpectomized teeth in dogs augmented the regeneration of pulp tissue. The combinatorial trophic effects of MDPSCs and G-CSF on cell migration, antiapoptosis, immunosuppression, and neurite outgrowth were also confirmed in vitro. Furthermore, MDPSCs from the aged donors were as potent as the young donors. It is noteworthy that there were no significant age-related changes in biological properties such as stability, regenerative potential, and expression of the senescence markers in MDPSCs. On the other hand, autologous transplantation of MDPSCs with G-CSF induced less regenerated pulp tissue in the aged dogs compared with the young dogs. In conclusion, the preclinical safety, feasibility, and efficacy of pulp regeneration by MDPSCs and G-CSF were established. Therefore, the standardization and establishment of regulatory guidelines for stem cell therapy in clinical endodontics is now a reality. Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  12. Influence of different types of pulp treatment during isolation in the obtention of human dental pulp stem cells.

    PubMed

    Viña-Almunia, J; Borras, C; Gambini, J; El Alamy, M; Peñarrocha, M; Viña, J

    2016-05-01

    Different methods have been used in order to isolate dental pulp stem cells. The aim of this study was to study the effect of different types of pulp treatment during isolation, under 3% O2 conditions, in the time needed and the efficacy for obtaining dental pulp stem cells. One hundred and twenty dental pulps were used to isolate dental pulp stem cells treating the pulp tissue during isolation using 9 different methods, using digestive, disgregation, or mechanical agents, or combining them. The cells were positive for CD133, Oct4, Nestin, Stro-1, CD34 markers, and negative for the hematopoietic cell marker CD-45, thus confirming the presence of mesenchymal stem cells. The efficacy of dental pulp stem cells obtention and the minimum time needed to obtain such cells comparing the 9 different methods was analyzed. Dental pulp stem cells were obtained from 97 of the 120 pulps used in the study, i.e. 80.8% of the cases. They were obtained with all the methods used except with mechanical fragmentation of the pulp, where no enzymatic digestion was performed. The minimum time needed to isolate dental pulp stem cells was 8 hours, digesting with 2mg/ml EDTA for 10 minutes, 4mg/ml of type I collagenase, 4mg/ml of type II dispase for 40 minutes, 13ng/ml of thermolysine for 40 minutes and sonicating the culture for one minute. Dental pulp stem cells were obtained in 97 cases from a series of 120 pulps. The time for obtaining dental pulp stem cells was reduced maximally, without compromising the obtention of the cells, by combining digestive, disgregation, and mechanical agents.

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

    PubMed

    Zhang, Xuexin; Li, Hui; Sun, Jingjing; Luo, Xiangyou; Yang, Hefeng; Xie, Li; Yang, Bo; Guo, Weihua; Tian, Weidong

    2017-10-01

    The function of the dental pulp is closely connected to the extracellular matrix (ECM) structure, and ECM has received significant attention due to its biological functions for regulating cells. As such, the interaction between the ECM niche and cells is worth exploring for potential clinical uses. In this study, dental pulp stem cell (DPSC)-derived ECM (DPM) was prepared through cell culture and decellularization to function as the cell niche, and changes in DPSC behaviour and histological analysis of dental pulp tissue regeneration were evaluated following the DPM culture. DPM promoted the replication of DPSCs and exhibited retention of their mineralization. Then, the DPM-based culture strategy under odontogenic culture medium was further investigated, and the mineralization-related markers showed that DPSCs were regulated towards odontogenic differentiation. Dental pulp-like tissue with well-arranged ECM was harvested after a 2-month subcutaneous implantation in nude mice with DPM application. Additionally, DPSCs cultured on the plastic culture surface showed the up-regulation of mineralization makers in vitro, but there was a disorder in matrix formation and mineralization when the cells were cultured in vivo. DPM-based cultivation could serve as a cell niche and modulate DPSC behaviour, and this method also provided an alternative to harvest tissue-specific ECM and provided a strategy for ECM-cell interaction. © 2017 John Wiley & Sons Ltd.

  14. Dentin regeneration using deciduous pulp stem/progenitor cells.

    PubMed

    Zheng, Y; Wang, X Y; Wang, Y M; Liu, X Y; Zhang, C M; Hou, B X; Wang, S L

    2012-07-01

    Reparative dentin formation is essential for maintaining the integrity of dentin structure during disease or trauma. In this study, we investigated stem/progenitor cell-based tissue engineering for dentin regeneration in a large animal model. Porcine deciduous pulp stem/progenitor cells (PDPSCs) were mixed with a beta-tricalcium phosphate (β-TCP) scaffold for dentin regeneration. Different concentrations of PDPSCs were tested to determine the optimal density for dentin regeneration. Aliquots of 5×10(5) PDPSCs in 1 mL resulted in the highest number of cells attached to the scaffold and the greatest alkaline phosphatase activity. We labeled PDPSCs with green fluorescent protein (GFP) and used the optimal cell numbers mixed with β-TCP to repair pulp chamber roof defects in the premolars of swine. Four weeks after transplantation, GFP-positive PDPSCs were observed in PDPSC-embedded scaffold constructs. At 16 weeks after transplantation, the PDPSCs mixed with β-TCP significantly regenerated the dentin-like structures and nearly completely restored the pulp chamber roof defects. This study demonstrated that the PDPSC/scaffold construct was useful in direct pulp-capping and provides pre-clinical evidence for stem/progenitor cell-based dentin regeneration.

  15. Regenerative medicine using dental pulp stem cells for liver diseases

    PubMed Central

    Ohkoshi, Shogo; Hara, Hajime; Hirono, Haruka; Watanabe, Kazuhiko; Hasegawa, Katsuhiko

    2017-01-01

    Acute liver failure is a refractory disease and its prognosis, if not treated using liver transplantation, is extremely poor. It is a good candidate for regenerative medicine, where stem cell-based therapies play a central role. Mesenchymal stem cells (MSCs) are known to differentiate into multiple cell lineages including hepatocytes. Autologous cell transplant without any foreign gene induction is feasible using MSCs, thereby avoiding possible risks of tumorigenesis and immune rejection. Dental pulp also contains an MSC population that differentiates into hepatocytes. A point worthy of special mention is that dental pulp can be obtained from deciduous teeth during childhood and can be subsequently harvested when necessary after deposition in a tooth bank. MSCs have not only a regenerative capacity but also act in an anti-inflammatory manner via paracrine mechanisms. Promising efficacies and difficulties with the use of MSC derived from teeth are summarized in this review. PMID:28217369

  16. Regenerative medicine using dental pulp stem cells for liver diseases.

    PubMed

    Ohkoshi, Shogo; Hara, Hajime; Hirono, Haruka; Watanabe, Kazuhiko; Hasegawa, Katsuhiko

    2017-02-06

    Acute liver failure is a refractory disease and its prognosis, if not treated using liver transplantation, is extremely poor. It is a good candidate for regenerative medicine, where stem cell-based therapies play a central role. Mesenchymal stem cells (MSCs) are known to differentiate into multiple cell lineages including hepatocytes. Autologous cell transplant without any foreign gene induction is feasible using MSCs, thereby avoiding possible risks of tumorigenesis and immune rejection. Dental pulp also contains an MSC population that differentiates into hepatocytes. A point worthy of special mention is that dental pulp can be obtained from deciduous teeth during childhood and can be subsequently harvested when necessary after deposition in a tooth bank. MSCs have not only a regenerative capacity but also act in an anti-inflammatory manner via paracrine mechanisms. Promising efficacies and difficulties with the use of MSC derived from teeth are summarized in this review.

  17. A Novel Combinatorial Therapy With Pulp Stem Cells and Granulocyte Colony-Stimulating Factor for Total Pulp Regeneration

    PubMed Central

    Iohara, Koichiro; Murakami, Masashi; Takeuchi, Norio; Osako, Yohei; Ito, Masataka; Ishizaka, Ryo; Utunomiya, Shinji; Nakamura, Hiroshi; Matsushita, Kenji

    2013-01-01

    Treatment of deep caries with pulpitis is a major challenge in dentistry. Stem cell therapy represents a potential strategy to regenerate the dentin-pulp complex, enabling conservation and restoration of teeth. The objective of this study was to assess the efficacy and safety of pulp stem cell transplantation as a prelude for the impending clinical trials. Clinical-grade pulp stem cells were isolated and expanded according to good manufacturing practice conditions. The absence of contamination, abnormalities/aberrations in karyotype, and tumor formation after transplantation in an immunodeficient mouse ensured excellent quality control. After autologous transplantation of pulp stem cells with granulocyte-colony stimulating factor (G-CSF) in a dog pulpectomized tooth, regenerated pulp tissue including vasculature and innervation completely filled in the root canal, and regenerated dentin was formed in the coronal part and prevented microleakage up to day 180. Transplantation of pulp stem cells with G-CSF yielded a significantly larger amount of regenerated dentin-pulp complex compared with transplantation of G-CSF or stem cells alone. Also noteworthy was the reduction in the number of inflammatory cells and apoptotic cells and the significant increase in neurite outgrowth compared with results without G-CSF. The transplanted stem cells expressed angiogenic/neurotrophic factors. It is significant that G-CSF together with conditioned medium of pulp stem cells stimulated cell migration and neurite outgrowth, prevented cell death, and promoted immunosuppression in vitro. Furthermore, there was no evidence of toxicity or adverse events. In conclusion, the combinatorial trophic effects of pulp stem cells and G-CSF are of immediate utility for pulp/dentin regeneration, demonstrating the prerequisites of safety and efficacy critical for clinical applications. PMID:23761108

  18. Preliminary study on dental pulp stem cell-mediated pulp regeneration in canine immature permanent teeth.

    PubMed

    Wang, Yuanyuan; Zhao, Yuming; Jia, Weiqian; Yang, Jie; Ge, Lihong

    2013-02-01

    The health of human teeth depends on the integrity of the hard tissue and the activity of the pulp and periodontal tissues, which are responsible for nutritional supply. Without the nourishing of the pulp tissue, the possibility of tooth fracture can increase. In immature permanent teeth, root development may be influenced as well. This study explored the potential of using autologous dental pulp stem cells (DPSCs) to achieve pulp regeneration in a canine pulpless model. The establishment of the pulpless animal model involved pulp extirpation and root canal preparation of young permanent incisor teeth in beagles. Autologous DPSCs were obtained from extracted first molars and expanded ex vivo to obtain a larger number of cells. The biological characteristics of canine DPSCs (cDPSCs) were analyzed both in vitro and in vivo by using the same method as used in human DPSCs. cDPSCs were transplanted into the pulpless root canal with Gelfoam as the scaffold, and root development was evaluated by radiographic and histologic analyses. cDPSCs with rapid proliferation, multiple differentiation capacity, and development potential were successfully isolated and identified both in vitro and in vivo. After they were transplanted into the pulpless root canal with Gelfoam as the scaffold, DPSCs were capable of generating pulp-like tissues containing blood vessels and dentin-like tissue. Thickening of the root canal wall was also observed. This study demonstrates the feasibility of using stem cell-mediated tissue engineering to realize pulp regeneration in immature teeth. Copyright © 2013 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  19. Stem cells in dental pulp of deciduous teeth.

    PubMed

    Kerkis, Irina; Caplan, Arnold I

    2012-04-01

    Dental pulp from deciduous (baby) teeth, which are discarded after exfoliation, represents an advantageous source of young stem cells. Herein, we discuss the methods of deciduous teeth stem cell (DTSC) isolation and cultivation. We show that based on these methods, at least three different stem cell populations can be identified: a population similar to bone marrow-derived mesenchymal stem cells, an epithelial stem-like cells, and/or a mixed population composed of both cell types. We analyzed the embryonic origin and stem cell niche of DTSCs with respect to the advantages they can provide for their future use in cell therapies and regenerative medicine. In vitro and in vivo differentiation of the DTSC populations, their developmental potential, immunological compatibility, tissue engineering, and transplantation use in studies in animal models are also the focus of the current report. We briefly describe the derivation of induced pluripotent stem (iPS) cells from DTSCs, which can be obtained more easily and efficiently in comparison with human fibroblasts. These iPS cells represent an interesting model for the investigation of pediatric diseases and disorders. The importance of DTSC banking is also discussed.

  20. In Vivo Experiments with Dental Pulp Stem Cells for Pulp-Dentin Complex Regeneration.

    PubMed

    Kim, Sunil; Shin, Su-Jung; Song, Yunjung; Kim, Euiseong

    2015-01-01

    In recent years, many studies have examined the pulp-dentin complex regeneration with DPSCs. While it is important to perform research on cells, scaffolds, and growth factors, it is also critical to develop animal models for preclinical trials. The development of a reproducible animal model of transplantation is essential for obtaining precise and accurate data in vivo. The efficacy of pulp regeneration should be assessed qualitatively and quantitatively using animal models. This review article sought to introduce in vivo experiments that have evaluated the potential of dental pulp stem cells for pulp-dentin complex regeneration. According to a review of various researches about DPSCs, the majority of studies have used subcutaneous mouse and dog teeth for animal models. There is no way to know which animal model will reproduce the clinical environment. If an animal model is developed which is easier to use and is useful in more situations than the currently popular models, it will be a substantial aid to studies examining pulp-dentin complex regeneration.

  1. In Vivo Experiments with Dental Pulp Stem Cells for Pulp-Dentin Complex Regeneration

    PubMed Central

    Kim, Sunil; Shin, Su-Jung; Song, Yunjung; Kim, Euiseong

    2015-01-01

    In recent years, many studies have examined the pulp-dentin complex regeneration with DPSCs. While it is important to perform research on cells, scaffolds, and growth factors, it is also critical to develop animal models for preclinical trials. The development of a reproducible animal model of transplantation is essential for obtaining precise and accurate data in vivo. The efficacy of pulp regeneration should be assessed qualitatively and quantitatively using animal models. This review article sought to introduce in vivo experiments that have evaluated the potential of dental pulp stem cells for pulp-dentin complex regeneration. According to a review of various researches about DPSCs, the majority of studies have used subcutaneous mouse and dog teeth for animal models. There is no way to know which animal model will reproduce the clinical environment. If an animal model is developed which is easier to use and is useful in more situations than the currently popular models, it will be a substantial aid to studies examining pulp-dentin complex regeneration. PMID:26688616

  2. Human dental pulp stem cells: Applications in future regenerative medicine.

    PubMed

    Potdar, Pravin D; Jethmalani, Yogita D

    2015-06-26

    Stem cells are pluripotent cells, having a property of differentiating into various types of cells of human body. Several studies have developed mesenchymal stem cells (MSCs) from various human tissues, peripheral blood and body fluids. These cells are then characterized by cellular and molecular markers to understand their specific phenotypes. Dental pulp stem cells (DPSCs) are having a MSCs phenotype and they are differentiated into neuron, cardiomyocytes, chondrocytes, osteoblasts, liver cells and β cells of islet of pancreas. Thus, DPSCs have shown great potentiality to use in regenerative medicine for treatment of various human diseases including dental related problems. These cells can also be developed into induced pluripotent stem cells by incorporation of pluripotency markers and use for regenerative therapies of various diseases. The DPSCs are derived from various dental tissues such as human exfoliated deciduous teeth, apical papilla, periodontal ligament and dental follicle tissue. This review will overview the information about isolation, cellular and molecular characterization and differentiation of DPSCs into various types of human cells and thus these cells have important applications in regenerative therapies for various diseases. This review will be most useful for postgraduate dental students as well as scientists working in the field of oral pathology and oral medicine.

  3. Human dental pulp stem cells: Applications in future regenerative medicine

    PubMed Central

    Potdar, Pravin D; Jethmalani, Yogita D

    2015-01-01

    Stem cells are pluripotent cells, having a property of differentiating into various types of cells of human body. Several studies have developed mesenchymal stem cells (MSCs) from various human tissues, peripheral blood and body fluids. These cells are then characterized by cellular and molecular markers to understand their specific phenotypes. Dental pulp stem cells (DPSCs) are having a MSCs phenotype and they are differentiated into neuron, cardiomyocytes, chondrocytes, osteoblasts, liver cells and β cells of islet of pancreas. Thus, DPSCs have shown great potentiality to use in regenerative medicine for treatment of various human diseases including dental related problems. These cells can also be developed into induced pluripotent stem cells by incorporation of pluripotency markers and use for regenerative therapies of various diseases. The DPSCs are derived from various dental tissues such as human exfoliated deciduous teeth, apical papilla, periodontal ligament and dental follicle tissue. This review will overview the information about isolation, cellular and molecular characterization and differentiation of DPSCs into various types of human cells and thus these cells have important applications in regenerative therapies for various diseases. This review will be most useful for postgraduate dental students as well as scientists working in the field of oral pathology and oral medicine. PMID:26131314

  4. Isolation of Mesenchymal Stem Cells from Human Deciduous Teeth Pulp

    PubMed Central

    Tsai, Aileen I.; Hong, Hsiang-Hsi; Fu, Jen-Fen; Chang, Chih-Chun; Wang, I-Kuan; Huang, Wen-Hung; Weng, Cheng-Hao; Hsu, Ching-Wei

    2017-01-01

    This study aimed to identify predictors of success rate of mesenchymal stem cell (MSC) isolation from human deciduous teeth pulp. A total of 161 deciduous teeth were extracted at the dental clinic of Chang Gung Memorial Hospital. The MSCs were isolated from dental pulps using a standard protocol. In total, 128 colonies of MSCs were obtained and the success rate was 79.5%. Compared to teeth not yielding MSCs successfully, those successfully yielding MSCs were found to have less severe dental caries (no/mild-to-moderate/severe: 63.3/24.2/12.5% versus 12.5/42.4/42.4%, P < 0.001) and less frequent pulpitis (no/yes: 95.3/4.7% versus 51.5/48.5%, P < 0.001). In a multivariate regression model, it was confirmed that the absence of dental caries (OR = 4.741, 95% CI = 1.564–14.371, P = 0.006) and pulpitis (OR = 9.111, 95% CI = 2.921–28.420, P < 0.001) was significant determinants of the successful procurement of MSCs. MSCs derived from pulps with pulpitis expressed longer colony doubling time than pulps without pulpitis. Furthermore, there were higher expressions of proinflammatory cytokines, interleukin- (IL-) 6 and monocyte chemoattractant protein- (MCP-) 1, P < 0.01, and innate immune response [toll-like receptor 1 (TLR1) and TLR8, P < 0.05; TLR2, TLR3, and TLR6, P < 0.01] in the inflamed than noninflamed pulps. Therefore, a carious deciduous tooth or tooth with pulpitis was relatively unsuitable for MSC processing and isolation. PMID:28377925

  5. Mesenchymal Stem Cells Derived from Dental Pulp: A Review

    PubMed Central

    Santiago-Osorio, Edelmiro

    2016-01-01

    The mesenchymal stem cells of dental pulp (DPSCs) were isolated and characterized for the first time more than a decade ago as highly clonogenic cells that were able to generate densely calcified colonies. Now, DPSCs are considered to have potential as stem cell source for orthopedic and oral maxillofacial reconstruction, and it has been suggested that they may have applications beyond the scope of the stomatognathic system. To date, most studies have shown that, regardless of their origin in third molars, incisors, or exfoliated deciduous teeth, DPSCs can generate mineralized tissue, an extracellular matrix and structures type dentin, periodontal ligament, and dental pulp, as well as other structures. Different groups worldwide have designed and evaluated new efficient protocols for the isolation, expansion, and maintenance of clinically safe human DPSCs in sufficient numbers for various therapeutics protocols and have discussed the most appropriate route of administration, the possible contraindications to their clinical use, and the parameters to be considered for monitoring their clinical efficacy and proper biological source. At present, DPSC-based therapy is promising but because most of the available evidence was obtained using nonhuman xenotransplants, it is not a mature technology. PMID:26779263

  6. Angiogenic properties of human dental pulp stem cells.

    PubMed

    Bronckaers, Annelies; Hilkens, Petra; Fanton, Yanick; Struys, Tom; Gervois, Pascal; Politis, Constantinus; Martens, Wendy; Lambrichts, Ivo

    2013-01-01

    Angiogenesis, the formation of capillaries from pre-existing blood vessels, is a key process in tissue engineering. If blood supply cannot be established rapidly, there is insufficient oxygen and nutrient transport and necrosis of the implanted tissue will occur. Recent studies indicate that the human dental pulp contains precursor cells, named dental pulp stem cells (hDPSC) that show self-renewal and multilineage differentiation capacity. Since these cells can be easily isolated, cultured and cryopreserved, they represent an attractive stem cell source for tissue engineering. Until now, only little is known about the angiogenic abilities and mechanisms of the hDPSC. In this study, the angiogenic profile of both cell lysates and conditioned medium of hDPSC was determined by means of an antibody array. Numerous pro-and anti-angiogenic factors such as vascular endothelial growth factor (VEGF), monocyte chemotactic protein-1 (MCP-1), plasminogen activator inhibitor-1 (PAI-1) and endostatin were found both at the mRNA and protein level. hDPSC had no influence on the proliferation of the human microvascular endothelial cells (HMEC-1), but were able to significantly induce HMEC-1 migration in vitro. Addition of the PI3K-inhibitor LY294002 and the MEK-inhibitor U0126 to the HMEC-1 inhibited this effect, suggesting that both Akt and ERK pathways are involved in hDPSC-mediated HMEC-1 migration. Antibodies against VEGF also abolished the chemotactic actions of hDPSC. Furthermore, in the chicken chorioallantoic membrane (CAM) assay, hDPSC were able to significantly induce blood vessel formation. In conclusion, hDPSC have the ability to induce angiogenesis, meaning that this stem cell population has a great clinical potential, not only for tissue engineering but also for the treatment of chronic wounds, stroke and myocardial infarctions.

  7. Human dental pulp stem cells: from biology to clinical applications.

    PubMed

    d'Aquino, Riccardo; De Rosa, Alfredo; Laino, Gregorio; Caruso, Filippo; Guida, Luigi; Rullo, Rosario; Checchi, Vittorio; Laino, Luigi; Tirino, Virginia; Papaccio, Gianpaolo

    2009-07-15

    Dental pulp stem cells (DPSCs) can be found within the "cell rich zone" of dental pulp. Their embryonic origin, from neural crests, explains their multipotency. Up to now, two groups have studied these cells extensively, albeit with different results. One group claims that these cells produce a "dentin-like tissue", whereas the other research group has demonstrated that these cells are capable of producing bone, both in vitro and in vivo. In addition, it has been reported that these cells can be easily cryopreserved and stored for long periods of time and still retain their multipotency and bone-producing capacity. Moreover, recent attention has been focused on tissue engineering and on the properties of these cells: several scaffolds have been used to promote 3-D tissue formation and studies have demonstrated that DPSCs show good adherence and bone tissue formation on microconcavity surface textures. In addition, adult bone tissue with good vascularization has been obtained in grafts. These results enforce the notion that DPSCs can be used successfully for tissue engineering. (c) 2008 Wiley-Liss, Inc.

  8. JAB1 accelerates odontogenic differentiation of dental pulp stem cells.

    PubMed

    Lian, Min; Zhang, Ye; Shen, Qijie; Xing, Jing; Lu, Xiaohui; Huang, Dan; Cao, Peipei; Shen, Shuling; Zheng, Ke; Zhang, Jinlong; Chen, Jie; Wang, Yi; Feng, Guijuan; Feng, Xingmei

    2016-06-01

    Jun activation domain-binding protein 1 (JAB1) is a multifunctional protein that participates in the control of cell proliferation and the stability of multiple proteins. JAB1 regulates several key proteins, and thereby produces varied effects on cell cycle progression, genome stability and cell survival. Some studies have shown that the loss of JAB1 in osteochondral progenitor cells severely impairs embryonic limb development in mice. However, the biological significance of JAB1 activity in the odontogenic differentiation of dental pulp stem cells (DPSCs) remains unclear. This study aimed to determine the role of JAB1, a key player in tooth development, in reparative dentin formation, especially odontogenic differentiation. We found that increased expression of JAB1 promoted odontogenic differentiation of DPSCs via Wnt/β-catenin signaling. The role of JAB1 in the odontogenic differentiation of DPSCs was further confirmed by knocking down JAB1. Our findings provide novel insights on odontogenic differentiation of DPSCs.

  9. Wnt/β-catenin Inhibits Dental Pulp Stem Cell Differentiation

    PubMed Central

    Scheller, E.L.; Chang, J.; Wang, C.Y.

    2010-01-01

    Dental pulp stem cells (DPSCs) are a unique precursor population isolated from post-natal human dental pulp and have the ability to regenerate a reparative dentin-like complex. Canonical Wnt signaling plays a critical role in tooth development and stem cell self-renewal through β-catenin. In this study, the regulation of odontoblast-like differentiation of DPSCs by canonical Wnt signaling was examined. DPSCs were stably transduced with canonical Wnt-1 or the active form of β-catenin, with retrovirus-mediated infection. Northern blot analysis found that Wnt-1 strongly induced the expression of matricellular protein osteopontin, and modestly enhanced the expression of type I collagen in DPSCs. Unexpectedly, Wnt-1 inhibited alkaline phosphatase (ALP) activity and the formation of mineralized nodules in DPSCs. Moreover, over-expression of β-catenin was also sufficient to suppress the differentiation and mineralization of DPSCs. In conclusion, our results suggest that canonical Wnt signaling negatively regulates the odontoblast-like differentiation of DPSCs. PMID:18218837

  10. Mesenchymal stem cell characteristics of dental pulp and periodontal ligament stem cells after in vivo transplantation.

    PubMed

    Lei, Ming; Li, Kun; Li, Bei; Gao, Li-Na; Chen, Fa-Ming; Jin, Yan

    2014-08-01

    Mesenchymal stem cells (MSCs) isolated from human postnatal dental pulp and periodontal ligament (PDL) tissues can give rise to multilineage differentiation in vitro and generate related dental tissues in vivo. However, the cell properties of human dental pulp stem cells (DPSCs) and PDL stem cells (PDLSCs) after in vivo implantation remain largely unidentified. In this study, cells were re-isolated from in vivo-generated dental pulp-like and PDL-like tissues (termed re-DPCs and re-PDLCs, respectively) as a result of ectopic transplantation of human DPSC and PDLSC sheets. The cell characteristics in terms of colony-forming ability, cell surface antigens and multi-differentiation potentials were all evaluated before and after implantation. It was found that re-DPCs and re-PDLCs were of human and mesenchymal origin and positive for MSC markers such as STRO-1, CD146, CD29, CD90 and CD105; and, to some extent, re-DPCs could maintain their colony forming abilities. Moreover, both cell types were able to form mineral deposits and differentiate into adipocytes and chondrocytes; however, quantitative analysis and related gene expression determination showed that the osteo-/chondro-differentiation capabilities of re-DPCs and re-PDLCs were significantly reduced compared to those of DPSCs and PDLSCs, respectively (P < 0.05); re-PDLCs showed a greater reduction potential than re-DPCs. We conclude that DPSCs and PDLSCs may maintain their MSC characteristics after in vivo implantation and, compared to PDLSCs, DPSCs appear much more stable under in vivo conditions. These findings provide additional cellular and molecular evidence that supports expanding the use of dental tissue-derived stem cells in cell therapy and tissue engineering.

  11. Differentiation of dental pulp stem cells into neuron-like cells in serum-free medium.

    PubMed

    Zainal Ariffin, Shahrul Hisham; Kermani, Shabnam; Zainol Abidin, Intan Zarina; Megat Abdul Wahab, Rohaya; Yamamoto, Zulham; Senafi, Sahidan; Zainal Ariffin, Zaidah; Abdul Razak, Mohamad

    2013-01-01

    Dental pulp tissue contains dental pulp stem cells (DPSCs). Dental pulp cells (also known as dental pulp-derived mesenchymal stem cells) are capable of differentiating into multilineage cells including neuron-like cells. The aim of this study was to examine the capability of DPSCs to differentiate into neuron-like cells without using any reagents or growth factors. DPSCs were isolated from teeth extracted from 6- to 8-week-old mice and maintained in complete medium. The cells from the fourth passage were induced to differentiate by culturing in medium without serum or growth factors. RT-PCR molecular analysis showed characteristics of Cd146(+) , Cd166(+) , and Cd31(-) in DPSCs, indicating that these cells are mesenchymal stem cells rather than hematopoietic stem cells. After 5 days of neuronal differentiation, the cells showed neuron-like morphological changes and expressed MAP2 protein. The activation of Nestin was observed at low level prior to differentiation and increased after 5 days of culture in differentiation medium, whereas Tub3 was activated only after 5 days of neuronal differentiation. The proliferation of the differentiated cells decreased in comparison to that of the control cells. Dental pulp stem cells are induced to differentiate into neuron-like cells when cultured in serum- and growth factor-free medium.

  12. Human dental pulp mesenchymal stem cells isolation and osteoblast differentiation.

    PubMed

    Alkhalil, Moustafa; Smajilagić, Amer; Redžić, Amira

    2015-02-01

    This study was focused on the isolation and characterization of mesenchymal stem cells (MSCs) from human dental pulp (DPSC). The study was performed in the Department for Oral and Cranio-Maxillo- Facial Surgey Hamad Medical Corporation, Doha, Qatar and Weill Cornell Medical Colleague Doha, Qatar, in period 2010-2011. Dental pulp was extracted from premolars and third molars of 19 healthy patients. The pulp was digested in a solution of 3 mg/mL collagenase type I and 4 mg/mL dispase for 1 hour at 37C. After filtration, cells were cultured in Dulbecco's Modified Eagle Medium (DMEM Low Glucoses) with 20% Fetal Bovine Serum (FBS), 2mM L-glutamine and antibiotics (100 U/mL penicillin, 100 ug/mL streptomycin) at 37 °C under 5% CO2. Cultures were treated with osteoinductive medium for differentiation MSC in to the osteoblast cell line. Staining with Alizarin red were used for the detection of the osteoblast production and calcification new formed tissue. On the total of three out of 19 patients it was possible to isolate DPMSCs after 2 to 3 weeks: in one patient it was not possible to expand MSCs because of infection, and in other two patients positive Alizarin red staining reaction showed osteogenic differentiation capability and strong mineralization in vitro. The main advantage of using DPSC is absence of morbidity. MSCs could be isolated noninvasively from teeth, routinely extracted in the clinic and discarded as medical waste. Standardization of clinical and laboratory protocols for DPMSCs isolation and team work coordination could lead to significantly improved result.

  13. Transplantation of dental pulp stem cells and platelet-rich plasma for pulp regeneration.

    PubMed

    Zhu, Xiaofei; Zhang, Chengfei; Huang, George T-J; Cheung, Gary S P; Dissanayaka, Waruna Lakmal; Zhu, Wenhao

    2012-12-01

    The loss of dental pulp may weaken teeth, rendering them susceptible to reinfection, fracture, and subsequent tooth loss. Therefore, regeneration of pulp is considered an ideal treatment to preserve teeth. The aim of this study was to explore the capacity of dental pulp stem cells (DPSCs) and platelet-rich plasma (PRP) to regenerate dental pulp in canine mature permanent teeth. Pulpectomy with apical foramen enlarged to a #80 file was performed in 16 upper premolars of 4 beagle dogs. Four experimental groups were randomly established: (1) the blood clot group, (2) the autologous DPSCs group, (3) the PRP group, and (4) the DP + PRP group (a mixture of DPSCs and PRP). Four lower premolars without any further treatment after pulpectomy were used as the control group. All teeth were sealed with mineral trioxide aggregate and composite. Twelve weeks after transplantation, the teeth were subjected to radiographic and histologic examination. Twenty-four of 32 experimental root canals gained newly formed tissues. All canals with an introduction of a blood clot showed histologic evidence of vital tissue formation. Cementum-like and periodontal ligament-like tissues along the internal root canal walls were typical structures in most cases. There is no significant difference between groups with or without autologous DPSC transplantation (exact chi-square test, P < .05). New vital tissues can be regenerated in permanent canine teeth after pulpectomy and enlargement of the apical foramen. Histologically, transplantation of DPSCs and/or PRP into root canals showed no enhancement in new tissue formation compared with inducement of a blood clot into the root canals alone. Copyright © 2012 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  14. Characterization of stem and progenitor cells in the dental pulp of erupted and unerupted murine molars

    PubMed Central

    Balic, Anamaria; Aguila, H. Leonardo; Caimano, Melissa J.; Francone, Victor P.; Mina, Mina

    2010-01-01

    In the past few years there have been significant advances in the identification of putative stem cells also referred to as “mesenchymal stem cells” (MSC) in dental tissues including the dental pulp. It is thought that MSC in dental pulp share certain similarities with MSC isolated from other tissues. However, cells in dental pulp are still poorly characterized. This study focused on the characterization of progenitor and stem cells in dental pulps of erupted and unerupted mice molars. Our study showed that dental pulps from unerupted molars contain a significant number of cells expressing CD90+/CD45-, CD117+/CD45-, Sca-1+/CD45- and little if any CD45+ cells. Our in vitro functional studies showed that dental pulp cells from unerupted molars displayed extensive osteo-dentinogenic potential but were unable to differentiate into chondrocytes and adipocytes. Dental pulp from erupted molars displayed a reduced number of cells, contained higher percentage of CD45+ and lower percentage of cells expressing CD90+/CD45-, CD117+/CD45- as compared to unerupted molars. In vitro functional assays demonstrated the ability of a small fraction of cells to differentiate into odontoblasts, osteoblasts, adipocytes and chondrocytes. There was a significant reduction in the osteo-dentinogenic potential of the pulp cells derived from erupted molars compared to unerupted molars. Furthermore, the adipogenic and chondrogenic differentiation of pulp cells from erupted molars was dependent on a long induction period and infrequent. Based on these findings we propose that the dental pulp of the erupted molars contain a small population of multipotent cells, whereas the dental pulp of the unerupted molars does not contain multipotent cells but is enriched in osteo-dentinogenic progenitors engaged in the formation of coronal and radicular odontoblasts. PMID:20193787

  15. Regenerative potential of dental pulp mesenchymal stem cells harvested from high caries patient's teeth.

    PubMed

    Rajendran, Ramesh; Gopal, Sushruth; Masood, Huda; Vivek, Purushottam; Deb, Kaushik

    2013-01-01

    Dental pulp are known to contains stem cells or dentinogenic progenitors that are responsible for dentin repair. Dental pulp Stem cells from Human Exfoliated Deciduous teeth (SHED) represent a population of postnatal stem cells capable of extensive proliferation and multipotential or multilineage differentiations. This potential for tissue regeneration has become the current basis for dental pulp stem cell banking. Here, we have attempted to develop a protocol for harvesting stem cells from patients with High Caries tooth, which are most often electively discarded. We have characterized the stem cells with mesenchymal stem cell markers and have compared their potential to grow in culture, doubling times, and differentiate into different lineages, with normal bone marrow mesenchymal stem cells (MSCs). We observed that the MSCs from dental pulp grew faster, with lower doubling time, and had equal efficiency in differentiating to various lineages, when subjected to standard directed differentiation protocols. This paper establishes that discarded High Carries Tooth can be a good source for regenerative medicine and also could be a potential source for MSCs and dental pulp MSC banking.

  16. Spontaneous Differentiation of Dental Pulp stem cells on Dental polymers

    NASA Astrophysics Data System (ADS)

    Bherwani, Aneel; Suarato, Giulia; Qin, Sisi; Chang, Chung-Cheh; Akhavan, Aaron; Spiegel, Joseph; Jurukovski, Vladimir; Rafailovich, Miriam; Simon, Marcia

    2012-02-01

    Dental pulp stem cells were plated on two dentally relevant materials i.e. PMMA commonly used for denture and Titanium used for implants. In both cases, we probed for the role of surface interaction and substrate morphology. Different films of PMMA were spun cast directly onto Si wafers; PMMA fibers of different diameters were electro spun onto some of these substrates. Titanium metal was evaporated onto Si surfaces using an electron beam evaporator. In addition, on some surfaces, P4VP nanofibers were spun cast. DPSC were grown in alpha-MEM supplemented with 10% fetal bovine serum, 0.2mM L-ascorbic acid 2-phosphate, 2mm glutamine and 10mM beta-glycerol phosphate either with or without 10nM dexamethasone. After 21 days samples were examined using confocal microscopy of cells and by scanning electron microscopy (SEM) and Energy dispersive X-ray Analysis (EDAX). In the case of Titanium biomineralization was observed independent of dexamethasone, where the deposits were templated along the fibers. Minimal biomineralization was observed on flat Titanium and PMMA samples. Markers of osteogenesis and specific signaling pathways are being evaluated by RT-PCR, which are up regulated on each surface, to understand the fundamental manner in which surfaces interact with cell differentiation.

  17. Isolation and morphology of Stem Cells from Deciduous Tooth (SHED) and Human Dental Pulp Stem Cells (hDPSC)

    NASA Astrophysics Data System (ADS)

    Ariffin, Shahrul Hisham Zainal; Manogaran, Thanaletchumi; Abidin, Intan Zarina Zainol; Senafi, Sahidan; Wahab, Rohaya Megat Abdul

    2016-11-01

    Dental pulp is a tissue obtained from pulp chamber of deciduous and permanent tooth which contain stem cells. Stem cell isolation procedure is performed to obtain cells from tissue using enzymatic digestion. The aim of this study is to isolate and observe the morphology of stem cells during passage 0 and passage 3. Dental pulp from deciduous and permanent tooth was enzymatically digested using collagenase Type I and cells obtained were cultured in DMEM-KO that contains 10% fetal bovine serum, 1% antibiotic-antimycotic solution and 0.001× GlutaMax®. During culture, cell morphology was observed under the microscope on day 3, 16 and 33 and captured using cellB software. Giemsa staining was conducted on cells at passage 3. Cells attached at the bottom of the flask on day 3 and started forming small colonies. Cells became confluent after approximately 4 weeks. Both Stem Cells from Deciduous Tooth (SHED) and Human Dental Pulp Stem Cells (hDPSC) exhibited fibroblast-like morphology during passage 0 and passage 3. Meanwhile, Giemsa staining at passage 3 revealed single intact nucleus surrounded by fibroblastic cytoplasm structure. It can be concluded that SHED and hDPSC showed consistent fibroblast-like morphology throughout culture period.

  18. Role of CD146 Enrichment in Purification of Stem Cells Derived from Dental Pulp Polyp

    PubMed Central

    Tavangar, Maryam Sadat; Hosseini, Seyed-Mojtaba; Dehghani-Nazhvani, Ali; Monabati, Ahmad

    2017-01-01

    Introduction: Hyperplastic pulpitis (pulp polyp) tissues contains cells with stem cell properties similar to that of the dental pulp stem cells (DPSCs). It has also been shown that CD146 enrichment can homogenize the cultures of DPSCs and enhance the colony forming potentials of their cultures. This study determines whether CD146 enrichment can help purifying the stem cells from heterogeneous cultures of the pulp polyp derived stem cells (PPSCs). Methods and Materials: Healthy dental pulps and pulp polyp tissues were enzymatically digested and the harvested single cells were sorted according to the presence of CD146 marker. The sorted cells were seeded directly for colony forming unit (CFU) assays of the negative and positive portions. Flowcytometric antigen panel and differentiation assays were used to see if these cells conform with mesenchymal stems cells (MSCs) definition. Differences between the between groups was assessed using independent t-test. The level of significance was set at 0.05. Results: Normal pulp tissue derived cells formed higher colonies (42.5±16.8 per 104 cells) than the pulp polyp (17.75±8.9 per 104 cells) (P=0.015). The CD146 positive portion of the polyp derived cells formed an average of 91.5±29.7 per 104 cells per CFU. On the other hand, CD146 negative portion did not show any colonies (P<0.001). Both resources showed cells with flowcytometric antigen panel and differentiation potentials conforming to MSC definition. Conclusion: The entire CFU of PPSCs were formed within CD146 enriched portion. It seems that CD146 enrichment may reduce the number of possible fibroblasts of the pulp polyps and may further homogenize the culture of the PPSCs. PMID:28179933

  19. Strategies to rescue mesenchymal stem cells (MSCs) and dental pulp stem cells (DPSCs) from NK cell mediated cytotoxicity.

    PubMed

    Jewett, Anahid; Arasteh, Aida; Tseng, Han-Ching; Behel, Armin; Arasteh, Hobie; Yang, Wendy; Cacalano, Nicholas A; Paranjpe, Avina

    2010-03-31

    The aim of this paper is to study the function of allogeneic and autologous NK cells against Dental Pulp Stem Cells (DPSCs) and Mesenchymal Stem Cells (MSCs) and to determine the function of NK cells in a three way interaction with monocytes and stem cells. We demonstrate here that freshly isolated untreated or IL-2 treated NK cells are potent inducers of cell death in DPSCs and MSCs, and that anti-CD16 antibody which induces functional split anergy and apoptosis in NK cells inhibits NK cell mediated lysis of DPSCs and MSCs. Monocytes co-cultured with either DPSCs or MSCs decrease lysis of stem cells by untreated or IL-2 treated NK cells. Monocytes also prevent NK cell apoptosis thereby raising the overall survival and function of NK cells, DPSCs or MSCs. Both total population of monocytes and those depleted of CD16(+) subsets were able to prevent NK cell mediated lysis of MSCs and DPSCs, and to trigger an increased secretion of IFN-gamma by IL-2 treated NK cells. Protection of stem cells from NK cell mediated lysis was also seen when monocytes were sorted out from stem cells before they were added to NK cells. However, this effect was not specific to monocytes since the addition of T and B cells to stem cells also protected stem cells from NK cell mediated lysis. NK cells were found to lyse monocytes, as well as T and B cells. By increasing the release of IFN-gamma and decreasing the cytotoxic function of NK cells monocytes are able to shield stem cells from killing by the NK cells, resulting in an increased protection and differentiation of stem cells. More importantly studies reported in this paper indicate that anti-CD16 antibody can be used to prevent NK cell induced rejection of stem cells.

  20. Interferon Gamma-treated Dental Pulp Stem Cells Promote Human Mesenchymal Stem Cell Migration In Vitro.

    PubMed

    Strojny, Chelsee; Boyle, Michael; Bartholomew, Amelia; Sundivakkam, Premanand; Alapati, Satish

    2015-08-01

    Chronic inflammation disrupts dental pulp regeneration by disintegrating the recruitment process of progenitors for repair. Bone marrow-derived mesenchymal stem cells (BM-MSCs) share the common features with dental pulp stem cells (DPSCs). The aim of the study was to investigate the migration of BM-MSCs toward DPSCs in response to inflammatory chemoattractants. Additionally, our studies also delineated the signaling mechanisms from BM-MSCs in mediating the proliferation and differentiation of DPSCs in vitro. Human DPSCs and BM-MSCs between passages 2 and 4 were used and were grown in odontogenic differentiation medium. Mineralization was determined by alizarin red staining analysis. Migration was assessed using crystal violet staining in cells grown in Boyden chamber Transwell inserts (Corning Inc Foundation, Tewksbury, MA). The mineralization potential of DPSCs was evaluated using alkaline phosphatase activity assay. Real-time polymerase chain reaction analysis was performed to assess the gene expression profile of chemokine (C-X-C motif) ligand (Cxcl) 3, 5, 6, 10, 11, 12, 14, and 16; stromal cell-derived factor (SDF) α; vascular endothelial growth factor; and fibroblast growth factor. Interferon gamma (FN-γ) treatment significantly abrogated the differentiation potential of DPSCs as shown by using alizarin red and alkaline phosphatase activity analysis. An increase in the migration of BM-MSCs was documented when cocultured with IFN-γ-treated DPSCs. RNA expression studies showed an increase in the levels of Cxcl6 and Cxcl12 in BM-MSCs when cocultured with IFN-γ-treated DPSCs. Additionally, an up-regulation of proangiogenic factors vascular endothelial growth factor and fibroblast growth factor were observed in DPSCs exposed to IFN-γ. Our findings indicate that inflamed IFN-γ-treated DPSCs release factors (presumably Cxcl6 and 12) that contribute to the homing of MSCs. This model might provide a potential research tool for studying MSC-DPSC cross talk and

  1. Electroporation for Transfection and Differentiation of Dental Pulp Stem Cells

    PubMed Central

    Rabie, Bakr M.

    2013-01-01

    Abstract Target gene delivery is needed to induce cellular differentiation or a specific therapeutic effect. Electroporation is a relatively safe and simple technique to deliver nucleic acids to the cell that acts by rendering cells transiently permeable using short periods of high voltage. In stem cell research, human dental pulp stem cells (hDPSCS) are highly accessible, and they exhibit broad differentiation potential. Until now, no studies have attempted to optimize electroporation parameters for DPSCs with respect to transfection efficiency and viability. In this study, we aimed to optimize transfection of DPSCs through varying different electroporation parameters, including voltage, mode of pulsation, and the number of pulses. As positive control, we used commonly utilized the chemical transfection reagents Lipofectamine 2000 and FuGene 6. In addition, we used our newly optimized transfection conditions to transfect hDPSCs with a functional chondrogenic transgene. We obtained higher transfection efficiency and cell viability with these electroporation conditions compared to controls. The highest transfection efficiency (63.81±4.72%) was achieved with 100 V, 20 msec, one-pulse square-wave condition. Among chemical transfection groups, FuGene 6 showed the highest cell viability at all tested transfection ratios, while Lipofectamine 2000 showed the highest transfection efficiency (19.23±3.19%) using 1:1 DNA (μg):Lipofectamine (μL). Transfected DPSCs functionally expressed the transforming growth factor β-3 chondrogenic transgene on the mRNA level as detected by real-time polymerase chain reaction and on the protein level as detected by Western blot analysis. An increase in various chondrogenic markers was also found when studying mRNA expression in transfected cells. In conclusion, the results of our study demonstrate optimal electroporation and chemical transfection reagent conditions for hDPSCs, and, subsequently, we provide proof of concept for

  2. Fluorapatite-modified scaffold on dental pulp stem cell mineralization.

    PubMed

    Guo, T; Li, Y; Cao, G; Zhang, Z; Chang, S; Czajka-Jakubowska, A; Nör, J E; Clarkson, B H; Liu, J

    2014-12-01

    In previous studies, fluorapatite (FA) crystal-coated surfaces have been shown to stimulate the differentiation and mineralization of human dental pulp stem cells (DPSCs) in two-dimensional cell culture. However, whether the FA surface can recapitulate these properties in three-dimensional culture is still unknown. This study examined the differences in behavior of human DPSCs cultured on electrospun polycaprolactone (PCL) NanoECM nanofibers with or without the FA crystals. Under near-physiologic conditions, the FA crystals were synthesized on the PCL nanofiber scaffolds. The FA crystals were evenly distributed on the scaffolds. DPSCs were cultured on the PCL+FA or the PCL scaffolds for up to 28 days. Scanning electron microscope images showed that DPSCs attached well to both scaffolds after the initial seeding. However, it appeared that more multicellular aggregates formed on the PCL+FA scaffolds. After 14 days, the cell proliferation on the PCL+FA was slower than that on the PCL-only scaffolds. Interestingly, even without any induction of mineralization, from day 7, the upregulation of several pro-osteogenic molecules (dmp1, dspp, runx2, ocn, spp1, col1a1) was detected in cells seeded on the PCL+FA scaffolds. A significant increase in alkaline phosphatase activity was also seen on FA-coated scaffolds compared with the PCL-only scaffolds at days 14 and 21. At the protein level, osteocalcin expression was induced only in the DPSCs on the PCL+FA surfaces at day 21 and then significantly enhanced at day 28. A similar pattern was observed in those specimens stained with Alizarin red and Von Kossa after 21 and 28 days. These data suggest that the incorporation of FA crystals within the three-dimensional PCL nanofiber scaffolds provided a favorable extracellular matrix microenvironment for the growth, differentiation, and mineralization of human DPSCs. This FA-modified PCL nanofiber scaffold shows promising potential for future bone, dental, and orthopedic regenerative

  3. Biomimetic extracellular matrix mediated somatic stem cell differentiation: applications in dental pulp tissue regeneration.

    PubMed

    Ravindran, Sriram; George, Anne

    2015-01-01

    Dental caries is one of the most widely prevalent infectious diseases in the world. It affects more than half of the world's population. The current treatment for necrotic dental pulp tissue arising from dental caries is root canal therapy. This treatment results in loss of tooth sensitivity and vitality making it prone for secondary infections. Over the past decade, several tissue-engineering approaches have attempted regeneration of the dental pulp tissue. Although several studies have highlighted the potential of dental stem cells, none have transitioned into a clinical setting owing to limited availability of dental stem cells and the need for growth factor delivery systems. Our strategy is to utilize the intact ECM of pulp cells to drive lineage specific differentiation of bone marrow derived mesenchymal stem cells. From a clinical perspective, pulp ECM scaffolds can be generated using cell lines and patient specific somatic stem cells can be used for regeneration. Our published results have shown the feasibility of using pulp ECM scaffolds for odontogenic differentiation of non-dental mesenchymal cells. This focused review discusses the issues surrounding dental pulp tissue regeneration and the potential of our strategy to overcome these issues.

  4. Biomimetic extracellular matrix mediated somatic stem cell differentiation: applications in dental pulp tissue regeneration

    PubMed Central

    Ravindran, Sriram; George, Anne

    2015-01-01

    Dental caries is one of the most widely prevalent infectious diseases in the world. It affects more than half of the world's population. The current treatment for necrotic dental pulp tissue arising from dental caries is root canal therapy. This treatment results in loss of tooth sensitivity and vitality making it prone for secondary infections. Over the past decade, several tissue-engineering approaches have attempted regeneration of the dental pulp tissue. Although several studies have highlighted the potential of dental stem cells, none have transitioned into a clinical setting owing to limited availability of dental stem cells and the need for growth factor delivery systems. Our strategy is to utilize the intact ECM of pulp cells to drive lineage specific differentiation of bone marrow derived mesenchymal stem cells. From a clinical perspective, pulp ECM scaffolds can be generated using cell lines and patient specific somatic stem cells can be used for regeneration. Our published results have shown the feasibility of using pulp ECM scaffolds for odontogenic differentiation of non-dental mesenchymal cells. This focused review discusses the issues surrounding dental pulp tissue regeneration and the potential of our strategy to overcome these issues. PMID:25954205

  5. Dental stem cell therapy with calcium hydroxide in dental pulp capping.

    PubMed

    Ji, Young-Min; Jeon, Soung Hoo; Park, Joo-Young; Chung, Jong-Hoon; Choung, Yun-Hoon; Choung, Pill-Hoon

    2010-06-01

    Calcium hydroxide has been extensively and steadily used for direct pulp capping in modern clinical dentistry. As it was known to have potential to induce hard tissue repair, this chemical has been applied to the exposed dental pulp and the hard tissue is expected to be regenerated above the pulp. During the reparative process of exposed pulp, primary odontoblasts that were lost as a result of extensive damage are replaced with newly differentiated odontoblast-like cells. This process is known to follow the sequential steps of proliferation, migration, and differentiation of progenitor cells. This research will examine the relationship between calcium hydroxide and the recruitment, proliferation, and mineralization of postnatal dental stem cells, obtained from an immature dental tissue of beagle dogs. Immunocytochemical staining and reverse transcriptase-polymerase chain reaction were used to identify the putative stem cell markers. Immunoblot analysis, wound healing assay, cell migration assay, and alizarin red staining were used to evaluate proliferation, migration, and mineralization capacity of the calcium hydroxide-treated stem cells. As an in vivo study, a combination of calcium hydroxide and autologous dental pulp stem cells (DPSCs) was applied for the treatment of intentionally created tooth defects on the premolars and the molars in beagle dogs to observe dentin regeneration. Ex vivo expanded DPSCs and periodontal ligament stem cells expressed STRO-1 and CD146, the mesenchymal stem cell markers. It was evident that calcium hydroxide increased recruitment, migration, proliferation, and mineralization of the DPSCs and periodontal ligament stem cells. Such results are valuable for future availability of DPSCs, which are recently focused as the stem cell reservoir for regeneration of dentin upon tooth injury, as well as for elucidation of the role of calcium hydroxide in pulp capping therapy.

  6. [Comparison of sorting of fluorescently and magnetically labelled dental pulp stem cells].

    PubMed

    Kerényi, Farkas; Tarapcsák, Szabolcs; Hrubi, Edit; Baráthne, Szabó Ágnes; Hegedüs, Viktória; Balogh, Sára; Bágyi, Kinga; Varga, Gábor; Hegedüs, Csaba

    2016-03-01

    Stem cells are present in many tissues, such as dental pulp. Stem cells can be easily isolated from dental pulp because third molars are often removed from patients. Stem cells could be separated from the tissue derived heterogeneous cell population. There are two main methods to separate a cell type from the other ones: the fluorescence activated cell sorting (FACS) and the magnetic activated cell sorting (MACS). The aim of this study was to compare these methods' effect on cell surviving and population growth after sorting on dental pulp cells. The anti-STRO-1 antibody was used as primary antibody to specifically label stem cells. Two secondary antibodies were used: magnetic or fluorescent labelled. We sorted the cells by MACS or by FACS or by combination of both (MACS-FACS). Our results show that the effectivity of MACS and FACS sorting are comparable while of MACS-FACS was significantly higher (MACS 79.53 ± 5.78%, FACS 88.27 ± 3.70%, MACS-FACS 98.43 ± 0.67%). The cell surviving and the post-sorting population growth, on the contrary, are very different. The cell population is growing on first week after MACS but after FACS did not. Moreover, after MACS-FACS, on first week the cell number of population decreased. Taken together, our results suggest to use MACS instead of FACS, at least in case of sorting dental pulp stem cells with anti-STRO-1 antibody.

  7. Effects of hTERT immortalization on osteogenic and adipogenic differentiation of dental pulp stem cells.

    PubMed

    Ikbale, El-Ayachi; Goorha, Sarita; Reiter, Lawrence T; Miranda-Carboni, Gustavo A

    2016-03-01

    These data relate to the differentiation of human dental pulp stem cells (DPSC) and DPSC immortalized by constitutively expressing human telomerase reverse transcriptase (hTERT) through both osteogenic and adipogenic lineages (i.e. to make bone producing and fat producing cells from these dental pulp stem cells). The data augment another study to characterize immortalized DPSC for the study of neurogenetic "Characterization of neurons from immortalized dental pulp stem cells for the study of neurogenetic disorders" [1]. Two copies of one typical control cell line (technical replicates) were used in this study. The data represent the differentiation of primary DPSC into osteoblast cells approximately 60% more effectively than hTERT immortalized DPSC. Conversely, both primary and immortalized DPSC are poorly differentiated into adipocytes. The mRNA expression levels for both early and late adipogenic and osteogenic gene markers are shown.

  8. Mystery Inside the Tooth: The Dental Pulp Stem Cells

    PubMed Central

    Masthan, K.M.K.; Sankari, S. Leena; Babu, N. Aravindha; Gopalakrishnan, T.

    2013-01-01

    Stem cells are distinguished by their ability to differentiate into different types of cells in the body and to self-replicate. During the recent years, stem cells have been used extensively in the field of medicine for the repair and regeneration of defective tissues and organs. However, the knowledge on the stem cell technology is increasing quickly in all medical disciplines and it dictates the need for new protective approaches in all fields, which include reparative dentistry. Stem cell therapy constitutes a common challenge for dentists as well as for biotechnologists. The aim of this study was to review the knowledge which was related to stem cells and to consider the possibility of use of stem cell populations and their technology in the future clinical applications, to cure diseases like Parkinsonism, Juvenile diabetes, certain forms of cancer, spinal injuries and heart problems. PMID:23814752

  9. Role of angiogenesis in endodontics: contributions of stem cells and proangiogenic and antiangiogenic factors to dental pulp regeneration.

    PubMed

    Saghiri, Mohammad Ali; Asatourian, Armen; Sorenson, Christine M; Sheibani, Nader

    2015-06-01

    Dental pulp regeneration is a part of regenerative endodontics, which includes isolation, propagation, and re-transplantation of stem cells inside the prepared root canal space. The formation of new blood vessels through angiogenesis is mandatory to increase the survival rate of re-transplanted tissues. Angiogenesis is defined as the formation of new blood vessels from preexisting capillaries, which has great importance in pulp regeneration and homeostasis. Here the contribution of human dental pulp stem cells and proangiogenic and antiangiogenic factors to angiogenesis process and regeneration of dental pulp is reviewed. A search was performed on the role of angiogenesis in dental pulp regeneration from January 2005 through April 2014. The recent aspects of the relationship between angiogenesis, human dental pulp stem cells, and proangiogenic and antiangiogenic factors in regeneration of dental pulp were assessed. Many studies have indicated an intimate relationship between angiogenesis and dental pulp regeneration. The contribution of stem cells and mechanical and chemical factors to dental pulp regeneration has been previously discussed. Angiogenesis is an indispensable process during dental pulp regeneration. The survival of inflamed vital pulp and engineered transplanted pulp tissue are closely linked to the process of angiogenesis at sites of application. However, the detailed regulatory mechanisms involved in initiation and progression of angiogenesis in pulp tissue require investigation. Published by Elsevier Inc.

  10. Role of Angiogenesis in Endodontics: Contributions of Stem Cells and Proangiogenic and Antiangiogenic Factors to Dental Pulp Regeneration

    PubMed Central

    Saghiri, Mohammad Ali; Asatourian, Armen; Sorenson, Christine M.; Sheibani, Nader

    2016-01-01

    Introduction Dental pulp regeneration is a part of regenerative endodontics, which includes isolation, propagation, and re-transplantation of stem cells inside the prepared root canal space. The formation of new blood vessels through angiogenesis is mandatory to increase the survival rate of re-transplanted tissues. Angiogenesis is defined as the formation of new blood vessels from preexisting capillaries, which has great importance in pulp regeneration and homeostasis. Here the contribution of human dental pulp stem cells and proangiogenic and antiangiogenic factors to angiogenesis process and regeneration of dental pulp is reviewed. Methods A search was performed on the role of angiogenesis in dental pulp regeneration from January 2005 through April 2014. The recent aspects of the relationship between angiogenesis, human dental pulp stem cells, and proangiogenic and antiangiogenic factors in regeneration of dental pulp were assessed. Results Many studies have indicated an intimate relationship between angiogenesis and dental pulp regeneration. The contribution of stem cells and mechanical and chemical factors to dental pulp regeneration has been previously discussed. Conclusions Angiogenesis is an indispensable process during dental pulp regeneration. The survival of inflamed vital pulp and engineered transplanted pulp tissue are closely linked to the process of angiogenesis at sites of application. However, the detailed regulatory mechanisms involved in initiation and progression of angiogenesis in pulp tissue require investigation. PMID:25649306

  11. Dental pulp stem cells from traumatically exposed pulps exhibited an enhanced osteogenic potential and weakened odontogenic capacity.

    PubMed

    Wang, Yanping; Yan, Ming; Wang, Zhanwei; Wu, Jintao; Wang, Zilu; Zheng, Yangyu; Yu, Jinhua

    2013-11-01

    Traumatic pulp exposure can bring about some permanent damages to tooth tissues including dental pulps. This study was designed to evaluate the effects of traumatic pulp exposure on the osteo/odontogenic capacity of dental pulp stem cells (DPSCs). Rat incisors were artificially fractured and dental pulps were exposed to the oral environment for 48 h. Then, multi-colony-derived DPSCs from the injured pulps (iDPSCs) were isolated. Their osteo/odontogenic differentiation and the involvement of NF-κB pathway were subsequently investigated. iDPSCs presented a lower proliferative capacity than normal DPSCs (nDPSCs), as indicated by MTT and FCM assay. ALP levels in iDPSCs were significantly higher (P<0.01) than those in nDPSCs. Alizarin red staining revealed that iDPSCs exhibited an increased capacity of calcium deposition. Moreover, iDPSCs expressed stronger osteogenic markers (Runx2/RUNX2 and Ocn/OCN) and less odontogenic gene/protein (Dspp/DSP) than nDPSCs in vitro. In vivo transplantation showed that nDPSCs implants generated the typical dentine-pulp complex while all iDPSCs pellets formed the osteodentin-like tissues which were immunopositive for OCN. Mechanistically, iDPSCs expressed the higher levels of cytoplasmic phosphorylated IκBα/P65 and nuclear P65 than nDPSCs, indicating an active cellular NF-κB pathway in iDPSCs. After the inhibition of NF-κB pathway, the osteogenic potential in iDPSCs was significantly down-regulated while odontogenic differentiation was up-regulated, as indicated by the decreased Alp/Runx2/Ocn and uprised Dspp expression. Pulp exposure for 48 h decreased the odontogenic capacity and enhanced the osteogenic potential of DPSCs via the NF-κB signalling pathway. Copyright © 2013 Elsevier Ltd. All rights reserved.

  12. Imperative role of dental pulp stem cells in regenerative therapies: a systematic review.

    PubMed

    Kabir, Ramchandra; Gupta, Manish; Aggarwal, Avanti; Sharma, Deepak; Sarin, Anurag; Kola, Mohammed Zaheer

    2014-01-01

    Stem cells are primitive cells that can differentiate and regenerate organs in different parts of the body such as heart, bones, muscles and nervous system. This has been a field of great clinical interest with immense possibilities of using the stem cells in regeneration of human organ those are damaged due to disease, developmental defects and accident. The knowledge of stem cell technology is increasing quickly in all medical specialties and in dental field too. Stem cells of dental origin appears to hold the key to various cell-based therapies in regenerative medicine, but most avenues are in experimental stages and many procedures are undergoing standardization and validation. Long-term preservation of SHED cells or DPSC is becoming a popular consideration, similar to the banking of umbilical cord blood. Dental pulp stem cells (DPSCs) are the adult multipotent cells that reside in the cell rich zone of the dental pulp. The multipotent nature of these DPSCs may be utilized in both dental and medical applications. A systematic review of the literature was performed using various internet based search engines (PubMed, Medline Plus, Cochrane, Medknow, Ebsco, Science Direct, Hinari, WebMD, IndMed, Embase) using keywords like "dental pulp stem cells", "regeneration", "medical applications", "tissue engineering". DPSCs appears to be a promising innovation for the re-growth of tissues however, long term clinical studies need to be carried out that could establish some authentic guidelines in this perspective.

  13. Comparison of human mesenchymal stem cells derived from bone marrow, synovial fluid, adult dental pulp, and exfoliated deciduous tooth pulp.

    PubMed

    Isobe, Y; Koyama, N; Nakao, K; Osawa, K; Ikeno, M; Yamanaka, S; Okubo, Y; Fujimura, K; Bessho, K

    2016-01-01

    Populations of pluripotent stem cells were isolated from bone marrow, synovial fluid, adult dental pulp, and exfoliated deciduous teeth and their multipotentiality properties compared. Osteogenic, chondrogenic, adipogenic, and neurogenic differentiation potentials were examined. Bone marrow mesenchymal stem cells (BMMSCs) and synovial fluid-derived cells (SFCs) showed the highest levels of osteogenesis as expressed by alkaline phosphatase (ALP) activity (0.54±0.094 U/mg protein and 0.57±0.039 U/mg protein, respectively; P=0.60) and by osteocalcin (BGLAP; determined by real-time RT-PCR). SFCs showed the highest levels of chondrogenesis as expressed by ALP activity (1.75±0.097 U/mg protein) and of COL2A1 and COL10A1 by real-time PCR. In terms of adipogenesis, lipid vesicles were observed in the BMMSCs and SFCs. Dental pulp stem cells (DPSCs) and stem cells from human exfoliated deciduous teeth (SHED) exhibited neurogenesis potential, as shown by increases in expression of class III β-tubulin (TUBB3) and microtubule-associated protein 2 (MAP2) on RT-PCR. Variability was found in the differentiation potential corresponding to the tendency of the original tissue to differentiate. It is suggested that the cell type should be selected depending on the regenerative treatment regimen. Copyright © 2015 International Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.

  14. Imperative Role of Dental Pulp Stem Cells in Regenerative Therapies: A Systematic Review

    PubMed Central

    Kabir, Ramchandra; Gupta, Manish; Aggarwal, Avanti; Sharma, Deepak; Sarin, Anurag; Kola, Mohammed Zaheer

    2014-01-01

    Stem cells are primitive cells that can differentiate and regenerate organs in different parts of the body such as heart, bones, muscles and nervous system. This has been a field of great clinical interest with immense possibilities of using the stem cells in regeneration of human organ those are damaged due to disease, developmental defects and accident. The knowledge of stem cell technology is increasing quickly in all medical specialties and in dental field too. Stem cells of dental origin appears to hold the key to various cell-based therapies in regenerative medicine, but most avenues are in experimental stages and many procedures are undergoing standardization and validation. Long-term preservation of SHED cells or DPSC is becoming a popular consideration, similar to the banking of umbilical cord blood. Dental pulp stem cells (DPSCs) are the adult multipotent cells that reside in the cell rich zone of the dental pulp. The multipotent nature of these DPSCs may be utilized in both dental and medical applications. A systematic review of the literature was performed using various internet based search engines (PubMed, Medline Plus, Cochrane, Medknow, Ebsco, Science Direct, Hinari, WebMD, IndMed, Embase) using keywords like “dental pulp stem cells”, “regeneration”, “medical applications”, “tissue engineering”. DPSCs appears to be a promising innovation for the re-growth of tissues however, long term clinical studies need to be carried out that could establish some authentic guidelines in this perspective. PMID:24665194

  15. Dental pulp stem cells (DPSCs) differentiation study by confocal Raman microscopy

    NASA Astrophysics Data System (ADS)

    Salehi, H.; Collart-Dutilleul, P.-Y.; Gergely, C.; Cuisinier, F. J. G.

    2014-03-01

    Regenerative medicine brings a huge application for Mesenchymal stem cells such as Dental Pulp Stem Cells (DPSCs). Confocal Raman microscopy, a non-invasive, label free , real time and high spatial resolution imaging technique is used to study osteogenic differentiation of DPSCs. Integrated Raman intensities in the 2800-3000 cm-1 region (C-H stretching) and 960 cm-1 peak (phosphate PO4 3-) were collected. In Dental Pulp Stem Cells 21st day differentiated in buffer solution, phosphate peaks ν1 PO4 3- (first vibrational mode) at 960cm-1 and ν2 PO4 3- at 430cm-1 and ν4 PO4 3- at 585cm-1 are obviously present. Confocal Raman microscopy enables the detection of cell differentiation and it can be used to investigate clinical stem cell research.

  16. Multipotent Differentiation of Human Dental Pulp Stem Cells: a Literature Review.

    PubMed

    Nuti, N; Corallo, C; Chan, B M F; Ferrari, M; Gerami-Naini, B

    2016-10-01

    The advent of regenerative medicine has brought us the opportunity to regenerate, modify and restore human organs function. Stem cells, a key resource in regenerative medicine, are defined as clonogenic, self-renewing, progenitor cells that can generate into one or more specialized cell types. Stem cells have been classified into three main groups: embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs) and adult/postnatal stem cells (ASCs). The present review focused the attention on ASCs, which have been identified in many perioral tissues such as dental pulp, periodontal ligament, follicle, gingival, alveolar bone and papilla. Human dental pulp stem cells (hDPSCs) are ectodermal-derived stem cells, originating from migrating neural crest cells and possess mesenchymal stem cell properties. During last decade, hDPSCs have received extensive attention in the field of tissue engineering and regenerative medicine due to their accessibility and ability to differentiate in several cell phenotypes. In this review, we have carefully described the potential of hDPSCs to differentiate into odontoblasts, osteocytes/osteoblasts, adipocytes, chondrocytes and neural cells.

  17. Dental Pulp Stem Cells: Their Potential in Reinnervation and Angiogenesis by Using Scaffolds.

    PubMed

    Lambrichts, Ivo; Driesen, Ronald B; Dillen, Yörg; Gervois, Pascal; Ratajczak, Jessica; Vangansewinkel, Tim; Wolfs, Esther; Bronckaers, Annelies; Hilkens, Petra

    2017-09-01

    Dental pulp is a highly vascularized and innervated tissue containing a heterogeneous stem cell population with multilineage differentiation potential. Current endodontic treatments focus on the preservation of the pulp tissue and the regeneration of dental pulp after pathological insults. Human dental pulp stem cells (hDPSCs) are currently investigated as stem cell-based therapy for pulp regeneration and for peripheral nerve injury in which neurons and Schwann cells display limited regenerative capacity. We have developed a neuronal differentiation protocol for hDPSCs that requires neurosphere formation before neuronal maturation. Moreover, Schwann cell differentiation of hDPSCs in our group revealed that differentiated hDPSCs have acquired the ability to myelinate and guide neurites from dorsal root ganglia. Besides their dynamic differentiation capacity, hDPSCs were shown to exert a paracrine effect on neural and endothelial cells. Analysis of hDPSC conditioned medium revealed the secretion of a broad spectrum of growth factors including brain-derived neurotrophic factor, nerve growth factor, vascular endothelial growth factor, and glial-derived neurotrophic factor. Application of the conditioned medium to endothelial cells promoted cell migration and tubulogenesis, indicating a paracrine proangiogenic effect. This hypothesis was enforced by the enhanced formation of blood vessels in the chorioallantoic membrane assay in the presence of hDPSCs. In addition, transplantation of 3-dimensional-printed hydroxyapatite scaffolds containing peptide hydrogels and hDPSCs into immunocompromised mice revealed blood vessel ingrowth, pulplike tissue formation, and osteodentin deposition suggesting osteogenic/odontogenic differentiation of hDPSCs. Future studies in our research group will focus on the pulp regeneration capacity of hDPSCs and the role of fibroblasts within the pulp extracellular matrix. Copyright © 2017 American Association of Endodontists. Published by

  18. Effect of Biodentine™ on the proliferation, migration and adhesion of human dental pulp stem cells.

    PubMed

    Luo, Zhirong; Li, Dongmei; Kohli, Meetu R; Yu, Qing; Kim, Syngcuk; He, Wen-Xi

    2014-04-01

    To investigate the proliferative, migratory and adhesion effect of Biodentine™, a new tricalcium silicate cement formulation, on the human dental pulp stem cells (hDPSCs). The cell cultures of hDPSCs obtained from impacted third molars were treated with Biodentine™ extract at four different concentrations: Biodentine™ 0.02mg/ml (BD 0.02), Biodentine™ 0.2mg/ml (BD 0.2), Biodentine™ 2mg/ml (BD 2) and Biodentine™ 20mg/ml (BD 20). Human dental pulp stem cells proliferation was evaluated by MTT (3-(4,5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide) and BrdU (5-bromo-2'-deoxyuridine) viability analysis at different times. Migration was investigated by microphotographs of wound healing and transwell migration assays. Adhesion assay was performed as well in presence of BD 0.2, BD 2 and blank control, while qRT-PCR (quantitative real-time reverse-transcriptase polymerase chain) was used for further analysis of the mRNA expression of chemokine and adhesion molecules in hDPSCs. Biodentine™ significantly increased proliferation of stem cells at BD 0.2 and BD 2 concentrations while decreased significantly at higher concentration of BD 20. BD 0.2 concentration had a statistically significant increased migration and adhesion abilities. In addition, qRT-PCR results showed that BD 0.2 could have effect on the mRNA expression of chemokines and adhesion molecules in human dental pulp stem cells. The data imply that Biodentine™ is a bioactive and biocompatible material capable of enhancing hDPSCs proliferation, migration and adhesion abilities. Biodentine™ when placed in direct contact with the pulp during pulp exposure can positively influence healing by enhancing the proliferation, migration and adhesion of human dental pulp stem cells. Copyright © 2014 Elsevier Ltd. All rights reserved.

  19. Cell Surface Proteome of Dental Pulp Stem Cells Identified by Label-Free Mass Spectrometry

    PubMed Central

    Niehage, Christian; Karbanová, Jana; Steenblock, Charlotte

    2016-01-01

    Multipotent mesenchymal stromal cells (MSCs) are promising tools for regenerative medicine. They can be isolated from different sources based on their plastic-adherence property. The identification of reliable cell surface markers thus becomes the Holy Grail for their prospective isolation. Here, we determine the cell surface proteomes of human dental pulp-derived MSCs isolated from single donors after culture expansion in low (2%) or high (10%) serum-containing media. Cell surface proteins were tagged on intact cells using cell impermeable, cleavable sulfo-NHS-SS-biotin, which allows their enrichment by streptavidin pull-down. For the proteomic analyses, we first compared label-free methods to analyze cell surface proteomes i.e. composition, enrichment and proteomic differences, and we developed a new mathematical model to determine cell surface protein enrichment using a combinatorial gene ontology query. Using this workflow, we identified 101 cluster of differentiation (CD) markers and 286 non-CD cell surface proteins. Based on this proteome profiling, we identified 14 cell surface proteins, which varied consistently in abundance when cells were cultured under low or high serum conditions. Collectively, our analytical methods provide a basis for identifying the cell surface proteome of dental pulp stem cells isolated from single donors and its evolution during culture or differentiation. Our data provide a comprehensive cell surface proteome for the precise identification of dental pulp-derived MSC populations and their isolation for potential therapeutic intervention. PMID:27490675

  20. Novel Cryopreservation Method for the Effective Collection of Dental Pulp Stem Cells.

    PubMed

    Takebe, Yusuke; Tatehara, Seiko; Fukushima, Tatsuhiro; Tokuyama-Toda, Reiko; Yasuhara, Rika; Mishima, Kenji; Satomura, Kazuhito

    2017-03-17

    Dental pulp stem cells (DPSCs) are an attractive cell source for use in cell-based therapy, regenerative medicine, and tissue engineering because DPSCs have high cell proliferation ability and multi-differentiation capacity. However, several problems are associated with the collection and preservation of DPSCs for future cell-based therapy. In addition, the isolation of DPSCs for cryopreservation is time-consuming and expensive. In this study, we developed a novel cryopreservation method for dental pulp tissues that can isolate suitable DPSCs after thawing cryopreserved tissue. Using the novel cryopreservation method, dental pulp tissues were cultured on adhesion culture dishes for 5 days and then cryopreserved. After thawing, the cryopreserved dental pulp tissue fragments exhibited cell migration. We evaluated each property of DPSCs isolated using the novel cryopreservation method (DPSCs-NCM) and the explant method alone without cryopreservation (DPSCs-C). DPSCs-NCM had the same proliferation capacity as DPSCs-C. FACS analysis indicated that both DPSCs-NCM and DPSCs-C were positive for mesenchymal stem cell markers at the same level but negative for hematopoietic cell markers. Moreover, both DPSCs-NCM and DPSCs-C could differentiate into osteogenic, chondrogenic, and adipogenic cells during culture in each induction medium. These results suggest that DPSCs-NCM may be mesenchymal stem cells. Therefore, our novel method might allow for easy and less expensive cryopreservation of DPSCs, thereby providing suitable DPSCs for use in patients in future cell-based therapies.

  1. Isolation, Characterization, and Differentiation of Dental Pulp Stem Cells in Ferrets.

    PubMed

    Homayounfar, Negar; Verma, Prashant; Nosrat, Ali; El Ayachi, Ikbale; Yu, Zongdong; Romberg, Elaine; Huang, George T-J; Fouad, Ashraf F

    2016-03-01

    The ferret canine tooth has been introduced as a suitable model for studying dental pulp regeneration. The aim of this study was to isolate and characterize ferret dental pulp stem cells (fDPSCs) and their differentiation potential. Dental pulp stem cells were isolated from freshly extracted ferret canine teeth. The cells were examined for the expression of stem cell markers STRO-1, CD90, CD105, and CD146. The osteo/odontogenic and adipogenic differentiation potential of fDPSCs was evaluated. Osteogenic and odontogenic marker genes were evaluated using quantitative real-time polymerase chain reaction (qRT-PCR) on days 1, 4, and 8 after osteo/odontogenic induction of fDPSCs including dentin sialophosphoprotein (DSPP), dentin matrix protein-1, osteopontin, and alkaline phosphatase. Human dental pulp cells were used as the control. The results were analyzed using 3-way analysis of variance. fDPSCs were positive for STRO1, CD90, and CD105 and negative for CD146 markers with immunohistochemistry. fDPSCs showed strong osteogenic and weak adipogenic potential. The overall expression of DSPP was not significantly different between fDPSCs and human dental pulp cells. The expression of DSPP in osteo/odontogenic media was significantly higher in fDPSCs on day 4 (P < .01). The overall expression of dentin matrix protein-1, osteopontin, and alkaline phosphatase was significantly higher in fDPSCs (P = .0005). fDPSCs were positive for several markers of dental pulp stem cells resembling human DPSCs and appeared to show a stronger potential to differentiate to osteoblastic rather than odontoblastic lineage. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  2. Dental Pulp Stem Cells Differentiation Reveals New Insights in Oct4A Dynamics

    PubMed Central

    D'Aurizio, Federica; Puppato, Elisa; Pandolfi, Maura; Beltrami, Antonio Paolo; Cesselli, Daniela; Falini, Giuseppe; Beltrami, Carlo Alberto; Curcio, Francesco

    2012-01-01

    Although the role played by the core transcription factor network, which includes c-Myc, Klf4, Nanog, and Oct4, in the maintenance of embryonic stem cell (ES) pluripotency and in the reprogramming of adult cells is well established, its persistence and function in adult stem cells are still debated. To verify its persistence and clarify the role played by these molecules in adult stem cell function, we investigated the expression pattern of embryonic and adult stem cell markers in undifferentiated and fully differentiated dental pulp stem cells (DPSC). A particular attention was devoted to the expression pattern and intracellular localization of the stemness-associated isoform A of Oct4 (Oct4A). Our data demonstrate that: Oct4, Nanog, Klf4 and c-Myc are expressed in adult stem cells and, with the exception of c-Myc, they are significantly down-regulated following differentiation. Cell differentiation was also associated with a significant reduction in the fraction of DPSC expressing the stem cell markers CD10, CD29 and CD117. Moreover, a nuclear to cytoplasm shuttling of Oct4A was identified in differentiated cells, which was associated with Oct4A phosphorylation. The present study would highlight the importance of the post-translational modifications in DPSC stemness maintenance, by which stem cells balance self-renewal versus differentiation. Understanding and controlling these mechanisms may be of great importance for stemness maintenance and stem cells clinical use, as well as for cancer research. PMID:22844522

  3. Carious deciduous teeth are a potential source for dental pulp stem cells.

    PubMed

    Werle, Stefanie Bressan; Lindemann, Daniele; Steffens, Daniela; Demarco, Flávio Fernando; de Araujo, Fernando Borba; Pranke, Patrícia; Casagrande, Luciano

    2016-01-01

    The objectives of this study are to isolate, cultivate, and characterize stem cells from the pulp of carious deciduous teeth (SCCD) and compare them to those retrieved from sound deciduous teeth (SHED--stem cells from human exfoliated deciduous teeth). Cells were obtained of dental pulp collected from sound (n = 10) and carious (n = 10) deciduous human teeth. Rate of isolation, proliferation assay (0, 1, 3, 5, and 7 days), STRO-1, mesenchymal (CD29, CD73, and CD90) and hematopoietic surface marker expression (CD14, CD34, CD45, HLA-DR), and differentiation capacity were evaluated. Isolation success rates were 70 and 80 % from the carious and sound groups, respectively. SCCD and SHED presented similar proliferation rate. There were no statistical differences between the groups for the tested surface markers. The cells from sound and carious deciduous teeth were positive for CD29, CD73, and CD90 and negative for CD14, CD34, CD45, and HLA-DR and were capable of differentiating into osteogenic, chondrogenic, and adipogenic lineages. SCCD demonstrated a similar pattern of proliferation, immunophenotypical characteristics, and differentiation ability as those obtained from sound deciduous teeth. These SCCD represent a feasible source of stem cells. Decayed deciduous teeth have been usually discarded once the pulp tissue could be damaged and the activity of stem cells compromised. These findings show that stem cells from carious deciduous teeth can be applicable source for cell-based therapies in tissue regeneration.

  4. Therapeutic potential of dental pulp stem cells in regenerative medicine: An overview.

    PubMed

    Verma, Kavita; Bains, Rhythm; Bains, Vivek Kumar; Rawtiya, Manjusha; Loomba, Kapil; Srivastava, Shrish Charan

    2014-05-01

    The purpose of this review is to gain an overview of the applications of the dental pulp stem cells (DPSCs) in the treatment of various medical diseases. Stem cells have the capacity to differentiate and regenerate into various tissues. DPSCs are the adult stem cells that reside in the cell rich zone of the dental pulp. These are the multipotent cells that can be explained by their embryonic origin from the neural crest. Owing to this multipotency, these DPSCs can be used in both dental and medical applications. A review of literature has been performed using electronic and hand-searching methods for the medical applications of DPSCs. On the basis of the available information, DPSCs appear to be a promising alternative for the regeneration of tissues and treatment of various diseases, although, long-term clinical trials and studies are needed to confirm their efficacy.

  5. Stem/progenitor cells from inflamed human dental pulp retain tissue regeneration potential

    PubMed Central

    Alongi, Dominick J; Yamaza, Takayoshi; Song, Yingjie; Fouad, Ashraf F; Romberg, Elaine E; Shi, Songtao; Tuan, Rocky S; Huang, George T-J

    2011-01-01

    Background Potent stem/progenitor cells have been isolated from normal human dental pulps termed dental pulp stem cells (DPSCs). However, it is unknown whether these cells exist in inflamed pulps (IPs). Aims To determine whether DPSCs can be identified and isolated from IPs; and if they can be successfully cultured, whether they retain tissue regeneration potential in vivo. Materials & methods DPSCs from freshly collected normal pulps (NPs) and IPs were characterized in vitro and their tissue regeneration potential tested using an in vivo study model. Results The immunohistochemical analysis showed that IPs expressed higher levels of mesenchymal stem cell markers STRO-1, CD90, CD105 and CD146 compared with NPs (p < 0.05). Flow cytometry analysis showed that DPSCs from both NPs and IPs expressed moderate to high levels of CD146, stage-specific embryonic antigen-4, CD73 and CD166. Total population doubling of DPSCs-IPs (44.6 ± 2.9) was lower than that of DPSCs-NPs (58.9 ± 2.5) (p < 0.05), and DPSCs-IPs appeared to have a decreased osteo/dentinogenic potential compared with DPSCs-NPs based on the mineral deposition in cultures. Nonetheless, DPSCs-IPs formed pulp/dentin complexes similar to DPSCs-NPs when transplanted into immunocompromised mice. Conclusion DPSCs-IPs can be isolated and their mesenchymal stem cell marker profiles are similar to those from NPs. Although some stem cell properties of DPSCs-IPs were altered, cells from some samples remained potent in tissue regeneration in vivo. PMID:20465527

  6. Ionizing radiation induces senescence and differentiation of human dental pulp stem cells.

    PubMed

    Havelek, R; Soukup, T; Ćmielová, J; Seifrtová, M; Suchánek, J; Vávrová, J; Mokrý, J; Muthná, D; Řezáčová, M

    2013-01-01

    Head and neck cancer is one of the most common cancers in Europe. Many current anti-cancer treatments, including ionizing radiation, induce apoptosis via DNA damage. Unfortunately, such treatments are non-selective to cancer cells and produce similar toxicity in normal cells, including adult stem cells. One of the fundamental properties of an adult stem cell is that it does not have any tissue-specific structures that allow it to perform specialized functions. However, under certain stimuli, unspecialized adult stem cells can give rise to specialized cells to generate replacements for cells that are lost during one's life or due to injury or disease. Nevertheless, specialization of stem cells must be controlled by specific milieu and also initiated at the proper time, making the entire process beneficial for tissue recovery and maintaining it for a long time. In this paper we assess whether irradiated dental pulp stem cells have maintained open their options to mature into specialized cells, or whether they have lost their unspecialized (immature) state following irradiation. Our findings showed radiation-induced premature differentiation of dental pulp stem cells towards odonto-/osteoblast lineages in vitro. Matrix calcification was visualized from Day 6 or Day 9 following irradiation of cells expressing low or high levels of CD146, respectively.

  7. Effects of cryopreservation of intact teeth on the isolated dental pulp stem cells.

    PubMed

    Lee, Sheng-Yang; Chiang, Pao-Chang; Tsai, Yu-Hui; Tsai, Shih-Ying; Jeng, Jiiang-Huei; Kawata, Toshitsugu; Huang, Haw-Ming

    2010-08-01

    Human dental pulp stem cells (DPSCs) have been reported to be useful material for future regenerative medicine. Clinically, cryopreservation of intact teeth can successfully preserve the periodontal ligament for future autotransplantation; however, the effects of cryopreservation procedure on the properties of DPSCs are still unclear. The aim of this study was to test whether DPSCs isolated from cryopreserved teeth can express stem cell-specific markers. In this study, a novel programmable freezer coupled to a magnetic field was used to perform the cryopreservation experiments. The tested DPSCs were isolated from magnetically cryopreserved and non-cryopreserved fresh teeth with an enzyme digestion procedure. The success rate of isolation, growth curves, morphology, stem cell-specific markers, and the differentiation capacity of the isolated cells were evaluated and compared. The isolation rate of dental pulp cells from magnetically cryopreserved teeth was 73%. After culture for 5 generations, there was no significant difference in cell viability between cells isolated from magnetically cryopreserved teeth and those isolated from fresh teeth. There were also no visible differences between the 2 groups of dental pulp cells in morphology, expression of stem cell markers, or osteogenic and adipogenic differentiations. The results suggest that cryopreserved whole teeth can be used for autotransplantation and provide a viable source of DPSCs. Copyright 2010 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  8. Cytogenetic instability of dental pulp stem cell lines.

    PubMed

    Duailibi, Monica Talarico; Kulikowski, Leslie Domenici; Duailibi, Silvio Eduardo; Lipay, Monica Vannucci Nunes; Melaragno, Maria Isabel; Ferreira, Lydia Masako; Vacanti, Joseph Phillip; Yelick, Pamela Crotty

    2012-02-01

    Human adult stem cells (hASCs) offer a potentially renewable source of cell types that are easily isolated and rapidly expanded for use in regenerative medicine and cell therapies without the complicating ethical problems that are associated with embryonic stem cells. However, the eventual therapeutic use of hASCs requires that these cells and their derivatives maintain their genomic stability. There is currently a lack of systematic studies that are aimed at characterising aberrant chromosomal changes in cultured ASCs over time. However, the presence of mosaicism and accumulation of karyotypic abnormalities within cultured cell subpopulations have been reported. To investigate cytogenetic integrity of cultured human dental stem cell (hDSC) lines, we analysed four expanded hDSC cultures using classical G banding and fluorescent in situ hybridisation (FISH) with X chromosome specific probe. Our preliminary results revealed that about 70% of the cells exhibited karyotypic abnormalities including polyploidy, aneuploidy and ring chromosomes. The heterogeneous spectrum of abnormalities indicates a high frequency of chromosomal mutations that continuously arise upon extended culture. These findings emphasise the need for the careful analysis of the cytogenetic stability of cultured hDSCs before they can be used in clinical therapies.

  9. Adult human dental pulp stem cells promote blood-brain barrier permeability through vascular endothelial growth factor-a expression.

    PubMed

    Winderlich, Joshua N; Kremer, Karlea L; Koblar, Simon A

    2016-06-01

    Stem cell therapy is a promising new treatment option for stroke. Intravascular administration of stem cells is a valid approach as stem cells have been shown to transmigrate the blood-brain barrier. The mechanism that causes this effect has not yet been elucidated. We hypothesized that stem cells would mediate localized discontinuities in the blood-brain barrier, which would allow passage into the brain parenchyma. Here, we demonstrate that adult human dental pulp stem cells express a soluble factor that increases permeability across an in vitro model of the blood-brain barrier. This effect was shown to be the result of vascular endothelial growth factor-a. The effect could be amplified by exposing dental pulp stem cell to stromal-derived factor 1, which stimulates vascular endothelial growth factor-a expression. These findings support the use of dental pulp stem cell in therapy for stroke. © The Author(s) 2015.

  10. Deciphering the Epigenetic Code in Embryonic and Dental Pulp Stem Cells.

    PubMed

    Bayarsaihan, Dashzeveg

    2016-12-01

    A close cooperation between chromatin states, transcriptional modulation, and epigenetic modifications is required for establishing appropriate regulatory circuits underlying self-renewal and differentiation of adult and embryonic stem cells. A growing body of research has established that the epigenome topology provides a structural framework for engaging genes in the non-random chromosomal interactions to orchestrate complex processes such as cell-matrix interactions, cell adhesion and cell migration during lineage commitment. Over the past few years, the functional dissection of the epigenetic landscape has become increasingly important for understanding gene expression dynamics in stem cells naturally found in most tissues. Adult stem cells of the human dental pulp hold great promise for tissue engineering, particularly in the skeletal and tooth regenerative medicine. It is therefore likely that progress towards pulp regeneration will have a substantial impact on the clinical research. This review summarizes the current state of knowledge regarding epigenetic cues that have evolved to regulate the pluripotent differentiation potential of embryonic stem cells and the lineage determination of developing dental pulp progenitors.

  11. Deciphering the Epigenetic Code in Embryonic and Dental Pulp Stem Cells

    PubMed Central

    Bayarsaihan, Dashzeveg

    2016-01-01

    A close cooperation between chromatin states, transcriptional modulation, and epigenetic modifications is required for establishing appropriate regulatory circuits underlying self-renewal and differentiation of adult and embryonic stem cells. A growing body of research has established that the epigenome topology provides a structural framework for engaging genes in the non-random chromosomal interactions to orchestrate complex processes such as cell-matrix interactions, cell adhesion and cell migration during lineage commitment. Over the past few years, the functional dissection of the epigenetic landscape has become increasingly important for understanding gene expression dynamics in stem cells naturally found in most tissues. Adult stem cells of the human dental pulp hold great promise for tissue engineering, particularly in the skeletal and tooth regenerative medicine. It is therefore likely that progress towards pulp regeneration will have a substantial impact on the clinical research. This review summarizes the current state of knowledge regarding epigenetic cues that have evolved to regulate the pluripotent differentiation potential of embryonic stem cells and the lineage determination of developing dental pulp progenitors. PMID:28018144

  12. Human dental pulp stem cell behavior using natural nanotolith/bacterial cellulose scaffolds for regenerative medicine.

    PubMed

    Olyveira, Gabriel Molina; Acasigua, Gerson Arisoly Xavier; Costa, Ligia Maria Manzine; Scher, Cristiane Regina; Xavier Filho, Lauro; Pranke, Patricia Helena Lucas; Basmaji, Pierre

    2013-08-01

    Adhesion and Viability study with human dental pulp stem cell using natural nanotolith/bacterial cellulose scaffolds for regenerative medicine are presented at first time in this work. Nanotolith, are osteoinductors, i.e., they stimulate bone regeneration, enabling higher cells migration for bone tissue regeneration formation. This is mainly because nanotoliths are rich minerals present in the internal ear of bony fish. In addition, are part of a system which acts as a depth sensor and balance, acting as a sound vibrations detector and considered essential for the bone mineralization process, as in hydroxiapatites. Nanotoliths influence in bacterial cellulose was analyzed using transmission infrared spectroscopy (FTIR). Results shows that fermentation process and nanotoliths agglomeration decrease initial human dental pulp stem cell adhesion however tested bionanocomposite behavior has cell viability increase over time.

  13. Porcine tooth germ cell conditioned medium can induce odontogenic differentiation of human dental pulp stem cells.

    PubMed

    Wang, Yin-Xiong; Ma, Zhao-Feng; Huo, Na; Tang, Liang; Han, Chun; Duan, Yin-Zhong; Jin, Yan

    2011-05-01

    It is suggested that the differentiation of tooth-derived stem cells is modulated by the local microenvironment in which they reside. Previous studies have indicated that tooth germ cell-conditioned medium (TGC-CM) holds the potential to induce dental pulp stem cells (DPSCs) to differentiate into the odontogenic lineage. Nevertheless, human TGC-CM (hTGC-CM) is not feasible in practical application, so we conjectured that xenogenic TGC-CM might exert a similar influence on human dental stem cells. In this study, we chose swine as the xenogenic origin and compared the effect of porcine tooth germ cell-conditioned medium (pTGC-CM) with its human counterpart on human DPSCs. Morphological appearance, colony-forming assay, in vitro multipotential ability, protein and gene expression of the odontogenic phenotype and the in vivo differentiation capacity of DPSCs were evaluated. The results showed that pTGC-CM exerted a similar effect to hTGC-CM in inducing human DPSCs to present odontogenic changes, which were indicated by remarkable morphological changes, higher multipotential capability and the expression of some odontogenic markers in gene and protein levels. Besides, the in vivo results showed that pTGC-CM-treated DPSCs, similar to hTGC-CM-treated DPSCs, could form a more regular dentine-pulp complex. Our data provided the first evidence that pTGC-CM is able to exert almost the same effect on DPSCs with hTGC-CM. The observations suggest that the application of xenogenic TGC-CM may facilitate generating bioengineered teeth from tooth-derived stem cells in future.

  14. The response of human ectomesenchymal dental pulp stem cells to cisplatin treatment.

    PubMed

    Seifrtová, M; Havelek, R; Cmielová, J; Jiroutová, A; Soukup, T; Brůčková, L; Mokrý, J; English, D; Rezáčová, M

    2012-05-01

    To determine the response of dental pulp stem cells (DPSCs) to DNA-damaging cytostatic cisplatin and compare it with the response of normal human dermal fibroblasts (HDFs). Dental pulp stem cells were exposed to 5, 10, 20 or 40 μmol L(-1) of cisplatin. The proliferation of affected cells was assessed by a Z2 Counter and viability was assessed by means of a Vi-Cell XR using Trypan blue exclusion staining. Cell cycle analysis and induction of apoptosis were performed by flow cytometry. Induction of apoptosis was determined by monitoring the activities of caspases. The expression of proteins was detected by electrophoresis and Western blotting. The descriptive statistics of the results was analyzed by Student's t-test. Dental pulp stem cells had a greater genotoxic stress response to cisplatin compared to HDFs. All three main Mitogen-activated protein kinases (MAPK) families - extracellular signal-regulated kinases (ERK), c-Jun-N-terminal kinase (JNK) and p38 were activated after treatment of DPSCs with cisplatin. The activation of MAPK pathways was not observed in HDFs exposed to cisplatin. The exposure of DPSCs and HDFs to cisplatin provoked an increase in p53 and p21 expression and p53 phosphorylation of serine 15. Higher concentrations of cisplatin reduced the viability of DPSCs and HDFs and induced the activation of caspases 3/7 and 9. Dental pulp stem cells had a greater genotoxic stress response to cisplatin compared to HDFs. Cisplatin in higher concentrations triggered activation of MAPK and apoptosis in DPSCs but not in HDFs. © 2011 International Endodontic Journal.

  15. EDTA conditioning of dentine promotes adhesion, migration and differentiation of dental pulp stem cells.

    PubMed

    Galler, K M; Widbiller, M; Buchalla, W; Eidt, A; Hiller, K-A; Hoffer, P C; Schmalz, G

    2016-06-01

    To evaluate the effect of dentine conditioning on migration, adhesion and differentiation of dental pulp stem cells. Dentine discs prepared from extracted human molars were pre-treated with EDTA (10%), NaOCl (5.25%) or H2 O. Migration of dental pulp stem cells towards pre-treated dentine after 24 and 48 h was assessed in a modified Boyden chamber assay. Cell adhesion was evaluated indirectly by measuring cell viability. Expression of mineralization-associated genes (COL1A1, ALP, BSP, DSPP, RUNX2) in cells cultured on pre-treated dentine for 7 days was determined by RT-qPCR. Nonparametric statistical analysis was performed for cell migration and cell viability data to compare different groups and time-points (Mann-Whitney U-test, α = 0.05). Treatment of dentine with H2 O or EDTA allowed for cell attachment, which was prohibited by NaOCl with statistical significance (P = 0.000). Furthermore, EDTA conditioning induced cell migration towards dentine. The expression of mineralization-associated genes was increased in dental pulp cells cultured on dentine after EDTA conditioning compared to H2 O-pre-treated dentine discs. EDTA conditioning of dentine promoted the adhesion, migration and differentiation of dental pulp stem cells towards or onto dentine. A pre-treatment with EDTA as the final step of an irrigation protocol for regenerative endodontic procedures has the potential to act favourably on new tissue formation within the root canal. © 2015 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  16. Stage-specific embryonic antigen-4 identifies human dental pulp stem cells.

    PubMed

    Kawanabe, Noriaki; Murata, Satoko; Fukushima, Hiroaki; Ishihara, Yoshihito; Yanagita, Takeshi; Yanagita, Emmy; Ono, Mitsuaki; Kurosaka, Hiroshi; Kamioka, Hiroshi; Itoh, Tomoo; Kuboki, Takuo; Yamashiro, Takashi

    2012-03-10

    Embryonic stem cell-associated antigens are expressed in a variety of adult stem cells as well as embryonic stem cells. In the present study, we investigated whether stage-specific embryonic antigen (SSEA)-4 can be used to isolate dental pulp (DP) stem cells. DP cells showed plastic adherence, specific surface antigen expression, and multipotent differentiation potential, similar to mesenchymal stem cells (MSC). SSEA-4+ cells were found in cultured DP cells in vitro as well as in DP tissue in vivo. Flow cytometric analysis demonstrated that 45.5% of the DP cells were SSEA-4+. When the DP cells were cultured in the presence of all-trans-retinoic acid, marked downregulation of SSEA-3 and SSEA-4 and the upregulation of SSEA-1 were observed. SSEA-4+ DP cells showed a greater telomere length and a higher growth rate compared to ungated and SSEA-4- cells. A clonal assay demonstrated that 65.5% of the SSEA-4+ DP cells had osteogenic potential, and the SSEA-4+ clonal DP cells showed multilineage differentiation potential toward osteoblasts, chondrocytes, and neurons in vitro. In addition, the SSEA-4+ DP cells had the capacity to form ectopic bone in vivo. Thus, our results suggest that SSEA-4 is a specific cell surface antigen that can be used to identify DP stem cells.

  17. Dental pulp of the third molar: a new source of pluripotent-like stem cells.

    PubMed

    Atari, Maher; Gil-Recio, Carlos; Fabregat, Marc; García-Fernández, Dani; Barajas, Miguel; Carrasco, Miguel A; Jung, Han-Sung; Alfaro, F Hernández; Casals, Nuria; Prosper, Felipe; Ferrés-Padró, Eduard; Giner, Luis

    2012-07-15

    Dental pulp is particularly interesting in regenerative medicine because of the accessibility and differentiation potential of the tissue. Dental pulp has an early developmental origin with multi-lineage differentiation potential as a result of its development during childhood and adolescence. However, no study has previously identified the presence of stem cell populations with embryonic-like phenotypes in human dental pulp from the third molar. In the present work, we describe a new population of dental pulp pluripotent-like stem cells (DPPSCs) that were isolated by culture in medium containing LIF, EGF and PDGF. These cells are SSEA4(+), OCT3/4(+), NANOG(+), SOX2(+), LIN28(+), CD13(+), CD105(+), CD34(-), CD45(-), CD90(+), CD29(+), CD73(+), STRO1(+) and CD146(-), and they show genetic stability in vitro based on genomic analysis with a newly described CGH technique. Interestingly, DPPSCs were able to form both embryoid-body-like structures (EBs) in vitro and teratoma-like structures that contained tissues derived from all three embryonic germ layers when injected in nude mice. We examined the capacity of DPPSCs to differentiate in vitro into tissues that have similar characteristics to mesoderm, endoderm and ectoderm layers in both 2D and 3D cultures. We performed a comparative RT-PCR analysis of GATA4, GATA6, MIXL1, NANOG, OCT3/4, SOX1 and SOX2 to determine the degree of similarity between DPPSCs, EBs and human induced pluripotent stem cells (hIPSCs). Our analysis revealed that DPPSCs, hIPSC and EBs have the same gene expression profile. Because DPPSCs can be derived from healthy human molars from patients of different sexes and ages, they represent an easily accessible source of stem cells, which opens a range of new possibilities for regenerative medicine.

  18. Magnetic resonance imaging of human dental pulp stem cells in vitro and in vivo.

    PubMed

    Struys, T; Ketkar-Atre, A; Gervois, P; Leten, C; Hilkens, P; Martens, W; Bronckaers, A; Dresselaers, T; Politis, C; Lambrichts, I; Himmelreich, U

    2013-01-01

    Recent advances in stem cell research have shown the promising nature of mesenchymal stem cells as plausible candidates for cell-based regenerative medicine. Many studies reported the use of human dental pulp stem cells (hDPSCs), which possess self-renewal capacity, high proliferation potential, and the ability to undergo multilineage differentiation. Together with this therapeutic approach, development of effective, noninvasive and nontoxic imaging techniques for visualizing and tracking the cells in vivo is crucial for the evaluation and improvement of stem cell therapy. Magnetic resonance imaging (MRI) is one of the most powerful diagnostic imaging techniques currently available for in vivo diagnosis and has been proposed as the most attractive modality for monitoring stem cell migration. The aim of this study was to investigate the labeling efficiency of hDPSCs using superparamagnetic iron oxide (SPIO) particles in order to allow visualization using in vitro and in vivo MRI without influencing cellular metabolism. MRI and transmission electron microscopy (TEM) showed optimal uptake with low SPIO concentrations of 15 µg/ml in combination with 0.75 µg/ml poly-L-lysine (PLL) resulting in more than 13 pg iron/cell and an in vitro detection limit of 50 labeled cells/µl. Very low SPIO concentrations in the culture medium resulted in extremely high labeling efficiency not reported before. For these conditions, tetrazolium salt assays showed no adverse effects on cell viability. Furthermore, in vivo MRI was performed to detect labeled hDPSCs transplanted into the brain of Rag 2-γ C immune-deficient mice. Transplanted cells did not show any signs of tumorgenecity or teratoma formation during the studied time course. We have reported on a labeling and imaging strategy to visualize human dental pulp stem cells in vivo using MRI. These data provide a solid base to allow cell tracking in future regenerative studies in the brain longitudinally.

  19. Differentiation of Human Dental Pulp Stem Cells into Dopaminergic Neuron-like Cells in Vitro.

    PubMed

    Chun, So Young; Soker, Shay; Jang, Yu-Jin; Kwon, Tae Gyun; Yoo, Eun Sang

    2016-02-01

    We investigated the potential of human dental pulp stem cells (hDPSCs) to differentiate into dopaminergic neurons in vitro as an autologous stem cell source for Parkinson's disease treatment. The hDPSCs were expanded in knockout-embryonic stem cell (KO-ES) medium containing leukemia inhibitory factor (LIF) on gelatin-coated plates for 3-4 days. Then, the medium was replaced with KO-ES medium without LIF to allow the formation of the neurosphere for 4 days. The neurosphere was transferred into ITS medium, containing ITS (human insulin-transferrin-sodium) and fibronectin, to select for Nestin-positive cells for 6-8 days. The cells were then cultured in N-2 medium containing basic fibroblast growth factor (FGF), FGF-8b, sonic hedgehog-N, and ascorbic acid on poly-l-ornithine/fibronectin-coated plates to expand the Nestin-positive cells for up to 2 weeks. Finally, the cells were transferred into N-2/ascorbic acid medium to allow for their differentiation into dopaminergic neurons for 10-15 days. The differentiation stages were confirmed by morphological, immunocytochemical, flow cytometric, real-time PCR, and ELISA analyses. The expressions of mesenchymal stem cell markers were observed at the early stages. The expressions of early neuronal markers were maintained throughout the differentiation stages. The mature neural markers showed increased expression from stage 3 onwards. The percentage of cells positive for tyrosine hydroxylase was 14.49%, and the amount was 0.526 ± 0.033 ng/mL at the last stage. hDPSCs can differentiate into dopaminergic neural cells under experimental cell differentiation conditions, showing potential as an autologous cell source for the treatment of Parkinson's disease.

  20. Single CD271 marker isolates mesenchymal stem cells from human dental pulp.

    PubMed

    Alvarez, Ruth; Lee, Hye-Lim; Hong, Christine; Wang, Cun-Yu

    2015-12-18

    Mesenchymal stem cells (MSCs) are a promising tool in regenerative medicine due to their capacity to differentiate into multiple lineages. In addition to MSCs isolated from bone marrow (BMSCs), adult MSCs are isolated from craniofacial tissues including dental pulp tissues (DPs) using various stem cell surface markers. However, there has been a lack of consensus on a set of surface makers that are reproducibly effective at isolating putative multipotent dental mesenchymal stem cells (DMSCs). In this study, we used different combinations of surface markers (CD51/CD140α, CD271, and STRO-1/CD146) to isolate homogeneous populations of DMSCs from heterogeneous dental pulp cells (DPCs) obtained from DP and compared their capacity to undergo multilineage differentiation. Fluorescence-activated cell sorting revealed that 27.3% of DPCs were CD51(+)/CD140α(+), 10.6% were CD271(+), and 0.3% were STRO-1(+)/CD146(+). Under odontogenic conditions, all three subsets of isolated DMSCs exhibited differentiation capacity into odontogenic lineages. Among these isolated subsets of DMSCs, CD271(+) DMSCs demonstrated the greatest odontogenic potential. While all three combinations of surface markers in this study successfully isolated DMSCs from DPCs, the single CD271 marker presents the most effective stem cell surface marker for identification of DMSCs with high odontogenic potential. Isolated CD271(+) DMSCs could potentially be utilized for future clinical applications in dentistry and regenerative medicine.

  1. Dynamic hydrostatic pressure promotes differentiation of human dental pulp stem cells.

    PubMed

    Yu, V; Damek-Poprawa, M; Nicoll, S B; Akintoye, S O

    2009-09-04

    The masticatory apparatus absorbs high occlusal forces, but uncontrolled parafunctional or orthodontic forces damage periodontal ligament (PDL), cause pulpal calcification, pulp necrosis and tooth loss. Morphology and functional differentiation of connective tissue cells can be controlled by mechanical stimuli but effects of uncontrolled forces on intra-pulpal homeostasis and ability of dental pulp stem cells (DPSCs) to withstand direct external forces are unclear. Using dynamic hydrostatic pressure (HSP), we tested the hypothesis that direct HSP disrupts DPSC survival and odontogenic differentiation. DPSCs from four teenage patients were subjected to HSP followed by assessment of cell adhesion, survival and recovery capacity based on odontogenic differentiation, mineralization and responsiveness to bone morphogenetic protein-2 (BMP-2). HSP down-regulated DPSC adhesion and survival but promoted differentiation by increasing mineralization, in vivo hard tissue regeneration and BMP-2 responsiveness despite reduced cell numbers. HSP-treated DPSCs displayed enhanced odontogenic differentiation, an indication of favorable recovery from HSP-induced cellular stress.

  2. Isolation and characterization of equine dental pulp stem cells derived from Thoroughbred wolf teeth

    PubMed Central

    ISHIKAWA, Shingo; HORINOUCHI, Chie; MURATA, Daiki; MATSUZAKI, Shota; MISUMI, Kazuhiro; IWAMOTO, Yohei; KOROSUE, Kenji; HOBO, Seiji

    2016-01-01

    Mesenchymal stem cells (MSCs) are adult multipotent stem cells that are capable of self-renewal and differentiation into multiple cell lineages. Methods for cell therapy using MSCs have been developed in equine medicine. Recently, human dental pulp stem cells (DPSCs) have drawn much attention owing to their trophic factor producing ability and minimally invasive collection methods. However, there have been no reports on equine dental pulp-derived cells (eDPCs). Therefore, the aim of this study was to isolate and characterize the eDPCs from discarded wolf teeth. Plastic-adherent spindle-shaped cells were isolated from wolf teeth. The doubling time of the isolated eDPCs was approximately 1 day. Differentiation assays using induction medium eDPCs differentiated into osteogenic, chondrogenic and adipogenic lineages. The eDPCs expressed mesenchymal makers (CD11a/18, CD44, CD90 CD105 and MHC class I and II), but did not express hematopoietic markers (CD34 and CD45). Taken together, the results show that eDPCs can be isolated from discarded wolf teeth, and they satisfy the minimal criteria for MSCs. Thus, these eDPCs can be referred to as equine DPSCs (eDPSCs). These eDPSCs may become a new source for cell therapy. PMID:27818457

  3. Confocal Raman microscopy to monitor extracellular matrix during dental pulp stem cells differentiation

    NASA Astrophysics Data System (ADS)

    Salehi, Hamideh; Collart-Dutilleul, Pierre-Yves; Gergely, Csilla; Cuisinier, Frédéric J. G.

    2015-07-01

    Regenerative medicine brings promising applications for mesenchymal stem cells, such as dental pulp stem cells (DPSCs). Confocal Raman microscopy, a noninvasive technique, is used to study osteogenic differentiation of DPSCs. Integrated Raman intensities in the 2800 to 3000 cm-1 region (C-H stretching) and the 960 cm-1 peak (ν1 PO43-) were collected (to image cells and phosphate, respectively), and the ratio of two peaks 1660 over 1690 cm-1 (amide I bands) to measure the collagen cross-linking has been calculated. Raman spectra of DPSCs after 21 days differentiation reveal several phosphate peaks: ν1 (first stretching mode) at 960 cm-1, ν2 at 430 cm-1, and ν4 at 585 cm-1 and collagen cross-linking can also be calculated. Confocal Raman microscopy enables monitoring osteogenic differentiation in vitro and can be a credible tool for clinical stem cell based research.

  4. Epigenetic modulation of dental pulp stem cells: implications for regenerative endodontics.

    PubMed

    Duncan, H F; Smith, A J; Fleming, G J P; Cooper, P R

    2016-05-01

    Dental pulp stem cells (DPSCs) offer significant potential for use in regenerative endodontics, and therefore, identifying cellular regulators that control stem cell fate is critical to devising novel treatment strategies. Stem cell lineage commitment and differentiation are regulated by an intricate range of host and environmental factors of which epigenetic influence is considered vital. Epigenetic modification of DNA and DNA-associated histone proteins has been demonstrated to control cell phenotype and regulate the renewal and pluripotency of stem cell populations. The activities of the nuclear enzymes, histone deacetylases, are increasingly being recognized as potential targets for pharmacologically inducing stem cell differentiation and dedifferentiation. Depending on cell maturity and niche in vitro, low concentration histone deacetylase inhibitor (HDACi) application can promote dedifferentiation of several post-natal and mouse embryonic stem cell populations and conversely increase differentiation and accelerate mineralization in DPSC populations, whilst animal studies have shown an HDACi-induced increase in stem cell marker expression during organ regeneration. Notably, both HDAC and DNA methyltransferase inhibitors have also been demonstrated to dramatically increase the reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) for use in regenerative therapeutic procedures. As the regulation of cell fate will likely remain the subject of intense future research activity, this review aims to describe the current knowledge relating to stem cell epigenetic modification, focusing on the role of HDACi on alteration of DPSC phenotype, whilst presenting the potential for therapeutic application as part of regenerative endodontic regimens. © 2015 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  5. Regeneration of dental pulp following pulpectomy by fractionated stem/progenitor cells from bone marrow and adipose tissue.

    PubMed

    Ishizaka, Ryo; Iohara, Koichiro; Murakami, Masashi; Fukuta, Osamu; Nakashima, Misako

    2012-03-01

    Pulp stem/progenitor cells can induce complete pulp regeneration. However, due to the limited availability of pulp tissue with age, there is a need to examine other sources for fractions of side population (SP) cells. In the present investigation bone marrow and adipose tissues of the same individual were evaluated as alternate sources. Pulp CD31(-) SP cells have higher migration activity and higher expression of angiogenic/neurotrophic factors than bone marrow and adipose CD31(-) SP cells. Adipose tissue CD31(-) SP cell transplantation yielded the same amount of regenerated tissue as pulp derived cells. However, bone marrow CD31(-) SP cell transplantation yielded significantly less regenerated tissue in pulpectomized root canals in dogs. The rate of matrix formation was much higher in adipose CD31(-) SP cell transplantation compared to pulp CD31(-) SP cell transplantation on day 28. Microarray analysis demonstrated similar qualitative and quantitative patterns of mRNA expression characteristic of pulp in the regenerated tissues from all three cell sources. Expression of many angiogenic/neurotrophic factors in the transplanted cells demonstrated trophic effects. Our results demonstrate that bone marrow and adipose CD31(-) SP cells might be suitable alternative cell sources for pulp regeneration.

  6. Amniotic fluid-derived mesenchymal stem cells lead to bone differentiation when cocultured with dental pulp stem cells.

    PubMed

    De Rosa, Alfredo; Tirino, Virginia; Paino, Francesca; Tartaglione, Antonella; Mitsiadis, Thimios; Feki, Anis; d'Aquino, Riccardo; Laino, Luigi; Colacurci, Nicola; Papaccio, Gianpaolo

    2011-03-01

    Mesenchymal stem cells are present in many tissues of the human body, including amniotic fluid (AF) and dental pulp (DP). Stem cells of both AF and DP give rise to a variety of differentiated cells. In our experience, DP stem cells (DPSCs) display a high capacity to produce bone. Therefore, our aim was to investigate if AF-derived stem cells (AFSCs) were able to undergo bone differentiation in the presence of DPSCs. AFSCs were seeded under three different conditions: (i) cocultured with DPSCs previously differentiated into osteoblasts; (ii) cultured in the conditioned medium of osteoblast-differentiated DPSCs; (iii) cultured in the osteogenic medium supplemented with vascular endothelial growth factor and bone morphogenetic protein-2 (BMP-2). Results showed that AFSCs were positive for mesenchymal markers, and expressed high levels of Tra1-60, Tra1-80, BMPR1, BMPR2, and BMP-2. In contrast, AFSCs were negative for epithelial and hematopoietic/endothelial markers. When AFSCs were cocultured with DPSCs-derived osteoblasts, they differentiated into osteoblasts. A similar effect was observed when AFSCs were cultured in the presence of a conditioned medium originated from DPSCs. We found that osteoblasts derived from DPSCs released large amounts of BMP-2 and vascular endothelial growth factor into the culture medium and that those morphogens significantly upregulate RUNX-2 gene, stimulating osteogenesis. This study highlights the mechanisms of osteogenesis and strongly suggests that the combination of AFSCs with DPSCs may provide a rich source of soluble proteins useful for bone engineering purposes.

  7. Isolation and in vitro characterisation of dental pulp stem cells from natal teeth.

    PubMed

    Karaöz, Erdal; Doğan, Burcu Nur; Aksoy, Ayça; Gacar, Gülçin; Akyüz, Serap; Ayhan, Selda; Genç, Zehra Seda; Yürüker, Sinan; Duruksu, Gökhan; Demircan, Pinar Cetinalp; Sariboyaci, Ayla Eker

    2010-01-01

    Dental pulp stem cells were primarily derived from the pulp tissues of exfoliated deciduous teeth, primary incisors and permanent third molar teeth. The aim of this study was to isolate and extensively characterise SCs derived from human natal dental pulp (hNDP). For characterisation, proliferation capacity, phenotypic properties, ultrastructural and differentiation characteristics and gene expression profiles were utilised. A comparison was done between the properties of NDP-SCs and the properties of mesenchymal stem cells (MSCs) from bone marrow (BM) of the human. Stem cells isolated from hNDP and hBM were analysed by flow cytometry, reverse transcriptase-PCR, Real Time-PCR, and immunocytochemistry. Both cell lines were directionally differentiated towards adipogenic, osteogenic chondrogenic, myogenic and neurogenic lineages. hNDP-SCs and hBM-MSCs expressed CD13, CD44, CD90, CD146 and CD166, but not CD3, CD8, CD11b, CD14, CD15, CD19, CD33, CD34, CD45, CD117, and HLA-DR. Ultrastructural characteristics of hNDP-SCs showed more developed and metabolically active cells. hNDP-SCs and hBM-MSCs expressed some adipogenic (leptin, adipophilin and PPARgamma), myogenic (desmin, myogenin, myosinIIa, and alpha-SMA), neurogenic (gamma-enolase, MAP2a,b, c-fos, nestin, NF-H, NF-L, GFAP and betaIII tubulin), osteogenic (osteonectin, osteocalcin, osteopontin, Runx-2, and type I collagen) and chondrogenic (type II collagen, SOX9) markers without any stimulation towards differentiation under basal conditions. Embryonic stem cell markers Oct4, Rex-1, FoxD-3, Sox2, and Nanog were also identified. The differentiation potential of hNDP-SCs and hBM-MSCs to adipogenic, osteogenic, chondrogenic, myogenic and neurogenic was shown. This report described the first successful isolation and characterisation of hNDP-SCs.

  8. Investigation of dental pulp stem cells isolated from discarded human teeth extracted due to aggressive periodontitis.

    PubMed

    Sun, Hai-Hua; Chen, Bo; Zhu, Qing-Lin; Kong, Hui; Li, Qi-Hong; Gao, Li-Na; Xiao, Min; Chen, Fa-Ming; Yu, Qing

    2014-11-01

    Recently, human dental pulp stem cells (DPSCs) isolated from inflamed dental pulp tissue have been demonstrated to retain some of their pluripotency and regenerative potential. However, the effects of periodontal inflammation due to periodontitis and its progression on the properties of DPSCs within periodontally compromised teeth remain unknown. In this study, DPSCs were isolated from discarded human teeth that were extracted due to aggressive periodontitis (AgP) and divided into three experimental groups (Groups A, B and C) based on the degree of inflammation-induced bone resorption approaching the apex of the tooth root before tooth extraction. DPSCs derived from impacted or non-functional third molars of matched patients were used as a control. Mesenchymal stem cell (MSC)-like characteristics, including colony-forming ability, proliferation, cell cycle, cell surface antigens, multi-lineage differentiation capability and in vivo tissue regeneration potential, were all evaluated in a patient-matched comparison. It was found that STRO-1- and CD146-positive DPSCs can be isolated from human teeth, even in very severe cases of AgP. Periodontal inflammation and its progression had an obvious impact on the characteristics of DPSCs isolated from periodontally affected teeth. Although all the isolated DPSCs in Groups A, B and C showed decreased colony-forming ability and proliferation rate (P < 0.05), the decreases were not consistent with the degree of periodontitis. Furthermore, the cells did not necessarily show significantly diminished in vitro multi-differentiation potential. Only DPSCs from Group A and the Control group formed dentin-like matrix in vivo when cell-seeded biomaterials were transplanted directly into an ectopic transplantation model. However, when cell-seeded scaffolds were placed in the root fragments of human teeth, all the cells formed significant dentin- and pulp-like tissues. The ability of DPSCs to generate dental tissues decreased when the

  9. The Biomineralization of a Bioactive Glass-Incorporated Light-Curable Pulp Capping Material Using Human Dental Pulp Stem Cells

    PubMed Central

    Jun, Soo-Kyung; Lee, Hae-Hyoung

    2017-01-01

    The aim of this study was to investigate the biomineralization of a newly introduced bioactive glass-incorporated light-curable pulp capping material using human dental pulp stem cells (hDPSCs). The product (Bioactive® [BA]) was compared with a conventional calcium hydroxide-incorporated (Dycal [DC]) and a light-curable (Theracal® [TC]) counterpart. Eluates from set specimens were used for investigating the cytotoxicity and biomineralization ability, determined by alkaline phosphatase (ALP) activity and alizarin red staining (ARS). Cations and hydroxide ions in the extracts were measured. An hDPSC viability of less than 70% was observed with 50% diluted extract in all groups and with 25% diluted extract in the DC. Culturing with 12.5% diluted BA extract statistically lowered ALP activity and biomineralization compared to DC (p < 0.05), but TC did not (p > 0.05). Ca (~110 ppm) and hydroxide ions (pH 11) were only detected in DC and TC. Ionic supplement-added BA, which contained similar ion concentrations as TC, showed similar ARS mineralization compared to TC. In conclusion, the BA was similar to, yet more cytotoxic to hDPSCs than, its DC and TC. The BA was considered to stimulate biomineralization similar to DC and TC only when it released a similar amount of Ca and hydroxide ions. PMID:28232937

  10. The Biomineralization of a Bioactive Glass-Incorporated Light-Curable Pulp Capping Material Using Human Dental Pulp Stem Cells.

    PubMed

    Jun, Soo-Kyung; Lee, Jung-Hwan; Lee, Hae-Hyoung

    2017-01-01

    The aim of this study was to investigate the biomineralization of a newly introduced bioactive glass-incorporated light-curable pulp capping material using human dental pulp stem cells (hDPSCs). The product (Bioactive® [BA]) was compared with a conventional calcium hydroxide-incorporated (Dycal [DC]) and a light-curable (Theracal® [TC]) counterpart. Eluates from set specimens were used for investigating the cytotoxicity and biomineralization ability, determined by alkaline phosphatase (ALP) activity and alizarin red staining (ARS). Cations and hydroxide ions in the extracts were measured. An hDPSC viability of less than 70% was observed with 50% diluted extract in all groups and with 25% diluted extract in the DC. Culturing with 12.5% diluted BA extract statistically lowered ALP activity and biomineralization compared to DC (p < 0.05), but TC did not (p > 0.05). Ca (~110 ppm) and hydroxide ions (pH 11) were only detected in DC and TC. Ionic supplement-added BA, which contained similar ion concentrations as TC, showed similar ARS mineralization compared to TC. In conclusion, the BA was similar to, yet more cytotoxic to hDPSCs than, its DC and TC. The BA was considered to stimulate biomineralization similar to DC and TC only when it released a similar amount of Ca and hydroxide ions.

  11. [Human stem cells from apical papilla can regenerate dentin-pulp complex].

    PubMed

    Xiong, Huacui; Chen, Ke; Huang, Yibin; Liu, Caiqi

    2013-10-01

    To regenerate dentin-pulp complex by tissue engineering with human stem cells from apical papilla cells (SCAP) as the seed cells. SCAP was separated from from normal human impacted third molars with immature roots by outgrowth culture. The cells were then cultured in the differentiation medium for 3 weeks or in normal medium for 60 days, and analyzed for mineralization potential by Alizarin red staining. The osteo/odontogenic markers including alkaline phosphatase (ALP), bone sialoprotein (BSP), osteocalcin (OC) and dentin sialoprotein (DSP) were investigated by immunofluorescence staining and reverse transcription-polymerase chain reaction. The co-cultured mixture of SCAP and HA/TCP, or HA/TCP alone was implanted subcutaneously on the back of nude mice for 8 weeks, and the implants were collected and examined by HE and immunohistochemical staining. Round alizarin red-positive nodules formed in the isolated cells after cell culture in the differentiation medium for 3 weeks or in normal medium for 60 days with positive staining for osteo/odontogenic markers. SCAP with HA/TCP could regenerate pulp-dentin complex-like tissue in nude mice. The cells near the dentin-like tissue were positive for DSP. No mineral tissue was found in mice receiving HA/TCP implantation. SCAP may serve as a promising seed cell for dentin-pulp complex tissue engineering.

  12. Biocompatibility of Accelerated Mineral Trioxide Aggregate on Stem Cells Derived from Human Dental Pulp.

    PubMed

    Kulan, Pinar; Karabiyik, Ozge; Kose, Gamze T; Kargul, Betul

    2016-02-01

    The aim of this study was to evaluate the effects of several additives on the setting time and cytotoxicity of accelerated-set mineral trioxide aggregate (MTA) on stem cells of human dental pulp. ProRoot white MTA (WMTA) (Dentsply Tulsa Dental, Johnson City, TN) was mixed with various additives including distilled water, 2.5% disodium hydrogen phosphate (Na2HPO4) (Merck, Darmstadt, Germany), K-Y Jelly (Johnson & Johnson, Markham, ON, Canada), and 5% and 10% calcium chloride (CaCl2) (Merck). The setting times were evaluated using a Vicat apparatus (Alsa Lab, Istanbul, Turkey). Human dental pulp stem cells were isolated and seeded into 48-well plates at 2 × 10(3) cells per well and incubated with MTA samples for 24 hours, 3 days, and 7 days. Cell viability was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay. MTA mixed with 10% CaCl2 showed the lowest setting time (P < .05). According to the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium results on the 1st, 3rd, and 7th days, a statistically significant difference was found (P < .05) between MTA groups and the control group. MTA mixed with K-Y Jelly in all groups showed the lowest cell viability at all time points (P < .05). The cell viability of MTA mixed with distilled water, 5% CaCl2, 10% CaCl2, and Na2HPO4 increased significantly through time (P < .05). This in vitro study found MTA mixed with 5% and 10% CaCl2 and Na2HPO4 is biocompatible with dental pulp stem cells in terms of cell viability. Further in vitro and in vivo investigations are required to prove the clinical applications of MTA mixed with various additives.

  13. The Interplay of Dental Pulp Stem Cells and Endothelial Cells in an Injectable Peptide Hydrogel on Angiogenesis and Pulp Regeneration In Vivo

    PubMed Central

    Dissanayaka, Waruna Lakmal; Hargreaves, Kenneth M.; Jin, Lijian; Samaranayake, Lakshman P.

    2015-01-01

    Securing an adequate blood supply for the survival of cell transplants is critical for a successful outcome in tissue engineering. Interactions between endothelial and progenitor/stem cells are important for vascularization of regenerating tissue. Recently, self-assembling peptide nanofibers were described as a promising environment for pulp regeneration due to their synthetic nature and controlled physicochemical properties. In this study, the peptide hydrogel PuraMatrix™ was used as a scaffold system to investigate the role of dental pulp stem cells (DPSCs) in triggering angiogenesis and the potential for regenerating vascularized pulp in vivo. Human umbilical vein endothelial cells (HUVECs), DPSCs, or cocultures of both cell types were encapsulated in three-dimensional PuraMatrix. The peptide nanofiber microenvironment supported cell survival, cell migration, and capillary network formation in the absence of exogenous growth factors. DPSCs increased early vascular network formation by facilitating the migration of HUVECs and by increasing vascular endothelial growth factor (VEGF) expression. Both the DPSC-monoculture and coculture groups exhibited vascularized pulp-like tissue with patches of osteodentin after transplantation in mice. The cocultured groups exhibited more extracellular matrix, vascularization, and mineralization than the DPSC-monocultures in vivo. The DPSCs play a critical role in initial angiogenesis, whereas coordinated efforts by the HUVECs and DPSCs are required to achieve a balance between extracellular matrix deposition and mineralization. The findings of this study also highlighted the importance of a microenvironment that supports cell–cell interactions and cell migration, which contribute to successful dental pulp regeneration. PMID:25203774

  14. Hydrogen sulphide increases hepatic differentiation of human tooth pulp stem cells compared with human bone marrow stem cells.

    PubMed

    Okada, M; Ishkitiev, N; Yaegaki, K; Imai, T; Tanaka, T; Fukuda, M; Ono, S; Haapasalo, M

    2014-12-01

    To determine the differences in stem cell properties, in hepatic differentiation and in the effects of hydrogen sulphide (H2 S) on hepatic differentiation between human bone marrow stem cells (hBMC) and stem cells from human exfoliated primary tooth pulp (SHED). CD117(+) cells were magnetically separated and subjected to hepatic differentiation. CD117(+) cell lineages were characterized for transcription factors indicative of stem cells by qRT-PCR. For the last 9 days of the differentiation, the test cells were exposed to 0.1 ng mL(-1) H2 S. Immunocytochemistry and flow cytometry of albumin, alpha-fetoprotein and carbamoyl phosphate synthetase were carried out after differentiation. Urea concentration and glycogen synthesis were also determined. Genes expressed in SHED were also expressed in BMC. No difference in expression level of hepatic markers was shown by immunofluorescence. SHED showed more positive cells than hBMC (P < 0.01). H2 S increased the number of positive cells in both cultures (P < 0.01). Urea concentration and glycogen synthesis increased significantly after H2 S exposure (P < 0.001 and P < 0.05, respectively). Real-time PCR data were analysed by RT(2) profiler RT-PCR Array Data Analysis version 3.5 (Qiagen), and ELISA data were analysed by Bonferroni's multiple comparison using Windows spss version 16 (SPSS Inc, Chicago, IL, USA). Bonferroni's multiple comparison test was also carried out after angle transformation for the percentage data of flow cytometer using Windows spss(®) version 16 (SPSS Inc). Statistical significance was accepted at P < 0.05. Stem cells from human exfoliated primary tooth pulp and BMC have similar properties. The level of hepatic differentiation in SHED compared with BMC was the same or higher. H2 S increased the level of hepatic differentiation. © 2014 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  15. Multifaceted neuro-regenerative activities of human dental pulp stem cells for functional recovery after spinal cord injury.

    PubMed

    Yamamoto, Akihito; Sakai, Kiyoshi; Matsubara, Kohki; Kano, Fumiya; Ueda, Minoru

    2014-01-01

    Spinal cord injury (SCI) often leads to persistent functional deficits due to the loss of neurons and glia and to limited axonal regeneration after such injury. Recently, three independent groups have reported marked recovery of hindlimb locomotor function after the transplantation of human adult dental pulp stem cells (DPSCs) and stem cells from human exfoliated deciduous teeth (SHEDs) into rats or mice with acute, sub-acute or chronic SCI. This review summarizes the primary characteristics of human dental pulp stem cells and their therapeutic benefits for treating SCI. Experimental data from multiple preclinical studies suggest that pulp stem cells may promote functional recovery after SCI through multifaceted neuro-regenerative activities. Copyright © 2013 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.

  16. [Experiment on inducing human dental pulp stem cells into neural-like cells].

    PubMed

    He, Hui-xia; Jin, Yan; Shi, Jun-nan; Luo, Yu-qing; Zhou, Yan-ni; Peng, Zhi; Xu, Yu-he

    2007-08-01

    To explore the multi-differentiated capability of human dental pulp stem cells (hDPSCs) obtained by cell-clone culture approach and to determine the appropriate induced medium. The cloned isolation and expansion of hDPSCs were preinduced for 24 h, and were subsequently replaced with neural-inductive medium containing certain concentration of dimethylsulfoxide (DMSO), butylated hydroxyanisode (BHA), forskolin, P-mercaptoethanol (p-ME) and hydrocortisone for 4 days. Then induced cells were analyzed by morphological observation, immnocytochemical staining for non-specific esterase (NSE) and glial fibrillary acidic protein (GFAP) expression, RT-PCR for GFAP mRNA. Meanwhile, the uninduced hDPSCs were used as negative control. The morphology of induced cells changed at the initial 12 h, and displayed a typical neuron-like cells at 24 h. There was a gradual increase in the number of these neuronal differentiated cells with continuous induction. Furthermore, immnocytochemical staining showed that the induced cell expressed NSE and GFAP, two marked enzymes of neuron cell. The GFAP mRNA was also detected in induced cells by RT-PCR assay. In contrast, the uninduced cells maintained its original appearance and had no expression on them. hDPSCs may possess potential of multiple-differentiation and may differentiate into neuron-like cells on certain inductive condition.

  17. Characterization of dental pulp stem/stromal cells of Huntington monkey tooth germs.

    PubMed

    Snyder, Brooke R; Cheng, Pei-Hsun; Yang, Jinjing; Yang, Shang-Hsun; Huang, Anderson H C; Chan, Anthony W S

    2011-09-12

    Dental pulp stem/stromal cells (DPSCs) are categorized as adult stem cells (ASCs) that retain multipotent differentiation capabilities. DPSCs can be isolated from individuals at any age and are considered to be true personal stem cells, making DPSCs one of the potential options for stem cell therapy. However, the properties of DPSCs from individuals with an inherited genetic disorder, such as Huntington's disease (HD), have not been fully investigated. To examine if mutant huntingtin (htt) protein impacts DPSC properties, we have established DPSCs from tooth germ of transgenic monkeys that expressed both mutant htt and green fluorescent protein (GFP) genes (rHD/G-DPSCs), and from a monkey that expressed only the GFP gene (rG-DPSCs), which served as a control. Although mutant htt and oligomeric htt aggregates were overtly present in rHD/G-DPSCs, all rHD/G-DPSCs and rG-DPSCs shared similar characteristics, including self-renewal, multipotent differentiation capabilities, expression of stemness and differentiation markers, and cell surface antigen profile. Our results suggest that DPSCs from Huntington monkeys retain ASC properties. Thus DPSCs derived from individuals with genetic disorders such as HD could be a potential source of personal stem cells for therapeutic purposes.

  18. Dental Stem Cells in Pulp Regeneration: Near Future or Long Road Ahead?

    PubMed

    Hilkens, Petra; Meschi, Nastaran; Lambrechts, Paul; Bronckaers, Annelies; Lambrichts, Ivo

    2015-07-15

    Although regenerative endodontic procedures have yielded an impressive body of favorable outcomes, the treatment of necrotic immature permanent teeth in particular remains to be a challenge. Recent advances in dental stem cell (DSC) research have gained increasing insight in their regenerative potential and prospective use in the formation of viable dental tissues. Numerous studies have already reported successful dental pulp regeneration following application of dental pulp stem cells, stem cells from the apical papilla, or dental follicle precursor cells in different in vivo models. Next to responsive cells, dental tissue engineering also requires the support of an appropriate scaffold material, ranging from naturally occurring polymers to treated dentin matrix components. However, the routine use and banking of DSCs still holds some major challenges, such as culture-associated differences, patient-related variability, and the effects of culture medium additives. Only in-depth evaluation of these problems and the implementation of standardized models and protocols will effectively lead to better alternatives for patients who no longer benefit from current treatment protocols.

  19. Properties of Dental Pulp-derived Mesenchymal Stem Cells and the Effects of Culture Conditions.

    PubMed

    Kawashima, Nobuyuki; Noda, Sonoko; Yamamoto, Mioko; Okiji, Takashi

    2017-09-01

    Dental pulp mesenchymal stem cells (DPMSCs) highly express mesenchymal stem cell markers and possess the potential to differentiate into neural cells, osteoblasts, adipocytes, and chondrocytes. Thus, DPMSCs are considered suitable for tissue regeneration. The colony isolation method has commonly been used to collect relatively large amounts of heterogeneous DPMSCs. Homogenous DPMSCs can be isolated by fluorescence-activated cell sorting using antibodies against mesenchymal stem cell markers, although this method yields a limited number of cells. Both quality and quantity of DPMSCs are critical to regenerative therapy, and cell culture methods need to be improved. We thus investigated the properties of DPMSCs cultured with different methods. DPMSCs in a three-dimensional spheroid culture system, which is similar to the hanging drop culture for differentiation of embryonic stem cells, showed upregulation of odonto-/osteoblastic markers and mineralized nodule formation. This suggests that this three-dimensional spheroid culturing system for DPMSCs may be suitable for inducing hard tissues. We further examined the effect of cell culture density on the properties of DPMSCs because the properties of stem cells can be altered depending on the cell density. DPMSCs cultured under the confluent cell density condition showed slight downregulation of some mesenchymal stem cell markers compared with those under the sparse condition. The ability of DPMSCs to differentiate into hard tissue-forming cells was found to be enhanced in the confluent condition, suggesting that the confluent culture condition may not be suitable for maintaining the stemness of DPMSCs. When DPMSCs are to be used for hard tissue regeneration, dense followed by sparse cell culture conditions may be a better alternative strategy. Copyright © 2017 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  20. A short-term treatment with tumor necrosis factor-alpha enhances stem cell phenotype of human dental pulp cells.

    PubMed

    Ueda, Mayu; Fujisawa, Takuo; Ono, Mitsuaki; Hara, Emilio Satoshi; Pham, Hai Thanh; Nakajima, Ryu; Sonoyama, Wataru; Kuboki, Takuo

    2014-02-28

    During normal pulp tissue healing, inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α) or interleukins, act in the initial 48 hours (inflammatory phase) and play important roles not only as chemo-attractants of inflammatory cells and stem/progenitor cells but also in inducing a cascade of reactions toward tissue regeneration or reparative dentin formation or both. Previous reports have shown that inflammatory cytokines regulate the differentiation capacity of dental pulp stem/progenitor cells (DPCs), but none has interrogated the impact of these cytokines on the stem cell phenotype of stem/progenitor cells. This study investigated the effects of a short-term treatment with TNF-α on the stem cell phenotype and differentiation ability of human DPCs. An in vivo mouse model of pulp exposure was performed for analysis of expression of the mesenchymal stem cell marker CD146 in DPCs during the initial stage of inflammatory response. For in vitro studies, human DPCs were isolated and incubated with TNF-α for 2 days and passaged to eliminate TNF-α completely. Analysis of stem cell phenotype was performed by quantification of cells positive for mesenchymal stem cell markers SSEA-4 (stage-specific embryonic antigen 4) and CD146 by flow cytometry as well as by quantitative analysis of telomerase activity and mRNA levels of OCT-4 and NANOG. Cell migration, colony-forming ability, and differentiation toward odontogenesis and adipogenesis were also investigated. The pulp exposure model revealed a strong staining for CD146 during the initial inflammatory response, at 2 days after pulp exposure. In vitro experiments demonstrated that a short-term (2-day) treatment of TNF-α increased by twofold the percentage of SSEA-4+ cells. Accordingly, STRO-1, CD146, and SSEA-4 protein levels as well as OCT-4 and NANOG mRNA levels were also significantly upregulated upon TNF-α treatment. A short-term TNF-α treatment also enhanced DPC function, including the ability to

  1. Postnatal stem/progenitor cells derived from the dental pulp of adult chimpanzee

    PubMed Central

    Cheng, Pei-Hsun; Snyder, Brooke; Fillos, Dimitri; Ibegbu, Chris C; Huang, Anderson Hsien-Cheng; Chan, Anthony WS

    2008-01-01

    Background Chimpanzee dental pulp stem/stromal cells (ChDPSCs) are very similar to human bone marrow derived mesenchymal stem/stromal cells (hBMSCs) as demonstrated by the expression pattern of cell surface markers and their multipotent differentiation capability. Results ChDPSCs were isolated from an incisor and a canine of a forty-seven year old female chimpanzee. A homogenous population of ChDPSCs was established in early culture at a high proliferation rate and verified by the expression pattern of thirteen cell surface markers. The ChDPSCs are multipotent and were capable of differentiating into osteogenic, adipogenic and chondrogenic lineages under appropriate in vitro culture conditions. ChDPSCs also express stem cell (Sox-2, Nanog, Rex-1, Oct-4) and osteogenic (Osteonectin, osteocalcin, osteopontin) markers, which is comparable to reported results of rhesus monkey BMSCs (rBMSCs), hBMSCs and hDPSCs. Although ChDPSCs vigorously proliferated during the initial phase and gradually decreased in subsequent passages, the telomere length indicated that telomerase activity was not significantly reduced. Conclusion These results demonstrate that ChDPSCs can be efficiently isolated from post-mortem teeth of adult chimpanzees and are multipotent. Due to the almost identical genome composition of humans and chimpanzees, there is an emergent need for defining the new role of chimpanzee modeling in comparative medicine. Teeth are easy to recover at necropsy and easy to preserve prior to the retrieval of dental pulp for stem/stromal cells isolation. Therefore, the establishment of ChDPSCs would preserve and maximize the applications of such a unique and invaluable animal model, and could advance the understanding of cellular functions and differentiation control of adult stem cells in higher primates. PMID:18430234

  2. Concise Review: Dental Pulp Stem Cells: A Novel Cell Therapy for Retinal and Central Nervous System Repair.

    PubMed

    Mead, Ben; Logan, Ann; Berry, Martin; Leadbeater, Wendy; Scheven, Ben A

    2017-01-01

    Dental pulp stem cells (DPSC) are neural crest-derived ecto-mesenchymal stem cells that can relatively easily and non-invasively be isolated from the dental pulp of extracted postnatal and adult teeth. Accumulating evidence suggests that DPSC have great promise as a cellular therapy for central nervous system (CNS) and retinal injury and disease. The mode of action by which DPSC confer therapeutic benefit may comprise multiple pathways, in particular, paracrine-mediated processes which involve a wide array of secreted trophic factors and is increasingly regarded as the principal predominant mechanism. In this concise review, we present the current evidence for the use of DPSC to repair CNS damage, including recent findings on retinal ganglion cell neuroprotection and regeneration in optic nerve injury and glaucoma. Stem Cells 2017;35:61-67. © 2016 AlphaMed Press.

  3. TOOTH (The Open study Of dental pulp stem cell Therapy in Humans): Study protocol for evaluating safety and feasibility of autologous human adult dental pulp stem cell therapy in patients with chronic disability after stroke.

    PubMed

    Nagpal, Anjali; Kremer, Karlea L; Hamilton-Bruce, Monica A; Kaidonis, Xenia; Milton, Austin G; Levi, Christopher; Shi, Songtao; Carey, Leeanne; Hillier, Susan; Rose, Miranda; Zacest, Andrew; Takhar, Parabjit; Koblar, Simon A

    2016-07-01

    Stroke represents a significant global disease burden. As of 2015, there is no chemical or biological therapy proven to actively enhance neurological recovery during the chronic phase post-stroke. Globally, cell-based therapy in stroke is at the stage of clinical translation and may improve neurological function through various mechanisms such as neural replacement, neuroprotection, angiogenesis, immuno-modulation, and neuroplasticity. Preclinical evidence in a rodent model of middle cerebral artery ischemic stroke as reported in four independent studies indicates improvement in neurobehavioral function with adult human dental pulp stem cell therapy. Human adult dental pulp stem cells present an exciting potential therapeutic option for improving post-stroke disability. TOOTH (The Open study Of dental pulp stem cell Therapy in Humans) will investigate the use of autologous stem cell therapy for stroke survivors with chronic disability, with the following objectives: (a) determine the maximum tolerable dose of autologous dental pulp stem cell therapy; (b) define that dental pulp stem cell therapy at the maximum tolerable dose is safe and feasible in chronic stroke; and (c) estimate the parameters of efficacy required to design a future Phase 2/3 clinical trial. TOOTH is a Phase 1, open-label, single-blinded clinical trial with a pragmatic design that comprises three stages: Stage 1 will involve the selection of 27 participants with middle cerebral artery ischemic stroke and the commencement of autologous dental pulp stem cell isolation, growth, and testing in sequential cohorts (n = 3). Stage 2 will involve the transplantation of dental pulp stem cell in each cohort of participants with an ascending dose and subsequent observation for a 6-month period for any dental pulp stem cell-related adverse events. Stage 3 will investigate the neurosurgical intervention of the maximum tolerable dose of autologous dental pulp stem cell followed by 9 weeks of intensive task

  4. An extracellular matrix culture system for induced pluripotent stem cells derived from human dental pulp cells.

    PubMed

    Chen, Y; Zheng, Y-L; Qiu, D-B; Sun, Y-P; Kuang, S-J; Xu, Y; He, F; Gong, Y-H; Zhang, Z-G

    2015-11-01

    Induced pluripotent stem cells (iPSCs) have emerged as a promising tool for treating incurable diseases. The current challenges are to avoid potential xenopathogenic transmission and immune rejection potentially caused by exposure of iPSCs to animal-derived products. In addition, an efficient feeder cell-free culture condition will be required for minimizing batch-to-batch variation and facilitating scale-up. Therefore, establishing an efficient extracellular matrix (ECM) culture system is considered as a prerequisite for the future clinical application of iPSC-based cell therapies. In this study, we evaluated the feasibility of culturing iPSCs in ECM derived from human dental pulp cells (hDCP). iPSCs growing in Matrigel were transferred to ECM or Matrigel and cultured in mTeSRTM1 medium. The number of adherent cells in the ECM group was higher than that in the Matrigel group after incubation for 8, 12, and 24 h, indicating that the ECM could enhance cell adherence. The adhesion of cells to ECM not only depends on simple physical attachment with ECM, but also mediated by fibronectin in the ECM. The hDPC-iPSCs showed orderly growth in the ECM, suggesting that the ECM could promote the growth and proliferation of hDPC-iPSCs. We also observed that stem cells growed along to avoid contact inhibition. The iPSCs maintained undifferentiated state when cultured in ECM when the iPSCs and ECM are of the same cell origin. ECM and mTeSRTM1, can both be used as new culture medium for iPSCs that facilitates the clinical application of iPSC-based cell therapies in the future.

  5. Biological Characteristics of Fluorescent Superparamagnetic Iron Oxide Labeled Human Dental Pulp Stem Cells

    PubMed Central

    Li, Ming-wei; Bai, Yu; Guo, Hui-hui

    2017-01-01

    Tracking transplanted stem cells is necessary to clarify cellular properties and improve transplantation success. In this study, we investigate the effects of fluorescent superparamagnetic iron oxide particles (SPIO) (Molday ION Rhodamine-B™, MIRB) on biological properties of human dental pulp stem cells (hDPSCs) and monitor hDPSCs in vitro and in vivo using magnetic resonance imaging (MRI). Morphological analysis showed that intracellular MIRB particles were distributed in the cytoplasm surrounding the nuclei of hDPSCs. 12.5–100 μg/mL MIRB all resulted in 100% labeling efficiency. MTT showed that 12.5–50 μg/mL MIRB could promote cell proliferation and MIRB over 100 μg/mL exhibited toxic effect on hDPSCs. In vitro MRI showed that 1 × 106 cells labeled with various concentrations of MIRB (12.5–100 μg/mL) could be visualized. In vivo MRI showed that transplanted cells could be clearly visualized up to 60 days after transplantation. These results suggest that 12.5–50 μg/mL MIRB is a safe range for labeling hDPSCs. MIRB labeled hDPSCs cell can be visualized by MRI in vitro and in vivo. These data demonstrate that MIRB is a promising candidate for hDPSCs tracking in hDPSCs based dental pulp regeneration therapy. PMID:28298928

  6. Exosomes from Human Dental Pulp Stem Cells Suppress Carrageenan-Induced Acute Inflammation in Mice.

    PubMed

    Pivoraitė, Ugnė; Jarmalavičiūtė, Akvilė; Tunaitis, Virginijus; Ramanauskaitė, Giedrė; Vaitkuvienė, Aida; Kašėta, Vytautas; Biziulevičienė, Genė; Venalis, Algirdas; Pivoriūnas, Augustas

    2015-10-01

    The primary goal of this study was to examine the effects of human dental pulp stem cell-derived exosomes on the carrageenan-induced acute inflammation in mice. Exosomes were purified by differential ultracentrifugation from the supernatants of stem cells derived from the dental pulp of human exfoliated deciduous teeth (SHEDs) cultivated in serum-free medium. At 1 h post-carrageenan injection, exosomes derived from supernatants of 2 × 10(6) SHEDs were administered by intraplantar injection to BALB/c mice; 30 mg/kg of prednisolone and phosphate-buffered saline (PBS) were used as positive and negative controls, respectively. Edema was measured at 6, 24, and 48 h after carrageenan injection. For the in vivo imaging experiments, AngioSPARK750, Cat B 750 FAST, and MMPSense 750 FAST were administered into the mouse tail vein 2 h post-carrageenan injection. Fluorescence images were acquired at 6, 24, and 48 h after edema induction by IVIS Spectrum in vivo imaging system. Exosomes significantly reduced the carrageenan-induced edema at all the time points studied (by 39.5, 41.6, and 25.6% at 6, 24, and 48 h after injection, respectively), to similar levels seen with the positive control (prednisolone). In vivo imaging experiments revealed that, both exosomes and prednisolone suppress activities of cathepsin B and matrix metalloproteinases (MMPs) at the site of carrageenan-induced acute inflammation, showing more prominent effects of prednisolone at the early stages, while exosomes exerted their suppressive effects gradually and at later time points. Our study demonstrates for the first time that exosomes derived from human dental pulp stem cells suppress carrageenan-induced acute inflammation in mice.

  7. Proteome of Human Stem Cells from Periodontal Ligament and Dental Pulp

    PubMed Central

    Sulpizio, Marilisa; Di Giuseppe, Fabrizio; Pierdomenico, Laura; Marchisio, Marco; Giancola, Raffaella; Giammaria, Gianluigi; Miscia, Sebastiano; Caputi, Sergio; Di Ilio, Carmine; Angelucci, Stefania

    2013-01-01

    Background Many adult tissues contain a population of stem cells with the ability to regenerate structures similar to the microenvironments from which they are derived in vivo and represent a promising therapy for the regeneration of complex tissues in the clinical disorder. Human adult stem cells (SCs) including bone marrow stem cells (BMSCs), dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PDLSCs) have been characterized for their high proliferative potential, expression of characteristic SC-associated markers and for the plasticity to differentiate in different lineage in vitro. Methodology/Principal Findings The aim of this study is to define the molecular features of stem cells from oral tissue by comparing the proteomic profiles obtained with 2-DE followed by MALDI-TOF/TOF of ex-vivo cultured human PDLSCs, DPSCs and BMSCs. Our results showed qualitative similarities in the proteome profiles among the SCs examined including some significant quantitative differences. To enrich the knowledge of oral SCs proteome we performed an analysis in narrow range pH 4–7 and 6–9, and we found that DPSCs vs PDLSCs express differentially regulated proteins that are potentially related to growth, regulation and genesis of neuronal cells, suggesting that SCs derived from oral tissue source populations may possess the potential ability of neuronal differentiation which is very consistent with their neural crest origin. Conclusion/Significance This study identifies some differentially expressed proteins by using comparative analysis between DPSCs and PDLSCs and BMSCs and suggests that stem cells from oral tissue could have a different cell lineage potency compared to BMSCs. PMID:23940696

  8. Modulation of Dental Pulp Stem Cell Odontogenesis in a Tunable PEG-Fibrinogen Hydrogel System.

    PubMed

    Lu, Qiqi; Pandya, Mirali; Rufaihah, Abdul Jalil; Rosa, Vinicius; Tong, Huei Jinn; Seliktar, Dror; Toh, Wei Seong

    2015-01-01

    Injectable hydrogels have the great potential for clinical translation of dental pulp regeneration. A recently developed PEG-fibrinogen (PF) hydrogel, which comprises a bioactive fibrinogen backbone conjugated to polyethylene glycol (PEG) side chains, can be cross-linked after injection by photopolymerization. The objective of this study was to investigate the use of this hydrogel, which allows tuning of its mechanical properties, as a scaffold for dental pulp tissue engineering. The cross-linking degree of PF hydrogels could be controlled by varying the amounts of PEG-diacrylate (PEG-DA) cross-linker. PF hydrogels are generally cytocompatible with the encapsulated dental pulp stem cells (DPSCs), yielding >85% cell viability in all hydrogels. It was found that the cell morphology of encapsulated DPSCs, odontogenic gene expression, and mineralization were strongly modulated by the hydrogel cross-linking degree and matrix stiffness. Notably, DPSCs cultured within the highest cross-linked hydrogel remained mostly rounded in aggregates and demonstrated the greatest enhancement in odontogenic gene expression. Consistently, the highest degree of mineralization was observed in the highest cross-linked hydrogel. Collectively, our results indicate that PF hydrogels can be used as a scaffold for DPSCs and offers the possibility of influencing DPSCs in ways that may be beneficial for applications in regenerative endodontics.

  9. Cytotoxic effects of glass ionomer cements on human dental pulp stem cells correlate with fluoride release.

    PubMed

    Kanjevac, Tatjana; Milovanovic, Marija; Volarevic, Vladislav; Lukic, Miodrag L; Arsenijevic, Nebojsa; Markovic, Dejan; Zdravkovic, Nebojsa; Tesic, Zivoslav; Lukic, Aleksandra

    2012-01-01

    Glass ionomer cements (GICs) are commonly used as restorative materials. Responses to GICs differ among cell types and it is therefore of importance to thoroughly investigate the influence of these restorative materials on pulp stem cells that are potential source for dental tissue regeneration. Eight biomaterials were tested: Fuji I, Fuji II, Fuji VIII, Fuji IX, Fuji Plus, Fuji Triage, Vitrebond and Composit. We compared their cytotoxic activity on human dental pulp stem cells (DPSC) and correlated this activity with the content of Fluoride, Aluminium and Strontium ions in their eluates. Elution samples of biomaterials were prepared in sterile tissue culture medium and the medium was tested for toxicity by an assay of cell survival/proliferation (MTT test) and apoptosis (Annexin V FITC Detection Kit). Concentrations of Fluoride, Aluminium and Strontium ions were tested by appropriate methods in the same eluates. Cell survival ranged between 79.62% (Fuji Triage) to 1.5% (Fuji Plus) and most dead DPSCs were in the stage of late apoptosis. Fluoride release correlated with cytotoxicity of GICs, while Aluminium and Strontium ions, present in significant amount in eluates of tested GICs did not. Fuji Plus, Vitrebond and Fuji VIII, which released fluoride in higher quantities than other GICs, were highly toxic to human DPSCs. Opposite, low levels of released fluoride correlated to low cytotoxic effect of Composit, Fuji I and Fuji Triage.

  10. The effects of LPS on adhesion and migration of human dental pulp stem cells in vitro.

    PubMed

    Li, Dongmei; Fu, Lei; Zhang, Yaqing; Yu, Qing; Ma, Fengle; Wang, Zhihua; Luo, Zhirong; Zhou, Zeyuan; Cooper, Paul R; He, Wenxi

    2014-10-01

    The aim of the present study was to investigate the effects of lipopolysaccharide (LPS) on the migration and adhesion of human dental pulp stem cells (hDPSCs) and the associated intracellular signalling pathways. hDPSCs obtained from impacted third molars were exposed to LPS and in vitro cell adhesion and migration were evaluated. The effects of LPS on gene expression of adhesion molecules and chemotactic factors were investigated using quantitative real-time reverse-transcriptase polymerase chain (qRT-PCR). The potential involvement of nuclear factor NF-kappa-B (NF-κB) or mitogen-activated protein kinase (MAPK) signalling pathways in the migration and adhesion of hDPSCs induced by LPS was assessed using a transwell cell migration assay and qRT-PCR. LPS promoted the adhesion of hDPSCs at 1μg/mL and 10μg/mL concentrations, 1μg/mL LPS showing the greater effect. Transwell cell migration assay demonstrated that LPS increased migration of hDPSCs at 1μg/mL concentration while decreasing it significantly at 10μg/mL. The mRNA expressions of adhesion molecules and chemotactic factors were enhanced significantly after stimulation with 1μg/mL LPS. Specific inhibitors for NF-κB and extracellular signal regulated kinases (ERK), c-Jun N-terminal kinase (JNK), and P38, markedly antagonised LPS-induced adhesion and migration of hDPSCs and also significantly abrogated LPS-induced up-regulation of adhesion molecules and chemotactic factors. In addition, specific inhibitors of SDF-1/CXCR4, AMD3100 significantly diminished LPS-induced migration of hDPSCs. LPS at specific concentrations can promote cell adhesion and migration in hDPSCs via the NF-κB and MAPK pathways by up-regulating the expression of adhesion molecules and chemotactic factors. LPS may influence pulp healing through enhancing the adhesion and migration of human dental pulp stem cells when it enters into pulp during pulp exposure or deep caries. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. The Role of ORAI1 in the Odontogenic Differentiation of Human Dental Pulp Stem Cells.

    PubMed

    Sohn, S; Park, Y; Srikanth, S; Arai, A; Song, M; Yu, B; Shin, K-H; Kang, M K; Wang, C; Gwack, Y; Park, N-H; Kim, R H

    2015-11-01

    Pulp capping, or placing dental materials directly onto the vital pulp tissues of affected teeth, is a dental procedure that aims to regenerate reparative dentin. Several pulp capping materials are clinically being used, and calcium ion (Ca(2+)) released from these materials is known to mediate reparative dentin formation. ORAI1 is an essential pore subunit of store-operated Ca(2+) entry (SOCE), which is a major Ca(2+) influx pathway in most nonexcitable cells. Here, we evaluated the role of ORAI1 in mediating the odontogenic differentiation and mineralization of dental pulp stem cells (DPSCs). During the odontogenic differentiation of DPSCs, the expression of ORAI1 increased in a time-dependent manner. DPSCs knocked down with ORAI1 shRNA (DPSC/ORAI1sh) or overexpressed with dominant negative mutant ORAI1(E106Q) (DPSC/E106Q) exhibited the inhibition of Ca(2+) influx and suppression of odontogenic differentiation and mineralization as demonstrated by alkaline phosphatase (ALP) activity/staining as well as alizarin red S staining when compared with DPSCs of their respective control groups (DPSC/CTLsh and DPSC/CTL). The gene expression for odontogenic differentiation markers such as osteocalcin, bone sialoprotein, and dentin matrix protein 1 (DMP1) was also suppressed. When DPSC/CTL or DPSC/E106Q cells were subcutaneously transplanted into nude mice, DPSC/CTL cells induced mineralized tissue formation with significant increases in ALP and DMP1 staining in vivo, whereas DPSC/E106Q cells did not. Collectively, our data showed that ORAI1 plays critical roles in the odontogenic differentiation and mineralization of DPSCs by regulating Ca(2+) influx and that ORAI1 may be a therapeutic target to enhance reparative dentin formation. © International & American Associations for Dental Research 2015.

  12. Retracted: Effects of pro-inflammatory cytokines on mineralization potential of rat dental pulp stem cells.

    PubMed

    2011-10-01

    The following article from the Journal of Tissue Engineering and Regenerative Medicine, 'Effects of Pro-inflammatory Cytokines on Mineralization Potential of Rat Dental Pulp Stem Cells' by Yang X, Walboomers XF, Bian Z, Jansen JA, Fan M, published online on 11 July 2011 in Wiley Online Library (onlinelibrary.wiley.com), has been retracted by agreement between the authors, the journal Editor-in-Chief, and John Wiley & Sons, Ltd. The retraction has been agreed due to two authors (Walboomers XF, and Jansen JA) not having been involved in the research described, nor made aware of their names being listed on the manuscript, nor told of its submission to the journal.

  13. A Hydrogel Scaffold That Maintains Viability and Supports Differentiation of Dental Pulp Stem Cells

    PubMed Central

    Cavalcanti, Bruno N.; Zeitlin, Benjamin D.; Nör, Jacques E.

    2012-01-01

    Objectives The clinical translation of stem cell-based Regenerative Endodontics demands further development of suitable injectable scaffolds. Puramatrix™ is a defined, self-assembling peptide hydrogel which instantaneously polymerizes under normal physiological conditions. Here, we assessed the compatibility of Puramatrix™ with dental pulp stem cell (DPSC) growth and differentiation. Methods DPSC cells were grown in 0.05 to 0.25% Puramatrix™. Cell viability was measured colorimetrically using the WST-1 assay. Cell morphology was observed in 3-D modeling using confocal microscopy. In addition, we used the human tooth slice model with Puramatrix™ to verify DPSC differentiation into odontoblast-like cells, as measured by expression of DSPP and DMP-1. Results DPSC survived and proliferated in Puramatrix™ for at least three weeks in culture. Confocal microscopy revealed that cells seeded in Puramatrix™ presented morphological features of healthy cells, and some cells exhibited cytoplasmic elongations. Notably, after 21 days in tooth slices containing Puramatrix™, DPSC cells expressed DMP-1 and DSPP, putative markers of odontoblastic differentiation. Significance Collectively, these data suggest that self-assembling peptide hydrogels might be useful injectable scaffolds for stem cell-based Regenerative Endodontics. PMID:22901827

  14. Adhesion and proliferation of human mesenchymal stem cells from dental pulp on porous silicon scaffolds.

    PubMed

    Collart-Dutilleul, Pierre-Yves; Secret, Emilie; Panayotov, Ivan; Deville de Périère, Dominique; Martín-Palma, Raúl J; Torres-Costa, Vicente; Martin, Marta; Gergely, Csilla; Durand, Jean-Olivier; Cunin, Frédérique; Cuisinier, Frédéric J

    2014-02-12

    In regenerative medicine, stem-cell-based therapy often requires a scaffold to deliver cells and/or growth factors to the injured site. Porous silicon (pSi) is a promising biomaterial for tissue engineering as it is both nontoxic and bioresorbable. Moreover, surface modification can offer control over the degradation rate of pSi and can also promote cell adhesion. Dental pulp stem cells (DPSC) are pluripotent mesenchymal stem cells found within the teeth and constitute a readily source of stem cells. Thus, coupling the good proliferation and differentiation capacities of DPSC with the textural and chemical properties of the pSi substrates provides an interesting approach for therapeutic use. In this study, the behavior of human DPSC is analyzed on pSi substrates presenting pores of various sizes, 10 ± 2 nm, 36 ± 4 nm, and 1.0 ± 0.1 μm, and undergoing different chemical treatments, thermal oxidation, silanization with aminopropyltriethoxysilane (APTES), and hydrosilylation with undecenoic acid or semicarbazide. DPSC adhesion and proliferation were followed for up to 72 h by fluorescence microscopy, scanning electron microscopy (SEM), enzymatic activity assay, and BrdU assay for mitotic activity. Porous silicon with 36 nm pore size was found to offer the best adhesion and the fastest growth rate for DPSC compared to pSi comporting smaller pore size (10 nm) or larger pore size (1 μm), especially after silanization with APTES. Hydrosilylation with semicarbazide favored cell adhesion and proliferation, especially mitosis after cell adhesion, but such chemical modification has been found to led to a scaffold that is stable for only 24-48 h in culture medium. Thus, semicarbazide-treated pSi appeared to be an appropriate scaffold for stem cell adhesion and immediate in vivo transplantation, whereas APTES-treated pSi was found to be more suitable for long-term in vitro culture, for stem cell proliferation and differentiation.

  15. In Vitro Differentiation and Mineralization of Dental Pulp Stem Cells on Enamel-Like Fluorapatite Surfaces

    PubMed Central

    Wang, Xiaodong; Jin, Taocong; Chang, Syweren; Zhang, Zhaocheng; Czajka-Jakubowska, Agata; Nör, Jacques E.; Clarkson, Brian H.

    2012-01-01

    Our previous studies have shown good biocompatibility of fluorapatite (FA) crystal surfaces in providing a favorable environment for functional cell–matrix interactions of human dental pulp stem cells (DPSCs) and also in supporting their long-term growth. The aim of the current study was to further investigate whether this enamel-like surface can support the differentiation and mineralization of DPSCs, and, therefore, act as a potential model for studying the enamel/dentin interface and, perhaps, dentine/pulp regeneration in tooth tissue engineering. The human pathway-focused osteogenesis polymerase chain reaction (PCR) array demonstrated that the expression of osteogenesis-related genes of human DPSCs was increased on FA surfaces compared with that on etched stainless steel (SSE). Consistent with the PCR array, FA promoted mineralization compared with the SSE surface with or without the addition of a mineralization promoting supplement (MS). This was confirmed by alkaline phosphatase (ALP) staining, Alizarin red staining, and tetracycline staining for mineral formation. In conclusion, FA crystal surfaces, especially ordered (OR) FA surfaces, which mimicked the physical architecture of enamel, provided a favorable extracellular matrix microenvironment for the cells. This resulted in the differentiation of human DPSCs and mineralized tissue formation, and, thus, demonstrated that it may be a promising biomimetic model for dentin-pulp tissue engineering. PMID:22563788

  16. Three-dimensional culture of dental pulp stem cells in direct contact to tricalcium silicate cements.

    PubMed

    Widbiller, M; Lindner, S R; Buchalla, W; Eidt, A; Hiller, K-A; Schmalz, G; Galler, K M

    2016-03-01

    Calcium silicate cements are biocompatible dental materials applicable in contact with vital tissue. The novel tricalcium silicate cement Biodentine™ offers properties superior to commonly used mineral trioxide aggregate (MTA). Objective of this study was to evaluate its cytocompatibility and ability to induce differentiation and mineralization in three-dimensional cultures of dental pulp stem cells after direct contact with the material. Test materials included a new tricalcium silicate (Biodentine™, Septodont, Saint-Maur-des-Fossés, France), MTA (ProRoot® MTA, DENSPLY Tulsa Dental Specialities, Johnson City, TN, USA), glass ionomer (Ketac™ Molar Aplicap™, 3M ESPE, Seefeld, Germany), human dentin disks and polystyrene. Magnetic activated cell sorting for to the surface antigen STRO-1 was performed to gain a fraction enriched with mesenchymal stem cells. Samples were allowed to set and dental pulp stem cells in collagen carriers were placed on top. Scanning electron microscopy of tricalcium silicate cement surfaces with and without cells was conducted. Cell viability was measured for 14 days by MTT assay. Alkaline phosphatase activity was evaluated (days 3, 7, and 14) and expression of mineralization-associated genes (COL1A1, ALP, DSPP, and RUNX2) was quantified by real-time quantitative PCR. Nonparametric statistical analysis for cell viability and alkaline phosphatase data was performed to compare different materials as well as time points (Mann-Whitney U test, α = 0.05). Cell viability was highest on tricalcium silicate cement, followed by MTA. Viability on glass ionomer cement and dentin disks was significantly lower. Alkaline phosphatase activity was lower in cells on new tricalcium silicate cement compared to MTA, whereas expression patterns of marker genes were alike. Increased cell viability and similar levels of mineralization-associated gene expression in three-dimensional cell cultures on the novel tricalcium silicate cement and mineral

  17. Trophic Effects and Regenerative Potential of Mobilized Mesenchymal Stem Cells From Bone Marrow and Adipose Tissue as Alternative Cell Sources for Pulp/Dentin Regeneration.

    PubMed

    Murakami, Masashi; Hayashi, Yuki; Iohara, Koichiro; Osako, Yohei; Hirose, Yujiro; Nakashima, Misako

    2015-01-01

    Dental pulp stem cell (DPSC) subsets mobilized by granulocyte-colony-stimulating factor (G-CSF) are safe and efficacious for complete pulp regeneration. The supply of autologous pulp tissue, however, is very limited in the aged. Therefore, alternative sources of mesenchymal stem/progenitor cells (MSCs) are needed for the cell therapy. In this study, DPSCs, bone marrow (BM), and adipose tissue (AD)-derived stem cells of the same individual dog were isolated using G-CSF-induced mobilization (MDPSCs, MBMSCs, and MADSCs). The positive rates of CXCR4 and G-CSFR in MDPSCs were similar to MADSCs and were significantly higher than those in MBMSCs. Trophic effects of MDPSCs on angiogenesis, neurite extension, migration, and antiapoptosis were higher than those of MBMSCs and MADSCs. Pulp-like loose connective tissues were regenerated in all three MSC transplantations. Significantly higher volume of regenerated pulp and higher density of vascularization and innervation were observed in response to MDPSCs compared to MBMSC and MADSC transplantation. Collagenous matrix containing dentin sialophosphoprotein (DSPP)-positive odontoblast-like cells was the highest in MBMSCs and significantly higher in MADSCs compared to MDPSCs. MBMSCs and MADSCs, therefore, have potential for pulp regeneration, although the volume of regenerated pulp tissue, angiogenesis, and reinnervation, were less. Thus, in conclusion, an alternative cell source for dental pulp/dentin regeneration are stem cells from BM and AD tissue.

  18. Dental pulp stem cells: function, isolation and applications in regenerative medicine.

    PubMed

    Tatullo, Marco; Marrelli, Massimo; Shakesheff, Kevin M; White, Lisa J

    2015-11-01

    Dental pulp stem cells (DPSCs) are a promising source of cells for numerous and varied regenerative medicine applications. Their natural function in the production of odontoblasts to create reparative dentin support applications in dentistry in the regeneration of tooth structures. However, they are also being investigated for the repair of tissues outside of the tooth. The ease of isolation of DPSCs from discarded or removed teeth offers a promising source of autologous cells, and their similarities with bone marrow stromal cells (BMSCs) suggest applications in musculoskeletal regenerative medicine. DPSCs are derived from the neural crest and, therefore, have a different developmental origin to BMSCs. These differences from BMSCs in origin and phenotype are being exploited in neurological and other applications. This review briefly highlights the source and functions of DPSCs and then focuses on in vivo applications across the breadth of regenerative medicine. © 2014 The Authors. Journal of Tissue Engineering and Regenerative Medicine published by John Wiley & Sons, Ltd.

  19. Interferon-gamma improves impaired dentinogenic and immunosuppressive functions of irreversible pulpitis-derived human dental pulp stem cells

    PubMed Central

    Sonoda, Soichiro; Yamaza, Haruyoshi; Ma, Lan; Tanaka, Yosuke; Tomoda, Erika; Aijima, Reona; Nonaka, Kazuaki; Kukita, Toshio; Shi, Songtao; Nishimura, Fusanori; Yamaza, Takayoshi

    2016-01-01

    Clinically, irreversible pulpitis is treated by the complete removal of pulp tissue followed by replacement with artificial materials. There is considered to be a high potential for autologous transplantation of human dental pulp stem cells (DPSCs) in endodontic treatment. The usefulness of DPSCs isolated from healthy teeth is limited. However, DPSCs isolated from diseased teeth with irreversible pulpitis (IP-DPSCs) are considered to be suitable for dentin/pulp regeneration. In this study, we examined the stem cell potency of IP-DPSCs. In comparison with healthy DPSCs, IP-DPSCs expressed lower colony-forming capacity, population-doubling rate, cell proliferation, multipotency, in vivo dentin regeneration, and immunosuppressive activity, suggesting that intact IP-DPSCs may be inadequate for dentin/pulp regeneration. Therefore, we attempted to improve the impaired in vivo dentin regeneration and in vitro immunosuppressive functions of IP-DPSCs to enable dentin/pulp regeneration. Interferon gamma (IFN-γ) treatment enhanced in vivo dentin regeneration and in vitro T cell suppression of IP-DPSCs, whereas treatment with tumor necrosis factor alpha did not. Therefore, these findings suggest that IFN-γ may be a feasible modulator to improve the functions of impaired IP-DPSCs, suggesting that autologous transplantation of IFN-γ-accelerated IP-DPSCs might be a promising new therapeutic strategy for dentin/pulp tissue engineering in future endodontic treatment. PMID:26775677

  20. Interferon-gamma improves impaired dentinogenic and immunosuppressive functions of irreversible pulpitis-derived human dental pulp stem cells.

    PubMed

    Sonoda, Soichiro; Yamaza, Haruyoshi; Ma, Lan; Tanaka, Yosuke; Tomoda, Erika; Aijima, Reona; Nonaka, Kazuaki; Kukita, Toshio; Shi, Songtao; Nishimura, Fusanori; Yamaza, Takayoshi

    2016-01-18

    Clinically, irreversible pulpitis is treated by the complete removal of pulp tissue followed by replacement with artificial materials. There is considered to be a high potential for autologous transplantation of human dental pulp stem cells (DPSCs) in endodontic treatment. The usefulness of DPSCs isolated from healthy teeth is limited. However, DPSCs isolated from diseased teeth with irreversible pulpitis (IP-DPSCs) are considered to be suitable for dentin/pulp regeneration. In this study, we examined the stem cell potency of IP-DPSCs. In comparison with healthy DPSCs, IP-DPSCs expressed lower colony-forming capacity, population-doubling rate, cell proliferation, multipotency, in vivo dentin regeneration, and immunosuppressive activity, suggesting that intact IP-DPSCs may be inadequate for dentin/pulp regeneration. Therefore, we attempted to improve the impaired in vivo dentin regeneration and in vitro immunosuppressive functions of IP-DPSCs to enable dentin/pulp regeneration. Interferon gamma (IFN-γ) treatment enhanced in vivo dentin regeneration and in vitro T cell suppression of IP-DPSCs, whereas treatment with tumor necrosis factor alpha did not. Therefore, these findings suggest that IFN-γ may be a feasible modulator to improve the functions of impaired IP-DPSCs, suggesting that autologous transplantation of IFN-γ-accelerated IP-DPSCs might be a promising new therapeutic strategy for dentin/pulp tissue engineering in future endodontic treatment.

  1. Effects of cryopreservation on the characteristics of dental pulp stem cells of intact deciduous teeth.

    PubMed

    Lindemann, Daniele; Werle, Stefanie B; Steffens, Daniela; Garcia-Godoy, Franklin; Pranke, Patricia; Casagrande, Luciano

    2014-09-01

    The aim of this study was to isolate and cultivate cells from the pulp of 7-day-cryopreserved intact deciduous human teeth and evaluate the effect of cryopreservation on dental pulp stem cell (DPSC) characteristics. Twenty-six deciduous teeth were collected and allocated in two groups: immediate cell isolation (non-cryopreserved group) and intact cryopreserved (cryopreserved group). The teeth were cryopreserved in dimethylsulfoxide solution and recovered after 7 days. The success rate of isolation, proliferation, surface markers (CD14, CD29, CD34, CD45, CD73, CD90, and HLA-DR), differentiation capacity, and morphology were evaluated. Isolation success rate was 61% and 30% for the non-cryopreserved and cryopreserved groups, respectively. There were no statistical differences between the groups for the tested surface markers. The cells in both groups were capable of differentiating into three mesenchymal lineages. No statistical differences between the groups were observed through the time course proliferation assay (0, 1, 3, 5, and 7 days); however, the mean time between isolation and the fifth passage was shorter for the non-cryopreserved group (p=0.035). The morphology of the cells was considered altered in the cryopreserved group. DPSCs were obtained from cryopreserved intact deciduous teeth without changes in the immunophenotypical characteristics and differentiation ability; however, lower culture rates, proliferation potential, and morphological alterations were observed in relation to the control group. Copyright © 2014. Published by Elsevier Ltd.

  2. Low-intensity laser phototherapy enhances the proliferation of dental pulp stem cells under nutritional deficiency.

    PubMed

    Moura-Netto, Cacio; Ferreira, Leila Soares; Maranduba, Carlos Magno; Mello-Moura, Anna Carolina Volpi; Marques, Marcia Martins

    2016-05-31

    Dental trauma in immature permanent teeth can damage pulp vascularization, which leads to necrosis and cessation of apexogenesis. Studies on tissue engineering using stem cells from human exfoliated deciduous teeth (SHEDs) have yielded promising results. Laser phototherapy (LPT) is able to influence the proliferation and differentiation of these cells, which could improve tissue engineering. SHEDs (eighth passage) were seeded into 96-well culture plates (103 cells/well) and were grown in culture medium supplemented with 15% defined fetal bovine serum (FBS) for 12 h. After determining the appropriate nutrition deficiency status (5% FBS), the cells were assigned into four groups: 1) G1 - 15% FBS (positive control); 2) G2 - 5% FBS (negative control); 3) G3 - 5% FBS+LPT 3 J/cm2; and 4) G4 - 5% FBS+LPT 5 J/cm2. For the LPT groups, two laser irradiations at 6 h intervals were performed using a continuous wave InGaAlP diode laser (660 nm, with a spot size of 0.028 cm2, 10 mW) in punctual and contact mode. Cell viability was assessed via an MTT reduction assay immediately after the second laser irradiation (0 h) and 24, 48, and 72 h later. We found that G3 and G4 presented a significantly higher cell growth rate when compared with G2 (p < 0.01). Moreover, G4 exhibited a similar cell growth rate as G1 throughout the entire experiment (p > 0.05). These findings indicate that LPT with 5 J/cm2 can enhance the growth of SHEDs during situations of nutritional deficiency. Therefore, LPT could be a valuable adjunct treatment in tissue engineering when using stem cells derived from the dental pulp of primary teeth.

  3. Dental Pulp Stem Cells Model Early Life and Imprinted DNA Methylation Patterns.

    PubMed

    Dunaway, Keith; Goorha, Sarita; Matelski, Lauren; Urraca, Nora; Lein, Pamela J; Korf, Ian; Reiter, Lawrence T; LaSalle, Janine M

    2017-04-01

    Early embryonic stages of pluripotency are modeled for epigenomic studies primarily with human embryonic stem cells (ESC) or induced pluripotent stem cells (iPSCs). For analysis of DNA methylation however, ESCs and iPSCs do not accurately reflect the DNA methylation levels found in preimplantation embryos. Whole genome bisulfite sequencing (WGBS) approaches have revealed the presence of large partially methylated domains (PMDs) covering 30%-40% of the genome in oocytes, preimplantation embryos, and placenta. In contrast, ESCs and iPSCs show abnormally high levels of DNA methylation compared to inner cell mass (ICM) or placenta. Here we show that dental pulp stem cells (DPSCs), derived from baby teeth and cultured in serum-containing media, have PMDs and mimic the ICM and placental methylome more closely than iPSCs and ESCs. By principal component analysis, DPSC methylation patterns were more similar to two other neural stem cell types of human derivation (EPI-NCSC and LUHMES) and placenta than were iPSCs, ESCs or other human cell lines (SH-SY5Y, B lymphoblast, IMR90). To test the suitability of DPSCs in modeling epigenetic differences associated with disease, we compared methylation patterns of DPSCs derived from children with chromosome 15q11.2-q13.3 maternal duplication (Dup15q) to controls. Differential methylation region (DMR) analyses revealed the expected Dup15q hypermethylation at the imprinting control region, as well as hypomethylation over SNORD116, and novel DMRs over 147 genes, including several autism candidate genes. Together these data suggest that DPSCs are a useful model for epigenomic and functional studies of human neurodevelopmental disorders. Stem Cells 2017;35:981-988. © 2016 AlphaMed Press.

  4. Effect of an Experimental Direct Pulp-capping Material on the Properties and Osteogenic Differentiation of Human Dental Pulp Stem Cells

    NASA Astrophysics Data System (ADS)

    Yu, Fan; Dong, Yan; Yang, Yan-Wei; Lin, Ping-Ting; Yu, Hao-Han; Sun, Xiang; Sun, Xue-Fei; Zhou, Huan; Huang, Li; Chen, Ji-Hua

    2016-10-01

    Effective pulp-capping materials must have antibacterial properties and induce dentin bridge formation; however, many current materials do not satisfy clinical requirements. Accordingly, the effects of an experiment pulp-capping material (Exp) composed of an antibacterial resin monomer (MAE-DB) and Portland cement (PC) on the viability, adhesion, migration, and differentiation of human dental pulp stem cells (hDPSCs) were examined. Based on a Cell Counting Kit-8 assay, hDPSCs exposed to Exp extracts showed limited viability at 24 and 48 h, but displayed comparable viability to the control at 72 h. hDPSC treatment with Exp extracts enhanced cellular adhesion and migration according to in vitro scratch wound healing and Transwell migration assays. Exp significantly upregulated the expression of osteogenesis-related genes. The hDPSCs cultured with Exp exhibited higher ALP activity and calcium deposition in vitro compared with the control group. The novel material showed comparable cytocompatibility to control cells and promoted the adhesion, migration, and osteogenic differentiation of hDPSCs, indicating excellent biocompatibility. This new direct pulp-capping material containing MAE-DB and PC shows promise as a potential alternative to conventional materials for direct pulp capping.

  5. Effect of an Experimental Direct Pulp-capping Material on the Properties and Osteogenic Differentiation of Human Dental Pulp Stem Cells.

    PubMed

    Yu, Fan; Dong, Yan; Yang, Yan-Wei; Lin, Ping-Ting; Yu, Hao-Han; Sun, Xiang; Sun, Xue-Fei; Zhou, Huan; Huang, Li; Chen, Ji-Hua

    2016-10-04

    Effective pulp-capping materials must have antibacterial properties and induce dentin bridge formation; however, many current materials do not satisfy clinical requirements. Accordingly, the effects of an experiment pulp-capping material (Exp) composed of an antibacterial resin monomer (MAE-DB) and Portland cement (PC) on the viability, adhesion, migration, and differentiation of human dental pulp stem cells (hDPSCs) were examined. Based on a Cell Counting Kit-8 assay, hDPSCs exposed to Exp extracts showed limited viability at 24 and 48 h, but displayed comparable viability to the control at 72 h. hDPSC treatment with Exp extracts enhanced cellular adhesion and migration according to in vitro scratch wound healing and Transwell migration assays. Exp significantly upregulated the expression of osteogenesis-related genes. The hDPSCs cultured with Exp exhibited higher ALP activity and calcium deposition in vitro compared with the control group. The novel material showed comparable cytocompatibility to control cells and promoted the adhesion, migration, and osteogenic differentiation of hDPSCs, indicating excellent biocompatibility. This new direct pulp-capping material containing MAE-DB and PC shows promise as a potential alternative to conventional materials for direct pulp capping.

  6. Effect of an Experimental Direct Pulp-capping Material on the Properties and Osteogenic Differentiation of Human Dental Pulp Stem Cells

    PubMed Central

    Yu, Fan; Dong, Yan; Yang, Yan-wei; Lin, Ping-ting; Yu, Hao-han; Sun, Xiang; Sun, Xue-fei; Zhou, Huan; Huang, Li; Chen, Ji-hua

    2016-01-01

    Effective pulp-capping materials must have antibacterial properties and induce dentin bridge formation; however, many current materials do not satisfy clinical requirements. Accordingly, the effects of an experiment pulp-capping material (Exp) composed of an antibacterial resin monomer (MAE-DB) and Portland cement (PC) on the viability, adhesion, migration, and differentiation of human dental pulp stem cells (hDPSCs) were examined. Based on a Cell Counting Kit-8 assay, hDPSCs exposed to Exp extracts showed limited viability at 24 and 48 h, but displayed comparable viability to the control at 72 h. hDPSC treatment with Exp extracts enhanced cellular adhesion and migration according to in vitro scratch wound healing and Transwell migration assays. Exp significantly upregulated the expression of osteogenesis-related genes. The hDPSCs cultured with Exp exhibited higher ALP activity and calcium deposition in vitro compared with the control group. The novel material showed comparable cytocompatibility to control cells and promoted the adhesion, migration, and osteogenic differentiation of hDPSCs, indicating excellent biocompatibility. This new direct pulp-capping material containing MAE-DB and PC shows promise as a potential alternative to conventional materials for direct pulp capping. PMID:27698421

  7. Characterization of Neurons from Immortalized Dental Pulp Stem Cells for the Study of Neurogenetic Disorders

    PubMed Central

    Urraca, Nora; Memon, Rawaha; El-Iyachi, Ikbale; Goorha, Sarita; Valdez, Colleen; Tran, Quynh T.; Scroggs, Reese; Miranda-Carboni, Gustavo A.; Donaldson, Martin; Bridges, Dave; Reiter, Lawrence T.

    2015-01-01

    A major challenge to the study and treatment of neurogenetic syndromes is accessing live neurons for study from affected individuals. Although several sources of stem cells are currently available, acquiring these involve invasive procedures, may be difficult or expensive to generate and are limited in number. Dental pulp stem cells (DPSC) are multipotent stem cells that reside deep the pulp of shed teeth. To investigate the characteristics of DPSC that make them a valuable resource for translational research, we performed a set of viability, senescence, immortalization and gene expression studies on control DPSC and derived neurons. We investigated the basic transport conditions and maximum passage number for primary DPSC. We immortalized control DPSC using human telomerase reverse transcriptase (hTERT) and evaluated neuronal differentiation potential and global gene expression changes by RNA-seq. We show that neurons from immortalized DPSC share morphological and electrophysiological properties with non-immortalized DPSC. We also show that differentiation of DPSC into neurons significantly alters gene expression for 1305 transcripts. Here we show that these changes in gene expression are concurrent with changes in protein levels of the transcriptional repressor REST/NSRF, which is known to be involved in neuronal differentiation. Immortalization significantly altered the expression of 183 genes after neuronal differentiation, 94 of which also changed during differentiation. Our studies indicate that viable DPSC can be obtained from teeth stored for ≥72hrs, these can then be immortalized and still produce functional neurons for in vitro studies, but that constitutive hTERT immortalization is not be the best approach for long term use of patient derived DPSC for the study of disease. PMID:26599327

  8. The effects of hypoxia on the stemness properties of human dental pulp stem cells (DPSCs)

    PubMed Central

    Ahmed, Nermeen El-Moataz Bellah; Murakami, Masashi; Kaneko, Satoru; Nakashima, Misako

    2016-01-01

    Recent studies have demonstrated that culture under hypoxia has beneficial effects on mesenchymal stem cells (MSCs). However, there are limitations to achieving a stable condition in conventional hypoxic CO2 incubators. DPSCs are a unique type of MSCs which are promising in many regenerative therapies. In this study, we investigated the ideal hypoxic culture environment for DPSCs using a new system that can provide controlled O2 environment. The effects of hypoxia (3%, 5%) on the stemness properties of DPSCs. Their morphology, proliferation rate, expression of stem cell markers, migration ability, mRNA expression of angiogenic/neurotrophic factors and immunomodulatory genes were evaluated and compared. Additionally, the effect of the discrete secretome on proliferation, migration, and neurogenic induction was assessed. Hypoxic DPSCs were found to be smaller in size and exhibited larger nuclei. 5% O2 significantly increased the proliferation rate, migration ability, expression of stem cell markers (CXCR4 and G-CSFR), and expression of SOX2, VEGF, NGF, and BDNF genes of DPSCs. Moreover, secretome collected from 5%O2 cultures displayed higher stimulatory effects on proliferation and migration of NIH3T3 cells and on neuronal differentiation of SH-SY5Y cells. These results demonstrate that 5%O2 may be ideal for enhancing DPSCs growth, stem cell properties, and secretome trophic effect. PMID:27739509

  9. Variation in human dental pulp stem cell ageing profiles reflect contrasting proliferative and regenerative capabilities.

    PubMed

    Alraies, Amr; Alaidaroos, Nadia Y A; Waddington, Rachel J; Moseley, Ryan; Sloan, Alastair J

    2017-02-02

    Dental pulp stem cells (DPSCs) are increasingly being recognized as a viable cell source for regenerative medicine. Although significant variations in their ex vivo expansion are well-established, DPSC proliferative heterogeneity remains poorly understood, despite such characteristics influencing their regenerative and therapeutic potential. This study assessed clonal human DPSC regenerative potential and the impact of cellular senescence on these responses, to better understand DPSC functional behaviour. All DPSCs were negative for hTERT. Whilst one DPSC population reached >80 PDs before senescence, other populations only achieved <40 PDs, correlating with DPSCs with high proliferative capacities possessing longer telomeres (18.9 kb) than less proliferative populations (5-13 kb). High proliferative capacity DPSCs exhibited prolonged stem cell marker expression, but lacked CD271. Early-onset senescence, stem cell marker loss and positive CD271 expression in DPSCs with low proliferative capacities were associated with impaired osteogenic and chondrogenic differentiation, favouring adipogenesis. DPSCs with high proliferative capacities only demonstrated impaired differentiation following prolonged expansion (>60 PDs). This study has identified that proliferative and regenerative heterogeneity is related to contrasting telomere lengths and CD271 expression between DPSC populations. These characteristics may ultimately be used to selectively screen and isolate high proliferative capacity/multi-potent DPSCs for regenerative medicine exploitation.

  10. Human dental pulp stem cells produce mineralized matrix in 2D and 3D cultures

    PubMed Central

    Riccio, M.; Resca, E.; Maraldi, T.; Pisciotta, A.; Ferrari, A.; Bruzzesi, G.; De Pol, A.

    2010-01-01

    The aim of this study was to characterize the in vitro osteogenic differentiation of dental pulp stem cells (DPSCs) in 2D cultures and 3D biomaterials. DPSCs, separated from dental pulp by enzymatic digestion, and isolated by magnetic cell sorting were differentiated toward osteogenic lineage on 2D surface by using an osteogenic medium. During differentiation process, DPSCs express specific bone proteins like Runx-2, Osx, OPN and OCN with a sequential expression, analogous to those occurring during osteoblast differentiation, and produce extracellular calcium deposits. In order to differentiate cells in a 3D space that mimes the physiological environment, DPSCs were cultured in two distinct bioscaffolds, Matrigel™ and Collagen sponge. With the addition of a third dimension, osteogenic differentiation and mineralized extracellular matrix production significantly improved. In particular, in Matrigel™ DPSCs differentiated with osteoblast/osteocyte characteristics and connected by gap junction, and therefore formed calcified nodules with a 3D intercellular network. Furthermore, DPSCs differentiated in collagen sponge actively secrete human type I collagen micro-fibrils and form calcified matrix containing trabecular-like structures. These neo-formed DPSCs-scaffold devices may be used in regenerative surgical applications in order to resolve pathologies and traumas characterized by critical size bone defects. PMID:21263745

  11. Mechanical Changes in Human Dental Pulp Stem Cells during Early Odontogenic Differentiation

    PubMed Central

    Jones, Taneka D.; Naimipour, Hamed; Sun, Shan; Cho, Michael; Alapati, Satish B.

    2014-01-01

    Cell adhesion and migration in bioactive scaffolds require actin cytoskeleton remodeling and focal adhesion formation. Additionally, human dental pulp stem cells (hDPSCs) undergo several changes in their mechanical properties during odontogenic differentiation. The effect of factors essential for odontogenesis on the actin stress fiber elasticity and focal adhesion formation is not known. Methods Live hDPSCs cultured in odontogenic media were imaged for cytoskeleton changes using an atomic force microscope. The Young’s modulus (kPa) of the cytoskeleton was recorded as a function of culture medium for 10 days. Focal adhesion formation was assessed using immunofluorescence. Cultured hDPSCs were incubated with a monoclonal vinculin antibody and filamentous actins were visualized using 0.5 µM phalloidin. Results Cytoskeletal elasticity significantly increased in response to odontogenic media. Both the number and physical size of focal adhesions in hDPSCs also increased. Up-regulation of vinculin expression was evident. The increase in the formation of focal adhesions was consistent with actin remodeling to stress fibers. Conclusions Our findings suggest that hDPSCs firmly attach to the glass substrate in response to odontogenic media. Successful regeneration of pulp-dentin tissue using biomimetic scaffolds will likely require cell-extracellular matrix (ECM) interactions influenced by biochemical induction factors. PMID:25241024

  12. Mechanical changes in human dental pulp stem cells during early odontogenic differentiation.

    PubMed

    Jones, Taneka D; Naimipour, Hamed; Sun, Shan; Cho, Michael; Alapati, Satish B

    2015-01-01

    Cell adhesion and migration in bioactive scaffolds require actin cytoskeleton remodeling and focal adhesion formation. Additionally, human dental pulp stem cells (hDPSCs) undergo several changes in their mechanical properties during odontogenic differentiation. The effect of factors essential for odontogenesis on actin stress fiber elasticity and focal adhesion formation is not known. Live hDPSCs cultured in odontogenic media were imaged for cytoskeleton changes using an atomic force microscope. The Young's modulus (kPa) of the cytoskeleton was recorded as a function of culture medium for 10 days. Focal adhesion formation was assessed using immunofluorescence. Cultured hDPSCs were incubated with a monoclonal vinculin antibody, and filamentous actins were visualized using 0.5 μmol/L phalloidin. Cytoskeletal elasticity significantly increased in response to odontogenic media. Both the number and physical size of focal adhesions in hDPSCs also increased. Up-regulation of vinculin expression was evident. The increase in the formation of focal adhesions was consistent with actin remodeling to stress fibers. Our findings suggest that hDPSCs firmly attach to the glass substrate in response to odontogenic media. Successful regeneration of pulp-dentin tissue using biomimetic scaffolds will likely require cell-extracellular matrix interactions influenced by biochemical induction factors. Copyright © 2015 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  13. The relationship between cell proliferation and differentiation and mapping of putative dental pulp stem/progenitor cells during mouse molar development by chasing BrdU-labeling.

    PubMed

    Ishikawa, Yuko; Ida-Yonemochi, Hiroko; Nakakura-Ohshima, Kuniko; Ohshima, Hayato

    2012-04-01

    Human dental pulp contains adult stem cells. Our recent study demonstrated the localization of putative dental pulp stem/progenitor cells in the rat developing molar by chasing 5-bromo-2'-deoxyuridine (BrdU)-labeling. However, there are no available data on the localization of putative dental pulp stem/progenitor cells in the mouse molar. This study focuses on the mapping of putative dental pulp stem/progenitor cells in addition to the relationship between cell proliferation and differentiation in the developing molar using BrdU-labeling. Numerous proliferating cells appeared in the tooth germ and the most active cell proliferation in the mesenchymal cells occurred in the prenatal stages, especially on embryonic Day 15 (E15). Cell proliferation in the pulp tissue dramatically decreased in number by postnatal Day 3 (P3) when nestin-positive odontoblasts were arranged in the cusped areas and disappeared after postnatal Week 1 (P1W). Root dental papilla included numerous proliferating cells during P5 to P2W. Three to four intraperitoneal injections of BrdU were given to pregnant ICR mice and revealed slow-cycling long-term label-retaining cells (LRCs) in the mature tissues of postnatal animals. Numerous dense LRCs postnatally decreased in number and reached a plateau after P1W when they mainly resided in the center of the dental pulp, associating with blood vessels. Furthermore, numerous dense LRCs co-expressed mesenchymal stem cell markers such as STRO-1 and CD146. Thus, dense LRCs in mature pulp tissues were believed to be dental pulp stem/progenitor cells harboring in the perivascular niche surrounding the endothelium.

  14. Effects of SOX2 on Proliferation, Migration and Adhesion of Human Dental Pulp Stem Cells.

    PubMed

    Liu, Pengfei; Cai, Jinglei; Dong, Delu; Chen, Yaoyu; Liu, Xiaobo; Wang, Yi; Zhou, Yulai

    2015-01-01

    As a key factor for cell pluripotent and self-renewing phenotypes, SOX2 has attracted scientists' attention gradually in recent years. However, its exact effects in dental pulp stem cells (DPSCs) are still unclear. In this study, we mainly investigated whether SOX2 could affect some biological functions of DPSCs. DPSCs were isolated from the dental pulp of human impacted third molar. SOX2 overexpressing DPSCs (DPSCs-SOX2) were established through retroviral infection. The effect of SOX2 on cell proliferation, migration and adhesion ability was evaluated with CCK-8, trans-well system and fibronectin-induced cell attachment experiment respectively. Whole genome expression of DPSCs-SOX2 was analyzed with RNA microarray. Furthermore, a rescue experiment was performed with SOX2-siRNA in DPSC-SOX2 to confirm the effect of SOX2 overexpression in DPSCs. We found that SOX2 overexpression could result in the enhancement of cell proliferation, migration, and adhesion in DPSCs obviously. RNA microarray analysis indicated that some key genes in the signal pathways associated with cell cycle, migration and adhesion were upregulated in different degree, and the results were further confirmed with qPCR and western-blot. Finally, DPSC-SOX2 transfected with SOX2-siRNA showed a decrease of cell proliferation, migration and adhesion ability, which further confirmed the biological effect of SOX2 in human DPSCs. This study indicated that SOX2 could improve the cell proliferation, migration and adhesion ability of DPSCs through regulating gene expression about cell cycle, migration and adhesion, and provided a novel strategy to develop seed cells with strong proliferation, migration and adhesion ability for tissue engineering.

  15. Differentiation of human dental pulp stem cells into neuronal by resveratrol.

    PubMed

    Geng, Ya-Wei; Zhang, Zhen; Liu, Ming-Yue; Hu, Wei-Ping

    2017-08-07

    Dental pulp stem cells (DPSCs) have been proposed as a promising source of stem cells in nerve regeneration due to their close embryonic origin and ease of harvest. Resveratrol (RSV) is a natural polyphenolic and possesses many biological functions such as anti-inflammatory activity and protection against atherosclerosis and neuroprotective activities. There is increasing evidence showing that RSV plays a pivotal role in neuron protection and neuronal differentiation. In this study, we isolated DPSCs from impacted third molars and investigated whether RSV induces neuronal differentiation of DPSCs. To avoid loss of DPSCs multipotency, all the experiments were conducted on cells at early passages. RT-PCR results showed that RSV-treated DPSCs (RSV-DPSCs) significantly increased the expression of the neuroprogenitor marker Nestin. When RSV-DPSCs were differentiated with neuronal induction media (RSV-dDPSCs), they showed a cell morphology similar to neurons. The expression of neuronal-specific marker genes Nestin, Musashi, and NF-M in RSV-dDPSCs was significantly increased. Immunocytochemical staining and Western blot analysis showed that the expression of neuronal marker proteins, Nestin, and NF-M, was significantly increased in RSV-dDPSCs. Therefore, we have shown that RSV treatment, along with the use of neuronal induction media, effectively promotes neuronal cell differentiation of DPSCs. © 2017 International Federation for Cell Biology.

  16. Differentiation and Behavior of Dental Pulp Stem Cells in Hydrogel Scaffolds of Various Stiffnesses

    NASA Astrophysics Data System (ADS)

    Bhatnagar, Divya; Jurukovski, Vladimir; Rafailovich, Miriam; Simon, Marcia

    2011-03-01

    Dental Pulp Stem Cells (DPSCs) are known to differentiate in bone, dentine, or nerve tissue through different environment signals. This work investigates whether differentiation could occur in the absence of chemical induction and through mechanical stimuli only. For this study, we chose enzymatically cross-linked gelatin hydrogels as our substrates. Rheological studies carried out by oscillatory shear rheometry indicated that the modulus of the hardest hydrogel was of the order of 8kPa where as the medium and the softest hydrogel had modulus of the order of 1kPa and 100Pa respectively. DPSC were then plated on all three substrates and cultured with and without dexamethasone induction media. After 21 days of incubation, SEM analysis indicated that the cells cultured in the induction media produced biomineralized deposits on hard, medium as well as soft hydrogels. On the other hand, the cells cultured without the induction media also produced large amounts of biomineralized deposits.The modulus of the cells was also measured using AFM. En mass cell migration was also studied to determine the average velocity of migration of DPSCs. We also investigated whether stem cells that are induced to differentiate by their scaffold environment would continue to differentiate and biomineralize when removed from the inducing scaffold.

  17. Chronic Inflammation and Angiogenic Signaling Axis Impairs Differentiation of Dental-Pulp Stem Cells

    PubMed Central

    Boyle, Michael; Chun, Crystal; Strojny, Chelsee; Narayanan, Raghuvaran; Bartholomew, Amelia; Sundivakkam, Premanand; Alapati, Satish

    2014-01-01

    Dental-pulp tissue is often exposed to inflammatory injury. Sequested growth factors or angiogenic signaling proteins that are released following inflammatory injury play a pivotal role in the formation of reparative dentin. While limited or moderate angiogenesis may be helpful for dental pulp maintenance, the induction of significant level of angiogenesis is probably highly detrimental. Hitherto, several studies have addressed the effects of proinflammatory stimuli on the survival and differentiation of dental-pulp stem cells (DPSC), in vitro. However, the mechanisms communal to the inflammatory and angiogenic signaling involved in DPSC survival and differentiation remain unknown. Our studies observed that short-term exposure to TNF-α (6 and 12 hours [hrs]) induced apoptosis with an upregulation of VEGF expression and NF-κB signaling. However, long-term (chronic) exposure (14 days) to TNF-α resulted in an increased proliferation with a concomitant shortening of the telomere length. Interestingly, DPSC pretreated with Nemo binding domain (NBD) peptide (a cell permeable NF-κB inhibitor) significantly ameliorated TNF-α- and/or VEGF-induced proliferation and the shortening of telomere length. NBD peptide pretreatment significantly improved TNF-α-induced downregulation of proteins essential for differentiation, such as bone morphogenic proteins (BMP)-1 & 2, BMP receptor isoforms-1&2, trasnforming growth factor (TGF), osteoactivin and osteocalcin. Additionally, inhibition of NF-κB signaling markedly increased the mineralization potential, a process abrogated by chronic exposure to TNF-α. Thus, our studies demonstrated that chronic inflammation mediates telomere shortening via NF-κB signaling in human DPSC. Resultant chromosomal instability leads to an emergence of increased proliferation of DPSC, while negatively regulating the differentiation of DPSC, in vitro. PMID:25427002

  18. Potential dental pulp revascularization and odonto-/osteogenic capacity of a novel transplant combined with dental pulp stem cells and platelet-rich fibrin.

    PubMed

    Chen, Yong-Jin; Zhao, Yin-Hua; Zhao, Ya-Juan; Liu, Nan-Xia; Lv, Xin; Li, Qiang; Chen, Fa-Ming; Zhang, Min

    2015-08-01

    Our aim is to investigate the cytobiological effects of autologous platelet-rich fibrin (PRF) on dental pulp stem cells (DPSCs) and to explore the ectopic and orthotopic possibilities of dental pulp revascularization and pulp-dentin complex regeneration along the root canal cavities of the tooth by using a novel tissue-engineered transplant composed of cell-sheet fragments of DPSCs and PRF granules. Canine DPSCs were isolated and characterized by assaying their colony-forming ability and by determining their cell surface markers and osteogenic/adipogenic differentiation potential. The biological effects of autologous PRF on DPSCs, including cell proliferation, alkaline phosphatase (ALP) activity and odonto-/osteogenic gene expression, were then investigated and quantified. A novel transplant consisting of cell-sheet fragments of DPSCs and PRF granules was adopted to regenerate pulp-dentin-like tissues in the root canal, both subcutaneously in nude mice and in the roots of canines. PRF promoted the proliferation of DPSCs in a dose- and time-dependent manner and induced the differentiation of DPSCs to odonto-/osteoblastic fates by increasing the expression of the Alp, Dspp, Dmp1 and Bsp genes. Transplantation of the DPSC/PRF construct led both to a favorable regeneration of homogeneous and compact pulp-like tissues with abundantly distributed blood capillaries and to the deposition of regenerated dentin along the intracanal walls at 8 weeks post-operation. Thus, the application of DPSC/PRF tissue constructs might serve as a potential therapy in regenerative endodontics for pulp revitalization or revascularization.

  19. Attachment and proliferation of dental pulp stem cells on dentine treated with different regenerative endodontic protocols.

    PubMed

    Alghilan, M A; Windsor, L J; Palasuk, J; Yassen, G H

    2017-07-01

    To investigate the attachment and proliferation of dental pulp stem cells (DPSC) on dentine treated with various endodontic regeneration protocols. Standardized dentine samples were irrigated with sodium hypochlorite (1.5% NaOCl) and ethylenediaminetetraacetic acid (17% EDTA) and randomized into four treatment groups and two control groups. The treatment groups were treated with a clinically used concentration of triple antibiotic paste (TAP), double antibiotic paste (DAP), calcium hydroxide (Ca(OH)2 ) or diluted TAP in a methylcellulose system (DTAP) for 1 week. Each sample in the treatment groups was then irrigated with EDTA. The two control groups were treated with EDTA or received no treatment. Dental pulp stem cells were seeded on each dentine specimen (10 000 cells). Lactate dehydrogenase activity assays were then performed to evaluate the attached DPSC after 1 day of incubation. Water-soluble tetrazolium assays were used to determine DPSC proliferation after three additional days of incubation. Friedman's test followed by least significant difference were used for statistical analyses (α = 0.05). Triple antibiotic paste and DTAP regeneration protocols, as well as EDTA-treated dentine, caused significant increases in DPSC attachment to dentine. Triple antibiotic paste, DAP and Ca(OH)2 regeneration protocols caused significant reductions in DPSC proliferation on dentine. However, the DTAP regeneration protocol did not have any significant negative effects on DPSC proliferation. The clinically used endodontic regeneration protocols that include the use of TAP, DAP or Ca(OH)2 medicament negatively affected DPSC proliferation on dentine. However, the use of DTAP medicament during regenerative endodontic treatment may not adversely affect the proliferation of DPSC. © 2016 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  20. Epidermal growth factor enhances osteogenic differentiation of dental pulp stem cells in vitro.

    PubMed

    Del Angel-Mosqueda, Casiano; Gutiérrez-Puente, Yolanda; López-Lozano, Ada Pricila; Romero-Zavaleta, Ricardo Emmanuel; Mendiola-Jiménez, Andrés; Medina-De la Garza, Carlos Eduardo; Márquez-M, Marcela; De la Garza-Ramos, Myriam Angélica

    2015-09-03

    Epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) play an important role in extracellular matrix mineralization, a complex process required for proper bone regeneration, one of the biggest challenges in dentistry. The purpose of this study was to evaluate the osteogenic potential of EGF and bFGF on dental pulp stem cells (DPSCs). Human DPSCs were isolated using CD105 magnetic microbeads and characterized by flow cytometry. To induce osteoblast differentiation, the cells were cultured in osteogenic medium supplemented with EGF or bFGF at a low concentration. Cell morphology and expression of CD146 and CD10 surface markers were analyzed using fluorescence microscopy. To measure mineralization, an alizarin red S assay was performed and typical markers of osteoblastic phenotype were evaluated by RT-PCR. EGF treatment induced morphological changes and suppression of CD146 and CD10 markers. Additionally, the cells were capable of producing calcium deposits and increasing the mRNA expression to alkaline phosphatase (ALP) and osteocalcin (OCN) in relation to control groups (p < 0.001). However, bFGF treatment showed an inhibitory effect. These data suggests that DPSCs in combination with EGF could be an effective stem cell-based therapy for bone tissue engineering applications in periodontics and oral implantology.

  1. Promising cell-based therapy for bone regeneration using stem cells from deciduous teeth, dental pulp, and bone marrow.

    PubMed

    Yamada, Yoichi; Ito, Kenji; Nakamura, Sayaka; Ueda, Minoru; Nagasaka, Tetsuro

    2011-01-01

    We attempted to regenerate bone in a significant osseous defect with various stem cells from deciduous teeth, extracted from puppies, and grafted them into a parent canine mandible as an allograft, parent dental pulp, and bone marrow by tissue engineering and regenerative medicine technology using platelet-rich plasma as an autologous scaffold and signal molecules. Initially, teeth were extracted from a child and parent hybrid canine mandible region and bone marrow (canine mesenchymal stem cells; cMSCs), and parent teeth (canine dental pulp stem cells; cDPSCs), and stem cells were extracted from deciduous teeth (puppy deciduous teeth stem cells; pDTSCs). After 4 weeks, bone defects were prepared on both sides of the mandible with a trephine bar. Graft materials were implanted into these defects: 1) control (defect only), 2) platelet-rich plasma (PRP), 3) cMSCs/PRP, 4) cDPSCs/PRP, and 5) pDTSCs/PRP to investigate the effect of stem cells. The newly formed bones were evaluated by histology and histomorphometric analysis in the defects at 2, 4, and 8 weeks. According to histological observations, the cMSCs/PRP, cDPSCs/PRP, and pDTSCs/PRP groups had well-formed mature bone and neovascularization compared with the control (defect only) and PRP groups at 4 and 8 weeks, respectively, and the mineralized tissues in cMSCs/PRP, cDPSCs/PRP, and pDTSCs/PRP specimens were positive for osteocalcin at 8 weeks. Histometrically, newly formed bone areas were 19.0 ± 2.9% (control), 19.7 ± 6.0% (PRP), 52.8 ± 3.5% (cMSCs/PRP), 61.6 ± 1.3% (cDPSCs/PRP), and 54.7 ± 2.2% (pDTSCs/PRP) at 8 weeks. There were significant differences between cMSCs, cDPSCs, pDTSCs/PRP, and control and PRP groups. These results demonstrate that stem cells from deciduous teeth, dental pulp, and bone marrow with PRP have the ability to form bone, and bone formation with DTSCs might have the potential to generate a graft between a child and parent. This preclinical study could pave the way for stem cell

  2. Role of lipid rafts in neuronal differentiation of dental pulp-derived stem cells.

    PubMed

    Mattei, Vincenzo; Santacroce, Costantino; Tasciotti, Vincenzo; Martellucci, Stefano; Santilli, Francesca; Manganelli, Valeria; Piccoli, Luca; Misasi, Roberta; Sorice, Maurizio; Garofalo, Tina

    2015-12-10

    Human dental pulp-derived stem cells (hDPSCs) are characterized by a typical fibroblast-like morphology. They express specific markers for mesenchymal stem cells and are capable of differentiation into osteoblasts, adipoblasts and neurons in vitro. Previous studies showed that gangliosides are involved in the induction of early neuronal differentiation of hDPSCs. This study was undertaken to investigate the role of lipid rafts in this process. Lipid rafts are signaling microdomains enriched in glycosphingolipids, cholesterol, tyrosine kinase receptors, mono- or heterotrimeric G proteins and GPI-anchored proteins. We preliminary showed that established cells expressed multipotent mesenchymal stromal-specific surface antigens. Then, we analyzed the distribution of lipid rafts, revealing plasma membrane microdomains with GM2 and EGF-R enrichment. Following stimulation with EGF/bFGF, neuronal differentiation was observed. To analyze the functional role of lipid rafts in EGF/bFGF-induced hDPSCs differentiation, cells were preincubated with lipid raft affecting agents, i.e. [D]-PDMP or methyl-β-cyclodextrin. These compounds significantly prevented neuronal-specific antigen expression, as well as Akt and ERK 1/2 phosphorylation, induced by EGF/bFGF, indicating that lipid raft integrity is essential for EGF/bFGF-induced hDPSCs differentiation. These results suggest that lipid rafts may represent specific chambers, where multimolecular signaling complexes, including lipids (gangliosides, cholesterol) and proteins (EGF-R), play a role in hDPSCs differentiation.

  3. Growth kinetics and characterization of human dental pulp stem cells: Comparison between third molar and first premolar teeth

    PubMed Central

    Mehrabani, Davood; Mahdiyar, Parisa; Robati, Reza; Zare, Shahrokh; Dianatpour, Mehdi; Tamadon, Amin

    2017-01-01

    Background Dental pulp stem cells (DPSCs) play an important role in tissue regeneration. This study compares the growth kinetics and characterization of third molar and first premolar human DPSCs. Material and Methods Dental pulp tissues were isolated from human first premolar and third molar teeth and were digested by treating them with collagenase type I. Single-cell suspensions from each dental pulp were seeded in T25 culture flasks and the media were replaced every 3 days until 70% confluence. The cells were enumerated to determine the population doubling time (PDT). Cells were characterized using flow cytometry, RT-PCR and osteogenic medium for differentiation of DPSCs. Karyotyping assay was also performed till passage 7th. Results The DPSCs had spindle-shaped morphology. There was an increase in PDT in third molar DPSCs when compared to first premolar teeth. Positive expression of CD44, CD73, and CD90 and negative expression of CD34 and CD45 were illustrated. A normal karyotype was visible for all seven passages. The Alizarin red staining was positive for osteogenic induction of DPSCs. Conclusions When DPSCs are needed, third molar teeth can be a good and convenient candidate for cell transplantation, yielding high number of cells with mesenchymal characteristics. They can be a source for further investigations in vitro and work on tissue engineering protocols. Key words:Stem cells, dental pulp, growth kinetics, characterization. PMID:28210430

  4. Comparative analysis of proliferation and differentiation potentials of stem cells from inflamed pulp of deciduous teeth and stem cells from exfoliated deciduous teeth.

    PubMed

    Yu, Shi; Diao, Shu; Wang, Jinsong; Ding, Gang; Yang, Dongmei; Fan, Zhipeng

    2014-01-01

    Stem cells isolated from exfoliated deciduous teeth (SHEDs) are highly capable of proliferation and differentiation, and they represent good cell sources for mesenchymal stem cell- (MSC-) mediated dental tissue regeneration, but the supply of SHEDs is limited. A previous study found that stem cells could be isolated from inflamed tissues, but it is unknown whether primary dental pulp diagnosed with irreversible pulpitis might contain stem cells with appropriate tissue regeneration capacity. In this study, we aimed to isolate stem cells from both inflamed pulps of deciduous teeth (SCIDs) and SHEDs from Chinese children and to compare their proliferation and differentiation potentials. Our results showed that SCIDs were positive for cell surface markers, including CD105, CD90, and CD146, and they had high proliferation ability and osteogenic, adipogenic, and chondrogenic differentiation potentials. There was no significant difference in proliferation and differentiation potentials between SCIDs and SHEDs. The mRNA of inflammatory factors, including IL-1β, IL-6, and TNF-α, was expressed at similar levels in SCIDs and SHEDs, but SCIDs secreted more TNF-α protein. In conclusion, our in vitro results showed that SCIDs have proliferation and differentiation potentials similar to those of SHEDs. Thus, SCIDs represent a new potentially applicable source for MSC mediated tissue regeneration.

  5. Effect of dentin treatment on proliferation and differentiation of human dental pulp stem cells

    PubMed Central

    Park, Minjeong; Pang, Nan-Sim

    2015-01-01

    Objectives Sodium hypochlorite (NaOCl) is an excellent bactericidal agent, but it is detrimental to stem cell survival, whereas intracanal medicaments such as calcium hydroxide (Ca[OH]2) promote the survival and proliferation of stem cells. This study evaluated the effect of sequential NaOCl and Ca[OH]2 application on the attachment and differentiation of dental pulp stem cells (DPSCs). Materials and Methods DPSCs were obtained from human third molars. All dentin specimens were treated with 5.25% NaOCl for 30 min. DPSCs were seeded on the dentin specimens and processed with additional 1 mg/mL Ca[OH]2, 17% ethylenediaminetetraacetic acid (EDTA) treatment, file instrumentation, or a combination of these methods. After 7 day of culture, we examined DPSC morphology using scanning electron microscopy and determined the cell survival rate with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. We measured cell adhesion gene expression levels after 4 day of culture and odontogenic differentiation gene expression levels after 4 wk using quantitative real-time polymerase chain reaction. Results DPSCs did not attach to the dentin in the NaOCl-treated group. The gene expression levels of fibronectin-1 and secreted phosphoprotein-1 gene in both the Ca[OH]2- and the EDTA-treated groups were significantly higher than those in the other groups. All Ca[OH]2-treated groups showed higher expression levels of dentin matrix protein-1 than that of the control. The dentin sialophosphoprotein level was significantly higher in the groups treated with both Ca[OH]2 and EDTA. Conclusions The application of Ca[OH]2 and additional treatment such as EDTA or instrumentation promoted the attachment and differentiation of DPSCs after NaOCl treatment. PMID:26587415

  6. Dental pulp stem cells as a multifaceted tool for bioengineering and the regeneration of craniomaxillofacial tissues

    PubMed Central

    Aurrekoetxea, Maitane; Garcia-Gallastegui, Patricia; Irastorza, Igor; Luzuriaga, Jon; Uribe-Etxebarria, Verónica; Unda, Fernando; Ibarretxe, Gaskon

    2015-01-01

    Dental pulp stem cells, or DPSC, are neural crest-derived cells with an outstanding capacity to differentiate along multiple cell lineages of interest for cell therapy. In particular, highly efficient osteo/dentinogenic differentiation of DPSC can be achieved using simple in vitro protocols, making these cells a very attractive and promising tool for the future treatment of dental and periodontal diseases. Among craniomaxillofacial organs, the tooth and salivary gland are two such cases in which complete regeneration by tissue engineering using DPSC appears to be possible, as research over the last decade has made substantial progress in experimental models of partial or total regeneration of both organs, by cell recombination technology. Moreover, DPSC seem to be a particularly good choice for the regeneration of nerve tissues, including injured or transected cranial nerves. In this context, the oral cavity appears to be an excellent testing ground for new regenerative therapies using DPSC. However, many issues and challenges need yet to be addressed before these cells can be employed in clinical therapy. In this review, we point out some important aspects on the biology of DPSC with regard to their use for the reconstruction of different craniomaxillofacial tissues and organs, with special emphasis on cranial bones, nerves, teeth, and salivary glands. We suggest new ideas and strategies to fully exploit the capacities of DPSC for bioengineering of the aforementioned tissues. PMID:26528190

  7. Tooth storage, dental pulp stem cell isolation, and clinical scale expansion without animal serum.

    PubMed

    Eubanks, Emily J; Tarle, Susan A; Kaigler, Darnell

    2014-05-01

    Dental pulp stem cells (DPSCs) have therapeutic potential for dentin and dental pulp regeneration. For regenerative approaches to gain clinical acceptance, protocols are needed to determine feasible ways to store teeth, isolate DPSCs, and expand them to clinical scale numbers. In this study, 32 third molars were obtained from patients and immediately placed in saline or tissue culture medium followed by overnight storage at 4°C or immediate isolation of DPSCs. Upon isolation, cells were expanded in medium containing either fetal bovine serum (FBS) or human serum (HS). Cell proliferation (population doubling time [PDT]), cell surface marker expression, and multipotency were compared between DPSCs in FBS and DPSCs in HS. The time frame of storage and storage medium did not affect the ability to isolate DPSCs. However, using HS instead of FBS in the initial isolation of DPSCs significantly decreased (P < .01) the isolation success rate from 89% (FBS) to 23% (HS). Yet, incorporating fibronectin in the DPSC initial isolation (using HS) significantly (P < .01) increased the isolation success rate to 83%. Interestingly, it was found that the proliferation rate was significantly (P < .05) higher for DPSCs in HS (PDT = 1.59 ± 0.46) than that for DPSCs in FBS (PDT = 2.84 ± 2.5). Finally, there was no difference in the expression of CD73, CD90, CD105, or multipotency (as measured by osteogenic, adipogenic, and chondrogenic differentiation) between DPSCs in FBS and DPSCs in HS. These findings show a clinically feasible method of storing third molars for the isolation of DPSCs. Additionally, DPSCs can be isolated and expanded to clinical scale numbers in media devoid of FBS and still maintain their phenotypic properties. Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  8. Accelerated craniofacial bone regeneration through dense collagen gel scaffolds seeded with dental pulp stem cells

    NASA Astrophysics Data System (ADS)

    Chamieh, Frédéric; Collignon, Anne-Margaux; Coyac, Benjamin R.; Lesieur, Julie; Ribes, Sandy; Sadoine, Jérémy; Llorens, Annie; Nicoletti, Antonino; Letourneur, Didier; Colombier, Marie-Laure; Nazhat, Showan N.; Bouchard, Philippe; Chaussain, Catherine; Rochefort, Gael Y.

    2016-12-01

    Therapies using mesenchymal stem cell (MSC) seeded scaffolds may be applicable to various fields of regenerative medicine, including craniomaxillofacial surgery. Plastic compression of collagen scaffolds seeded with MSC has been shown to enhance the osteogenic differentiation of MSC as it increases the collagen fibrillary density. The aim of the present study was to evaluate the osteogenic effects of dense collagen gel scaffolds seeded with mesenchymal dental pulp stem cells (DPSC) on bone regeneration in a rat critical-size calvarial defect model. Two symmetrical full-thickness defects were created (5 mm diameter) and filled with either a rat DPSC-containing dense collagen gel scaffold (n = 15), or an acellular scaffold (n = 15). Animals were imaged in vivo by microcomputer tomography (Micro-CT) once a week during 5 weeks, whereas some animals were sacrificed each week for histology and histomorphometry analysis. Bone mineral density and bone micro-architectural parameters were significantly increased when DPSC-seeded scaffolds were used. Histological and histomorphometrical data also revealed significant increases in fibrous connective and mineralized tissue volume when DPSC-seeded scaffolds were used, associated with expression of type I collagen, osteoblast-associated alkaline phosphatase and osteoclastic-related tartrate-resistant acid phosphatase. Results demonstrate the potential of DPSC-loaded-dense collagen gel scaffolds to benefit of bone healing process.

  9. Accelerated craniofacial bone regeneration through dense collagen gel scaffolds seeded with dental pulp stem cells

    PubMed Central

    Chamieh, Frédéric; Collignon, Anne-Margaux; Coyac, Benjamin R.; Lesieur, Julie; Ribes, Sandy; Sadoine, Jérémy; Llorens, Annie; Nicoletti, Antonino; Letourneur, Didier; Colombier, Marie-Laure; Nazhat, Showan N.; Bouchard, Philippe; Chaussain, Catherine; Rochefort, Gael Y.

    2016-01-01

    Therapies using mesenchymal stem cell (MSC) seeded scaffolds may be applicable to various fields of regenerative medicine, including craniomaxillofacial surgery. Plastic compression of collagen scaffolds seeded with MSC has been shown to enhance the osteogenic differentiation of MSC as it increases the collagen fibrillary density. The aim of the present study was to evaluate the osteogenic effects of dense collagen gel scaffolds seeded with mesenchymal dental pulp stem cells (DPSC) on bone regeneration in a rat critical-size calvarial defect model. Two symmetrical full-thickness defects were created (5 mm diameter) and filled with either a rat DPSC-containing dense collagen gel scaffold (n = 15), or an acellular scaffold (n = 15). Animals were imaged in vivo by microcomputer tomography (Micro-CT) once a week during 5 weeks, whereas some animals were sacrificed each week for histology and histomorphometry analysis. Bone mineral density and bone micro-architectural parameters were significantly increased when DPSC-seeded scaffolds were used. Histological and histomorphometrical data also revealed significant increases in fibrous connective and mineralized tissue volume when DPSC-seeded scaffolds were used, associated with expression of type I collagen, osteoblast-associated alkaline phosphatase and osteoclastic-related tartrate-resistant acid phosphatase. Results demonstrate the potential of DPSC-loaded-dense collagen gel scaffolds to benefit of bone healing process. PMID:27934940

  10. Enhanced Chondrogenic Differentiation of Dental Pulp Stem Cells Using Nanopatterned PEG-GelMA-HA Hydrogels

    PubMed Central

    Nemeth, Cameron L.; Janebodin, Kajohnkiart; Yuan, Alex E.; Dennis, James E.

    2014-01-01

    We have examined the effects of surface nanotopography and hyaluronic acid (HA) on in vitro chondrogenesis of dental pulp stem cells (DPSCs). Ultraviolet-assisted capillary force lithography was employed to fabricate well-defined nanostructured scaffolds of composite PEG-GelMA-HA hydrogels that consist of poly(ethylene glycol) dimethacrylate (PEGDMA), methacrylated gelatin (GelMA), and HA. Using this microengineered platform, we first demonstrated that DPSCs formed three-dimensional spheroids, which provide an appropriate environment for in vitro chondrogenic differentiation. We also found that DPSCs cultured on nanopatterned PEG-GelMA-HA scaffolds showed a significant upregulation of the chondrogenic gene markers (Sox9, Alkaline phosphatase, Aggrecan, Procollagen type II, and Procollagen type X), while downregulating the pluripotent stem cell gene, Nanog, and epithelial–mesenchymal genes (Twist, Snail, Slug) compared with tissue culture polystyrene-cultured DPSCs. Immunocytochemistry showed more extensive deposition of collagen type II in DPSCs cultured on the nanopatterned PEG-GelMA-HA scaffolds. These findings suggest that nanotopography and HA provide important cues for promoting chondrogenic differentiation of DPSCs. PMID:24749806

  11. Treatment of osteoradionecrosis of mandible with bone marrow concentrate and with dental pulp stem cells.

    PubMed

    Manimaran, K; Sankaranarayanan, S; Ravi, V R; Elangovan, S; Chandramohan, M; Perumal, S Mahendra

    2014-01-01

    Osteoradionecrosis (ORN) is a noninfectious, necrotic condition of the bone occurring as a complication of radiotherapy. Most cases occur following trauma or surgical manipulation of the irradiated site. Mandible is the most common bone to be affected following head and neck irradiation. The aim was to develop a successful therapeutic approach for ORN. A spectrum of treatment modalities is practiced for ORN with variable success rate that includes simple irrigation of the affected bone to the partial or complete resection of the jaw bone. In this paper, we present two cases which had successful therapeutic approach for ORN of mandible with autologous bone marrow concentrate stem cells and allogeneic dental pulp stem cells (DPSC) with platelet rich plasma (PRP) following failure of conventional methods. Autologous bone marrow aspirate concentrate (BMAC) was injected around the socket and into the periosteum for one case, and DPSC were mixed with tricalcium phosphate and inserted at the site of the defect in one case. The patient treated with BMAC remained asymptomatic and complete bone remodeling was noticed after 1 year. The extraoral sinus was excised, and healing was uneventful without recurrence in the patient treated with allogeneic DPSC and PRP. Periodic panoramic radiographs revealed an appreciable bone formation from the 2(nd) month onward. We have successfully treated two cases of ORN with BMAC and DPSC, respectively.

  12. Effects of equiaxial strain on the differentiation of dental pulp stem cells without using biochemical reagents.

    PubMed

    Tabatabaei, F S; Jazayeri, M; Ghahari, P; Haghighipour, N

    2014-09-01

    During orthodontic treatments, applied mechanical forces create strain and result in tooth movement through the alveolar bone. This response to mechanical strain is a fundamental biological reaction. The present study evaluated the effect of equiaxial strain within the range of orthodontic forces on the osteogenic differentiation of human dental pulp stem cells (hDPSCs). Following isolation and culture of hDPSCs, 3rd passage cells were transferred on a silicone membrane covered with collagen. Cell adhesion to the membrane was evaluated under scanning electron microscope (SEM). Cells were divided into three groups: the first group was placed in a conventional culture medium, transferred to an equiaxial stretching device (3% strain for 2 weeks). The positive control was placed in an osteogenic medium with no mechanical strain. The negative control group was placed in the conventional culture medium with no mechanical strain either. Study groups were evaluated for expression ofosteogenic markers (Alkaline phosphatase and Osteopontin) with immunofluorescence and real time PCR. SEM images revealed optimal adhesion of cells to the silicone membrane. Immunofluorescence study demonstrated that osteocalcin expression occurred after 2 weeks in the two groups under mechanical and chemical signals. After application of equiaxial strain, level of expression of osteogenic markers was significantly higher than in the negative and positive control groups. Based on the study results, static equiaxial strain which mimics the types of orthodontic forces can result in differentiation of hDPSCs to osteoblasts. The results obtained may be used in cell therapy and tissue engineering.

  13. Transplantation of Human Dental Pulp-Derived Stem Cells or Differentiated Neuronal Cells from Human Dental Pulp-Derived Stem Cells Identically Enhances Regeneration of the Injured Peripheral Nerve.

    PubMed

    Ullah, Imran; Park, Ju-Mi; Kang, Young-Hoon; Byun, June-Ho; Kim, Dae-Geon; Kim, Joo-Heon; Kang, Dong-Ho; Rho, Gyu-Jin; Park, Bong-Wook

    2017-09-01

    Human dental mesenchymal stem cells isolated from the dental follicle, pulp, and root apical papilla of extracted wisdom teeth have been known to exhibit successful and potent neurogenic differentiation capacity. In particular, human dental pulp-derived stem cells (hDPSCs) stand out as the most prominent source for in vitro neuronal differentiation. In this study, to evaluate the in vivo peripheral nerve regeneration potential of hDPSCs and differentiated neuronal cells from DPSCs (DF-DPSCs), a total of 1 × 10(6) hDPSCs or DF-hDPSCs labeled with PKH26 tracking dye and supplemented with fibrin glue scaffold and collagen tubulization were transplanted into the sciatic nerve resection (5-mm gap) of rat models. At 12 weeks after cell transplantation, both hDPSC and DF-hDPSC groups showed notably increased behavioral activities and higher muscle contraction forces compared with those in the non-cell transplanted control group. In immunohistochemical analysis of regenerated nerve specimens, specific markers for angiogenesis, axonal fiber, and myelin sheath increased in both the cell transplantation groups. Pretransplanted labeled PKH26 were also distinctly detected in the regenerated nerve tissues, indicating that transplanted cells were well-preserved and differentiated into nerve cells. Furthermore, no difference was observed in the nerve regeneration potential between the hDPSC and DF-hDPSC transplanted groups. These results demonstrate that dental pulp tissue is an excellent stem cell source for nerve regeneration, and in vivo transplantation of the undifferentiated hDPSCs could exhibit sufficient and excellent peripheral nerve regeneration potential.

  14. The effect of Aloe vera gel on viability of dental pulp stem cells.

    PubMed

    Sholehvar, Fatemeh; Mehrabani, Davood; Yaghmaei, Parichehr; Vahdati, Akbar

    2016-10-01

    Dental pulp stem cells (DPSCs) can play a prominent role in tissue regeneration. Aloe vera L. (Liliaceae) contains the polysaccharide of acemannan that was shown to be a trigger factor for cell proliferation, differentiation, mineralization, and dentin formation. This study sought to determine the viability of DPSCs in Aloe vera in comparison with Hank's balanced salt solution (HBSS). Twelve rabbits underwent anesthesia, and their incisor teeth were extracted; the pulp tissue was removed, chopped, treated with collagenase and plated in culture flasks. DPSCs from passage 3 were cultured in 24-well plates, and after 3 days, the culture media changed to 10, 25, 50, and 100% concentrations of Aloe vera at intervals of 45 and 90 min and 3 and 6 h. Distilled water was used as negative and HBSS as positive control for comparison. The cell morphology, viability, population doubling time (PDT), and growth kinetics were evaluated. RT-PCR was carried out for characterization and karyotyping for chromosomal stability. Aloe vera showed a significant higher viability than HBSS (74.74%). The 50% Aloe vera showed higher viability (97.73%) than other concentrations. PDT in 50% concentration was 35.1 h and for HBSS was 49.5 h. DPSCs were spindle shaped and were positive for CD73 and negative for CD34 and CD45. Karyotyping was normal. Aloe vera as an inexpensive and available herb can improve survival of avulsed or broken teeth in emergency cases as a transfer media. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  15. In vitro osteogenic and odontogenic differentiation of human dental pulp stem cells seeded on carboxymethyl cellulose-hydroxyapatite hybrid hydrogel

    PubMed Central

    Teti, Gabriella; Salvatore, Viviana; Focaroli, Stefano; Durante, Sandra; Mazzotti, Antonio; Dicarlo, Manuela; Mattioli-Belmonte, Monica; Orsini, Giovanna

    2015-01-01

    Stem cells from human dental pulp have been considered as an alternative source of adult stem cells in tissue engineering because of their potential to differentiate into multiple cell lineages. Recently, polysaccharide based hydrogels have become especially attractive as matrices for the repair and regeneration of a wide variety of tissues and organs. The incorporation of inorganic minerals as hydroxyapatite nanoparticles can modulate the performance of the scaffolds with potential applications in tissue engineering. The aim of this study was to verify the osteogenic and odontogenic differentiation of dental pulp stem cells (DPSCs) cultured on a carboxymethyl cellulose—hydroxyapatite hybrid hydrogel. Human DPSCs were seeded on carboxymethyl cellulose—hydroxyapatite hybrid hydrogel and on carboxymethyl cellulose hydrogel for 1, 3, 5, 7, 14, and 21 days. Cell viability assay and ultramorphological analysis were carried out to evaluate biocompatibility and cell adhesion. Real Time PCR was carried out to demonstrate the expression of osteogenic and odontogenic markers. Results showed a good adhesion and viability in cells cultured on carboxymethyl cellulose—hydroxyapatite hybrid hydrogel, while a low adhesion and viability was observed in cells cultured on carboxymethyl cellulose hydrogel. Real Time PCR data demonstrated a temporal up-regulation of osteogenic and odontogenic markers in dental pulp stem cells cultured on carboxymethyl cellulose—hydroxyapatite hybrid hydrogel. In conclusion, our in vitro data confirms the ability of DPSCs to differentiate toward osteogenic and odontogenic lineages in presence of a carboxymethyl cellulose—hydroxyapatite hybrid hydrogel. Taken together, our results provide evidence that DPSCs and carboxymethyl cellulose—hydroxyapatite hybrid hydrogel could be considered promising candidates for dental pulp complex and periodontal tissue engineering. PMID:26578970

  16. ReNCell VM conditioned medium enhances the induction of dental pulp stem cells into dopaminergic like cells.

    PubMed

    Gnanasegaran, Nareshwaran; Govindasamy, Vijayendran; Musa, Sabri; Abu Kasim, Noor Hayaty

    2016-03-01

    Among the debilitating diseases, neurological related diseases are the most challenging ones to be treated using cell replacement therapies. Recently, dental pulp stem cells (SHED) were found to be most suitable cell choice for neurological related diseases as evidenced with many preclinical studies. To enhance the neurological potential of SHED, we recapitulated one of the pharmacological therapeutic tools in cell replacement treatment, we pre-conditioned dental pulp stem cells (SHED) with culture medium of ReNCell VM, an immortalized neuron progenitor cell, prior to neurogenesis induction and investigated whether this practice enhances their neurogenesis potential especially towards dopaminergic neurons. We hypothesed that the integration of pharmacological practices such as co-administration of various drugs, a wide range of doses and duration as well as pre-conditioning into cell replacement may enhance the efficacy of stem cell therapy. In particular, pre-conditioning is shown to be involved in the protective effect from some membrano-tropic drugs, thereby improving the resistance of cell structures and homing capabilities. We found that cells pre-treated with ReNCell VM conditioned medium displayed bipolar structures with extensive branches resembling putative dopaminergic neurons as compared to non-treated cells. Furthermore, many neuronal related markers such as NES, NR4A2, MSI1, and TH were highly expressed (fold changes > 2; p < 0.05) in pre-treated cells. Similar observations were detected at the protein level. The results demonstrate for the first time that SHED pre-conditioning enhances neurological potential and we suggest that cells should be primed to their respective environment prior to transplantation.

  17. Dental Pulp Stem Cell Differentiation on Poly-4-vinyl-pyridine surfaces

    NASA Astrophysics Data System (ADS)

    Suarato, Giulia; Bherwani, Aneel; Chang, Chung-Chueh; Rafailovich, Miriam; Simon, Marcia

    2012-02-01

    In the regeneration of a natural tissue, the mechanics and the chemical properties of the artificial substrate play a critical role. In this study, the influence of poly-4-vinyl-pyridine scaffold morphology on dental pulp stem cell differentiation was analyzed. Cells were plated on spun cast films and electrospun fibers with diameters ranging from nano to micrometers. Confocal microscopy showed the presence of various cell morphologies: on microfibers cells conform precisely to the main axis of elongation, while on nanometric scaffolds they result spread and in contact with several fibers. Even if the surface chemistry was identical, a great variation in the curvature was present. From day 9 of incubation, spontaneous biomineralization in the absence of induction agents occurred only on the fibrous structures. The SEM revealed template deposits directly on the microfibers, while on the nanofibers large spherical islands were also present. EDAX determined hydroxyl apatite nature of the deposits. RT-PCR indicated upregulation of osteogenic markers, confirming differentiation. SEM also revealed the presence of ECM fibers covering the polymer structure, which may enhance the expression of focal adhesion sites on the cell membrane.

  18. Neurogenic potential of dental pulp stem cells isolated from murine incisors

    PubMed Central

    2014-01-01

    Introduction Interest in the use of dental pulp stem cells (DPSC) to enhance neurological recovery following stroke and traumatic injury is increasing following successful pre-clinical studies. A murine model of autologous neural stem cell transplantation would be useful for further pre-clinical investigation of the underlying mechanisms. However, while human-derived DPSC have been well characterised, the neurogenic potential of murine DPSC (mDPSC) has been largely neglected. In this study we demonstrate neuronal differentiation of DPSC from murine incisors in vitro. Methods mDPSC were cultured under neuroinductive conditions and assessed for neuronal and glial markers and electrophysiological functional maturation. Results mDPSC developed a neuronal morphology and high expression of neural markers nestin, ßIII-tubulin and GFAP. Neurofilament M and S100 were found in lower abundance. Differentiated cells also expressed protein markers for cholinergic, GABAergic and glutaminergic neurons, indicating a mixture of central and peripheral nervous system cell types. Intracellular electrophysiological analysis revealed the presence of voltage-gated L-type Ca2+ channels in a majority of cells with neuronal morphology. No voltage-gated Na+ or K+ currents were found and the cultures did not support spontaneous action potentials. Neuronal-like networks expressed the gap junction protein, connexin 43 but this was not associated with dye coupling between adjacent cells after injection of the low-molecular weight tracers Lucifer yellow or Neurobiotin. This indicated that the connexin proteins were not forming traditional gap junction channels. Conclusions The data presented support the differentiation of mDPSC into immature neuronal-like networks. PMID:24572146

  19. Mechanosensitivity of dental pulp stem cells is related to their osteogenic maturity.

    PubMed

    Kraft, David C E; Bindslev, Dorthe A; Melsen, Birte; Abdallah, Basem M; Kassem, Moustapha; Klein-Nulend, Jenneke

    2010-02-01

    For engineering bone tissue, mechanosensitive cells are needed for bone (re)modelling. Local bone mass and architecture are affected by mechanical loading, which provokes a cellular response via loading-induced interstitial fluid flow. We studied whether human dental pulp-derived mesenchymal stem cells (PDSCs) portraying mature (PDSC-mature) or immature (PDSC-immature) bone cell characteristics are responsive to pulsating fluid flow (PFF) in vitro. We also assessed bone formation by PDSCs on hydroxyapatite-tricalcium phosphate granules after subcutaneous implantation in mice. Cultured PDSC-mature exhibited higher osteocalcin and alkaline phosphatase gene expression and activity than PDSC-immature. Pulsating fluid flow (PFF) stimulated nitric oxide production within 5 min by PDSC-mature but not by PDSC-immature. In PDSC-mature, PFF induced prostaglandin E(2) production, and cyclooxygenase 2 gene expression was higher than in PDSC-immature. Implantation of PDSC-mature resulted in more osteoid deposition and lamellar bone formation than PDSC-immature. We conclude that PDSCs with a mature osteogenic phenotype are more responsive to pulsating fluid shear stress than osteogenically immature PDSCs and produce more bone in vivo. These data suggest that PDSCs with a mature osteogenic phenotype might be preferable for bone tissue engineering to restore, for example, maxillofacial defects, because they might be able to perform mature bone cell-specific functions during bone adaptation to mechanical loading in vivo.

  20. Cellular Responses in Human Dental Pulp Stem Cells Treated with Three Endodontic Materials

    PubMed Central

    Ibañez-Cabellos, José Santiago; de Cutanda, Sergio Bañuls-Sánchez; Berenguer-Pascual, Ester; Beltrán-García, Jesús; García-López, Eva; Pallardó, Federico V.; García-Giménez, José Luis; Pallarés-Sabater, Antonio; Zarzosa-López, Ignacio; Monterde, Manuel

    2017-01-01

    Human dental pulp stem cells (HDPSCs) are of special relevance in future regenerative dental therapies. Characterizing cytotoxicity and genotoxicity produced by endodontic materials is required to evaluate the potential for regeneration of injured tissues in future strategies combining regenerative and root canal therapies. This study explores the cytotoxicity and genotoxicity mediated by oxidative stress of three endodontic materials that are widely used on HDPSCs: a mineral trioxide aggregate (MTA-Angelus white), an epoxy resin sealant (AH-Plus cement), and an MTA-based cement sealer (MTA-Fillapex). Cell viability and cell death rate were assessed by flow cytometry. Oxidative stress was measured by OxyBlot. Levels of antioxidant enzymes were evaluated by Western blot. Genotoxicity was studied by quantifying the expression levels of DNA damage sensors such as ATM and RAD53 genes and DNA damage repair sensors such as RAD51 and PARP-1. Results indicate that AH-Plus increased apoptosis, oxidative stress, and genotoxicity markers in HDPSCs. MTA-Fillapex was the most cytotoxic oxidative stress inductor and genotoxic material for HDPSCs at longer times in preincubated cell culture medium, and MTA-Angelus was less cytotoxic and genotoxic than AH-Plus and MTA-Fillapex at all times assayed. PMID:28751918

  1. In Vitro and In Vivo Differentiation of Progenitor Stem Cells Obtained After Mechanical Digestion of Human Dental Pulp.

    PubMed

    Monti, Manuela; Graziano, Antonio; Rizzo, Silvana; Perotti, Cesare; Del Fante, Claudia; d'Aquino, Riccardo; Redi, Carlo Alberto; Rodriguez Y Baena, Ruggero

    2017-03-01

    Human population is facing a revolutionary change in the demographic structure with an increasing number of elderly people requiring an unmet need to ensure a smooth aging process and dental care is certainly an important aspect that has to be considered. To date, dentistry has been conservative and the need of transferring the scientific models of regenerative dentistry into clinical practice is becoming a necessity. The aim of this study was to characterize the differentiation commitment (in vitro) and the clinical grafting ability (in vivo) of a population of progenitor stem cells obtained after mechanical digestion of dental pulp with an innovative system recently developed. This approach was successfully used in previous studies to obtain a clinical-grade ready to use dental pulp fragments that could be grafted in autologous tissues to obtain bone. We are thus showing that micro grafts resulting from mechanical digestion contain stem cells with a mesenchymal phenotype, able to differentiate toward different cell types and to generate new bone in patients. We are providing data for the establishment of standardized and routinely oral surgery approaches, having outlined the cellular properties of human stem cells obtained from the dental pulp. This method can represent a valid tool for both regenerative medicine and tissue engineering purposes not only applicable to the cranio-maxillofacial region but, likely, to different bone pathologies for a fastening and healing recovering of patients. J. Cell. Physiol. 232: 548-555, 2017. © 2016 Wiley Periodicals, Inc.

  2. Stem cell regulatory gene expression in human adult dental pulp and periodontal ligament cells undergoing odontogenic/osteogenic differentiation.

    PubMed

    Liu, Lu; Ling, Junqi; Wei, Xi; Wu, Liping; Xiao, Yin

    2009-10-01

    During development and regeneration, odontogenesis and osteogenesis are initiated by a cascade of signals driven by several master regulatory genes. In this study, we investigated the differential expression of 84 stem cell-related genes in dental pulp cells (DPCs) and periodontal ligament cells (PDLCs) undergoing odontogenic/osteogenic differentiation. Our results showed that, although there was considerable overlap, certain genes had more differential expression in PDLCs than in DPCs. CCND2, DLL1, and MME were the major upregulated genes in both PDLCs and DPCs, whereas KRT15 was the only gene significantly downregulated in PDLCs and DPCs in both odontogenic and osteogenic differentiation. Interestingly, a large number of regulatory genes in odontogenic and osteogenic differentiation interact or crosstalk via Notch, Wnt, transforming growth factor beta (TGF-beta)/bone morphogenic protein (BMP), and cadherin signaling pathways, such as the regulation of APC, DLL1, CCND2, BMP2, and CDH1. Using a rat dental pulp and periodontal defect model, the expression and distribution of both BMP2 and CDH1 have been verified for their spatial localization in dental pulp and periodontal tissue regeneration. This study has generated an overview of stem cell-related gene expression in DPCs and PDLCs during odontogenic/osteogenic differentiation and revealed that these genes may interact through the Notch, Wnt, TGF-beta/BMP, and cadherin signaling pathways to play a crucial role in determining the fate of dental derived cell and dental tissue regeneration. These findings provided a new insight into the molecular mechanisms of the dental tissue mineralization and regeneration.

  3. Nanofibrous spongy microspheres enhance odontogenic differentiation of human dental pulp stem cells.

    PubMed

    Kuang, Rong; Zhang, Zhanpeng; Jin, Xiaobing; Hu, Jiang; Gupte, Melanie J; Ni, Longxing; Ma, Peter X

    2015-09-16

    Dentin regeneration is challenging due to its complicated anatomical structure and the shortage of odontoblasts. In this study, a novel injectable cell carrier, nanofibrous spongy microspheres (NF-SMS), is developed for dentin regeneration. Biodegradable and biocompatible poly(l-lactic acid)-block-poly(l-lysine) are synthesized and fabricated into NF-SMS using self-assembly and thermally induced phase separation techniques. It is hypothesized that NF-SMS with interconnected pores and nanofibers can enhance the proliferation and odontogenic differentiation of human dental pulp stem cells (hDPSCs), compared to nanofibrous microspheres (NF-MS) without pore structure and conventional solid microspheres (S-MS) with neither nanofibers nor pore structure. During the first 9 d in culture, hDPSCs proliferate significantly faster on NF-SMS than on NF-MS or S-MS (p < 0.05). Following in vitro odontogenic induction, all the examined odontogenic genes (alkaline phosphatase content, osteocalcin, bone sialoprotein, collagen 1, dentin sialophosphoprotein (DSPP)), calcium content, and DSPP protein content are found significantly higher in the NF-SMS group than in the control groups. Furthermore, 6 weeks after subcutaneous injection of hDPSCs and microspheres into nude mice, histological analysis shows that NF-SMS support superior dentin-like tissue formation compared to NF-MS or S-MS. Taken together, NF-SMS have great potential as an injectable cell carrier for dentin regeneration. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Human Adult Dental Pulp Stem Cells Enhance Poststroke Functional Recovery Through Non-Neural Replacement Mechanisms

    PubMed Central

    Leong, Wai Khay; Henshall, Tanya L.; Arthur, Agnes; Kremer, Karlea L.; Lewis, Martin D.; Helps, Stephen C.; Field, John; Hamilton-Bruce, Monica A.; Warming, Scott; Manavis, Jim; Vink, Robert; Gronthos, Stan

    2012-01-01

    Human adult dental pulp stem cells (DPSCs), derived from third molar teeth, are multipotent and have the capacity to differentiate into neurons under inductive conditions both in vitro and following transplantation into the avian embryo. In this study, we demonstrate that the intracerebral transplantation of human DPSCs 24 hours following focal cerebral ischemia in a rodent model resulted in significant improvement in forelimb sensorimotor function at 4 weeks post-treatment. At this time, 2.3 ± 0.7% of engrafted cells had survived in the poststroke brain and demonstrated targeted migration toward the stroke lesion. In the peri-infarct striatum, transplanted DPSCs differentiated into astrocytes in preference to neurons. Our data suggest that the dominant mechanism of action underlying DPSC treatment that resulted in enhanced functional recovery is unlikely to be due to neural replacement. Functional improvement is more likely to be mediated through DPSC-dependent paracrine effects. This study provides preclinical evidence for the future use of human DPSCs in cell therapy to improve outcome in stroke patients. PMID:23197777

  5. Interaction of dental pulp stem cells with Biodentine and MTA after exposure to different environments

    PubMed Central

    Agrafioti, Anastasia; Taraslia, Vasiliki; Chrepa, Vanessa; Lymperi, Stefania; Panopoulos, Panos; Anastasiadou, Ema; Kontakiotis, Evangelos G.

    2016-01-01

    ABSTRACT Objective: The aim of the present study was to evaluate and compare the cytotoxic effects of Biodentine and MTA on dental pulp stem cells (DPSCs) and to assess cell viability and adherence after material exposure to an acidic environment. Material and Methods: DPSCs were cultured either alone or in contact with either: Biodentine; MTA set for 1 hour; or MTA set for 24 hours. After 4 and 7 days, cell viability was measured using the MTT assay. Biodentine and MTA were also prepared and packed into standardized bovine dentin disks and divided into three groups according to the storage media (n=6/group): freshly mixed materials without storage medium (Group A); materials stored in saline (Group B); materials stored in citric acid buffered at pH 5.4 (Group C). After 24 hours, DPSCs were introduced in the wells and cell adherence, viability, and cellular morphology were observed via confocal microscopy after three days of culture. Cell viability was analyzed using repeated-measures analysis of variance test with Tukey's post hoc tests (α=0.05). Results: Biodentine expressed significantly higher cell viability compared with all other groups after 4 days, with no differences after 7 days. Notably, cell viability was significantly greater in 24-hour set MTA compared with 1-hour set MTA and control groups after 7 days. Material exposure to an acidic environment showed an increase in cell adherence and viability in both groups. Conclusions: Biodentine induced a significantly accelerated cell proliferation compared with MTA. Setting of these materials in the presence of citric acid enhanced DPSC viability and adherence. PMID:27812618

  6. Interaction of dental pulp stem cells with Biodentine and MTA after exposure to different environments.

    PubMed

    Agrafioti, Anastasia; Taraslia, Vasiliki; Chrepa, Vanessa; Lymperi, Stefania; Panopoulos, Panos; Anastasiadou, Ema; Kontakiotis, Evangelos G

    2016-01-01

    The aim of the present study was to evaluate and compare the cytotoxic effects of Biodentine and MTA on dental pulp stem cells (DPSCs) and to assess cell viability and adherence after material exposure to an acidic environment. DPSCs were cultured either alone or in contact with either: Biodentine; MTA set for 1 hour; or MTA set for 24 hours. After 4 and 7 days, cell viability was measured using the MTT assay. Biodentine and MTA were also prepared and packed into standardized bovine dentin disks and divided into three groups according to the storage media (n=6/group): freshly mixed materials without storage medium (Group A); materials stored in saline (Group B); materials stored in citric acid buffered at pH 5.4 (Group C). After 24 hours, DPSCs were introduced in the wells and cell adherence, viability, and cellular morphology were observed via confocal microscopy after three days of culture. Cell viability was analyzed using repeated-measures analysis of variance test with Tukey's post hoc tests (α=0.05). Biodentine expressed significantly higher cell viability compared with all other groups after 4 days, with no differences after 7 days. Notably, cell viability was significantly greater in 24-hour set MTA compared with 1-hour set MTA and control groups after 7 days. Material exposure to an acidic environment showed an increase in cell adherence and viability in both groups. Biodentine induced a significantly accelerated cell proliferation compared with MTA. Setting of these materials in the presence of citric acid enhanced DPSC viability and adherence.

  7. Differential expression of basal microRNAs' patterns in human dental pulp stem cells.

    PubMed

    Vasanthan, Punitha; Govindasamy, Vijayendran; Gnanasegaran, Nareshwaran; Kunasekaran, Wijenthiran; Musa, Sabri; Abu Kasim, Noor Hayaty

    2015-03-01

    MicroRNAs (miRNAs) are small non-coding RNAs that regulate translation of mRNA into protein and play a crucial role for almost all biological activities. However, the identification of miRNAs from mesenchymal stem cells (MSCs), especially from dental pulp, is poorly understood. In this study, dental pulp stem cells (DPSCs) were characterized in terms of their proliferation and differentiation capacity. Furthermore, 104 known mature miRNAs were profiled by using real-time PCR. Notably, we observed 19 up-regulated miRNAs and 29 significantly down-regulated miRNAs in DPSCs in comparison with bone marrow MSCs (BM-MSCs). The 19 up-regulated miRNAs were subjected to ingenuity analysis, which were composed into 25 functional networks. We have chosen top 2 functional networks, which comprised 10 miRNA (hsa-miR-516a-3p, hsa-miR-125b-1-3p, hsa-miR-221-5p, hsa-miR-7, hsa-miR-584-5p, hsa-miR-190a, hsa-miR-106a-5p, hsa-mir-376a-5p, hsa-mir-377-5p and hsa-let-7f-2-3p). Prediction of target mRNAs and associated biological pathways regulated by each of this miRNA was carried out. We paid special attention to hsa-miR-516a-3p and hsa-miR-7-5p as these miRNAs were highly expressed upon validation with qRT-PCR analysis. We further proceeded with loss-of-function analysis with these miRNAs and we observed that hsa-miR-516a-3p knockdown induced a significant increase in the expression of WNT5A. Likewise, the knockdown of hsa-miR-7-5p increased the expression of EGFR. Nevertheless, further validation revealed the role of WNT5A as an indirect target of hsa-miR-516a-3p. These results provide new insights into the dynamic role of miRNA expression in DPSCs. In conclusion, using miRNA signatures in human as a prediction tool will enable us to elucidate the biological processes occurring in DPSCs.

  8. Factors secreted from dental pulp stem cells show multifaceted benefits for treating experimental rheumatoid arthritis.

    PubMed

    Ishikawa, Jun; Takahashi, Nobunori; Matsumoto, Takuya; Yoshioka, Yutaka; Yamamoto, Noriyuki; Nishikawa, Masaya; Hibi, Hideharu; Ishigro, Naoki; Ueda, Minoru; Furukawa, Koichi; Yamamoto, Akihito

    2016-02-01

    Rheumatoid arthritis (RA) is an autoimmune disease characterized by synovial hyperplasia and chronic inflammation, which lead to the progressive destruction of cartilage and bone in the joints. Numerous studies have reported that administrations of various types of MSCs improve arthritis symptoms in animal models, by paracrine mechanisms. However, the therapeutic effects of the secreted factors alone, without the cell graft, have been uncertain. Here, we show that a single intravenous administration of serum-free conditioned medium (CM) from human deciduous dental pulp stem cells (SHED-CM) into anti-collagen type II antibody-induced arthritis (CAIA), a mouse model of rheumatoid arthritis (RA), markedly improved the arthritis symptoms and joint destruction. The therapeutic efficacy of SHED-CM was associated with an induction of anti-inflammatory M2 macrophages in the CAIA joints and the abrogation of RANKL expression. SHED-CM specifically depleted of an M2 macrophage inducer, the secreted ectodomain of sialic acid-binding Ig-like lectin-9 (ED-Siglec-9), exhibited a reduced ability to induce M2-related gene expression and attenuate CAIA. SHED-CM also inhibited the RANKL-induced osteoclastogenesis in vitro. Collectively, our findings suggest that SHED-CM provides multifaceted therapeutic effects for treating CAIA, including the ED-Siglec-9-dependent induction of M2 macrophage polarization and inhibition of osteoclastogenesis. Thus, SHED-CM may represent a novel anti-inflammatory and reparative therapy for RA. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Cytotoxic effect of silorane and methacrylate based composites on the human dental pulp stem cells and fibroblasts

    PubMed Central

    Shafiei, Fereshteh; Razmkhah, Mahboobeh; Attar, Armin; Alavi, Ali A.

    2014-01-01

    Objectives: The aim of this study was to compare the cytotoxic effect of a methacrylate-based and a silorane-based composite on the human dental pulp stem cells (DPSCs) versus human dental pulp fibroblasts (DPFs). Study Design: Samples of the Filtek Z250 and P90 were polymerized and immersed in the culture medium to obtain extracts after incubation for one, seven and 14 days. Magnetic cell sorting based on the CD146 expression was performed to purify DPSCs and DPFs. After incubation of both cells with the extracts, cytotoxicity was determined using the MTT test. Results: For the extracts of first and seventh day, both composites showed significantly lower cytotoxicity on DPSCs than DPFs (p=0.003). In addition, there was a significant difference in the time-group interaction of both materials indicating different cytotoxic behaviours (p=0.014). In contrast to Z250, exposure to the 14th day extract of P90 resulted in higher cell viability compared to that of day seven. Conclusions: DPSCs are less susceptible to the cytotoxic effect of the composites than DPFs. Compared to Z250, the cytotoxic effect of silorane-based composite decreases as the time passes on. This difference should be considered, particularly in deep cavities, in order to preserve the regenerative capacity of the pulp. Key words:Composite resins, Dental pulp, Mesenchymal Stromal Cells, Silorane, Toxicology. PMID:24608214

  10. Effects of extracts of Salvadora persica on proliferation and viability of human dental pulp stem cells

    PubMed Central

    Tabatabaei, Fahimeh sadat; Moezizadeh, Maryam; Javand, Fateme

    2015-01-01

    Objectives: Efficacy of an ideal antimicrobial agent depends on its ability to eliminate microorganisms while causing minimal toxicity to host cells. The purpose of this study was to assess the effect of ethanolic and water extracts of Salvadora persica (SP) on proliferation and viability of human dental pulp stem cells (hDPSCs). Materials and Methods: In this in-vitro study, the effects of seven concentrations of ethanolic and water extracts of SP (ranging from 5.75 mg/ml to 0.08 mg/ml) on hDPSCs were evaluated using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assay. The results were analyzed using one-way ANOVA and Tukey's post-hoc test. P < 0.05 was considered statistically significant. Results: Water extract of SP only had cytotoxic effect at 5.75 mg/ml concentration; and caused significant cell proliferation at 1.43-0.08 mg/ml concentrations at 24 h (P < 0.05). At 48 h, only 0.17 and 0.08 mg/ml concentrations caused significant cell proliferation (P < 0.05). Ethanolic extract of SP at 5.75-1.43 mg/ml concentrations showed severe cytotoxic effects at 24 and 48 h. Other concentrations had no significant effects on cells (P > 0.05). Conclusion: The highest concentrations of both water and ethanolic extracts of SP had cytotoxic effects on hDPSCs. Water extract of SP has favorable effects on cell proliferation at specific concentrations in a time-dependent manner. PMID:26180418

  11. Human dental pulp stem cells hook into biocoral scaffold forming an engineered biocomplex.

    PubMed

    Mangano, Carlo; Paino, Francesca; d'Aquino, Riccardo; De Rosa, Alfredo; Iezzi, Giovanna; Piattelli, Adriano; Laino, Luigi; Mitsiadis, Thimios; Desiderio, Vincenzo; Mangano, Francesco; Papaccio, Gianpaolo; Tirino, Virginia

    2011-04-11

    The aim of this study was to evaluate the behavior of human Dental Pulp Stem Cells (DPSCs), as well as human osteoblasts, when challenged on a Biocoral scaffold, which is a porous natural hydroxyapatite. For this purpose, human DPSCs were seeded onto a three-dimensional (3D) Biocoral scaffold or on flask surface (control). Either normal or rotative (3D) cultures were performed. Scanning electron microscopic analyses, at 8, 24 and 48 h of culture showed that cells did not adhere on the external surface, but moved into the cavities inside the Biocoral structure. After 7, 15 and 30 days of culture, morphological and molecular analyses suggested that the Biocoral scaffold leads DPSCs to hook into the cavities where these cells quickly start to secrete the extra cellular matrix (ECM) and differentiate into osteoblasts. Control human osteoblasts also moved into the internal cavities where they secreted the ECM. Histological sections revealed a diffuse bone formation inside the Biocoral samples seeded with DPSCs or human osteoblasts, where the original scaffold and the new secreted biomaterial were completely integrated and cells were found within the remaining cavities. In addition, RT-PCR analyses showed a significant increase of osteoblast-related gene expression and, above all, of those genes highly expressed in mineralized tissues, including osteocalcin, OPN and BSP. Furthermore, the effects on the interaction between osteogenesis and angiogenesis were observed and substantiated by ELISA assays. Taken together, our results provide clear evidence that DPSCs differentiated into osteoblasts, forming a biocomplex made of Biocoral, ECM and differentiated cells.

  12. Human Dental Pulp Stem Cells Hook into Biocoral Scaffold Forming an Engineered Biocomplex

    PubMed Central

    d'Aquino, Riccardo; De Rosa, Alfredo; Iezzi, Giovanna; Piattelli, Adriano; Laino, Luigi; Mitsiadis, Thimios; Desiderio, Vincenzo; Mangano, Francesco; Papaccio, Gianpaolo; Tirino, Virginia

    2011-01-01

    The aim of this study was to evaluate the behavior of human Dental Pulp Stem Cells (DPSCs), as well as human osteoblasts, when challenged on a Biocoral scaffold, which is a porous natural hydroxyapatite. For this purpose, human DPSCs were seeded onto a three-dimensional (3D) Biocoral scaffold or on flask surface (control). Either normal or rotative (3D) cultures were performed. Scanning electron microscopic analyses, at 8, 24 and 48 h of culture showed that cells did not adhere on the external surface, but moved into the cavities inside the Biocoral structure. After 7, 15 and 30 days of culture, morphological and molecular analyses suggested that the Biocoral scaffold leads DPSCs to hook into the cavities where these cells quickly start to secrete the extra cellular matrix (ECM) and differentiate into osteoblasts. Control human osteoblasts also moved into the internal cavities where they secreted the ECM. Histological sections revealed a diffuse bone formation inside the Biocoral samples seeded with DPSCs or human osteoblasts, where the original scaffold and the new secreted biomaterial were completely integrated and cells were found within the remaining cavities. In addition, RT-PCR analyses showed a significant increase of osteoblast-related gene expression and, above all, of those genes highly expressed in mineralized tissues, including osteocalcin, OPN and BSP. Furthermore, the effects on the interaction between osteogenesis and angiogenesis were observed and substantiated by ELISA assays. Taken together, our results provide clear evidence that DPSCs differentiated into osteoblasts, forming a biocomplex made of Biocoral, ECM and differentiated cells. PMID:21494568

  13. Transplantation of human dental pulp-derived stem cells protects against heatstroke in mice.

    PubMed

    Tseng, Ling-Shu; Chen, Sheng-Hsien; Lin, Mao-Tsun; Lin, Ying-Chu

    2015-01-01

    Stem cells from human exfoliated deciduous tooth pulp (SHED) is a promising approach for the treatment of stroke and spinal cord injury. In this study, we investigated the therapeutic effects of SHED for the treatment of multiple organ (including brain, particularly hypothalamus) injury in heatstroke mice. ICR male mice were exposed to whole body heating (WBH; 41.2°C, relative humidity 50-55%, for 1 h) and then returned to normal room temperature (26°C). We observed that intravenous administration of SHED immediately post-WBH exhibited the following therapeutic benefits for recovery after heatstroke: (a) inhibition of WBH-induced neurologic and thermoregulatory deficits; (b) reduction of WBH-induced ischemia, hypoxia, and oxidative damage to the brain (particularly the hypothalamus); (c) attenuation of WBH-induced increased plasma levels of systemic inflammatory response molecules, such as tumor necrosis factor-α and intercellular adhesion molecule-1; (d) improvement of WBH-induced hypothalamo-pituitary-adrenocortical (HPA) axis activity (as reflected by enhanced plasma levels of both adrenocorticotrophic hormone and corticosterone); and (e) attenuation of WBH-induced multiple organ apoptosis as well as lethality. In conclusion, post-WBH treatment with SHED reduced induction of proinflammatory cytokines and oxidative radicals, enhanced plasma induction of both adrenocorticotrophic hormone and corticosterone, and improved lethality in mouse heatstroke. The protective effect of SHED may be related to a decreased inflammatory response, decreased oxidative stress, and an increased HPA axis activity following the WBH injury.

  14. Modulation of the differentiation of dental pulp stem cells by different concentrations of β-glycerophosphate.

    PubMed

    Liu, Mingyue; Sun, Yao; Liu, Yang; Yuan, Mengtong; Zhang, Zhihui; Hu, Weiping

    2012-01-31

    Dentinogenesis is a necessary prerequisite for dental tissue engineering. One of the steps for dentinogenesis is to obtain large quantities of highly purified odontoblasts. Therefore, we have undertaken an experiment applying different concentrations of β-glycerophosphate (β-GP) to induce the differentiation of dental pulp stem cells (DPSCs) in a long-term 28-day culture. In the meanwhile, we have studied the time- and maturation-dependent expression of matrix extracellular phosphoglycoprotein (MEPE) and that of the odontoblast-like marker-dentin sialoprotein (DSP), in order to investigate an optimized mineralized condition. Western blot results revealed that the expression of DSP became lower when accompanied by the increase of the β-GP concentration, and there was also an influence on MEPE expression when different concentrations of β-GP were applied. Meanwhile, the mineralized groups had an inhibitory function on the expression of MEPE as compared with the control group. Above all, all experimental groups successfully generated mineralized nodules by Alizarin Red S and the 5 mM β-GP group formed more mineralized nodules quantitated using the CPC extraction method. In conclusion, there is a significant modulation of the β-GP during the differentiation of the DPSCs. The degree of odontoblast differentiation is β-glycerophosphate concentration dependent. A low concentration of β-GP (5 mM) has been shown to be the optimal concentration for stimulating the maturation of the DPSCs. Moreover, MEPE accompanied with DSP clearly demonstrates the degree of the differentiation.

  15. Zinc-modified titanium surface enhances osteoblast differentiation of dental pulp stem cells in vitro

    PubMed Central

    Yusa, Kazuyuki; Yamamoto, Osamu; Takano, Hiroshi; Fukuda, Masayuki; Iino, Mitsuyoshi

    2016-01-01

    Zinc is an essential trace element that plays an important role in differentiation of osteoblasts and bone modeling. This in vitro study aimed to evaluate the osteoblast differentiation of human dental pulp stem cells (DPSCs) on zinc-modified titanium (Zn-Ti) that releases zinc ions from its surface. Based on real-time PCR, alkaline phosphatase (ALP) activity and Western blot analysis data, we investigated osteoblast differentiation of DPSCs cultured on Zn-Ti and controls. DPSCs cultured on Zn-Ti exhibited significantly up-regulated gene expression levels of osteoblast-related genes of type I collagen (Col I), bone morphogenetic protein 2 (BMP2), ALP, runt-related transcription factor 2 (Runx2), osteopontin (OPN), and vascular endothelial growth factor A (VEGF A), as compared with controls. We also investigated extracellular matrix (ECM) mineralization by Alizarin Red S (ARS) staining and found that Zn-Ti significantly promoted ECM mineralization when compared with controls. These findings suggest that the combination of Zn-Ti and DPSCs provides a novel approach for bone regeneration therapy. PMID:27387130

  16. Zinc chloride for odontogenesis of dental pulp stem cells via metallothionein up-regulation.

    PubMed

    Lin, Chia-Yung; Lin, Hsin-Hua; Tsai, Mong-Hsun; Lin, Shau-Ping; Chen, Min-Huey

    2011-02-01

    Previous studies have shown that zinc chloride (ZnCl(2)) can induce metallthionein (MT) in the liver and kidney to protect tissues against toxicants and shows a better corneal wound healing than conventional drugs do. We hypothesized that ZnCl(2) can promote odontogenesis of dental pulp stem cells (DPSCs) via MT. The purpose of this study was to investigate the effects of ZnCl(2) on human DPSCs and the expression of MT. DPSCs were isolated by flow cytometry with selective surface marker CD146 and STRO-1. After they grew into confluence, DPSCs were induced into odontoblasts with or without ZnCl(2) supplemented in the culture medium for 21 days. The effect of ZnCl(2) on DPSCs differentiation was examined followed by alkaline phosphatase staining/activity and quantitative real-time polymerase chain reaction analysis. By treating DPSCs with ZnCl(2), the duration of mineralization was shortened and expressions of differentiation markers into odontoblasts were more significant than those without ZnCl(2) stimulation. Besides, the MT gene expression was increased with the increasing expressions of odontoblasts' markers after treated with ZnCl(2). This was the first report that ZnCl(2) could promote odontoblastic differentiation of DPSCs through the up-regulation of gene MT. Copyright © 2011 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  17. Concurrent Expression of Oct4 and Nanog Maintains Mesenchymal Stem-Like Property of Human Dental Pulp Cells

    PubMed Central

    Huang, Chuan-En; Hu, Fang-Wei; Yu, Chuan-Hang; Tsai, Lo-Lin; Lee, Tzu-Hsin; Chou, Ming-Yung; Yu, Cheng-Chia

    2014-01-01

    Human dental pulp stem cells (DPSCs), unique mesenchymal stem cells (MSCs) type, exhibit the characteristics of self-renewal and multi-lineage differentiation capacity. Oct4 and Nanog are pluripotent genes. The aim of this study was to determine the physiological functions of Oct4 and Nanog expression in DPSCs. Herein, we determined the critical role of an Oct4/Nanog axis modulating MSCs properties of DPSCs by lentiviral-mediated co-overexpression or co-knockdown of Oct4/Nanog in DPSCs. MSCs properties including osteogenic/chondrogenic/adipogenic induction differentiation was assayed for expression of osteogenic/chondrogenic/adipogenic markers by quantitative real-time RT-PCR analysis. Initially, we observed that the expression profile of Oct4 and Nanog in dental pulp cells, which exerted properties of MSCs, was significantly up-regulated compared to that of STRO-1−CD146− dental pulp cells. Down-regulation of Oct4 and Nanog co-expression significantly reduced the cell proliferation, osteogenic differentiation capability, STRO-1, CD146, and Alkaline phosphatase (ALP) activity of DPSCs. In contrast, co-overexpression of Oct4 and Nanog enhanced the expression level of STRO-1 and CD146, proliferation rate and osteogenic/chondrogenic/adipogenic induction differentiation capability, and expression of osteogenic/chondrogenic/adipogenic induction differentiation markers. Our results suggest that Oct4-Nanog signaling is a regulatory switch to maintain properties in DPSCs. PMID:25322154

  18. A new population of human adult dental pulp stem cells: a useful source of living autologous fibrous bone tissue (LAB).

    PubMed

    Laino, Gregorio; d'Aquino, Riccardo; Graziano, Antonio; Lanza, Vladimiro; Carinci, Francesco; Naro, Fabio; Pirozzi, Giuseppe; Papaccio, Gianpaolo

    2005-08-01

    Stem cells, derived from human adult dental pulp of healthy subjects 30-45 years of age, were cultured, and cells were selected using a FACSorter. A new c-kit+/CD34+/CD45- cell population of stromal bone producing cells (SBP/DPSCs) was selected, expanded, and cultured. These SBP/DPSCs are highly clonogenic and, in culture, differentiate into osteoblast precursors (CD44+/RUNX-2+), still capable of self-renewing, and then in osteoblasts, producing, in vitro, a living autologous fibrous bone (LAB) tissue, which is markedly positive for several bone antibodies. This tissue constitute an ideal source of osteoblasts and mineralized tissue for bone regeneration. In fact, after in vivo transplantation into immunocompromised rats, LAB formed lamellar bone-containing osteocytes. Recently it has been reported that human dental pulp stem cells (DPSCs) are detectable, in humans, only up to the age of 30 years and that they are able to produce in vitro only sporadic calcified nodules and to form, after transplantation in vivo, a mineralized tissue. Stem cells, derived from human adult dental pulp of healthy subjects 30-45 years of age, were cultured, and cells were selected using a FACSorter. Light microscope, histochemistry, immunofluorescence, and RT-PCR analyses were performed to study both stem and differentiating cells. A new c-kit+/CD34+/CD45- cell population of stromal bone producing cells (SBP/DPSCs) has been selected by FACSorting, expanded, and cultured. These SBP/DPSCs are highly clonogenic and, in culture, differentiate into osteoblast precursors (CD44+/RUNX-2+), still capable of self-renewing, and in osteoblasts, producing, in vitro, a living autologous fibrous bone (LAB) tissue. This new-formed tissue is markedly positive for several antibodies for bone, including osteonectin, bone sialoprotein, osteocalcin, fibronectin, collagen III, and bone alkaline phosphatase (BALP). Cells producing LAB can be stored at -80 degrees C for a long period of time and are an

  19. Dental pulp stem cell responses to novel antibiotic-containing scaffolds for regenerative endodontics.

    PubMed

    Kamocki, K; Nör, J E; Bottino, M C

    2015-12-01

    To evaluate both the drug-release profile and the effects on human dental pulp stem cells' (hDPSC) proliferation and viability of novel bi-mix antibiotic-containing scaffolds intended for use as a drug delivery system for root canal disinfection prior to regenerative endodontics. Polydioxanone (PDS)-based fibrous scaffolds containing both metronidazole (MET) and ciprofloxacin (CIP) at selected ratios were synthesized via electrospinning. Fibre diameter was evaluated based on scanning electron microscopy (SEM) images. Pure PDS scaffolds and a saturated CIP/MET solution (i.e. 50 mg of each antibiotic in 1 mL) (hereafter referred to as DAP) served as both negative (nontoxic) and positive (toxic) controls, respectively. High-performance liquid chromatography (HPLC) was performed to investigate the amount of drug(s) released from the scaffolds. WST-1(®) proliferation assay was used to evaluate the effect of the scaffolds on cell proliferation. LIVE/DEAD(®) assay was used to qualitatively assess cell viability. Data obtained from drug release and proliferation assays were statistically analysed at the 5% significance level. A burst release of CIP and MET was noted within the first 24 h, followed by a sustained maintenance of the drug(s) concentration for 14 days. A concentration-dependent trend was noticed upon hDPSCs' exposure to all CIP-containing scaffolds, where increasing the CIP concentration resulted in reduced cell proliferation (P < 0.05) and viability. In groups exposed to pure MET or pure PDS scaffolds, no changes in proliferation were observed. Synthesized antibiotic-containing scaffolds had significantly lower effects on hDPSCs proliferation when compared to the saturated CIP/MET solution (DAP). © 2014 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  20. Cryopreserved dental pulp tissues of exfoliated deciduous teeth is a feasible stem cell resource for regenerative medicine.

    PubMed

    Ma, Lan; Makino, Yusuke; Yamaza, Haruyoshi; Akiyama, Kentaro; Hoshino, Yoshihiro; Song, Guangtai; Kukita, Toshio; Nonaka, Kazuaki; Shi, Songtao; Yamaza, Takayoshi

    2012-01-01

    Human exfoliated deciduous teeth have been considered to be a promising source for regenerative therapy because they contain unique postnatal stem cells from human exfoliated deciduous teeth (SHED) with self-renewal capacity, multipotency and immunomodulatory function. However preservation technique of deciduous teeth has not been developed. This study aimed to evaluate that cryopreserved dental pulp tissues of human exfoliated deciduous teeth is a retrievable and practical SHED source for cell-based therapy. SHED isolated from the cryopreserved deciduous pulp tissues for over 2 years (25-30 months) (SHED-Cryo) owned similar stem cell properties including clonogenicity, self-renew, stem cell marker expression, multipotency, in vivo tissue regenerative capacity and in vitro immunomodulatory function to SHED isolated from the fresh tissues (SHED-Fresh). To examine the therapeutic efficacy of SHED-Cryo on immune diseases, SHED-Cryo were intravenously transplanted into systemic lupus erythematosus (SLE) model MRL/lpr mice. Systemic SHED-Cryo-transplantation improved SLE-like disorders including short lifespan, elevated autoantibody levels and nephritis-like renal dysfunction. SHED-Cryo amended increased interleukin 17-secreting helper T cells in MRL/lpr mice systemically and locally. SHED-Cryo-transplantation was also able to recover osteoporosis bone reduction in long bones of MRL/lpr mice. Furthermore, SHED-Cryo-mediated tissue engineering induced bone regeneration in critical calvarial bone-defect sites of immunocompromised mice. The therapeutic efficacy of SHED-Cryo transplantation on immune and skeletal disorders was similar to that of SHED-Fresh. These data suggest that cryopreservation of dental pulp tissues of deciduous teeth provide a suitable and desirable approach for stem cell-based immune therapy and tissue engineering in regenerative medicine.

  1. Intrafibrillar-silicified collagen scaffolds enhance the osteogenic capacity of human dental pulp stem cells.

    PubMed

    Niu, Li-na; Sun, Jia-qi; Li, Qi-hong; Jiao, Kai; Shen, Li-juan; Wu, Dan; Tay, Franklin; Chen, Ji-hua

    2014-07-01

    The present study investigated the effects of intrafibrillar-silicified collagen scaffolds (ISCS) on the osteogenic differentiation of human dental pulp stem cells (hDPSCs) in vitro and in vivo. The hDPSCs were co-cultured with ISCS or nonsilicified collagen scaffolds (NCS) in control medium (CM) or osteogenic differentiation medium (ODM). Cell cycle and cell apoptosis were analyzed with flow cytometry to measure the viability of hDPSCs. Reverse transcription-polymerase chain reaction (RT-PCR) and western blotting were used to evaluate the expression levels of osteogenic marker genes and proteins of hDPSCs. Alkaline phosphatase (ALP) staining and alizarin red S assay were used to evaluate the ALP activity of hDPSCs and their calcium deposition potential. In addition, hDPSCs and scaffolds were implanted subcutaneously in nude mice for 8 weeks. Harvested tissues were immunohistochemically stained for osteocalcin (OCN) expression from hDPSCs, and stained with alizarin red S for examination of their calcium deposition in vivo. The ISCS had no adverse effect on hDPSCs, promoted their proliferation, and significantly up-regulated the expression of osteogenesis-related genes and proteins. The hDPSCs co-cultured with ISCS in ODM exhibited the highest ALP activity and calcium deposition in vitro. The ISCS promoted the OCN expression and calcium deposition of hDPSCs after ectopic transplantation in vivo. Intrafibrillar-silicified collagen scaffolds significantly promoted the proliferation, osteogenic differentiation and mineralization of hDPSCs, when compared with NCS. This study demonstrates combining the use of hDPSCs and ISCS to promote bone-like tissue formation is a promising approach for clinical bone repair and regeneration. Copyright © 2014 Elsevier Ltd. All rights reserved.

  2. Dental pulp stem cell responses to novel antibiotic-containing scaffolds for regenerative endodontics

    PubMed Central

    Kamocki, K.; Nör, J. E.; Bottino, M. C.

    2014-01-01

    Aim To evaluate both the drug release profile and the effects on human dental pulp stem cells’ (hDPSC) proliferation and viability of novel bi-mix antibiotic-containing scaffolds intended for use as a drug-delivery system for root canal disinfection prior to regenerative endodontics. Methodology Polydioxanone (PDS)-based fibrous scaffolds containing both metronidazole (MET) and ciprofloxacin (CIP) at selected ratios were synthesized via electrospinning. Fibre diameter was evaluated based on scanning electron microscopy (SEM) images. Pure PDS scaffolds and a saturated CIP/MET solution (i.e. 50 mg of each antibiotic in 1 mL) (hereafter referred to as DAP) served as both negative (non-toxic) and positive (toxic) controls, respectively. High performance liquid chromatography (HPLC) was done to investigate the amount of drug(s) released from the scaffolds. WST-1® proliferation assay was used to evaluate the effect of the scaffolds on cell proliferation. LIVE/DEAD® assay was used to qualitatively assess cell viability. Data obtained from drug release and proliferation assays were statistically analysed at the 5% significance level. Results A burst release of CIP and MET was noted within the first 24 h, followed by a sustained maintenance of the drug(s) concentration for 14 days. A concentration-dependent trend was noticed upon hDPSCs’ exposure to all CIP-containing scaffolds, where increasing the CIP concentration resulted in reduced cell proliferation (P<0.05) and viability. In groups exposed to pure MET or pure PDS scaffolds, no changes in proliferation were observed. Conclusions Synthesized antibiotic-containing scaffolds had significantly lower effects on hDPSCs proliferation when compared to the saturated CIP/MET solution (DAP). PMID:25425048

  3. Comparison of osteo/odontogenic differentiation of human adult dental pulp stem cells and stem cells from apical papilla in the presence of platelet lysate.

    PubMed

    Abuarqoub, Duaa; Awidi, Abdalla; Abuharfeil, Nizar

    2015-10-01

    Human dental pulp cells (DPSCs) and stem cells from apical papilla have been used for the repair of damaged tooth tissues. Human platelet lysate (PL) has been suggested as a substitute for fetal bovine serum (FBS) for large scale expansion of dental stem cells. However, biological effects and optimal concentrations of PL for proliferation and differentiation of human dental stem cells remain to be elucidated. DPSCs and SCAP cells were isolated from impacted third molars of young healthy donors, at the stage of root development and identified by markers using flow cytometry. For comparison the cells were cultured in media containing PL (1%, 5% and 10%) and FBS, with subsequent induction for osteogenic/odontogenic differentiation. The cultures were analyzed for; morphology, growth characteristics, mineralization potential (Alizarin Red method) and differentiation markers using ELISA and real time -polymerase chain reaction (qPCR). The proliferation rates of DPSCs and SCAP significantly increased when cells were treated with 5% PL (7X doubling time) as compared to FBS. 5% PL also enhanced mineralized differentiation of DPSCs and SCAP, as indicated by the measurement of alkaline phosphatase activity, osteocalcin and osteopontin, calcium deposition and q-PCR. Our findings suggest that using 5% platelet lysate, proliferation and osteo/odontogenesis of DPSCs and SCAP for a short period of time (15 days), was significantly improved. This may imply its use as an optimum concentration for expansion of dental stem cells in bone regeneration. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Effect of Aminated Mesoporous Bioactive Glass Nanoparticles on the Differentiation of Dental Pulp Stem Cells

    PubMed Central

    Lee, Jung-Hwan; Kang, Min-Sil; Mahapatra, Chinmaya; Kim, Hae-Won

    2016-01-01

    Mesoporous bioactive nanoparticles (MBNs) have been developed as promising additives to various types of bone or dentin regenerative material. However, biofunctionality of MBNs as dentin regenerative additive to dental materials have rarely been studied. We investigated the uptake efficiency of MBNs-NH2 with their endocytosis pathway and the role of MBNs-NH2 in odontogenic differentiation to clarify inherent biofunctionality. MBNs were fabricated by sol-gel synthesis, and 3% APTES was used to aminate these nanoparticles (MBNs-NH2) to reverse their charge from negative to positive. To characterize the MBNs-NH2, TEM, XRD, FTIR, zeta(ξ)-potential measurements, and Brunauer–Emmett–Teller analysis were performed. After primary cultured rat dental pulp stem cells (rDPSCs) were incubated with various concentrations of MBNs-NH2, stem cell viability (24 hours) with or without differentiated media, internalization of MBNs-NH2 in rDPSCs (~4 hours) via specific endocytosis pathway, intra or extracellular ion concentration and odontoblastic differentiation (~28 days) were investigated. Incubation with up to 50 μg/mL of MBNs-NH2 had no effect on rDPSCs viability with differentiated media (p>0.05). The internalization of MBNs-NH2 in rDPSCs was determined about 92% after 4 hours of incubation. Uptake was significantly decreased with ATP depletion and after 1 hour of pre-treatment with the inhibitor of macropinocytosis (p<0.05). There was significant increase of intracellular Ca and Si ion concentration in MBNs-NH2 treated cells compared to no-treated counterpart (p<0.05). The expression of odontogenic-related genes (BSP, COL1A, DMP-1, DSPP, and OCN) and the capacity for biomineralization (based on alkaline phosphatase activity and alizarin red staining) were significantly upregulated with MBNs-NH2. These results indicate that MBNs-NH2 induce odontogenic differentiation of rDPSCs and may serve as a potential dentin regenerative additive to dental material for promoting

  5. Bioactive injectable aggregates with nanofibrous microspheres and human dental pulp stem cells: A translational strategy in dental endodontics.

    PubMed

    Garzón, I; Martin-Piedra, M A; Carriel, V; Alaminos, M; Liu, X; D'Souza, R N

    2017-01-12

    Regeneration of the pulp-dentin complex with stem cells is a potential alternative to conventional root canal treatments. Human dental pulp stem cells (hDPSCs) have been extensively studied because of their ability to proliferate and differentiate into mineralized dental and non-dental tissues. Here we combined hDPSCs with two types of injectable poly-l-lactic acid (PLLA) microsphere with a nanofibrous or smooth surface to form bioactive injectable aggregates, and examined their ability to promote pulp regeneration in the root canal in an in vivo model. We investigated the biocompatibility, biosafety and odontogenic potential of fibrous (F-BIM) and smooth bioactive injectable microspheres (S-BIM) in vitro and in vivo. Our results demonstrated that PLLA microspheres and hDPSCs were able to form bioactive injectable aggregates that promoted dentin regeneration in both in vitro and in vivo models. Our results suggest that F-BIM and S-BIM may induce dentinogenesis upon in vivo grafting, and propose that the potential usefulness of the microsphere-hDPSC aggregates described here should be evaluated in clinical settings. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

  6. Biophysical characterization of low-frequency ultrasound interaction with dental pulp stem cells

    PubMed Central

    2013-01-01

    Background Low-intensity ultrasound is considered an effective non-invasive therapy to stimulate hard tissue repair, in particular to accelerate delayed non-union bone fracture healing. More recently, ultrasound has been proposed as a therapeutic tool to repair and regenerate dental tissues. Our recent work suggested that low-frequency kilohertz-range ultrasound is able to interact with dental pulp cells which could have potential to stimulate dentine reparative processes and hence promote the viability and longevity of teeth. Methods In this study, the biophysical characteristics of low-frequency ultrasound transmission through teeth towards the dental pulp were explored. We conducted cell culture studies using an odontoblast-like/dental pulp cell line, MDPC-23. Half of the samples underwent ultrasound exposure while the other half underwent ‘sham treatment’ where the transducer was submerged into the medium but no ultrasound was generated. Ultrasound was applied directly to the cell cultures using a therapeutic ultrasound device at a frequency of 45 kHz with intensity settings of 10, 25 and 75 mW/cm2 for 5 min. Following ultrasound treatment, the odontoblast-like cells were detached from the culture using a 0.25% Trypsin/EDTA solution, and viable cell numbers were counted. Two-dimensional tooth models based on μ-CT 2D images of the teeth were analyzed using COMSOL as the finite element analysis platform. This was used to confirm experimental results and to demonstrate the potential theory that with the correct combination of frequency and intensity, a tooth can be repaired using small doses of ultrasound. Frequencies in the 30 kHz–1 MHz range were analyzed. For each frequency, pressure/intensity plots provided information on how the intensity changes at each point throughout the propagation path. Spatial peak temporal average (SPTA) intensity was calculated and related to existing optimal spatial average temporal average (SATA) intensity deemed effective

  7. In vitro proliferation and osteogenic differentiation of human dental pulp stem cells in injectable thermo-sensitive chitosan/β-glycerophosphate/hydroxyapatite hydrogel.

    PubMed

    Chen, Yantian; Zhang, Fengli; Fu, Qiang; Liu, Yong; Wang, Zejian; Qi, Nianmin

    2016-09-01

    Injectable thermo-sensitive hydrogels have a potential application in bone tissue engineering for their sensitivities and minimal invasive properties. Human dental pulp stem cells have been considered a promising tool for tissue reconstruction. The objective of this study was to investigate the proliferation and osteogenic differentiation of dental pulp stem cells in injectable thermo-sensitive chitosan/β-glycerophosphate/hydroxyapatite hydrogel in vitro. The chitosan /β-glycerophosphate hydrogel and chitosan/β-glycerophosphate/hydroxyapatite hydrogel were prepared using the sol-gel method. The injectability of chitosan /β-glycerophosphate hydrogel and chitosan/β-glycerophosphate/hydroxyapatite hydrogel was measured using a commercial disposable syringe. Scanning electron microscopy was used to observe the inner structure of hydrogels. Then dental pulp stem cells were seeded in chitosan /β-glycerophosphate hydrogel and chitosan/β-glycerophosphate/hydroxyapatite hydrogel, respectively. The growth of dental pulp stem cells was periodically observed under an inverted microscope. The proliferation of dental pulp stem cells was detected by using an Alamar Blue kit, while cell apoptosis was determined by using a Live/Dead Viability/Cytotoxicity kit. The osteogenic differentiations of dental pulp stem cells in chitosan /β-glycerophosphate hydrogel and chitosan/β-glycerophosphate/hydroxyapatite hydrogel were evaluated by alkaline phosphatase activity assay and mRNA expression of osteogenesis gene for 21 days in osteogenic medium. The results indicated that there was no significant difference between chitosan /β-glycerophosphate hydrogel and chitosan/β-glycerophosphate/hydroxyapatite hydrogel in injectability. Cells within the chitosan/β-glycerophosphate/hydroxyapatite hydrogel displayed a typical adherent cell morphology and rapid proliferation with high cellular viability after 14 days of culture. Dental pulp stem cells seeded in chitosan

  8. Non-viral bone morphogenetic protein 2 transfection of rat dental pulp stem cells using calcium phosphate nanoparticles as carriers.

    PubMed

    Yang, Xuechao; Walboomers, X Frank; van den Dolder, Juliette; Yang, Fang; Bian, Zhuan; Fan, Mingwen; Jansen, John A

    2008-01-01

    Calcium phosphate nanoparticles have shown potential as non-viral vectors for gene delivery. The aim of this study was to induce bone morphogenetic protein (Bmp)2 transfection in rat dental pulp stem cells using calcium phosphate nanoparticles as a gene vector and then to evaluate the efficiency and bioactivity of the transfection. We also intended to investigate the behavior of transfected cells when seeded on 3-dimensional titanium fiber mesh scaffolds. Nanoparticles of calcium phosphate encapsulating plasmid deoxyribonucleic acid (DNA) (plasmid enhanced green fluorescent protein-BMP2) were prepared. Then, STRO-1-selected rat dental pulp stem cells were transfected using these nanoparticles. Transfection and bioactivity of the secreted BMP2 were examined. Thereafter, the transfected cells were cultured on a fibrous titanium mesh. The cultures were investigated using scanning electron microscipy and evaluated for cell proliferation, alkaline phosphatase activity and calcium content. Finally, real-time polymerase chain reaction was performed for odontogenesis-related gene expression. The results showed that the size of the DNA-loaded particles was approximately 100 nm in diameter. Nanoparticles could protect the DNA encapsulated inside from external DNase and release the loaded DNA in a low-acid environment (pH 3.0). In vitro, nanoparticle transfection was shown to be effective and to accelerate or promote the odontogenic differentiation of rat dental pulp stem cells when cultured in the 3-dimensional scaffolds. Based on our results, plasmid DNA-loaded calcium phosphate nanoparticles appear to be an effective non-viral vector for gene delivery and functioned well for odontogenic differentiation through Bmp2 transfection.

  9. Human Dental Pulp Stem Cells and Gingival Fibroblasts Seeded into Silk Fibroin Scaffolds Have the Same Ability in Attracting Vessels

    PubMed Central

    Woloszyk, Anna; Buschmann, Johanna; Waschkies, Conny; Stadlinger, Bernd; Mitsiadis, Thimios A.

    2016-01-01

    Neovascularization is one of the most important processes during tissue repair and regeneration. Current healing approaches based on the use of biomaterials combined with stem cells in critical-size bone defects fail due to the insufficient implant vascularization and integration into the host tissues. Therefore, here we studied the attraction, ingrowth, and distribution of blood vessels from the chicken embryo chorioallantoic membrane into implanted silk fibroin scaffolds seeded with either human dental pulp stem cells or human gingival fibroblasts. Perfusion capacity was evaluated by non-invasive in vivo Magnetic Resonance Imaging while the number and density of blood vessels were measured by histomorphometry. Our results demonstrate that human dental pulp stem cells and gingival fibroblasts possess equal abilities in attracting vessels within silk fibroin scaffolds. Additionally, the prolonged in vitro pre-incubation period of these two cell populations favors the homogeneous distribution of vessels within silk fibroin scaffolds, which further improves implant survival and guarantees successful healing and regeneration. PMID:27148078

  10. Dental Stem Cell Migration on Pulp Ceiling Cavities Filled with MTA, Dentin Chips, or Bio-Oss

    PubMed Central

    Lymperi, Stefania; Taraslia, Vasiliki; Tsatsoulis, Ioannis N.; Samara, Athina; Agrafioti, Anastasia; Anastasiadou, Ema; Kontakiotis, Evangelos

    2015-01-01

    MTA, Bio-Oss, and dentin chips have been successfully used in endodontics. The aim of this study was to assess the adhesion and migration of dental stem cells on human pulp ceiling cavities filled with these endodontic materials in an experimental model, which mimics the clinical conditions of regenerative endodontics. Cavities were formed, by a homemade mold, on untouched third molars, filled with endodontic materials, and observed with electron microscopy. Cells were seeded on cavities' surface and their morphology and number were analysed. The phenomenon of tropism was assessed in a migration assay. All three materials demonstrated appropriate microstructures for cell attachment. Cells grew on all reagents, but they showed a differential morphology. Moreover, variations were observed when comparing cells numbers on cavity's filling versus the surrounding dentine disc. The highest number of cells was recorded on dentin chips whereas the opposite was true for Bio-Oss. This was confirmed in the migration assay where a statistically significant lower number of cells migrated towards Bio-Oss as compared to MTA and dentin chips. This study highlights that MTA and dentin chips have a greater potential compared to Bio-Oss regarding the attraction of dental stem cells and are good candidates for bioengineered pulp regeneration. PMID:26146613

  11. Dental Stem Cell Migration on Pulp Ceiling Cavities Filled with MTA, Dentin Chips, or Bio-Oss.

    PubMed

    Lymperi, Stefania; Taraslia, Vasiliki; Tsatsoulis, Ioannis N; Samara, Athina; Velentzas, Athanasios D; Agrafioti, Anastasia; Anastasiadou, Ema; Kontakiotis, Evangelos

    2015-01-01

    MTA, Bio-Oss, and dentin chips have been successfully used in endodontics. The aim of this study was to assess the adhesion and migration of dental stem cells on human pulp ceiling cavities filled with these endodontic materials in an experimental model, which mimics the clinical conditions of regenerative endodontics. Cavities were formed, by a homemade mold, on untouched third molars, filled with endodontic materials, and observed with electron microscopy. Cells were seeded on cavities' surface and their morphology and number were analysed. The phenomenon of tropism was assessed in a migration assay. All three materials demonstrated appropriate microstructures for cell attachment. Cells grew on all reagents, but they showed a differential morphology. Moreover, variations were observed when comparing cells numbers on cavity's filling versus the surrounding dentine disc. The highest number of cells was recorded on dentin chips whereas the opposite was true for Bio-Oss. This was confirmed in the migration assay where a statistically significant lower number of cells migrated towards Bio-Oss as compared to MTA and dentin chips. This study highlights that MTA and dentin chips have a greater potential compared to Bio-Oss regarding the attraction of dental stem cells and are good candidates for bioengineered pulp regeneration.

  12. Characterization of Neurogenic Potential of Dental Pulp Stem Cells Cultured in Xeno/Serum-Free Condition: In Vitro and In Vivo Assessment

    PubMed Central

    Jung, Jieun; Kim, Jong-Wan; Moon, Ho-Jin; Hong, Jin Young

    2016-01-01

    Neural stem cells (NSCs) have a high potency for differentiation to neurons and glial cells for replacement of damaged cells and paracrine effects for the regeneration and remyelination of host axons. Dental pulp is known to have a potential to differentiate into neural-like cells; therefore, dental pulp may be used as an autologous cell source for neural repair. In this study, we selectively expanded stem cells from human dental pulp in an initial culture using NSC media under xeno- and serum-free conditions. At the initial step of primary culture, human dental pulp was divided into two groups according to the culture media: 10% fetal bovine serum medium group (FBS group) and NSC culture medium group (NSC group). In the NSC group relative to the FBS group, the expression of NSC markers and the concentrations of leukemia inhibitory factor, nerve growth factor, and stem cell factor were higher, although their expression levels were lower than those of human fetal NSCs. The transplanted cells of the NSC group survived well within the normal brain and injured spinal cord of rats and expressed nestin and Sox2. Under the xeno- and serum-free conditions, autologous human dental pulp-derived stem cells might prove useful for clinical cell-based therapies to repair damaged neural tissues. PMID:27688776

  13. Isolation of dental pulp stem cells from a single donor and characterization of their ability to differentiate after 2 years of cryopreservation

    PubMed Central

    Alsulaimani, Reem S.; Ajlan, Sumaiah A.; Aldahmash, Abdullah M.; Alnabaheen, May S.; Ashri, Nahid Y.

    2016-01-01

    Objectives: To investigate the viability and differentiation capacity of dental pulp stem cells (DPSCs) isolated from single donors after two years of cryopreservation. Methods: This prospective study was conducted between October 2010 and February 2014 in the Stem Unit, College of Medicine, King Saud University, Riyadh, Saudi Arabia. Seventeen teeth extracted from 11 participants were processed separately to assess the minimum tissue weight needed to yield cells for culturing in vitro. Cell stemness was evaluated before passage 4 using the colony forming unit assay, immunofluorescence staining, and bi-lineage differentiation. Dental pulp stem cells were cryopreserved for 2 years. Post-thaw DPSCs were cultured until senescence and differentiated toward osteogenic, odontogenic, adipogenic, and chondrogenic lineages. Results: Viable cells were isolated successfully from 6 of the 11 participants. Three of these 6 cultured cell lines were identified as DPSCs. A minimum of 0.2 g of dental pulp tissue was required for successful isolation of viable cells from a single donor. Post-thaw DPSCs successfully differentiated towards osteogenic, odontogenic, chondrogenic, and adipogenic lineages. The post-thaw DPSCs were viable in vitro up to 70 days before senescence. There was no significant difference between the cells. Conclusion: Within the limitations of this investigation, viable cells from dental pulp tissue were isolated successfully from the same donor using a minimum of 2 extracted teeth. Not all isolated cells from harvested dental pulp tissue had the characteristics of DPSCs. Post-thaw DPSCs maintained their multi-lineage differentiation capacity. PMID:27146619

  14. The role of PIN1 on odontogenic and adipogenic differentiation in human dental pulp stem cells.

    PubMed

    Lee, Young-Man; Shin, Seung-Yun; Jue, Seong-Suk; Kwon, Il-Keun; Cho, Eun-Hee; Cho, Eui-Sic; Park, Sang-Hyuk; Kim, Eun-Cheol

    2014-03-15

    Recently, the involvement of PIN1, a peptidyl-prolyl cis/trans isomerase, has been reported in age-related bone homeostasis and adipogenesis. However, the role of PIN1 during odontogenic and adipogenic differentiation remains to be fully understood, particularly regarding human dental pulp stem cells (HDPSCs). Thus, in the present study, we have investigated the role of PIN1 in odontogenic and adipogenic differentiation of HDPSCs and signaling pathways possibly involved. PIN1 mRNA and protein level were upregulated in a time-dependent manner during adipogenic differentiation, increasing until 1 day of odontogenic induction and then steadily declined during odontogenic differentiation. Treatment of a known PIN1 inhibitor, juglone, significantly increased odontogenic differentiation as confirmed by alkaline phosphatase (ALP) activity, calcium deposition, and mRNAs induction of odontogenic markers [ALP, osteopontin (OPN), osteocalcin (OCN), dentin sialophosphoprotein (DSPP), and dentin matrix protein 1 (DMP-1)]. On the contrary, adipogenic differentiation was dramatically reduced upon juglone treatment, with concomitant downregulation of lipid droplet accumulation and adipogenic marker genes [peroxisome proliferation-activated receptor gamma (PPARγ), lipoprotein lipase (LPL), and adipocyte fatty acid-binding protein (AP2)]. In contrast to PIN1 inhibition, the overexpression of PIN1 via adenoviral infection (Ad-PIN1) in HDPSCs inhibited odontogenic differentiation but increased adipogenic differentiation, in which stem cell property markers such as stage-specific embryonic antigen-4 (SSEA-4) and STRO-1 were upregulated during odontogenic differentiation but downregulated in adiopogenic differentiation. Consistently, juglone-mediated inhibition of PIN1 augmented the osteogenic medium (OM)-induced activation of bone morphogenetic protein (BMP), Wnt/β-catenin, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and nuclear factor-kappa B (NF

  15. Functional recovery upon human dental pulp stem cell transplantation in a diabetic neuropathy rat model.

    PubMed

    Datta, Indrani; Bhadri, Naini; Shahani, Pradnya; Majumdar, Debanjana; Sowmithra, Sowmithra; Razdan, Rema; Bhonde, Ramesh

    2017-10-01

    Diabetic neuropathy (DN) is among the most debilitating complications of diabetes. Here, we investigated the effects of human dental pulp stem cell (DPSC) transplantation in Streptozotocin (STZ)-induced neuropathic rats. Six weeks after STZ injection, DPSCs were transplanted through two routes, intravenous (IV) or intramuscular (IM), in single or two repeat doses. Two weeks after transplantation, a significant improvement in hyperalgesia, grip-strength, motor coordination and nerve conduction velocity was observed in comparison with controls. A rapid improvement in neuropathic symptoms was observed for a single dose of DPSC IV; however, repeat dose of DPSC IV did not bring about added improvement. A single dose of DPSC IM showed steady improvement, and further recovery continued upon repeat IM administration. DPSC single dose IV showed greater improvement than DPSC single dose IM, but IM transplantation brought about better improvement in body weight. A marked reduction in tumor necrosis factor (TNF) α and C-reactive protein (CRP) levels was observed in the blood plasma for all treated groups, as compared with controls. With respect to inflammatory cytokines, repeat dose of DPSC IM showed further improvement, suggesting that a repeat dose is required to maintain the improved inflammatory state. Gene expression of inflammatory markers in liver confirmed amelioration in inflammation. Arachidonic acid level was unaffected by IV DPSC transplantation but showed noticeable increase through IM administration of a repeat dose. These results suggest that DPSC transplantation through both routes and dosage was beneficial for the retrieval of neuropathic parameters of DN; transplantation via the IM route with repeat dose was the most effective. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  16. Neurogenic differentiation of dental pulp stem cells to neuron-like cells in dopaminergic and motor neuronal inductive media.

    PubMed

    Chang, Chia-Chieh; Chang, Kai-Chun; Tsai, Shang-Jye; Chang, Hao-Hueng; Lin, Chun-Pin

    2014-12-01

    Dental pulp stem cells (DPSCs) have been proposed as a promising source of stem cells in nerve regeneration due to their close embryonic origin and ease of harvest. The aim of this study was to evaluate the efficacy of dopaminergic and motor neuronal inductive media on transdifferentiation of human DPSCs (hDPSCs) into neuron-like cells. Isolation, cultivation, and identification of hDPSCs were performed with morphological analyses and flow cytometry. The proliferation potential of DPSCs was evaluated with an XTT [(2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide)] assay. Media for the induction of dopaminergic and spinal motor neuronal differentiation were prepared. The efficacy of neural induction was evaluated by detecting the expression of neuron cell-specific cell markers in DPSCs by immunocytochemistry and quantitative real-time reverse transcription polymerase chain reaction (RT-PCR). In the XTT assay, there was a 2.6- or 2-fold decrease in DPSCs cultured in dopaminergic or motor neuronal inductive media, respectively. The proportions of βIII-tubulin (βIII-tub), glial fibrillary acidic protein (GFAP), and oligodendrocyte (O1)-positive cells were significantly higher in DPSCs cultured in both neuronal inductive media compared with those cultured in control media. Furthermore, hDPSC-derived dopaminergic and spinal motor neuron cells after induction expressed a higher density of neuron cell markers than those before induction. These findings suggest that in response to the neuronal inductive stimuli, a greater proportion of DPSCs stop proliferation and acquire a phenotype resembling mature neurons. Such neural crest-derived adult DPSCs may provide an alternative stem cell source for therapy-based treatments of neuronal disorders and injury. Copyright © 2014. Published by Elsevier B.V.

  17. Cytotoxic effect of silorane and methacrylate based composites on the human dental pulp stem cells and fibroblasts.

    PubMed

    Shafiei, Fereshteh; Tavangar, Maryam S; Razmkhah, Mahboobeh; Attar, Armin; Alavi, Ali-Asghar

    2014-07-01

    The aim of this study was to compare the cytotoxic effect of a methacrylate-based and a silorane-based composite on the human dental pulp stem cells (DPSCs) versus human dental pulp fibroblasts (DPFs). Samples of the Filtek Z250 and P90 were polymerized and immersed in the culture medium to obtain extracts after incubation for one, seven and 14 days. Magnetic cell sorting based on the CD146 expression was performed to purify DPSCs and DPFs. After incubation of both cells with the extracts, cytotoxicity was determined using the MTT test. For the extracts of first and seventh day, both composites showed significantly lower cytotoxicity on DPSCs than DPFs (p=0.003). In addition, there was a significant difference in the time-group interaction of both materials indicating different cytotoxic behaviours (p=0.014). In contrast to Z250, exposure to the 14th day extract of P90 resulted in higher cell viability compared to that of day seven. DPSCs are less susceptible to the cytotoxic effect of the composites than DPFs. Compared to Z250, the cytotoxic effect of silorane-based composite decreases as the time passes on. This difference should be considered, particularly in deep cavities, in order to preserve the regenerative capacity of the pulp.

  18. Elucidating the cellular actions of demineralised dentine matrix extract on a clonal dental pulp stem cell population in orchestrating dental tissue repair

    PubMed Central

    Lee, Chi P; Colombo, John S; Ayre, Wayne Nishio; Sloan, Alastair J

    2015-01-01

    Bioactive growth factors identified within the extracellular matrix of dentine have been proposed roles in regulating the naturally inherent regenerative dentine formation seen in teeth in response to trauma and infection, which may also be harnessed for novel clinical treatments in augmenting mineralised tissue repair. This study examined the specific biological action of demineralised dentine matrix extract on a clonal population of dental pulp stem cells in stimulating the prerequisite stages of wound healing associated with mineralised tissue repair. A clonal dental pulp stem cell population with sustained proliferative capacity and multi-potentiality towards osteogenic, adipogenic and chondrogenic lineages was isolated from the pulp of human third molars. Dentine was collected from human healthy teeth, powdered and treated with ethylenediaminetetraacetic acid to obtain a solubilised DDM protein extract. The influence of DDM on the DPSC clonal population was assessed in vitro. Exposure of cells to proteolytically degraded DDM or unsupplemented media served as controls. Compared to controls, DDM stimulated cell expansion, reduced apoptotic marker caspase 3, increased cell survival marker Akt1 and enhanced mineralised matrix deposition as determined by mineral deposition and increased expression of bone-related markers, alkaline phosphatase and osteopontin. Dental pulp stem cells successfully migrated into collagen gels supplemented with demineralised dentine matrix, with cells remaining viable and expanding in numbers over a 3-day period. Collectively, the results provide evidence that soluble proteins extracted from dentine matrix are able to exert a direct biological effect on dental pulp stem cells in promoting mineralised tissue repair mechanisms. PMID:26019808

  19. Dental pulp stem cells express tendon markers under mechanical loading and are a potential cell source for tissue engineering of tendon-like tissue

    PubMed Central

    Chen, Yu-Ying; He, Sheng-Teng; Yan, Fu-Hua; Zhou, Peng-Fei; Luo, Kai; Zhang, Yan-Ding; Xiao, Yin; Lin, Min-Kui

    2016-01-01

    Postnatal mesenchymal stem cells have the capacity to differentiate into multiple cell lineages. This study explored the possibility of dental pulp stem cells (DPSCs) for potential application in tendon tissue engineering. The expression of tendon-related markers such as scleraxis, tenascin-C, tenomodulin, eye absent homologue 2, collagens I and VI was detected in dental pulp tissue. Interestingly, under mechanical stimulation, these tendon-related markers were significantly enhanced when DPSCs were seeded in aligned polyglycolic acid (PGA) fibre scaffolds. Furthermore, mature tendon-like tissue was formed after transplantation of DPSC-PGA constructs under mechanical loading conditions in a mouse model. This study demonstrates that DPSCs could be a potential stem cell source for tissue engineering of tendon-like tissue. PMID:27811845

  20. Dental pulp stem cells express tendon markers under mechanical loading and are a potential cell source for tissue engineering of tendon-like tissue.

    PubMed

    Chen, Yu-Ying; He, Sheng-Teng; Yan, Fu-Hua; Zhou, Peng-Fei; Luo, Kai; Zhang, Yan-Ding; Xiao, Yin; Lin, Min-Kui

    2016-12-16

    Postnatal mesenchymal stem cells have the capacity to differentiate into multiple cell lineages. This study explored the possibility of dental pulp stem cells (DPSCs) for potential application in tendon tissue engineering. The expression of tendon-related markers such as scleraxis, tenascin-C, tenomodulin, eye absent homologue 2, collagens I and VI was detected in dental pulp tissue. Interestingly, under mechanical stimulation, these tendon-related markers were significantly enhanced when DPSCs were seeded in aligned polyglycolic acid (PGA) fibre scaffolds. Furthermore, mature tendon-like tissue was formed after transplantation of DPSC-PGA constructs under mechanical loading conditions in a mouse model. This study demonstrates that DPSCs could be a potential stem cell source for tissue engineering of tendon-like tissue.

  1. Effects of cell cycle phases on the induction of dental pulp stem cells toward dopaminergic- like cells.

    PubMed

    Gnanasegaran, Nareshwaran; Govindasamy, Vijayendran; Kathirvaloo, Premasangery; Musa, Sabri; Abu Kasim, Noor Hayaty

    2017-01-12

    Parkinson's disease (PD) is characterized by tremors and cognitive issues, and is due to the death of dopaminergic (DA-ergic) neurons in brain circuits that are responsible for producing neurotransmitter dopamine (DA). Currently, cell replacement therapies are underway to overcome existing therapeutic approaches such as drug treatments and electrical stimulation. Among the widely available sources, dental pulp stem cells from deciduous teeth (DPSC) have gained popularity due to their neural crest origin and inherent propensity toward neuronal lineage. Despite the various pre-clinical studies conducted, an important factor yet to be elucidated is the influence of growth phases in a typical trans-differentiation process. This study selected DPSCs at three distinct time points with variable growth phase proportions (G0/G1, S, and G2/M) for in vitro trans-differentiation into DA-ergic-like cells. Using commercially available PCR arrays, we identified distinct gene profiles pertaining to cell cycles in these phases. The differentiation outcomes were assessed in terms of morphology, and gene and protein expression as well as functional assays. It was noted that DPSC with highest G0/G1 phase were relatively the best representing at least a 2-fold up regulation (p < 0.05) of DA-ergic molecular cues compared to those from the remaining time points. Further investigations in terms of protein expression and DA-release assays also revealed a similar phenomenon (p < 0.05). These findings are expected to provide vital information for consideration in improving standard operating procedures in future cell transplantation work.

  2. The effects of human platelet lysate on dental pulp stem cells derived from impacted human third molars.

    PubMed

    Chen, Bo; Sun, Hai-Hua; Wang, Han-Guo; Kong, Hui; Chen, Fa-Ming; Yu, Qing

    2012-07-01

    Human platelet lysate (PL) has been suggested as a substitute for fetal bovine serum (FBS) in the large-scale expansion of dental pulp stem cells (DPSCs). However, the biological effects and the optimal concentrations of PL for the proliferation and differentiation of human DPSCs remain unexplored. We isolated and expanded stem cells from the dental pulp of extracted third molars and evaluated the effects of PL on the cells' proliferative capacity and differentiation potential in vitro and in vivo. Before testing, immunocytochemical staining and flow cytometry-based cell sorting showed that the cells derived from human dental pulp contained mesenchymal stem cell populations. Cells were grown on tissue culture plastic or on hydroxyapatite-tricalcium phosphate (HA/TCP) biomaterials and were incubated with either normal or odontogenic/osteogenic media in the presence or absence of various concentrations of human PL for further investigation. The proliferation of DPSCs was significantly increased when the cells were cultured in 5% PL under all testing conditions (P < 0.05). However, this enhancement was inconsistent when the cells were cultured in 1% PL or in 10% PL; 10% PL significantly inhibited cell proliferation and was therefore excluded from further differentiation testing. Culture medium containing 5% PL also significantly promoted the mineralized differentiation of DPSCs, as indicated by the measurement of alkaline phosphatase activity and calcium deposition under mineral-conditioned media (P < 0.05). Scanning electron microscopy and modified Ponceau trichrome staining showed that the cells treated with 5% PL and mineralizing media were highly capable of integrating with the HA/TCP biomaterials and had fully covered the surface of the scaffold with an extensive sheet-like structure 14 d after seeding. In addition, 5% PL showed significantly positive effects on tissue regeneration in two in vivo transplantation models. We conclude that the appropriate

  3. Decreasing NF-κB Expression Enhances Odontoblastic Differentiation and Collagen Expression in Dental Pulp Stem Cells Exposed to Inflammatory Cytokines

    PubMed Central

    Hozhabri, Neda S. T.; Benson, M. Douglas; Vu, Michael D.; Patel, Rinkesh H.; Martinez, Rebecca M.; Nakhaie, Fatemeh N.; Kim, Harry K. W.; Varanasi, Venu G.

    2015-01-01

    Inflammatory response in the dental pulp can alter the collagen matrix formation by dental pulp stem cells and lead to a delay or poor healing of the pulp. This inflammatory response is mediated by cytokines, including interleukin-1β and tumor necrosis factor-α. In this study, it is hypothesized that suppressing the actions of these inflammatory cytokines by knocking down the activity of transcription factor Nuclear Factor–κB will lead to dental pulp stem cell differentiation into odontoblasts and the production of collagen. Here, the role of Nuclear Factor–κB signaling and its reduction was examined during odontogenic behavior in the presence of these cytokines. The results showed a significant increase in Nuclear Factor–κB gene expression and p65 protein expression by interleukin-1β and tumor necrosis factor-α. Nuclear Factor–κB activation in the presence of these cytokines decreased significantly in a dose-dependent manner by a Nuclear Factor–κB inhibitor (MG132) and p65 siRNA. Down-regulation of Nuclear Factor–κB activity also enhanced the gene expression of the odontoblastic markers (dentin sialophosphoprotein, Nestin, and alkaline phosphatase) and displayed an odontoblastic cell morphology indicating the promotion of odontogenic differentiation of dental pulp stem cells. Finally, dental pulp stem cells exposed to reduced Nuclear Factor–κB activity resulted in a significant increase in collagen (I)-α1 expression in the presence of these cytokines. In conclusion, a decrease in Nuclear Factor-κB in dental pulp stem cells in the presence of inflammatory cytokines enhanced odontoblastic differentiation and collagen matrix formation. PMID:25629155

  4. Comparison of human mesenchymal stem cells derived from dental pulp, bone marrow, adipose tissue, and umbilical cord tissue by gene expression.

    PubMed

    Stanko, Peter; Kaiserova, Katarina; Altanerova, Veronika; Altaner, Cestmir

    2014-09-01

    Our aims were to characterize human mesenchymal stem cells isolated from various tissues by pluripotent stem cells gene expression profile. Four strains of dental pulp stem cells (DP-MSCs) were isolated from dental pulp tissue fragments adhered to plastic tissue culture dishes. Mesenchymal stem cells derived from umbilical cord tissue (UBC-MSCs) were isolated with the same technique. Bone marrow derived mesenchymal stem cells (BM-MSCs) were isolated from nucleated cells of bone marrow obtained by density gradient centrifugation. Human mesenchymal stem cells from adipose tissue (AT-MSCs) were isolated by collagenase digestion. All kinds of MSCs used in this study were cultivated in low glucose DMEM containing 5% or human platelet extract. All stem cell manipulation was performed in GMP conditions. Expression of 15 pluripotent stem cells genes on the level of proteins was measured by Proteome Profiler Human Pluripotent Stem Cell Array. Induction of MSCs to in vitro differentiation to adipocytes, osteoblasts, chondroblasts was achieved by cultivation of cells in appropriate differentiation medium. All MSCs tested were phenotypically similar and of fibroblastoid morphology. DP-MSCs and UBC-MSCs were more proliferative than bone marrow BM-MSCs and AT-MSCs. Protein expression of 15 genes typical for pluripotent stem cells distinguished them into two groups. While the gene expression profiles of BM-MSC, AT-MSCs and UBC-MSCs were similar, DP-MSCS differed in relative gene expression on the level of their products in several genes. Dental pulp mesenchymal stem cells cultivated in vitro under the same conditions as MSCs from bone marrow, adipose tissue and umbilical cord tissue can be distinguished by pluripotent stem cell gene expression profile.

  5. Dental pulp stem cells: state of the art and suggestions for a true translation of research into therapy.

    PubMed

    La Noce, Marcella; Paino, Francesca; Spina, Anna; Naddeo, Pasqualina; Montella, Roberta; Desiderio, Vincenzo; De Rosa, Alfredo; Papaccio, Gianpaolo; Tirino, Virginia; Laino, Luigi

    2014-07-01

    Stem cells have the ability to rescue and/or repair injured tissue. In humans, it is possible to isolate different types of stem cells from the body. Among these, dental pulp stem cells (DPSCs) are relatively easily obtainable and exhibit high plasticity and multipotential capabilities. In particular they represent a gold standard for neural-crest-derived bone reconstruction in humans and can be used for the repair of body defects in low-risk autologous therapeutic strategies. An electronic search was conducted on PubMed databases and supplemented with a manual study of relevant references. All research described in this review highlight that DPSCs are mesenchymal stem cells that could be used in clinical applications. Unfortunately, very few clinical trials have been reported. Major obstacles imposed on researchers are hindering the translation of potentially effective therapies to the clinic. Both researchers and regulatory institutions need to develop a new approach to this problem, drawing up a new policy for good manufacturing practice (GMP) procedures. We strongly suggest that only general rules be standardized rather than everything. Importantly, this would not have an effect on the safety of patients, but may very well affect the results, which cannot be identical for all patients, due to physiological diversity in the biology of each patient. Alternatively, it would be important to study the role of specific molecules that recruit endogenous stem cells for tissue regeneration. In this way, the clinical use of stem cells could be successfully developed. DPSCs are mesenchymal stem cells that differentiate into different tissues, maintain their characteristics after cryopreservation, differentiate into bone-like tissues when loaded on scaffolds in animal models, and regenerate bone in human grafts. In summary, all data reported up to now should encourage the development of clinical procedures using DPSCs. Copyright © 2014 The Authors. Published by Elsevier

  6. Comparative Analysis of Human Mesenchymal Stem Cells from Umbilical Cord, Dental Pulp, and Menstrual Blood as Sources for Cell Therapy.

    PubMed

    Ren, Huaijuan; Sang, Yunxia; Zhang, Fengli; Liu, Zhaoqing; Qi, Nianmin; Chen, Yantian

    2016-01-01

    Although mesenchymal stem cells (MSCs) based therapy has been considered as a promising tool for tissue repair and regeneration, the optimal cell source remains unknown. Umbilical cord (UC), dental pulp (DP), and menstrual blood (MB) are easily accessible sources, which make them attractive candidates for MSCs. The goal of this study was to compare the biological characteristics, including morphology, proliferation, antiapoptosis, multilineage differentiation capacity, and immunophenotype of UC-, DP-, and MB-MSCs in order to provide a theoretical basis for clinical selection and application of these cells. As a result, all UC-, DP-, and MB-MSCs have self-renewal capacity and multipotentiality. However, the UC-MSCs seemed to have higher cell proliferation ability, while DP-MSCs may have significant advantages for osteogenic differentiation, lower cell apoptosis, and senescence. These differences may be associated with the different expression level of cytokines, including vascular endothelial growth factor, fibroblast growth factor, keratinocyte growth factor, and hepatocyte growth factor in each of the MSCs. Comprehensively, our results suggest DP-MSCs may be a desired source for clinical applications of cell therapy.

  7. Proposing the use of dental pulp stem cells as a suitable biological model of neurofibromatosis type 1.

    PubMed

    Almeida, Paula Nascimento; Souza, Gustavo Torres; de Souza, Camila Maurmann; de Zanette, Rafaella Souza Salomão; Maranduba, Claudinéia Pereira; Rettore, João Vitor Paes; de Santos, Marcelo Oliveira; do Carmo, Antônio Márcio Resende; da Maranduba, Carlos Magno Costa; de Silva, Fernando Sá

    2015-01-01

    This study aims to propose the dental pulp stem cells (DPSCs) as a model for studying two features related to neurofibromatosis type 1 (NF1), i.e. augmented proliferative capacity and altered osteogenic differentiation. We isolated a DPSC from the pulp of deciduous teeth of a 6-year-old NF1 patient and two other healthy children of similar age. Cell proliferation was assayed by counting with a haemocytometer after successive cell re-plating. In order to compare osteogenic differentiation, we used osteoblast-differentiating medium and quantified alizarin stain, which relates to degree of calcification, and evaluated the expression of osteoblastic markers by reverse transcription polymerase chain reaction (RT-PCR). The DPSCs isolated from the NF1 patient displayed a greater rate of proliferation when compared to the control cells. Osteogenic differentiation occurred as expected for both NF1 and control, which concerned cell morphology and expression of osteoblast marker genes ALP, BMP2, BMP4, OCN and SPP1. However, alizarin staining denoted a markedly lower calcification level in the cells from the NF1-diagnosed child, considering that less calcium deposits were visualized under light microscopy and a smaller amount of alizarin could be quantified by spectrophotometry after extraction from the stained cells. DPSCs seem to be useful as a model for studying NF1 and predicting prognosis of patients, since their in vitro behaviour seems to mimic at least two features of this disorder: higher tendency to develop bone abnormalities and neoplastic cell proliferation.

  8. Human dental pulp stem cells cultured onto dentin derived scaffold can regenerate dentin-like tissue in vivo.

    PubMed

    Tran, Ha Le Bao; Doan, Vu Nguyen

    2015-12-01

    Regeneration of dentin tissues in the pulp space of teeth serves the ultimate goal of preserving teeth via endodontic approaches. In recent times, many studies suggested that human dentin scaffolds combined with dental stem cells was a potential strategy for the complete dentin tissue regeneration. In this study, human dental pulp stem cells (DPSCs) were isolated and cultured. Dentin specimens were prepared from human third molars and treated with ethylene diamine tetra-acetic acid and citric acid to remove the smear layer. Then, DPSCs were cultured onto human treated dentin (hTD) and implanted in mouse model for 4, 6 and 8 weeks. The resulting grafts were assessed by hematoxylin and eosin stain and immunohistochemical stains. As a result, DPSCs were supported and induced to regenerate of dentin-like tissues which expressed specific dentin markers such as dentin sialophosphoprotein and dentin matrix protein 1 by combination with hTD in vivo. Furthermore, cells existed in the newly-formed dentin-like tissues also expressed typical human mitochondria antibodies, demonstrated that new tissues originated from human. In conclusion, the obtain results extend hopefully newly-established therapy to apply in endodontics and traumatic dental hard tissues.

  9. Effects of decellularized matrices derived from periodontal ligament stem cells and SHED on the adhesion, proliferation and osteogenic differentiation of human dental pulp stem cells in vitro.

    PubMed

    Heng, Boon Chin; Zhu, Shaoyue; Xu, Jianguang; Yuan, Changyong; Gong, Ting; Zhang, Chengfei

    2016-04-01

    A major bottleneck to the therapeutic applications of dental pulp stem cells (DPSC) are their limited proliferative capacity ex vivo and tendency to undergo senescence. This may be partly due to the sub-optimal in vitro culture milieu, which could be improved by an appropriate extracellular matrix substratum. This study therefore examined decellularized matrix (DECM) from stem cells derived from human exfoliated deciduous teeth (SHED) and periodontal ligament stem cells (PDLSC), as potential substrata for DPSC culture. Both SHED-DECM and PDLSC-DECM promoted rapid adhesion and spreading of newly-seeded DPSC compared to bare polystyrene (TCPS), with vinculin immunocytochemistry showing expression of more focal adhesions by newly-adherent DPSC cultured on DECM versus TCPS. Culture of DPSC on SHED-DECM and PDLSC-DECM yielded higher proliferation of cell numbers compared to TCPS. The qRT-PCR data showed significantly higher expression of nestin by DPSC cultured on DECM versus the TCPS control. Osteogenic differentiation of DPSC was enhanced by culturing on PDLSC-DECM and SHED-DECM versus TCPS, as demonstrated by alizarin red S staining for mineralized calcium deposition, alkaline phosphatase assay and qRT-PCR analysis of key osteogenic marker expression. Hence, both SHED-DECM and PDLSC-DECM could enhance the ex vivo culture of DPSC under both non-inducing and osteogenic-inducing conditions. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Effects of Ciprofloxacin-Containing Antimicrobial Scaffolds on Dental Pulp Stem Cell Viability — In Vitro Studies

    PubMed Central

    Kamocki, Krzysztof; Nör, Jacques E.; Bottino, Marco C.

    2017-01-01

    Objective A combination of antibiotics, including but not limited to metronidazole (MET) and ciprofloxacin (CIP), has been indicated to eradicate bacteria in necrotic immature permanent teeth prior to regenerative procedures. It has been shown clinically that antibiotic pastes may lead to substantial stem cell death. The aim of this study was to synthesize scaffolds containing various concentrations of CIP to enhance cell viability while preserving antimicrobial properties. Design Polydioxanone (PDS)-based electrospun scaffolds were processed with decreasing CIP concentrations (25 – 1 wt.%) and morphologically evaluated using scanning electron microscopy (SEM). Cytotoxicity assays were performed to determine whether the amount of CIP released from the scaffolds would lead to human dental pulp stem cell (hDPSC) toxicity. Similarly, WST-1 assays were performed to evaluate the impact of CIP release on hDPSC proliferation. Pure PDS scaffolds and saturated double antibiotic solution MET/CIP (DAP) served as both positive and negative controls, respectively. Antibacterial efficacy against E. faecalis (Ef) was tested. Results A significant decrease in hDPSC’ viability at concentrations 5–25 wt.% was observed. However, concentrations below 5 wt.% did not impair cell viability. Data from the WST-1 assays indicated no detrimental impact on cell proliferation for scaffolds containing 2.5 wt.% CIP or less. Significant antimicrobial properties were seen for CIP-scaffolds at lower concentrations (i.e., 1 and 2.5 wt.%). Conclusion The obtained data demonstrated that a reduced concentration of CIP incorporated into PDS-based scaffolds maintains its antimicrobial properties while enhancing viability and proliferation of dental pulp stem cells. PMID:26042622

  11. Assessment of the Tumorigenic Potential of Spontaneously Immortalized and hTERT-Immortalized Cultured Dental Pulp Stem Cells

    PubMed Central

    Wilson, Ryan; Urraca, Nora; Skobowiat, Cezary; Hope, Kevin A.; Miravalle, Leticia; Chamberlin, Reed; Donaldson, Martin; Seagroves, Tiffany N.

    2015-01-01

    Dental pulp stem cells (DPSCs) provide an exciting new avenue to study neurogenetic disorders. DPSCs are neural crest-derived cells with the ability to differentiate into numerous tissues including neurons. The therapeutic potential of stem cell-derived lines exposed to culturing ex vivo before reintroduction into patients could be limited if the cultured cells acquired tumorigenic potential. We tested whether DPSCs that spontaneously immortalized in culture acquired features of transformed cells. We analyzed immortalized DPSCs for anchorage-independent growth, genomic instability, and ability to differentiate into neurons. Finally, we tested both spontaneously immortalized and human telomerase reverse transcriptase (hTERT)-immortalized DPSC lines for the ability to form tumors in immunocompromised animals. Although we observed increased colony-forming potential in soft agar for the spontaneously immortalized and hTERT-immortalized DPSC lines relative to low-passage DPSC, no tumors were detected from any of the DPSC lines tested. We noticed some genomic instability in hTERT-immortalized DPSCs but not in the spontaneously immortalized lines tested. We determined that immortalized DPSC lines generated in our laboratory, whether spontaneously or induced, have not acquired the potential to form tumors in mice. These data suggest cultured DPSC lines that can be differentiated into neurons may be safe for future in vivo therapy for neurobiological diseases. Significance This study demonstrated that immortalized dental pulp stem cells (DPSCs) do not form tumors in animals and that immortalized DPSCs can be differentiated into neurons in culture. These results lend support to the use of primary and immortalized DPSCs for future therapeutic approaches to treatment of neurobiological diseases. PMID:26032749

  12. IFN-γ regulates human dental pulp stem cells behavior via NF-κB and MAPK signaling

    PubMed Central

    He, Xinyao; Jiang, Wenkai; Luo, Zhirong; Qu, Tiejun; Wang, Zhihua; Liu, Ningning; Zhang, Yaqing; Cooper, Paul R.; He, Wenxi

    2017-01-01

    During caries, dental pulp expresses a range of pro-inflammatory cytokines in response to the infectious challenge. Interferon gamma (IFN-γ) is a dimerized soluble cytokine, which is critical for immune responses. Previous study has demonstrated that IFN-γ at relative high concentration (100 ng/mL) treatment improved the impaired dentinogenic and immunosuppressive regulatory functions of disease-derived dental pulp stem cells (DPSCs). However, little is known about the regulatory effects of IFN-γ at relative low concentration on healthy DPSC behavior (including proliferation, migration, and multiple-potential differentiation). Here we demonstrate that IFN-γ at relatively low concentrations (0.5 ng/mL) promoted the proliferation and migration of DPSCs, but abrogated odonto/osteogenic differentiation. Additionally, we identified that NF-κB and MAPK signaling pathways are both involved in the process of IFN-γ-regulated odonto/osteogenic differentiation of DPSCs. DPSCs treated with IFN-γ and supplemented with pyrrolidine dithiocarbamate (PDTC, an NF-κB inhibitor) or SB203580 (a MAPK inhibitor) showed significantly improved potential for odonto/osteogenic differentiation of DPSCs both in vivo and in vitro. These data provide important insight into the regulatory effects of IFN-γ on the biological behavior of DPSCs and indicate a promising therapeutic strategy for dentin/pulp tissue engineering in future endodontic treatment. PMID:28098169

  13. Control of proliferation and osteogenic differentiation of human dental-pulp-derived stem cells by distinct surface structures.

    PubMed

    Kolind, K; Kraft, D; Bøggild, T; Duch, M; Lovmand, J; Pedersen, F S; Bindslev, D A; Bünger, C E; Foss, M; Besenbacher, F

    2014-02-01

    The ability to control the behavior of stem cells provides crucial benefits, for example, in tissue engineering and toxicity/drug screening, which utilize the stem cell's capacity to engineer new tissues for regenerative purposes and the testing of new drugs in vitro. Recently, surface topography has been shown to influence stem cell differentiation; however, general trends are often difficult to establish due to differences in length scales, surface chemistries and detailed surface topographies. Here we apply a highly versatile screening approach to analyze the interplay of surface topographical parameters on cell attachment, morphology, proliferation and osteogenic differentiation of human mesenchymal dental-pulp-derived stem cells (DPSCs) cultured with and without osteogenic differentiation factors in the medium (ODM). Increasing the inter-pillar gap size from 1 to 6 μm for surfaces with small pillar sizes of 1 and 2 μm resulted in decreased proliferation and in more elongated cells with long pseudopodial protrusions. The same alterations of pillar topography, up to an inter-pillar gap size of 4 μm, also resulted in enhanced mineralization of DPSCs cultured without ODM, while no significant trend was observed for DPSCs cultured with ODM. Generally, cells cultured without ODM had a larger deposition of osteogenic markers on structured surfaces relative to the unstructured surfaces than what was found when culturing with ODM. We conclude that the topographical design of biomaterials can be optimized for the regulation of DPSC differentiation and speculate that the inclusion of ODM alters the ability of the cells to sense surface topographical cues. These results are essential in order to transfer the use of this highly proliferative, easily accessible stem cell into the clinic for use in cell therapy and regenerative medicine.

  14. DENTAL PULP STEM CELLS AND HUMAN PERIAPICAL CYST MESENCHYMAL STEM CELLS IN BONE TISSUE REGENERATION: COMPARISON OF BASAL AND OSTEOGENIC DIFFERENTIATED GENE EXPRESSION OF A NEWLY DISCOVERED MESENCHYMAL STEM CELL LINEAGE.

    PubMed

    Tatullo, M; Falisi, G; Amantea, M; Rastelli, C; Paduano, F; Marrelli, M

    2015-01-01

    Bone regeneration is an interesting field of biomedicine. The most recent studies are aimed to achieve a bone regeneration using mesenchymal stem cells (MSCs) taken from more accessible sites: oral and dental tissues have been widely investigated as a rich accessible source of MSCs. Dental Pulp Stem Cells (DPSCs) and human Periapical Cysts Mesenchymal Stem Cells (hPCy-MSCs) represent the new generation MSCs. The aim of this study is to compare the gene expression of these two innovative cell types to highlight the advantages of their use in bone regeneration. The harvesting, culturing and differentiating of cells isolated from dental pulp as well as from periapical cystic tissue were carried out as described in previously published reports. qRT-PCR analyses were performed on osteogenic genes in undifferentiated and osteogenic differentiated cells of DPSC and hPCy-MSC lineage. Real-time RT-PCR data suggested that both DPSCs and hPCy-MSCs cultured in osteogenic media are able to differentiate into osteoblast/odontoblast-like cells: however, some differences indicated that DPSCs seem to be directed more towards dentinogenesis, while hPCy-MSCs seem to be directed more towards osteogenesis.

  15. Strontium folate loaded biohybrid scaffolds seeded with dental pulp stem cells induce in vivo bone regeneration in critical sized defects.

    PubMed

    Martin-Del-Campo, Marcela; Rosales-Ibañez, Raul; Alvarado, Keila; Sampedro, Jose G; Garcia-Sepulveda, Christian A; Deb, Sanjukta; San Román, Julio; Rojo, Luis

    2016-10-18

    Strontium folate (SrFO) is a recently developed bone promoting agent with interest in medical and pharmaceutical fields due to its improved features in comparison to current strontium based therapies for osteoporosis and other bone diseases. In this work SrFO derivative was synthesized and loaded into biohybrid scaffolds obtained through lyophilisation of semi-interpenetrating networks of chitosan polyethylene glycol dimethacrylate and beta tri-calcium phosphate (βTCP) fabricated using free radical polymerization. The scaffolds were seeded with pluripotent stem cells obtained from human dental pulp and their potential to regenerate bone tissues were assessed using a critical sized defect model of calvaria in rats and compared with those obtained without SrFO. The results obtained both in vitro and in vivo demonstrated excellent cyto-compatibility with resorption of scaffolds in 4-6 weeks and a total regeneration of the defect, with a more rapid and dense bone formation in the group with SrFO. Thus, the use of stem cells sourced from human dental pulp in combination with SrFO are very promising systems for their application in compromised osseous tissue regeneration.

  16. Highly Efficient In Vitro Reparative Behaviour of Dental Pulp Stem Cells Cultured with Standardised Platelet Lysate Supplementation

    PubMed Central

    Palmieri, Francesca; Marrelli, Massimo

    2016-01-01

    Dental pulp is an accessible source of multipotent mesenchymal stromal cells (MSCs). The perspective role of dental pulp stem cells (DPSCs) in regenerative medicine demands an in vitro expansion and in vivo delivery which must deal with the safety issues about animal serum, usually required in cell culture practice. Human platelet lysate (PL) contains autologous growth factors and has been considered as valuable alternative to fetal bovine serum (FBS) in cell cultures. The optimum concentration to be added of such supplement is highly dependent on its preparation whose variability limits comparability of results. By in vitro experiments, we aimed to evaluate a standardised formulation of pooled PL. A low selected concentration of PL (1%) was able to support the growth and maintain the viability of the DPSCs. The use of PL in cell cultures did not impair cell surface signature typically expressed by MSCs and even upregulated the transcription of Sox2. Interestingly, DPSCs cultured in presence of PL exhibited a higher healing rate after injury and are less susceptible to toxicity mediated by exogenous H2O2 than those cultured with FBS. Moreover, PL addition was shown as a suitable option for protocols promoting osteogenic and chondrogenic differentiation of DPSCs. Taken together, our results indicated that PL is a valid substitute of FBS to culture and differentiate DPSCs for clinical-grade use. PMID:27774106

  17. Antimicrobial properties and dental pulp stem cell cytotoxicity using carboxymethyl cellulose-silver nanoparticles deposited on titanium plates.

    PubMed

    Laredo-Naranjo, Martha Alicia; Carrillo-Gonzalez, Roberto; De La Garza-Ramos, Myriam Angelica; Garza-Navarro, Marco Antonio; Torre-Martinez, Hilda H H; Del Angel-Mosqueda, Casiano; Mercado-Hernandez, Roberto; Carrillo-Fuentevilla, Roberto

    2016-12-01

    Objective: To evaluate the antimicrobial properties and dental pulp stem cells (DPSCs) cytotoxicity of synthesized carboxymethyl cellulose-silver nanoparticles impregnated on titanium plates. Material and methods: The antibacterial effect of silver nanoparticles in a carboxymethyl cellulose matrix impregnated on titanium plates (Ti-AgNPs) in three concentrations: 16%, 50% and 100% was determined by adding these to bacterial cultures of Streptococcus mutans and Porphyromonas gingivalis. The Ti-AgNPs cytotoxicity on DPSCs was determined using a fluorimetric cytotoxicity assay with 0.12% chlorhexidine as a positive control. Results: Silver nanoparticles in all concentrations were antimicrobial, with concentrations of 50% and 100% being more cytotoxic with 4% cell viability. Silver nanoparticles 16% had a cell viability of 95%, being less cytotoxic than 0.12% chlorhexidine. Conclusions: Silver nanoparticles are a promising structure because of their antimicrobial properties. These have high cell viability at a concentration of 16%, and are less toxic than chlorhexidine.

  18. Antimicrobial properties and dental pulp stem cell cytotoxicity using carboxymethyl cellulose-silver nanoparticles deposited on titanium plates

    PubMed Central

    Laredo-Naranjo, Martha Alicia; Carrillo-Gonzalez, Roberto; De La Garza-Ramos, Myriam Angelica; Garza-Navarro, Marco Antonio; Torre-Martinez, Hilda H. H.; Del Angel-Mosqueda, Casiano; Mercado-Hernandez, Roberto; Carrillo-Fuentevilla, Roberto

    2016-01-01

    Abstract Objective: To evaluate the antimicrobial properties and dental pulp stem cells (DPSCs) cytotoxicity of synthesized carboxymethyl cellulose-silver nanoparticles impregnated on titanium plates. Material and methods: The antibacterial effect of silver nanoparticles in a carboxymethyl cellulose matrix impregnated on titanium plates (Ti-AgNPs) in three concentrations: 16%, 50% and 100% was determined by adding these to bacterial cultures of Streptococcus mutans and Porphyromonas gingivalis. The Ti-AgNPs cytotoxicity on DPSCs was determined using a fluorimetric cytotoxicity assay with 0.12% chlorhexidine as a positive control. Results: Silver nanoparticles in all concentrations were antimicrobial, with concentrations of 50% and 100% being more cytotoxic with 4% cell viability. Silver nanoparticles 16% had a cell viability of 95%, being less cytotoxic than 0.12% chlorhexidine. Conclusions: Silver nanoparticles are a promising structure because of their antimicrobial properties. These have high cell viability at a concentration of 16%, and are less toxic than chlorhexidine. PMID:28642914

  19. Phenotypic and proteomic characteristics of human dental pulp derived mesenchymal stem cells from a natal, an exfoliated deciduous, and an impacted third molar tooth.

    PubMed

    Akpinar, Gurler; Kasap, Murat; Aksoy, Ayca; Duruksu, Gokhan; Gacar, Gulcin; Karaoz, Erdal

    2014-01-01

    The level of heterogeneity among the isolated stem cells makes them less valuable for clinical use. The purpose of this study was to understand the level of heterogeneity among human dental pulp derived mesenchymal stem cells by using basic cell biology and proteomic approaches. The cells were isolated from a natal (NDPSCs), an exfoliated deciduous (stem cells from human exfoliated deciduous (SHED)), and an impacted third molar (DPSCs) tooth of three different donors. All three stem cells displayed similar features related to morphology, proliferation rates, expression of various cell surface markers, and differentiation potentials into adipocytes, osteocytes, and chondrocytes. Furthermore, using 2DE approach coupled with MALDI-TOF/TOF, we have generated a common 2DE profile for all three stem cells. We found that 62.3 ± 7% of the protein spots were conserved among the three mesenchymal stem cell lines. Sixty-one of these conserved spots were identified by MALDI-TOF/TOF analysis. Classification of the identified proteins based on biological function revealed that structurally important proteins and proteins that are involved in protein folding machinery are predominantly expressed by all three stem cell lines. Some of these proteins may hold importance in understanding specific properties of human dental pulp derived mesenchymal stem cells.

  20. A computer-designed scaffold for bone regeneration within cranial defect using human dental pulp stem cells

    PubMed Central

    Yeon Kwon, Doo; Seon Kwon, Jin; Hun Park, Seung; Hun Park, Ji; Hee Jang, So; Yun Yin, Xiang; Yun, Jeong-Ho; Ho Kim, Jae; Hyun Min, Byoung; Hee Lee, Jun; Kim, Wan-Doo; Suk Kim, Moon

    2015-01-01

    A computer-designed, solvent-free scaffold offer several potential advantages such as ease of customized manufacture and in vivo safety. In this work, we firstly used a computer-designed, solvent-free scaffold and human dental pulp stem cells (hDPSCs) to regenerate neo-bone within cranial bone defects. The hDPSCs expressed mesenchymal stem cell markers and served as an abundant source of stem cells with a high proliferation rate. In addition, hDPSCs showed a phenotype of differentiated osteoblasts in the presence of osteogenic factors (OF). We used solid freeform fabrication (SFF) with biodegradable polyesters (MPEG-(PLLA-co-PGA-co-PCL) (PLGC)) to fabricate a computer-designed scaffold. The SFF technology gave quick and reproducible results. To assess bone tissue engineering in vivo, the computer-designed, circular PLGC scaffold was implanted into a full-thickness cranial bone defect and monitored by micro-computed tomography (CT) and histology of the in vivo tissue-engineered bone. Neo-bone formation of more than 50% in both micro-CT and histology tests was observed at only PLGC scaffold with hDPSCs/OF. Furthermore, the PLGC scaffold gradually degraded, as evidenced by the fluorescent-labeled PLGC scaffold, which provides information to tract biodegradation of implanted PLGC scaffold. In conclusion, we confirmed neo-bone formation within a cranial bone defect using hDPSCs and a computer-designed PLGC scaffold. PMID:26234712

  1. Long-term cryopreservation of dental pulp stem cells (SBP-DPSCs) and their differentiated osteoblasts: a cell source for tissue repair.

    PubMed

    Papaccio, Gianpaolo; Graziano, Antonio; d'Aquino, Riccardo; Graziano, Maria Francesca; Pirozzi, Giuseppe; Menditti, Dardo; De Rosa, Alfredo; Carinci, Francesco; Laino, Gregorio

    2006-08-01

    It is not known whether cells derived from stem cells retain their differentiation and morpho-functional properties after long-term cryopreservation. This information is of importance to evaluate their potential for long-term storage with a view to subsequent use in therapy. Here, we describe the morpho-functional properties of dental pulp stem cells (SBP-DPSCs), and of their differentiated osteoblasts, recovered after long-term cryopreservation. After storage for 2 years, we found that stem cells are still capable of differentiation, and that their differentiated cytotypes proliferate and produce woven bone tissue. In addition, cells still express all their respective surface antigens, confirming cellular integrity. In particular, SBP-DPSCs differentiated into pre-osteoblasts, showing diffuse positivity for ALP, BAP, RUNX-2, and calcein. Recovered osteoblasts expressed bone-specific markers and were easily recognizable ultrastructurally, with no alterations observed at this level. In addition, after in vivo transplantation, woven bone converted into a 3D lamellar bone type. Therefore, dental pulp stem cells and their osteoblast-derived cells can be long-term cryopreserved and may prove to be attractive for clinical applications.

  2. In vitro osteoblastic differentiation of human mesenchymal stem cells and human dental pulp stem cells on poly-L-lysine-treated titanium-6-aluminium-4-vanadium.

    PubMed

    Galli, Daniela; Benedetti, Laura; Bongio, Matilde; Maliardi, Valentina; Silvani, Giulia; Ceccarelli, Gabriele; Ronzoni, Flavio; Conte, Silvio; Benazzo, Francesco; Graziano, Antonio; Papaccio, Gianpaolo; Sampaolesi, Maurilio; De Angelis, Maria Gabriella Cusella

    2011-05-01

    Three-dimensional (3D) titanium-6-aluminium-4-vanadium (Ti6Al4V) is a widely used biomaterial for orthopedic prosthesis and dental implants; thanks to its very high-mechanical strength and resistance to corrosion. Human mesenchymal stem cells (hMSCs) and dental pulp stem cells (hDPSCs) are responsible for bone regeneration following colonization of prosthesis or dental implants. Both hMSCs and hDPSCs have lower ability to colonize this biomaterial in comparison with tissue culture-treated plastic. Both hMSCs and hDPSCs show lack of focal adhesion kinase (FAK) activation when grown on Ti6Al4V. This signal is restored in the presence of poly-L-lysine (poly-L-lys). Poly-L-lys has been used as part of organoapatite or together with zinc and calcium ions. Our results suggest that poly-L-lys alone induces FAK activation through β1-INTEGRIN, because the presence of β1-INTEGRIN blocking antibody avoided FAK autophosphorylation. Presence of poly-L-lys also increases expression of osteoblastic differentiation marker genes in hMSCs and hDPSCs grown on Ti6Al4V.

  3. Dental Pulp Cell Behavior in Biomimetic Environments.

    PubMed

    Smith, J G; Smith, A J; Shelton, R M; Cooper, P R

    2015-11-01

    There is emerging recognition of the importance of a physiologically relevant in vitro cell culture environment to promote maintenance of stem cells for tissue engineering and regenerative medicine purposes. In vivo, appropriate cellular cues are provided by local tissue extracellular matrix (ECM), and these are not currently recapitulated well in vitro using traditional cultureware. We therefore hypothesized that better replication of the in vivo environment for cell culture and differentiation could be achieved by culturing dental pulp cells with their associated ECM. Primary dental pulp cells were subsequently seeded onto pulp-derived ECM-coated cultureware. While at up to 24 h they exhibited the same level of adherence as those cells seeded on tissue culture-treated surfaces, by 4 d cell numbers and proliferation rates were significantly decreased in cells grown on pulp ECM compared with controls. Analysis of stem cell and differentiation marker transcripts, as well as Oct 3/4 protein distribution, supported the hypothesis that cells cultured on ECM better maintained a stem cell phenotype compared with those cultured on standard tissue culture-treated surfaces. Subsequent differentiation analysis of cells cultured on ECM demonstrated that they exhibited enhanced mineralization, as determined by alizarin red staining and mineralized marker expression. Supplementation of a 3% alginate hydrogel with pulp ECM components and dental pulp cells followed by differentiation induction in mineralization medium resulted in a time-dependent mineral deposition at the periphery of the construct, as demonstrated histologically and using micro-computed tomography analysis, which was reminiscent of tooth structure. In conclusion, data indicate that culture of pulp cells in the presence of ECM better replicates the in vivo environment, maintaining a stem cell phenotype suitable for downstream tissue engineering applications. © International & American Associations for Dental

  4. In Vivo Articular Cartilage Regeneration Using Human Dental Pulp Stem Cells Cultured in an Alginate Scaffold: A Preliminary Study

    PubMed Central

    Milian, Lara; Oliver, Maria; Zurriaga, Javier; Sancho-Tello, Maria; de Llano, Jose Javier Martin; Carda, Carmen

    2017-01-01

    Osteoarthritis is an inflammatory disease in which all joint-related elements, articular cartilage in particular, are affected. The poor regeneration capacity of this tissue together with the lack of pharmacological treatment has led to the development of regenerative medicine methodologies including microfracture and autologous chondrocyte implantation (ACI). The effectiveness of ACI has been shown in vitro and in vivo, but the use of other cell types, including bone marrow and adipose-derived mesenchymal stem cells, is necessary because of the poor proliferation rate of isolated articular chondrocytes. In this investigation, we assessed the chondrogenic ability of human dental pulp stem cells (hDPSCs) to regenerate cartilage in vitro and in vivo. hDPSCs and primary isolated rabbit chondrocytes were cultured in chondrogenic culture medium and found to express collagen II and aggrecan. Both cell types were cultured in 3% alginate hydrogels and implanted in a rabbit model of cartilage damage. Three months after surgery, significant cartilage regeneration was observed, particularly in the animals implanted with hDPSCs. Although the results presented here are preliminary, they suggest that hDPSCs may be useful for regeneration of articular cartilage. PMID:28951745

  5. N-Acetyl-l-cysteine enhances ex-vivo amplification of deciduous teeth dental pulp stem cells.

    PubMed

    Debeljak Martacic, Jasmina; Borozan, Suncica; Radovanovic, Anita; Popadic, Dusan; Mojsilovic, Slavko; Vucic, Vesna; Todorovic, Vera; Kovacevic Filipovic, Milica

    2016-10-01

    Obtaining high number of stem cells is of interest for cell based therapies. N-Acetyl-l-cysteine (NAC) acts as a source of sulfhydryl groups and an anti-oxidative agent. The aim of this study was to test different NAC concentration on proliferation and differentiation of deciduous teeth dental pulp stem cells (DTSCs) in vitro as well as to define the possible underlining mechanism of its effect. Number of viable, apoptotic and senescent DTSCs was determined after addition of NAC (0.1mM, 1.0mM, 2.0mM). Also, cell cycle analysis, HIF1-α expression, LDH isoenzymes, superoxide-dismutase (SOD) and catalase (CAT) activity, sulfhydryl groups content, the level of lipids' and proteins' oxidative damage and differentiation capacity of NAC treated DTSCs was determined. DTSCs expressed HIF-1α in all conditions. The lowest NAC dose (0.1mM) increased the number of DTSCs by one fifth comparing to the control, most likely stimulating entry of cells into S phase of cell cycle and enhancing the activity of LDH5 isoenzyme. The highest NAC dose (2mM) inhibited DTSCs proliferation. Also, DTSCs had the lowest level of oxidative damage with 0.1mM NAC. All tested NAC concentrations enhanced DTSCs osteo-chondrogenesis. The lowest NAC dose exerted significant positive effect on DTSCs proliferation as well as antioxidative protection creating beneficial environment for stem cells in vitro cultivation especially when their clinical use is important for stimulation of osteo-chondrogenesis. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Immature Dental Pulp Stem Cells Showed Renotropic and Pericyte-Like Properties in Acute Renal Failure in Rats

    PubMed Central

    Barros, Michele A.; Martins, João Flávio Panattoni; Maria, Durvanei Augusto; Wenceslau, Crisitiane Valverde; De Souza, Dener Madeiro; Kerkis, Alexandre; Câmara, Niels Olsen S.; Balieiro, Julio Cesar C.; Kerkis, Irina

    2015-01-01

    Acute renal failure (ARF) is a common renal disease that can lead to high mortality. Recovery from ARF occurs with the replacement of necrotic tubular cells by functional tubular epithelial cells and the normalization of microvascular endothelial cell function in the peritubular capillaries. Conventional therapeutic techniques are often ineffective against ARF. Hence, stem cell therapies, which act through multiple trophic and regenerative mechanisms, are encouraging. We investigated the homing of human immature dental pulp stem cells (IDPSCs) after endovenous (EV) or intraperitoneal (IP) injection, in immunocompetent Wistar rats with ARF induced by intramuscular injection of glycerol, without the use of immunosuppression. The cells, which had been cryopreserved for 6 years, were CD105+, CD73+, CD44+, and partly, STRO-1+ and CD146+, and presented unaltered mesoderm differentiation potential. The presence of these cells in the tubular region of the kidney and in the peritubular capillaries was demonstrated. These cells accelerate tubular epithelial cell regeneration through significant increase of Ki-67-immunoreactive cells in damaged kidney. Flow cytometry analysis confirmed that IDPSCs home to the kidneys (EV 34.10% and IP 33.25%); a lower percentage of cells was found in the liver (EV 19.05% and IP 9.10%), in the muscles (EV 6.30% and IP 1.35%), and in the lungs (EV 2.0% and IP 1.85%). After infusion into rat, these cells express pericyte markers, such as CD146+, STRO-1+, and vascular endothelial growth factor (VEGF+). We found that IDPSCs demonstrate renotropic and pericyte-like properties and contributed to restore renal tubule structure in an experimental rat ARF model. PMID:26858898

  7. TNF-alpha stimulation increases dental pulp stem cell migration in vitro through integrin alpha-6 subunit upregulation.

    PubMed

    Shi, Lei; Fu, Shanqi; Fahim, Sidra; Pan, Shuang; Lina, He; Mu, Xiaodan; Niu, Yumei

    2017-03-01

    The dissemination of stem cells into tissues requiring inflammatory and reparative response is fundamentally dependent upon their chemotactic migration. Expression of TNF-α is up regulated in inflamed pulps. Dental pulp cells are also known to express integrin α6 subunit. Expression of integrin subunit α6 has been linked to the acquisition of migratory potential in a wide variety of cell types in both pathological and physiological capacities. Therefore, in this study we examined the effects of a pleiotropic cytokine TNF-α on the migration of hDPSCs and investigated its relationship with expression of integrin α6 in hDPSCs during chemotactic migration. hDPSC cultures were established. Protein expression profile of α6 integrin subunit was determined. Effect of exogenous TNF-α (50ng/mL) on hDPSCs' migration potential was evaluated by transwell inserts and in vitro scratch assay. Upregulation/downregulation of TNF-α mediated migration was assayed in presence/absence of integrin α6 respectively. To suppress integrin α6 expression, cells were transfected with integrin α6 siRNA and then cell migration and cytoskeletal changes were evaluated. Our results showed significant increase of hDPSCs' migration after stimulation with TNF-α. By knockdown of integrin α6, which is upregulated by TNF-α, we observed a decrease in the TNF-α directed chemotaxis of hDPSCs. In this study, we show that activation of integrin α6 brought about by TNF-α led to an increase in migratory activity in DPSCs in vitro thus describing a novel association between a cytokine TNF-α and α6 chain of an adhesion receptor integrin in regulating migration of hDPSCs. Copyright © 2016. Published by Elsevier Ltd.

  8. EDTA soluble chemical components and the conditioned medium from mobilized dental pulp stem cells contain an inductive microenvironment, promoting cell proliferation, migration, and odontoblastic differentiation.

    PubMed

    Kawamura, Rei; Hayashi, Yuki; Murakami, Hiroshi; Nakashima, Misako

    2016-05-25

    The critical challenge in tissue engineering is to establish an optimal combination of stem cells, signaling morphogenetic molecules, and extracellular matrix scaffold/microenvironment. The extracellular matrix components of teeth may be reconstituted as an inductive microenvironment in an ectopic tooth transplantation bioassay. Thus, the isolation and identification of the chemical components of the inductive microenvironment in pulp/dentin regeneration will accelerate progress towards the goal of tissue engineering of the tooth. The teeth demineralized in 0.6 M hydrochloric acid were sequentially extracted by 4.0 M guanidine hydrochloride (GdnHCl), pH 7.4, and 0.5 M ethylenediaminetetraacetic acid (EDTA), pH 7.4. The extracted teeth were transplanted into an ectopic site in severe combined immunodeficiency (SCID) mice with mobilized dental pulp stem cells (MDPSCs). The unextracted tooth served as a positive control. Furthermore, the soluble components for the inductive microenvironment, the GdnHCl extracts, or the EDTA extracts together with or without MDPSC conditioned medium (CM) were reconstituted systematically with autoclaved teeth in which the chemical components were completely inactivated and only the physical microenvironment was preserved. Their pulp/dentin regenerative potential and angiogenic potential were compared 28 days after ectopic tooth transplantation by histomorphometry and real-time RT-PCR analysis. Expression of an odontoblastic marker, enamelysin, and a pulp marker, thyrotropin-releasing hormone degrading enzyme (TRH-DE), was lower, and expression of a periodontal cell marker, anti-asporin/periodontal ligament-associated protein 1 (PLAP-1), was higher in the transplant of the EDTA-extracted teeth compared with the GdnHCl-extracted teeth. The autoclaved teeth reconstituted with the GdnHCl extracts or the EDTA extracts have weak regenerative potential and minimal angiogenic potential, and the CM significantly increased this potential

  9. Evaluating adhesion and alignment of dental pulp stem cells to a spider silk substrate for tissue engineering applications.

    PubMed

    Hafner, Katherine; Montag, Dallas; Maeser, Hannah; Peng, Congyue; Marcotte, William R; Dean, Delphine; Kennedy, Marian S

    2017-12-01

    A proposed source of stem cells for nerve regeneration are dental pulp stem cells (DPSCs), based on their close embryonic origin to neurons and the ease with which DPSCs can be obtained from a donor. This study evaluated the response of human DPSCs to spider dragline silk fibers, a potential substrate material for tissue regeneration. The DPSCs' morphology and spread pattern were characterized after these cells were plated onto Nephila clavipes dragline fibers in media. In addition, the responses of two other well established cell lines, osteoblasts (7F2s), and fibroblasts (3T3s), were also studied under identical conditions. The inclusion of 3T3s and 7F2s in this study allowed for both direct comparisons to prior published work and a qualitative comparison to the morphology of the DPSCs. After twelve days, the DPSCs exhibited greater relative alignment and adherence to the spider dragline fibers than the 3T3s and 7F2s. The impact of a common sterilization method (ultraviolet light) on the spider dragline fiber surface and subsequent cell response to this modified surface was also characterized. Exposure of the silk to ultraviolet light did not have a measureable effect on cell alignment, but it did eliminate bacterial growth and changed fiber surface roughness. Spiders' exposure to stressful environments did not have an effect on silk to impair cell alignment or adhesion. Synthetic recombinant protein silk did not act as a substrate for cell adhesion or alignment but hydrogels with similar composition supported cell attachment, growth and proliferation. In all cases, natural drawn spider silk acted as an effective substrate for cellular adhesion and alignment of DPSCs and could be used in neural differentiation applications. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. The effects of dental pulp stem cells on bone regeneration in rat calvarial defect model: micro-computed tomography and histomorphometric analysis.

    PubMed

    Asutay, Fatih; Polat, Serkan; Gül, Mehmet; Subaşı, Cansu; Kahraman, Sevil Altundağ; Karaöz, Erdal

    2015-12-01

    Stem cell therapies may be applicable to all fields of medicine, including craniomaxillofacial surgery. Dental pulp stem cells also have significant osteogenic properties. This study aimed to evaluate the influence of dental pulp stem cells on bone regeneration and to ascertain whether or not there was any superiority over traditional methods. In this study, 15 non-immunodeficient Wistar albino rats were used. The rats were divided into three groups: (1) untreated control group; (2) hydroxyapatite tri-calcium-phosphate (HA/TCP) paste; (3) human dental pulp derived stem cells (DPSC) mixed with HA/TCP paste (HA/TCP+DSPC group, n=10). Two symmetrical full-thickness cranial defects were created on each parietal region (10 defects for each group). The animals were sacrificed 8 weeks post-surgery and samples were analyzed by microcomputer tomography (μ-CT) and histomorphometry. The calcification rate and bone mineral density (BMD) values in Group 3 were found to be significantly higher than in the other two groups. Radiographically, bone regeneration was greater in Group 2 compared with the control group. However, there was no significant difference between Groups 2 and 1 in respect of histological analysis. According to the results of the present study, DPSCs may be a suitable factor for bone tissue engineering because they can be easily obtained and differentiate into bone cells. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Self-renewal and multipotency coexist in a long-term cultured adult rat dental pulp stem cell line: an exception to the rule?

    PubMed

    Sprio, Andrea E; Di Scipio, Federica; Raimondo, Stefania; Salamone, Paolina; Pagliari, Francesca; Pagliari, Stefania; Folino, Anna; Forte, Giancarlo; Geuna, Stefano; Di Nardo, Paolo; Berta, Giovanni N

    2012-12-10

    The stemness state is characterized by self-renewal and differentiation properties. However, stem cells are not able to preserve these characteristics in long-term culture because of the intrinsic fragility of their phenotype easily undergoing senescence or neoplastic transformation. Furthermore, although isolated from the same original tissue using similar protocols, adult stem cells can display dissimilar phenotypes and important cell clone/species contamination. Finally, the lack of a clear standardization contributes to complicate the comprehension about the stemness condition. In this context, cell lines displaying a particularly stable phenotype must be identified to define one or multiple benchmarks against which other stem cell lines could be reliably assessed. The present paper demonstrates that it is possible to isolate from the rat dental pulp a stem cell line (MUR-1) that does not display neoplastic transformation in long-term culture. MUR-1 cells stably express a broad range of stemness markers and are able to differentiate into adipogenic, osteogenic, chondrogenic, neurogenic, and cardiomyogenic lineages independently of the culture passages. Moreover, serial in vitro passages have not changed their immunophenotype, proliferation capacity, or differentiation potential. The uniqueness of these characteristics candidates MUR-1 as a model to reliably improve the understanding of the mechanisms governing the stem cell fate in the same as well as in other stem cell populations.

  12. Immunohistochemical analysis of two stem cell markers of α-smooth muscle actin and STRO-1 during wound healing of human dental pulp.

    PubMed

    Yoshiba, Nagako; Yoshiba, Kunihiko; Ohkura, Naoto; Shigetani, Yoshimi; Takei, Erika; Hosoya, Akihiro; Nakamura, Hiroaki; Okiji, Takashi

    2012-10-01

    Recent studies have employed two markers, alpha-smooth muscle actin (α-SMA) and STRO-1, to detect cells with mesenchymal stem cell properties in dental pulp. The present study aimed to explore the expression profile of α-SMA and STRO-1 in intact dental pulp as well as during wound healing in adult dental pulp tissue. Healthy pulps were mechanically exposed and capped with the clinically used materials MTA (ProRoot White MTA) or Ca(OH)₂ to induce a mineralized barrier at the exposed surface. After 7-42 days, the teeth were extracted and processed for immunohistochemical analysis using antibodies against α-SMA, STRO-1 and nestin (a neurogenic cytoskeletal protein expressed in odontoblasts). In normal pulp, α-SMA was detected in vascular smooth muscle cells and pericytes. Double immunofluorescent staining with STRO-1 and α-SMA showed that STRO-1 was localized in vascular smooth muscle cells, pericytes and endothelial cells, in addition to nerve fibers. During the process of dental pulp healing, numerous α-SMA-positive cells emerged at the wound margin at 14 days, and the initially formed mineralized barrier was lined with α-SMA-positive cells similar in appearance to reparative odontoblasts, some of which co-expressed nestin. STRO-1 was abundant in nerve fibers. In the advanced stage of mineralized barrier formation at 42 days, cells lining the barrier were stained with nestin, and no staining of α-SMA was detected in those cells. These observations indicate that α-SMA-positive cells temporarily appear along the wound margin during the earlier phase of mineralized barrier formation and STRO-1 is confined in vascular and neuronal elements.

  13. Regeneration of dental pulp/dentine complex with a three-dimensional and scaffold-free stem-cell sheet-derived pellet.

    PubMed

    Na, Sijia; Zhang, Hao; Huang, Fang; Wang, Weiqi; Ding, Yin; Li, Dechao; Jin, Yan

    2016-03-01

    Dental pulp/dentine complex regeneration is indispensable to the construction of biotissue-engineered tooth roots and represents a promising approach to therapy for irreversible pulpitis. We used a tissue-engineering method based on odontogenic stem cells to design a three-dimensional (3D) and scaffold-free stem-cell sheet-derived pellet (CSDP) with the necessary physical and biological properties. Stem cells were isolated and identified and stem cells from root apical papilla (SCAPs)-based CSDPs were then fabricated and examined. Compact cell aggregates containing a high proportion of extracellular matrix (ECM) components were observed, and the CSDP culture time was prolonged. The expression of alkaline phosphatase (ALP), dentine sialoprotein (DSPP), bone sialoprotein (BSP) and runt-related gene 2 (RUNX2) mRNA was higher in CSDPs than in cell sheets (CSs), indicating that CSDPs have greater odonto/osteogenic potential. To further investigate this hypothesis, CSDPs and CSs were inserted into human treated dentine matrix fragments (hTDMFs) and transplanted into the subcutaneous space in the backs of immunodeficient mice, where they were cultured in vivo for 6 weeks. The root space with CSDPs was filled entirely with a dental pulp-like tissue with well-established vascularity, and a continuous layer of dentine-like tissue was deposited onto the existing dentine. A layer of odontoblast-like cells was found to express DSPP, ALP and BSP, and human mitochondria lined the surface of the newly formed dentine-like tissue. These results clearly indicate that SCAP-CSDPs with a mount of endogenous ECM have a strong capacity to form a heterotopic dental pulp/dentine complex in empty root canals; this method can be used in the fabrication of bioengineered dental roots and also provides an alternative treatment approach for pulp disease.

  14. The use of granulocyte-colony stimulating factor induced mobilization for isolation of dental pulp stem cells with high regenerative potential.

    PubMed

    Murakami, Masashi; Horibe, Hiroshi; Iohara, Koichiro; Hayashi, Yuki; Osako, Yohei; Takei, Yoshifumi; Nakata, Kazuhiko; Motoyama, Noboru; Kurita, Kenichi; Nakashima, Misako

    2013-12-01

    Human dental pulp stem cells (DPSCs) contain subsets of progenitor/stem cells with high angiogenic, neurogenic and regenerative potential useful for cell therapy. It is essential to develop a safe and efficacious method to isolate the clinical-grade DPSCs subsets from a small amount of pulp tissue without using conventional flow cytometry. Thus, a method for isolation of DPSCs subsets based on their migratory response to optimized concentration of 100 ng/ml of granulocyte-colony stimulating factor (G-CSF) was determined in this study. The DPSCs mobilized by G-CSF (MDPSCs) were enriched for CD105, C-X-C chemokine receptor type 4 (CXCR-4) and G-CSF receptor (G-CSFR) positive cells, demonstrating stem cell properties including high proliferation rate and stability. The absence of abnormalities/aberrations in karyotype and lack of tumor formation after transplantation in an immunodeficient mouse were demonstrated. The conditioned medium of MDPSCs exhibited anti-apoptotic activity, enhanced migration and immunomodulatory properties. Furthermore, transplantation of MDPSCs accelerated vasculogenesis in an ischemic hindlimb model and augmented regenerated pulp tissue in an ectopic tooth root model compared to that of colony-derived DPSCs, indicating higher regenerative potential of MDPSCs. In conclusion, this isolation method for DPSCs subsets is safe and efficacious, having utility for potential clinical applications to autologous cell transplantation. Copyright © 2013 Elsevier Ltd. All rights reserved.

  15. Eluted zinc ions stimulate osteoblast differentiation and mineralization in human dental pulp stem cells for bone tissue engineering.

    PubMed

    Yusa, Kazuyuki; Yamamoto, Osamu; Iino, Mitsuyoshi; Takano, Hiroshi; Fukuda, Masayuki; Qiao, Zhiwei; Sugiyama, Toshihiro

    2016-11-01

    Zinc is an essential element for proliferation, differentiation and survival in various cell types. In a previous study, we found that zinc ions released from zinc-modified titanium surfaces (eluted zinc ions; EZ) stimulate cell viability, osteoblast marker gene expression and calcium deposition in human bone marrow-derived mesenchymal cells (hBMCs). The aim of the present study was to investigate the effects of EZ on osteoblast differentiation among dental pulp stem cells (DPSCs) in vitro. In this study, we evaluated the effects of EZ on osteogenesis in DPSCs. Osteoblast and osteoclast marker gene expression was evaluated by real-time PCR. We also evaluated alkaline phosphatase (ALP) staining and calcium deposition. We found that EZ stimulated osteoblast marker gene (type I collagen, alkaline phosphatase (ALP), osteocalcin (OCN) and Runx2) expression, vascular endothelial growth factor A (VEGF-A), and TGF-beta signaling pathway-related gene expression after 7days of incubation. Osteoclastogenesis occurs in a receptor for activated nuclear-factor kappa B ligand (RANKL)/osteoprotegerin (OPG)-independent manner. Real-time PCR analysis revealed that EZ did not affect RANKL or OPG mRNA expression. It was also revealed that EZ induced alkaline phosphatase (ALP) staining and calcium deposition in DPSCs. Collectively, these results demonstrate the potential for clinical application to prospective treatment of bone diseases. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Transplantation of human dental pulp stem cells: enhance bone consolidation in mandibular distraction osteogenesis.

    PubMed

    Alkaisi, Amera; Ismail, Abd Rashid; Mutum, Samarendra S; Ahmad, Zainal A Rifin; Masudi, Sam'an; Abd Razak, Noor Hayati

    2013-10-01

    The main aim of the present study was to evaluate the capacity of stem cells from human exfoliated deciduous teeth (SHED) to enhance mandibular distraction osteogenesis (DO) in rabbits. A randomized controlled trial was conducted. Eighteen skeletally immature New Zealand white rabbits were divided into 2 groups, with 9 in the control group and 9 in the SHED group. The SHED were isolated, expanded, and characterized. Six million cells were transplanted into the distracted area during the osteotomy period. After a 4-day latency period, a total of 6 mm was distracted for 6 days. The newly formed bone was analyzed radiologically, histologically, and histomorphometrically at 2, 4, and 6 weeks postoperatively. Nonparametric analysis of variance (Kruskal-Wallis test) was used for data analysis, and P < .05 was considered statistically significant. The cell lineage was positive for the 2 mesenchymal stem cell markers tested (CD105 and CD166). More mature bone in the SHED transplanted group was observed radiographically and histologically. Histomorphologically, the percentage of newly formed bone after 2, 4, and 6 weeks was 18.41% and 41.53%, 31.68% and 59.78%, and 52.34% and 65.24% in the control and SHED groups, respectively. The difference between the groups was statistically significant (P = .012). The bone union and stage of bone maturity scores were significantly different between the control and SHED groups (P = .006 and P = .011, respectively). Our findings suggest that SHED can serve as an additional cell resource for DO enhancement in rabbits and might be a promising model for the reconstruction of large mandibular defects in human oral maxillofacial surgery. Copyright © 2013 American Association of Oral and Maxillofacial Surgeons. Published by Elsevier Inc. All rights reserved.

  17. Multiwall carbon nanotubes/polycaprolactone scaffolds seeded with human dental pulp stem cells for bone tissue regeneration.

    PubMed

    Flores-Cedillo, M L; Alvarado-Estrada, K N; Pozos-Guillén, A J; Murguía-Ibarra, J S; Vidal, M A; Cervantes-Uc, J M; Rosales-Ibáñez, R; Cauich-Rodríguez, J V

    2016-02-01

    Conventional approaches to bone regeneration rarely use multiwall carbon nanotubes (MWCNTs) but instead use polymeric matrices filled with hydroxyapatite, calcium phosphates and bioactive glasses. In this study, we prepared composites of MWCNTs/polycaprolactone (PCL) for bone regeneration as follows: (a) MWCNTs randomly dispersed on PCL, (b) MWCNTs aligned with an electrical field to determine if the orientation favors the growing of human dental pulp stem cells (HDPSCs), and (c) MWCNTs modified with β-glycerol phosphate (BGP) to analyze its osteogenic potential. Raman spectroscopy confirmed the presence of MWCNTs and BGP on PCL, whereas the increase in crystallinity by the addition of MWCNTs to PCL was confirmed by X-ray diffraction and differential scanning calorimetry. A higher elastic modulus (608 ± 4.3 MPa), maximum stress (42 ± 6.1 MPa) and electrical conductivity (1.67 × 10(-7) S/m) were observed in non-aligned MWCNTs compared with the pristine PCL. Cell viability at 14 days was similar in all samples according to the live/dead assay, but the 21 day cell proliferation, measured by MTT was higher in MWCNTs aligned with BGP. Von Kossa and Alizarin red showed larger amounts of mineral deposits on MWCNTs aligned with BGP, indicating that at 21 days, this scaffold promotes osteogenic differentiation of HDPSCs.

  18. Effect of low-level diode laser on proliferation and osteogenic differentiation of dental pulp stem cells

    NASA Astrophysics Data System (ADS)

    Tabatabaei, Fahimeh S.; Torshabi, Maryam; Mojahedi Nasab, Masoud; Khosraviani, Keikhosro; Khojasteh, Arash

    2015-09-01

    This study assessed the effect of low-level laser irradiation (LLLI) on the proliferation and osteogenic differentiation of dental pulp stem cells (DPSCs). DPSCs were exposed to 810 nm laser light (0.1, 0.2, or 0.3 J cm-2) for 7 d (60 s daily). The negative control group (cells in regular medium) and positive control group (cells in osteogenic medium (OM)) were not lased. One group of cells in OM was irradiated with laser operated at 0.2 J cm-2. Cell viability was evaluated at 24 h and one week after the last day of laser irradiation using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Osteogenic differentiation was assessed using real-time reverse transcriptase polymerase chain reaction (RT-PCR) and alizarin Red S staining. Cell proliferation was not affected by laser irradiation at 24 h except in one group (cells in OM exposed to laser at 0.2 J cm-2). However, one week after the last day of laser irradiation, it was significantly increased in groups exposed to laser at 0.1 or 0.2 J cm-2 and decreased in groups containing OM (P  <  0.05). Osteoblast marker expression was observed in groups containing OM. LLLI at 0.2 J cm-2 dramatically enhanced cell differentiation. Laser at 0.3 J cm-2 increased bone sialoprotein (BSP) and decreased alkaline phosphatase (ALP). Mineralized nodules were only observed in groups containing OM. Considering these findings, LLLI may be used as a novel approach for preconditioning of DPSCs in vitro prior to bone tissue engineering.

  19. SDF-1/CXCR4 axis induces human dental pulp stem cell migration through FAK/PI3K/Akt and GSK3β/β-catenin pathways.

    PubMed

    Li, Mingwei; Sun, Xuefei; Ma, Liang; Jin, Lu; Zhang, Wenfei; Xiao, Min; Yu, Qing

    2017-01-09

    SDF-1 (stromal cell derived factor-1) has been found to be widely expressed during dental pulp inflammation, while hDPSCs (human dental pulp stem cells) contribute to the repair of dental pulp. We showed that the migration of hDPSCs was induced by SDF-1 in a concentration-dependent manner and could be inhibited with siCXCR4 (C-X-C chemokine receptor type 4) and siCDC42 (cell division control protein 42), as well as drug inhibitors such as AMD3100 (antagonist of CXCR4), LY294002 (inhibitor of PI3K) and PF573228 (inhibitor of FAK). It was also confirmed that SDF-1 regulated the phosphorylation of FAK (focal adhesion kinases) on cell membranes and the translocation of β-catenin into the cell nucleus. Subsequent experiments confirmed that the expression of CXCR4 and β-catenin and the phosphorylation of FAK, PI3K (phosphoinositide 3-kinase), Akt and GSK3β (glycogen synthase kinase-3β) were altered significantly with SDF-1 stimulation. FAK and PI3K worked in coordination during this process. Our findings provide direct evidence that SDF-1/CXCR4 axis induces hDPSCs migration through FAK/PI3K/Akt and GSK3β/β-catenin pathways, implicating a novel mechanism of dental pulp repair and a possible application of SDF-1 for the treatment of pulpitis.

  20. SDF-1/CXCR4 axis induces human dental pulp stem cell migration through FAK/PI3K/Akt and GSK3β/β-catenin pathways

    PubMed Central

    Li, Mingwei; Sun, Xuefei; Ma, Liang; Jin, Lu; Zhang, Wenfei; Xiao, Min; Yu, Qing

    2017-01-01

    SDF-1 (stromal cell derived factor-1) has been found to be widely expressed during dental pulp inflammation, while hDPSCs (human dental pulp stem cells) contribute to the repair of dental pulp. We showed that the migration of hDPSCs was induced by SDF-1 in a concentration-dependent manner and could be inhibited with siCXCR4 (C-X-C chemokine receptor type 4) and siCDC42 (cell division control protein 42), as well as drug inhibitors such as AMD3100 (antagonist of CXCR4), LY294002 (inhibitor of PI3K) and PF573228 (inhibitor of FAK). It was also confirmed that SDF-1 regulated the phosphorylation of FAK (focal adhesion kinases) on cell membranes and the translocation of β-catenin into the cell nucleus. Subsequent experiments confirmed that the expression of CXCR4 and β-catenin and the phosphorylation of FAK, PI3K (phosphoinositide 3-kinase), Akt and GSK3β (glycogen synthase kinase-3β) were altered significantly with SDF-1 stimulation. FAK and PI3K worked in coordination during this process. Our findings provide direct evidence that SDF-1/CXCR4 axis induces hDPSCs migration through FAK/PI3K/Akt and GSK3β/β-catenin pathways, implicating a novel mechanism of dental pulp repair and a possible application of SDF-1 for the treatment of pulpitis. PMID:28067275

  1. Alcohol-induced suppression of KDM6B dysregulates the mineralization potential in dental pulp stem cells.

    PubMed

    Hoang, Michael; Kim, Jeffrey J; Kim, Yiyoung; Tong, Elizabeth; Trammell, Benjamin; Liu, Yao; Shi, Songtao; Lee, Chang-Ryul; Hong, Christine; Wang, Cun-Yu; Kim, Yong

    2016-07-01

    Epigenetic changes, such as alteration of DNA methylation patterns, have been proposed as a molecular mechanism underlying the effect of alcohol on the maintenance of adult stem cells. We have performed genome-wide gene expression microarray and DNA methylome analysis to identify molecular alterations via DNA methylation changes associated with exposure of human dental pulp stem cells (DPSCs) to ethanol (EtOH). By combined analysis of the gene expression and DNA methylation, we have found a significant number of genes that are potentially regulated by EtOH-induced DNA methylation. As a focused approach, we have also performed a pathway-focused RT-PCR array analysis to examine potential molecular effects of EtOH on genes involved in epigenetic chromatin modification enzymes, fibroblastic markers, and stress and toxicity pathways in DPSCs. We have identified and verified that lysine specific demethylase 6B (KDM6B) was significantly dysregulated in DPSCs upon EtOH exposure. EtOH treatment during odontogenic/osteogenic differentiation of DPSCs suppressed the induction of KDM6B with alterations in the expression of differentiation markers. Knockdown of KDM6B resulted in a marked decrease in mineralization from implanted DPSCs in vivo. Furthermore, an ectopic expression of KDM6B in EtOH-treated DPSCs restored the expression of differentiation-related genes. Our study has demonstrated that EtOH-induced inhibition of KDM6B plays a role in the dysregulation of odontogenic/osteogenic differentiation in the DPSC model. This suggests a potential molecular mechanism for cellular insults of heavy alcohol consumption that can lead to decreased mineral deposition potentially associated with abnormalities in dental development and also osteopenia/osteoporosis, hallmark features of fetal alcohol spectrum disorders.

  2. Runx2 modified dental pulp stem cells (DPSCs) enhance new bone formation during rapid distraction osteogenesis (DO).

    PubMed

    Feng, Guijuan; Zhang, Jinlong; Feng, Xingmei; Wu, Senbin; Huang, Dan; Hu, Jing; Zhu, Songsong; Song, Donghui

    Distraction osteogenesis (DO) remains a major challenge in orthopedic and craniofacial surgery. The transplantion of mesenchymal stem cells (MSCs) could reduce the treatment period and the associated complications by increasing new bone formation during long-bone DO. Runt-related transcription factor 2 (Runx2) encodes a nuclear protein that is a pivotal regulator of osteoblast differentiation. It significantly stimulates calcium accumulation and alkaline phosphatase (ALP) activity in dental pulp stem cells (DPSCs). In this study, we investigated the effects of gene therapy using Runx2 on new bone formation during tibia DO of rabbits. The distraction gap of the rabbits was injected with adenovirus (Adv)-Runx2-green fluorescent protein (GFP)-transfected DPSCs (overexpression group, Group OE) or Adv-GFP-transfected DPSCs (negative control group, Group NC). Rabbits in the control group (Groups CON) were injected with physiologic saline. The generation of new bone tissue in the distraction gap was studied by radiographic examination, micro-computed tomography (CT) evaluation, histological analyze, and Mechanical testing at weeks 8 in the consolidation period. Excellent bone formation in the distracted callus was observed in Group OE and Group NC. Moreover, the OE group showed better bone formation and the highest bone mineral density (BMD) and bone mineral content (BMC). Group CON animals showed inadequate bone formation in the distracted callus compared to the other groups. The results suggest that gene therapy using Runx2-modified DPSCs was more effective during bone deposition and new bone formation in tibia DO. Copyright © 2016 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

  3. Multifaceted Therapeutic Benefits of Factors Derived From Dental Pulp Stem Cells for Mouse Liver Fibrosis.

    PubMed

    Hirata, Marina; Ishigami, Masatoshi; Matsushita, Yoshihiro; Ito, Takanori; Hattori, Hisashi; Hibi, Hideharu; Goto, Hidemi; Ueda, Minoru; Yamamoto, Akihito

    2016-10-01

    : Chronic liver injury from various causes often results in liver fibrosis (LF). Although the liver possesses endogenous tissue-repairing activities, these can be overcome by sustained inflammation and excessive fibrotic scar formation. Advanced LF leads to irreversible cirrhosis and subsequent liver failure and/or hepatic cancer. Here, using the mouse carbon tetrachloride (CCl4)-induced LF model, we showed that a single intravenous administration of stem cells derived from human exfoliated deciduous teeth (SHEDs) or of SHED-derived serum-free conditioned medium (SHED-CM) resulted in fibrotic scar resolution. SHED-CM suppressed the gene expression of proinflammatory mediators, such as TNF-α, IL-1β, and iNOS, and eliminated activated hepatic stellate cells by inducing their apoptosis, but protected parenchymal hepatocytes from undergoing apoptosis. In addition, SHED-CM induced tissue-repairing macrophages that expressed high levels of the profibrinolytic factor, matrix metalloproteinase 13. Furthermore, SHED-CM suppressed the CCl4-induced apoptosis of primary cultured hepatocytes. SHED-CM contained a high level of hepatocyte growth factor (HGF). Notably, HGF-depleted SHED-CM (dHGF-CM) did not suppress the proinflammatory response or resolve fibrotic scarring. Furthermore, SHED-CM, but not dHGF-CM, inhibited CCl4-induced hepatocyte apoptosis. These results suggest that HGF plays a central role in the SHED-CM-mediated resolution of LF. Taken together, our findings suggest that SHED-CM provides multifaceted therapeutic benefits for the treatment of LF. This study demonstrated that a single intravenous administration of stem cells from human exfoliated deciduous teeth (SHEDs) or of the serum-free conditioned medium (CM) derived from SHEDs markedly improved mouse liver fibrosis (LF). SHED-CM suppressed chronic inflammation, eliminated activated hepatic stellate cells by inducing their apoptosis, protected hepatocytes from undergoing apoptosis, and induced

  4. The role of integrin-α5 in the proliferation and odontogenic differentiation of human dental pulp stem cells.

    PubMed

    Cui, Li; Xu, Shuaimei; Ma, Dandan; Gao, Jie; Liu, Ying; Yue, Jing; Wu, Buling

    2014-02-01

    It has been reported that integrin-α5 (ITGA5) activity is related to cell proliferation, differentiation, migration, and organ development. However, the involvement of ITGA5 in the biological functions of human dental pulp stem cells (hDPSCs) has not been explored. The aim of this study was to investigate the role of ITGA5 in the proliferation and odontogenic differentiation of hDPSCs. We knocked down ITGA5 in hDPSCs using lentivirus-mediated ITGA5 short hairpin RNA (shRNA). Changes in the proliferation in hDPSCs infected with lentiviruses expressing ITGA5-specific shRNA or negative control shRNA were examined using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and 5-ethynyl-2'-deoxyuridine labeling. Both ITGA5 knockdown cells and shMock cells were cultured in mineralization medium for 3 weeks, and the differentiation of cells was detected with alizarin red S staining. The expression of odontogenic differentiation-related molecular markers was assessed using real-time polymerase chain reaction and Western blot assays. The knockdown of ITGA5 decreased the proliferation capacity of hDPSCs. ITGA5 shRNA promoted odontogenic differentiation of hDPSCs with the enhanced formation of mineralized nodules. It also up-regulated the messenger RNA expression of multiple markers of odontogenesis and the expression of dentin sialophosphoprotein protein. These findings suggest that ITGA5 plays an important role in maintaining hDPSCs in a proliferative state. The inhibition of ITGA5 signaling promotes the odontogenic differentiation of hDPSCs. Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  5. Neurogenic Maturation of Human Dental Pulp Stem Cells Following Neurosphere Generation Induces Morphological and Electrophysiological Characteristics of Functional Neurons

    PubMed Central

    Struys, Tom; Hilkens, Petra; Bronckaers, Annelies; Ratajczak, Jessica; Politis, Constantinus; Brône, Bert; Lambrichts, Ivo; Martens, Wendy

    2015-01-01

    Cell-based therapies are emerging as an alternative treatment option to promote functional recovery in patients suffering from neurological disorders, which are the major cause of death and permanent disability. The present study aimed to differentiate human dental pulp stem cells (hDPSCs) toward functionally active neuronal cells in vitro. hDPSCs were subjected to a two-step protocol. First, neuronal induction was acquired through the formation of neurospheres, followed by neuronal maturation, based on cAMP and neurotrophin-3 (NT-3) signaling. At the ultrastructural level, it was shown that the intra-spheral microenvironment promoted intercellular communication. hDPSCs grew out of the neurospheres in vitro and established a neurogenic differentiated hDPSC culture (d-hDPSCs) upon cAMP and NT-3 signaling. d-hDPSCs were characterized by the increased expression of neuronal markers such as neuronal nuclei, microtubule-associated protein 2, neural cell adhesion molecule, growth-associated protein 43, synapsin I, and synaptophysin compared with nondifferentiated hDPSCs. Enzyme-linked immunosorbent assay demonstrated that the secretion of brain-derived neurotrophic factor, vascular endothelial growth factor, and nerve growth factor differed between d-hDPSCs and hDPSCs. d-hDPSCs acquired neuronal features, including multiple intercommunicating cytoplasmic extensions and increased vesicular transport, as shown by the electron microscopic observation. Patch clamp analysis demonstrated the functional activity of d-hDPSCs by the presence of tetrodotoxin- and tetraethyl ammonium-sensitive voltage-gated sodium and potassium channels, respectively. A subset of d-hDPSCs was able to fire a single action potential. The results reported in this study demonstrate that hDPSCs are capable of neuronal commitment following neurosphere formation, characterized by distinct morphological and electrophysiological properties of functional neuronal cells. PMID:25203005

  6. Immunohistochemical and histochemical analysis of newly formed tissues in root canal space transplanted with dental pulp stem cells plus platelet-rich plasma.

    PubMed

    Zhu, Xiaofei; Wang, Yu; Liu, Yuan; Huang, George T-J; Zhang, Chengfei

    2014-10-01

    Tissue regeneration in root canals after pulpectomy can be achieved by transplantation of autologous dental pulp stem cells and/or platelet-rich plasma. However, the identity of the newly formed tissue in the pulp space has been only examined by histologic analysis. This study aimed to apply immunohistochemistry and histochemistry to detect specific markers in the newly generated tissues after root canal regenerative treatment. In our previous study, 32 root canals in 4 mature dogs were treated with a pulp regeneration procedure after pulpectomy using either blood clot, transplantation of dental pulp stem cells, platelet-rich plasma, or a combination of cells and plasma. In the present study, the tissues were examined for the expression of periostin to detect periodontal ligament tissue, nestin and dentin sialoprotein for odontoblasts, and bone sialoprotein and osteocalcin for bone tissues. Samples were also stained for tartrate-resistant acid phosphatase (TRAP) as a marker for osteoclastic lineages. Continuous periostin-positive tissue was observed extending from the periodontal ligament into the inner canal surface in which the mineral islands were surrounded by weak periostin staining. There was also positive staining for TRAP, bone sialoprotein, and osteocalcin in the canal space, suggesting the presence of bone tissue. A layer of mineralized tissue along the inner surface of the root canal was negative for TRAP, suggesting the tissue likely to be cementum. In all samples, no nestin-positive reaction was observed, whereas dentin sialoprotein was detected in PDL, dentinal tubules, and intracanal fibrous tissues. There was no difference between any of the 4 groups. The tissues formed in the dog mature root canals after regenerative endodontic procedures are not pulp tissues but mainly periodontal tissues. Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  7. Isolation of a Stable Subpopulation of Mobilized Dental Pulp Stem Cells (MDPSCs) with High Proliferation, Migration, and Regeneration Potential Is Independent of Age

    PubMed Central

    Iohara, Koichiro; Hayashi, Yuki; Takeuchi, Norio; Takei, Yoshifumi; Kurita, Kenichi; Nakashima, Misako

    2014-01-01

    Insights into the understanding of the influence of the age of MSCs on their cellular responses and regenerative potential are critical for stem cell therapy in the clinic. We have isolated dental pulp stem cells (DPSCs) subsets based on their migratory response to granulocyte-colony stimulating factor (G-CSF) (MDPSCs) from young and aged donors. The aged MDPSCs were efficiently enriched in stem cells, expressing high levels of trophic factors with high proliferation, migration and anti-apoptotic effects compared to young MDPSCs. In contrast, significant differences in those properties were detected between aged and young colony-derived DPSCs. Unlike DPSCs, MDPSCs showed a small age-dependent increase in senescence-associated β-galactosidase (SA-β-gal) production and senescence markers including p16, p21, Interleukin (IL)-1β, -6, -8, and Groα in long-term culture. There was no difference between aged and young MDPSCs in telomerase activity. The regenerative potential of aged MDPSCs was similar to that of young MDPSCs in an ischemic hindlimb model and an ectopic tooth root model. These results demonstrated that the stem cell properties and the high regenerative potential of MDPSCs are independent of age, demonstrating an immense utility for clinical applications by autologous cell transplantation in dental pulp regeneration and ischemic diseases. PMID:24870376

  8. Increased proliferation and adhesion properties of human dental pulp stem cells in PLGA scaffolds via simulated microgravity.

    PubMed

    He, L; Pan, S; Li, Y; Zhang, L; Zhang, W; Yi, H; Song, C; Niu, Y

    2016-02-01

    To explore the possibility of utilizing a rotary cell culture system (RCCS) to model simulated microgravity and investigate its effects on the proliferation, adhesion, migration and cytoskeletal organization of human dental pulp stem cells (hDPSCs) on poly (lactic-co-glycolic acid) (PLGA) scaffolds. Isolated and identified hDPSCs grown on PLGA scaffolds were exposed to simulated microgravity (SMG) or normal gravity (NG) conditions for 3 days. MTT cell proliferation assays, BrdU incorporation assays, flow cytometry analysis and Western blotting were undertaken to identify the proliferation ability of hDPSCs under SMG conditions. Additionally, immunofluorescence detection, SEM observations and cell migration and adhesion assays were performed to compare adhesion, migration and cytoskeletal changes in hDPCSs subjected to SMG conditions. To further investigate the mechanisms, human pathway-focused matrix and adhesion PCR array analyses were performed. The Student's t-test was used for statistical analyses. SMG promoted proliferation and adhesion, decreased migration and reorganized the cytoskeletal organization of hDPSCs compared with the NG group. PCR array analyses revealed that following SMG treatment, ITGA6 (integrin alpha-6), ITGAV (integrin alpha-V), ITGB1 (integrin beta-1), LAMB1 (laminin beta-1) and TNC (tenascin-C) were significantly upregulated (P < 0.05). SMG may regulate the behaviour of hDPSCs grown in PLGA scaffolds in an integrin-mediated manner, which may contribute to tooth tissue engineering by increasing their expandability and scaffold adhesion. © 2015 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  9. Impact of isolation method on doubling time and the quality of chondrocyte and osteoblast differentiated from murine dental pulp stem cells

    PubMed Central

    Senafi, Sahidan; Zainol Abidin, Intan Zarina; Zainal Ariffin, Zaidah

    2017-01-01

    Background Stem cells are normally isolated from dental pulps using the enzymatic digestion or the outgrowth method. However, the effects of the isolation method on the quality of the isolated stem cells are not studied in detail in murine models. The aim of this study was to compare the matrices secreted by osteoblast and chondrocytes differentiated from dental pulp stem cells isolated through different means. Method DPSC from murine incisors were isolated through either the outgrowth (DPSC-OG) or the enzymatic digestion (DPSC-ED) method. Cells at passage 4 were used in this study. The cells were characterized through morphology and expression of cell surface markers. The cells’ doubling time when cultured using different seeding densities was calculated and analyzed using one-way ANOVA and Tukey’s multiple comparison post-test. The ability of cells to differentiate to chondrocyte and osteoblast was evaluated through staining and analysis on the matrices secreted. Results Gene expression analysis showed that DPSC-OG and DPSC-ED expressed dental pulp mesenchymal stem cell markers, but not hematopoietic stem cell markers. The least number of cells that could have been used to culture DPSC-OG and DPSC-ED with the shortest doubling time was 5 × 102 cells/cm2 (11.49 ± 2.16 h) and 1 × 102 cells/cm2 (10.55 h ± 0.50), respectively. Chondrocytes differentiated from DPSC-ED produced  2 times more proteoglycan and at a faster rate than DPSC-OG. FTIR revealed that DPSC-ED differentiated into osteoblast also secreted matrix, which more resembled a calvaria. Discussion Isolation approaches might have influenced the cell populations obtained. This, in turn, resulted in cells with different proliferation and differentiation capability. While both DPSC-OG and DPSC-ED expressed mesenchymal stem cell markers, the percentage of cells carrying each marker might have differed between the two methods. Regardless, enzymatic digestion clearly yielded cells with better

  10. Isolation and Characterization of Human Dental Pulp Derived Stem Cells by Using Media Containing Low Human Serum Percentage as Clinical Grade Substitutes for Bovine Serum

    PubMed Central

    Beltrami, Antonio Paolo; Cesselli, Daniela; Curcio, Francesco

    2012-01-01

    Adult stem cells have been proposed as an alternative to embryonic stem cells to study multilineage differentiation in vitro and to use in therapy. Current culture media for isolation and expansion of adult stem cells require the use of large amounts of animal sera, but animal-derived culture reagents give rise to some questions due to the real possibility of infections and severe immune reactions. For these reasons a clinical grade substitute to animal sera is needed. We tested the isolation, proliferation, morphology, stemness related marker expression, and osteoblastic differentiation potential of Dental Pulp Stem Cells (DPSC) in a chemically defined medium containing a low percentage of human serum, 1.25%, in comparison to a medium containing 10% Fetal Bovine Serum (FBS). DPSCs cultured in presence of our isolation/proliferation medium added with low HS percentage were obtained without immune-selection methods and showed high uniformity in the expression of stem cell markers, proliferated at higher rate, and demonstrated comparable osteoblastic potential with respect to DPSCs cultured in 10% FBS. In this study we demonstrated that a chemically defined medium added with low HS percentage, derived from autologous and heterologous sources, could be a valid substitute to FBS-containing media and should be helpful for adult stem cells clinical application. PMID:23155430

  11. Stem Cells

    MedlinePlus

    Stem cells are cells with the potential to develop into many different types of cells in the body. ... the body. There are two main types of stem cells: embryonic stem cells and adult stem cells. Stem ...

  12. Human dental pulp-derived stem cells promote locomotor recovery after complete transection of the rat spinal cord by multiple neuro-regenerative mechanisms.

    PubMed

    Sakai, Kiyoshi; Yamamoto, Akihito; Matsubara, Kohki; Nakamura, Shoko; Naruse, Mami; Yamagata, Mari; Sakamoto, Kazuma; Tauchi, Ryoji; Wakao, Norimitsu; Imagama, Shiro; Hibi, Hideharu; Kadomatsu, Kenji; Ishiguro, Naoki; Ueda, Minoru

    2012-01-01

    Spinal cord injury (SCI) often leads to persistent functional deficits due to loss of neurons and glia and to limited axonal regeneration after injury. Here we report that transplantation of human dental pulp stem cells into the completely transected adult rat spinal cord resulted in marked recovery of hind limb locomotor functions. Transplantation of human bone marrow stromal cells or skin-derived fibroblasts led to substantially less recovery of locomotor function. The human dental pulp stem cells exhibited three major neuroregenerative activities. First, they inhibited the SCI-induced apoptosis of neurons, astrocytes, and oligodendrocytes, which improved the preservation of neuronal filaments and myelin sheaths. Second, they promoted the regeneration of transected axons by directly inhibiting multiple axon growth inhibitors, including chondroitin sulfate proteoglycan and myelin-associated glycoprotein, via paracrine mechanisms. Last, they replaced lost cells by differentiating into mature oligodendrocytes under the extreme conditions of SCI. Our data demonstrate that tooth-derived stem cells may provide therapeutic benefits for treating SCI through both cell-autonomous and paracrine neuroregenerative activities.

  13. Effects of Nerve Growth Factor and Basic Fibroblast Growth Factor Promote Human Dental Pulp Stem Cells to Neural Differentiation.

    PubMed

    Zhang, Jinlong; Lian, Min; Cao, Peipei; Bao, Guofeng; Xu, Guanhua; Sun, Yuyu; Wang, Lingling; Chen, Jiajia; Wang, Yi; Feng, Guijuan; Cui, Zhiming

    2017-04-01

    Dental pulp stem cells (DPSCs) were the most widely used seed cells in the field of neural regeneration and bone tissue engineering, due to their easily isolation, lack of ethical controversy, low immunogenicity and low rates of transplantation rejection. The purpose of this study was to investigate the role of basic fibroblast growth factor (bFGF) and nerve growth factor (NGF) on neural differentiation of DPSCs in vitro. DPSCs were cultured in neural differentiation medium containing NGF and bFGF alone or combination for 7 days. Then neural genes and protein markers were analyzed using western blot and RT-PCR. Our study revealed that bFGF and NGF increased neural differentiation of DPSCs synergistically, compared with bFGF and NGF alone. The levels of Nestin, MAP-2, βIII-tubulin and GFAP were the most highest in the DPSCs + bFGF + NGF group. Our results suggested that bFGF and NGF signifiantly up-regulated the levels of Sirt1. After treatment with Sirt1 inhibitor, western blot, RT-PCR and immunofluorescence staining showed that neural genes and protein markers had markedly decreased. Additionally, the ERK and AKT signaling pathway played a key role in the neural differentiation of DPSCs stimulated with bFGF + NGF. These results suggested that manipulation of the ERK and AKT signaling pathway may be associated with the differentiation of bFGF and NGF treated DPSCs. Our date provided theoretical basis for DPSCs to treat neurological diseases and repair neuronal damage.

  14. Analysis of the Adherence of Dental Pulp Stem Cells on Two-Dimensional and Three-Dimensional Silk Fibroin-Based Biomaterials: Applications in Regenerative Dentistry.

    PubMed

    Pecci-Lloret, María Pilar; Vera-Sánchez, Mar; Aznar-Cervantes, Salvador; García-Bernal, David; Sánchez, Ricardo Oñate; Pecci-Lloret, Miguel Ramón; Moraleda, José María; Cenis, José Luis; Rodríguez-Lozano, Francisco Javier

    2017-02-22

    Among various biomaterials used as scaffolds in tissue engineering, silk fibroin is a highly attractive material. A scaffold should be biocompatible and nontoxic, with optimal physical features and mechanical properties. For this reason, tissue-engineering approaches in regenerative medicine have focused on investigating the biocompatibility of possible biomaterials by analyzing cell-scaffold interaction properties. The aim of the present study was to examine the biocompatibility of silk fibroin as a film (two-dimensional [2D]) and a scaffold (three-dimensional [3D]) after being cellularized with human dental pulp stem cells (hDPSCs). Human dental pulp stem cells were isolated from healthy patients aged between 18 and 31 years. Further, silk fibroin-based 2D films and 3D scaffolds were prepared. Human dental pulp stem cells were directly seeded onto the biomaterial surfaces and their proliferation, adherence, and cell morphology were analyzed after 24, 120, and 168 hours. Additionally, the characteristics of the silk fibroin 2D films and 3D scaffolds before and after cell seeding were analyzed by scanning electron microscopy. After the initial 24 hours, silk fibroin-based 3D scaffolds displayed more adhered cells with a suitable fibroblastic morphology than those displayed on the 2D films. After longer culture times, hDPSCs proliferated sufficiently to cover the entire surface of the 3D silk fibroin scaffold, whereas the 2D films were only partially covered. Our results indicate the good in vitro biocompatibility of silk fibroin-based biomaterials, especially when 3D scaffolds rather than 2D films are used.

  15. Tooth Tissue Engineering: The Importance of Blood Products as a Supplement in Tissue Culture Medium for Human Pulp Dental Stem Cells.

    PubMed

    Pisciolaro, Ricardo Luiz; Duailibi, Monica Talarico; Novo, Neil Ferreira; Juliano, Yara; Pallos, Debora; Yelick, Pamela Crotty; Vacanti, Joseph Phillip; Ferreira, Lydia Masako; Duailibi, Silvio Eduardo

    2015-11-01

    One of the goals in using cells for tissue engineering (TE) and cell therapy consists of optimizing the medium for cell culture. The present study compares three different blood product supplements for improved cell proliferation and protection against DNA damage in cultured human dental pulp stem cells for tooth TE applications. Human cells from dental pulp were first characterized as adult stem cells (ectomesenchymal mixed origin) by flow cytometry. Next, four different cell culture conditions were tested: I, supplement-free; II, supplemented with fetal bovine serum; III, allogeneic human serum; and IV, autologous human serum. Cultured cells were then characterized for cell proliferation, mineralized nodule formation, and colony-forming units (CFU) capability. After 28 days in culture, the comet assay was performed to assess possible damage in cellular DNA. Our results revealed that Protocol IV achieved higher cell proliferation than Protocol I (p = 0.0112). Protocols II and III resulted in higher cell proliferation than Protocol I, but no statistical differences were found relative to Protocol IV. The comet assay revealed less cell damage in cells cultured using Protocol IV as compared to Protocols II and III. The damage percentage observed on Protocol II was significantly higher than all other protocols. CFUs capability was highest using Protocol IV (p = 0.0018) and III, respectively, and the highest degree of mineralization was observed using Protocol IV as compared to Protocols II and III. Protocol IV resulted in significantly improved cell proliferation, and no cell damage was observed. These results demonstrate that human blood product supplements can be used as feasible supplements for culturing adult human dental stem cells.

  16. Insulin-like growth factor 1 promotes the proliferation and committed differentiation of human dental pulp stem cells through MAPK pathways.

    PubMed

    Lv, Taohong; Wu, Yongzheng; Mu, Chao; Liu, Genxia; Yan, Ming; Xu, Xiangqin; Wu, Huayin; Du, Jinyin; Yu, Jinhua; Mu, Jinquan

    2016-12-01

    Insulin-like growth factor 1 (IGF-1) is a broad-spectrum growth-promoting factor that plays a key role in natural tooth development. Human dental pulp stem cells (hDPSCs) are multipotent and can influence the reparative regeneration of dental pulp and dentin. This study was designed to evaluate the effects of IGF-1 on the proliferation and differentiation of human dental pulp stem cells. HDPSCs were isolated and purified from human dental pulps. The proliferation and osteo/odontogenic differentiation of hDPSCs treated with 100ng/ml exogenous IGF-1 were subsequently investigated. MTT assays revealed that IGF-1 enhanced the proliferation of hDPSCs. ALP activity in IGF-1-treated group was obviously enhanced compared to the control group from days 3 to 9. Alizarin red staining revealed that the IGF-1-treated cells contained a greater number of mineralization nodules and had higher calcium concentrations. Moreover, western blot and qRT-PCR analyses demonstrated that the expression levels of several osteogenic genes (e.g., RUNX2, OSX, and OCN) and an odontoblast-specific marker (DSPP) were significantly up-regulated in IGF-1-treated hDPSCs as compared with untreated cells (P<0.01). Interestingly, the expression of phospho-ERK and phospho-p38 were also up-regulated, indicating that the MAPK signaling pathway is activated during the differentiation of hDPSCs. IGF-1 can promote the proliferation and osteo/odontogenic differentiation of hDPSCs by activating MAPK pathways. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. OPTIMIZED CRYOPRESERVATION METHOD FOR HUMAN DENTAL PULP-DERIVED STEM CELLS AND THEIR TISSUES OF ORIGIN FOR BANKING AND CLINICAL USE

    PubMed Central

    Woods, Erik J.; Perry, Brandon C.; Hockema, J. Jeffrey; Larson, Lindsay; Zhou, Dan; Goebel, W. Scott

    2009-01-01

    Dental pulp is a promising source of mesenchymal stem cells with the potential for cell-mediated therapies and tissue engineering applications. We recently reported that isolation of dental pulp-derived stem cells (DPSC) is feasible for at least 120 hours after tooth extraction, and that cryopreservation of early-passage cultured DPSC leads to high-efficiency recovery post thaw. This study investigated additional processing and cryobiological characteristics of DPSC, ending with development of procedures for banking. First, we aimed to optimize cryopreservation of established DPSC cultures, with regards to optimizing the cryoprotective agent (CPA), the CPA concentration, the concentration of cells frozen, and storage temperatures. Secondly, we focused on determining cryopreservation characteristics of enzymatically digested tissue as a cell suspension. Lastly, we evaluated the growth, surface markers and differentiation properties of DPSC obtained from intact teeth and undigested, whole dental tissue frozen and thawed using the optimized procedures. In these experiments it was determined that Me2SO at a concentration between 1 and 1.5M was the ideal cryopreservative of the three studied. It was also determined that DPSC viability after cryopreservation is not limited by the concentration of cells frozen, at least up to 2 × 106 cells/mL. It was further established that DPSC can be stored at −85°C or −196°C for at least six months without loss of functionality. The optimal results with the least manipulation were achieved by isolating and cryopreserving the tooth pulp tissues, with digestion and culture performed post-thaw. A recovery of cells from >85% of the tissues frozen was achieved and cells isolated post thaw from tissue processed and frozen with a serum free, defined cryopreservation medium maintained morphological and developmental competence and demonstrated MSC-hallmark trilineage differentiation under the appropriate culture conditions. PMID

  18. Effects of Perivitelline Fluid Obtained from Horseshoe Crab on The Proliferation and Genotoxicity of Dental Pulp Stem Cells.

    PubMed

    Musa, Marahaini; Mohd Ali, Khadijah; Kannan, Thirumulu Ponnuraj; Azlina, Ahmad; Omar, Nor Shamsuria; Chatterji, Anil; Mokhtar, Khairani Idah

    2015-01-01

    Perivitelline fluid (PVF) of the horseshoe crab embryo has been reported to possess an important role during embryogenesis by promoting cell proliferation. This study aims to evaluate the effect of PVF on the proliferation, chromosome aberration (CA) and mutagenicity of the dental pulp stem cells (DPSCs). This is an in vitro experimental study. PVF samples were collected from horseshoe crabs from beaches in Malaysia and the crude extract was prepared. DPSCs were treated with different concentrations of PVF crude extract in an 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay (cytotoxicity test). We choose two inhibitory concentrations (IC50 and IC25) and two PVF concentrations which produced more cell viability compared to a negative control (100%) for further tests. Quantitative analysis of the proliferation activity of PVF was studied using the AlamarBlue®assay for 10 days. Population doubling times (PDTs) of the treatment groups were calculated from this assay. Genotoxicity was evaluated based on the CA and Ames tests. Statistical analysis was carried out using independent t test to calculate significant differences in the PDT and mitotic indices in the CA test between the treatment and negative control groups. Significant differences in the data were P<0.05. A total of four PVF concentrations retrieved from the MTT assay were 26.887 mg/ml (IC50), 14.093 mg/ml (IC25), 0.278 mg/ml (102% cell viability) and 0.019 mg/ml (102.5% cell viability). According to the AlamarBlue®assay, these PVF groups produced comparable proliferation activities compared to the negative (untreated) control. PDTs between PVF groups and the negative control were insignificantly different (P>0.05). No significant aberrations in chromosomes were observed in the PVF groups and the Ames test on the PVF showed the absence of significant positive results. PVF from horseshoe crabs produced insignificant proliferative activity on treated DPSCs. The PVF was non

  19. Effects of Perivitelline Fluid Obtained from Horseshoe Crab on The Proliferation and Genotoxicity of Dental Pulp Stem Cells

    PubMed Central

    Musa, Marahaini; Mohd Ali, Khadijah; Kannan, Thirumulu Ponnuraj; Azlina, Ahmad; Omar, Nor Shamsuria; Chatterji, Anil; Mokhtar, Khairani Idah

    2015-01-01

    Objective Perivitelline fluid (PVF) of the horseshoe crab embryo has been reported to possess an important role during embryogenesis by promoting cell proliferation. This study aims to evaluate the effect of PVF on the proliferation, chromosome aberration (CA) and mutagenicity of the dental pulp stem cells (DPSCs). Materials and Methods This is an in vitro experimental study. PVF samples were collected from horseshoe crabs from beaches in Malaysia and the crude extract was prepared. DPSCs were treated with different concentrations of PVF crude extract in an 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay (cytotoxicity test). We choose two inhibitory concentrations (IC50 and IC25) and two PVF concentrations which produced more cell viability compared to a negative control (100%) for further tests. Quantitative analysis of the proliferation activity of PVF was studied using the AlamarBlue®assay for 10 days. Population doubling times (PDTs) of the treatment groups were calculated from this assay. Genotoxicity was evaluated based on the CA and Ames tests. Statistical analysis was carried out using independent t test to calculate significant differences in the PDT and mitotic indices in the CA test between the treatment and negative control groups. Significant differences in the data were P<0.05. Results A total of four PVF concentrations retrieved from the MTT assay were 26.887 mg/ml (IC50), 14.093 mg/ml (IC25), 0.278 mg/ml (102% cell viability) and 0.019 mg/ml (102.5% cell viability). According to the AlamarBlue®assay, these PVF groups produced comparable proliferation activities compared to the negative (untreated) control. PDTs between PVF groups and the negative control were insignificantly different (P>0.05). No significant aberrations in chromosomes were observed in the PVF groups and the Ames test on the PVF showed the absence of significant positive results. Conclusion PVF from horseshoe crabs produced insignificant proliferative

  20. Behaviour of dental pulp stem cells on different types of innovative mesoporous and nanoporous silicon scaffolds with different functionalizations of the surfaces.

    PubMed

    Marrelli, M; Falisi, G; Apicella, A; Apicella, D; Amantea, M; Cielo, A; Bonanome, L; Palmieri, F; Santacroce, L; Giannini, S; Di Fabrizio, E; Rastelli, C; Gargari, M; Cuda, G; Paduano, F; Tatullo, M

    2015-01-01

    Dental pulp stem cells (DPSCs) are stem cells found in the dental pulp. The ability of DPSCs to differentiate towards odontoblastic and osteoblastic phenotype was reported first in the literature, then in the following years, numerous studies on odontogenesis were carried out, starting from mesenchymal stem cells isolated from tissues of dental and oral origin. The aim of this research was to evaluate the behaviour of DPSCs grown on silicon nanoporous and mesoporous matrices and differentiated towards the osteogenic phenotype, but also to investigate the use of DPSCs in pilot studies focused on the biological compatibility of innovative dental biomaterials. Twenty-eight silicon samples were created with standardized procedures. These scaffolds were divided into samples made of silicon bulk, nanoporous silicon, mesoporous silicon, nanoporous silicon functionalized with (3-Aminopropyl) Trimethoxysilane (APTMS) and methanol (MeOH), nanoporous silicon functionalized with (3-Aminopropyl) Trimethoxysilane (APTMS)/toluene, mesoporous silicon functionalized with (3-Aminopropyl) Trimethoxysilane (APTMS) and methanol (MeOH) andmesoporous silicon functionalized with (3-Aminopropyl) Trimethoxysilane (APTMS)/toluene. DPSC proliferation on the tested silicon scaffolds was analyzed at 3 and 5 days. The assay showed that DPSCs proliferated better on mesoporous scaffolds functionalized with APTMS/toluene compared to a silicon one. These results show that the functionalization of silicon scaffold with APTMS/toluene supports the growth of DPSCs and could be used for future applications in tissue engineering.

  1. Adhesion and growth of dental pulp stem cells on enamel-like fluorapatite surfaces.

    PubMed

    Liu, J; Jin, T C; Chang, S; Czajka-Jakubowska, A; Clarkson, B H

    2011-03-01

    To study how apatite crystal alignment of an enamel-like substrate affects DPSC cellular adhesion and growth as a precursor to produce an in vitro enamel/dentin superstructure for future studies. The cells were subcultured in 10% FBS DMEM up to seven weeks on the two surfaces. Specimens were observed under SEM, counted, and analyzed using the human pathway-focused matrix and adhesion PCR array. After three days, the cell number on ordered FA surface was significantly higher than on the disordered surface. Of the 84 focused pathway genes, a total of 20 genes were either up or down regulated in the cells on ordered FA surface compared to the disordered surface. More interestingly, of the cell-matrix adhesion molecules, integrin alpha 7 and 8 (ITGA 7 and 8), integrin beta 3 and 4 (ITGB3 and 4), and the vitronectin receptor-integrin alpha V (ITGAV) and the key adhesion protein-fibronectin1 (FN1) were up-regulated. In SEM, both surfaces showed good biocompatibility and supported long term growth of DPSC cells but with functional cell-matrix interaction on the ordered FA surfaces. The enhanced cellular response of DPSC cell to the ordered FA crystal surface involves a set of delicately regulated matrix and adhesion molecules which could be manipulated by treating the cells with a dentin extract, to produce a dentin/enamel superstructure. Copyright © 2010 Wiley Periodicals, Inc.

  2. Activation of p38, p21, and NRF-2 Mediates Decreased Proliferation of Human Dental Pulp Stem Cells Cultured under 21% O2

    PubMed Central

    El Alami, Marya; Viña-Almunia, Jose; Gambini, Juan; Mas-Bargues, Cristina; Siow, Richard C.M.; Peñarrocha, Miguel; Mann, Giovanni E.; Borrás, Consuelo; Viña, Jose

    2014-01-01

    Summary High rates of stem cell proliferation are important in regenerative medicine and in stem cell banking for clinical use. Ambient oxygen tensions (21% O2) are normally used for in vitro culture, but physiological levels in vivo range between 3% and 6% O2. We compared proliferation of human dental pulp stem cells (hDPSCs) cultured under 21% versus 3% O2. The rate of hDPSC proliferation is significantly lower at 21% O2 compared to physiological oxygen levels due to enhanced oxidative stress. Under 21% O2, increased p38 phosphorylation led to activation of p21. Increased generation of reactive oxygen species and p21 led to activation of the NRF-2 signaling pathway. The upregulation of NRF-2 antioxidant defense genes under 21% O2 may interact with cell-cycle-related proteins involved in regulating cell proliferation. Activation of p38/p21/NRF-2 in hDPSCs cultured under ambient oxygen tension inhibits stem cell proliferation and upregulates NRF-2 antioxidant defenses. PMID:25358785

  3. Priming Dental Pulp Stem Cells With Fibroblast Growth Factor-2 Increases Angiogenesis of Implanted Tissue-Engineered Constructs Through Hepatocyte Growth Factor and Vascular Endothelial Growth Factor Secretion

    PubMed Central

    Gorin, Caroline; Rochefort, Gael Y.; Bascetin, Rumeyza; Ying, Hanru; Lesieur, Julie; Sadoine, Jérémy; Beckouche, Nathan; Berndt, Sarah; Novais, Anita; Lesage, Matthieu; Hosten, Benoit; Vercellino, Laetitia; Merlet, Pascal; Le-Denmat, Dominique; Marchiol, Carmen; Letourneur, Didier; Nicoletti, Antonino; Vital, Sibylle Opsahl; Poliard, Anne; Salmon, Benjamin; Germain, Stéphane

    2016-01-01

    Tissue engineering strategies based on implanting cellularized biomaterials are promising therapeutic approaches for the reconstruction of large tissue defects. A major hurdle for the reliable establishment of such therapeutic approaches is the lack of rapid blood perfusion of the tissue construct to provide oxygen and nutrients. Numerous sources of mesenchymal stem cells (MSCs) displaying angiogenic potential have been characterized in the past years, including the adult dental pulp. Establishment of efficient strategies for improving angiogenesis in tissue constructs is nevertheless still an important challenge. Hypoxia was proposed as a priming treatment owing to its capacity to enhance the angiogenic potential of stem cells through vascular endothelial growth factor (VEGF) release. The present study aimed to characterize additional key factors regulating the angiogenic capacity of such MSCs, namely, dental pulp stem cells derived from deciduous teeth (SHED). We identified fibroblast growth factor-2 (FGF-2) as a potent inducer of the release of VEGF and hepatocyte growth factor (HGF) by SHED. We found that FGF-2 limited hypoxia-induced downregulation of HGF release. Using three-dimensional culture models of angiogenesis, we demonstrated that VEGF and HGF were both responsible for the high angiogenic potential of SHED through direct targeting of endothelial cells. In addition, FGF-2 treatment increased the fraction of Stro-1+/CD146+ progenitor cells. We then applied in vitro FGF-2 priming to SHED before encapsulation in hydrogels and in vivo subcutaneous implantation. Our results showed that FGF-2 priming is more efficient than hypoxia at increasing SHED-induced vascularization compared with nonprimed controls. Altogether, these data demonstrate that FGF-2 priming enhances the angiogenic potential of SHED through the secretion of both HGF and VEGF. Significance The results from the present study show that fibroblast growth factor-2 (FGF-2) priming is more

  4. The Effect of 1α,25(OH)2D3 on Osteogenic Differentiation of Stem Cells from Dental Pulp of Exfoliated Deciduous Teeth

    PubMed Central

    Mojarad, Farzad; Amiri, Iraj; Rafatjou, Rezvan; Janeshin, Atousa; Farhadian, Maryam

    2016-01-01

    Statement of the Problem: Stem cells from human exfoliated deciduous teeth (SHEDs) are a population of highly proliferative cells, being capable of differentiating into osteogenic, odontogenic, adipocytes, and neural cells. Vitamin D3 metabolites such as 1α, 25-dihydroxyvitamin D3 are key factors in the regulation of bone metabolism. Purpose: The aim of this study was to investigate the effect of 1α, 25-dihydroxyvitamin D3 on osteogenic differentiation (alkaline phosphatase activity and alizarin red staining) of stem cells of exfoliated deciduous teeth. Materials and Method: Dental pulp was removed from freshly extracted primary teeth and immersed in a digestive solution. Then, the dental pulp cells were immersed in α-MEM (minimum essential medium) to which 10% fetal bovine serum was added. After the third passage, the cells were isolated from the culture plate and were used for osteogenic differentiation. As a control group, the cells were cultured in osteogenic cell culture medium. As the case group, the cells were cultured in osteogenic culture medium supplemented with 100 nM 1α,25 (OH)2D3. The alkaline phosphatase (ALP) activity and alizarin red staining were analyzed to evaluate the osteogenic differentiation at day 21. The results were analyzed by using t-test. Results: Compared with the control group, significant increase was observed in ALP activity of SHEDs after being treated with 1α,25(OH)2D3 (p= 0.002). Alizarin red staining demonstrated that the cells exposed to 1α,25(OH)2D3 induced higher mineralized nodules (p< 0.001). Conclusion: Osteoblast differentiation in SHEDs was stimulated by 1α,25(OH) 2D3. It can be concluded that 1α,25(OH)2D3 can improve osteoblastic differentiation. PMID:27942551

  5. In-vivo Generation of Dental Pulp-Like Tissue Using Human Pulpal Stem Cells, a Collagen Scaffold and Dentin Matrix Protein 1 Following Subcutaneous Transplantation in Mice

    PubMed Central

    Prescott, Rebecca S.; Alsanea, Rajaa; Fayad, Mohamed I.; Johnson, Bradford R.; Wenckus, Christopher S.; Hao, Jianjun; John, Asha S.; George, Anne

    2008-01-01

    The presence of a perforation is known to significantly compromise the outcome of endodontic treatment. One potential use of regenerative endodontic therapy may be the repair of root canal perforations. In addition to nutrients and systemic in-situ interactions, the three main components believed to be essential for tissue regeneration are: stem cells, scaffold, and growth factors. This study investigated the role of each component of the tissue engineering triad in the organization and differentiation of Dental Pulp Stem Cells (DPSCs) in a simulated furcal perforation site using a mouse model. Collagen served as the scaffold and dentin matrix protein 1 (DMP1) was the growth factor. Materials were placed in simulated perforation sites in dentin slices. MTA was the control repair material. At six weeks, the animals were sacrificed and the perforation sites were evaluated by light microscopy and histological staining. Organization of newly derived pulp tissue was seen in the group containing the triad of DPSCs, a collagen scaffold, and DMP1. The other four groups did not demonstrate any apparent tissue organization. Under the conditions of the present study, it may be concluded that the triad of DPSCs, a collagen scaffold, and DMP1 can induce an organized matrix formation similar to that of pulpal tissue, which may lead to hard tissue formation. PMID:18358888

  6. Human dental pulp stem cells can differentiate into Schwann cells and promote and guide neurite outgrowth in an aligned tissue-engineered collagen construct in vitro.

    PubMed

    Martens, Wendy; Sanen, Kathleen; Georgiou, Melanie; Struys, Tom; Bronckaers, Annelies; Ameloot, Marcel; Phillips, James; Lambrichts, Ivo

    2014-04-01

    In the present study, we evaluated the differentiation potential of human dental pulp stem cells (hDPSCs) toward Schwann cells, together with their functional capacity with regard to myelination and support of neurite outgrowth in vitro. Successful Schwann cell differentiation was confirmed at the morphological and ultrastructural level by transmission electron microscopy. Furthermore, compared to undifferentiated hDPSCs, immunocytochemistry and ELISA tests revealed increased glial marker expression and neurotrophic factor secretion of differentiated hDPSCs (d-hDPSCs), which promoted survival and neurite outgrowth in 2-dimensional dorsal root ganglia cultures. In addition, neurites were myelinated by d-hDPSCs in a 3-dimensional collagen type I hydrogel neural tissue construct. This engineered construct contained aligned columns of d-hDPSCs that supported and guided neurite outgrowth. Taken together, these findings provide the first evidence that hDPSCs are able to undergo Schwann cell differentiation and support neural outgrowth in vitro, proposing them to be good candidates for cell-based therapies as treatment for peripheral nerve injury.

  7. Improvement of osteogenesis in dental pulp pluripotent-like stem cells by oligopeptide-modified poly(β-amino ester)s.

    PubMed

    Núñez-Toldrà, Raquel; Dosta, Pere; Montori, Sheyla; Ramos, Víctor; Atari, Maher; Borrós, Salvador

    2017-04-15

    Controlling pluripotent stem cell differentiation via genetic manipulation is a promising technique in regenerative medicine. However, the lack of safe and efficient delivery vehicles limits this application. Recently, a new family of poly(β-amino ester)s (pBAEs) with oligopeptide-modified termini showing high transfection efficiency of both siRNA and DNA plasmid has been developed. In this study, oligopeptide-modified pBAEs were used to simultaneously deliver anti-OCT3/4 siRNA, anti-NANOG siRNA, and RUNX2 plasmid to cells from the dental pulp with pluripotent-like characteristics (DPPSC) in order to promote their osteogenic differentiation. Results indicate that transient inhibition of the pluripotency marker OCT3/4 and the overexpression of RUNX2 at day 7 of differentiation markedly increased and accelerated the expression of osteogenic markers. Furthermore, terminally-differentiated cells exhibited higher matrix mineralization and alkaline phosphatase activity. Finally, cell viability and genetic stability assays indicate that this co-delivery system has high chromosomal stability and minimal cytotoxicity. Therefore, we conclude that such co-delivery strategy is a safe and a quick option for the improvement of DPPSC osteogenic differentiation. Controlling pluripotent stem cell differentiation via genetic manipulation is a promising technique in regenerative medicine. However, the lack of safe and efficient delivery vehicles limits this application. In this study, we propose the use of a new family of oligopeptide-modified pBAEs developed in our group to control the differentiation of dental pulp pluripotential stem cells (DPPSC). In order to promote their osteogenic differentiation. The strategy proposed markedly increased and accelerated the expression of osteogenic markers, cell mineralization and alkaline phosphatase activity. Finally, cell viability and genetic stability assays indicated that this co-delivery system has high chromosomal stability and

  8. Schwann-like cells differentiated from human dental pulp stem cells combined with a pulsed electromagnetic field can improve peripheral nerve regeneration.

    PubMed

    Hei, Wei-Hong; Kim, Soochan; Park, Joo-Cheol; Seo, Young-Kwon; Kim, Soung-Min; Jahng, Jeong Won; Lee, Jong-Ho

    2016-03-15

    The purpose of this study was to investigate the effect of Schwann-like cells combined with pulsed electromagnetic field (PEMF) on peripheral nerve regeneration. Schwann-like cells were derived from human dental pulp stem cells (hDPSCs) and verified with CD104, S100, glial fibrillary acidic protein (GFAP), laminin, and P75(NTR) immunocytochemistry. Gene expression of P75(NTR) and S100 were analyzed. Male Sprague-Dawley rats (200-250g, 6-week-old) were divided into seven groups (n = 10 each): control, sham, PEMF, hDPSCs, hDPSCs + PEMF, Schwann-like cells, Schwann-like cells + PEMF. Cells were transplanted (1 × 10(6) /10µl/rat) at crush-injury site or combined with PEMF (50 Hz, 1 h/day, 1 mT). Nerve regeneration was evaluated with functional test, histomorphometry and retrograde labelled neurons. Schwann-like cells expressed CD104, S100, GFAP, laminin, and p75 neurotrophin receptor (P75(NTR) ). P75(NTR) and S100 mRNA expression was highest in Schwann-like cells + PEMF group, which also showed increased Difference and Gap scores. Axons and retrograde labeled neurons increased in all treatment groups. Schwann-like cells, hDPSCs with or without PEMF, and PEMF only improved peripheral nerve regeneration. Schwann-like cells + PEMF showed highest regeneration ability; PEMF has additive effect on hDPSCs, Schwann-like cell in vitro and nerve regeneration ability after transplantation in vivo. Bioelectromagnetics. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  9. Role of p16(INK4a) and BMI-1 in oxidative stress-induced premature senescence in human dental pulp stem cells.

    PubMed

    Mas-Bargues, Cristina; Viña-Almunia, José; Inglés, Marta; Sanz-Ros, Jorge; Gambini, Juan; Ibáñez-Cabellos, José Santiago; García-Giménez, José Luis; Viña, José; Borrás, Consuelo

    2017-08-01

    Human dental pulp stem cells (hDPSCs) are a source for cell therapy. Before implantation, an in vitro expansion step is necessary, with the inconvenience that hDPSCs undergo senescence following a certain number of passages, loosing their stemness properties. Long-term in vitro culture of hDPSCs at 21% (ambient oxygen tension) compared with 3-6% oxygen tension (physiological oxygen tension) caused an oxidative stress-related premature senescence, as evidenced by increased β-galactosidase activity and increased lysil oxidase expression, which is mediated by p16(INK4a) pathway. Furthermore, hDPSCs cultured at 21% oxygen tension underwent a downregulation of OCT4, SOX2, KLF4 and c-MYC factors, which was recued by BMI-1 silencing. Thus, p16(INK4a) and BMI-1 might play a role in the oxidative stress-associated premature senescence. We show that it is important for clinical applications to culture cells at physiological pO2 to retain their stemness characteristics and to delay senescence. Copyright © 2017. Published by Elsevier B.V.

  10. Nano-Structured Gelatin/Bioactive Glass Hybrid Scaffolds for the Enhancement of Odontogenic Differentiation of Human Dental Pulp Stem Cells

    PubMed Central

    Qu, Tiejun; Liu, Xiaohua

    2013-01-01

    Tooth decay is one of the most common chronic disorders throughout the world. Regenerating decayed dentin/pulp structure requires the design of novel scaffolding materials that mimic the architecture of natural dental extracellular matrix (ECM) and provide suitable environments for the attachment, proliferation, differentiation, and biomineralization of dental pulp stem cells (DPSCs). In this work, we developed an approach to prepare three-dimensional (3D) nano-fibrous gelatin/silica bioactive glass (NF-gelatin/SBG) hybrid scaffolds that mimic the nano-structured architecture and chemical composition of natural dental ECM. This approach involved the combination of a thermally induced phase separation, sol-gel, and porogen leaching process, and synthesized hybrid scaffolds possessing natural ECM-like architecture, high porosity, well-defined pore size and interconnectivity, and improved mechanical strength. An in vitro cell culture study showed that human DPSCs had a significantly higher proliferation rate on NF-gelatin/SBG scaffolds compared to NF-gelatin scaffolds under the same conditions. Furthermore, the integration of SBG into the hybrid scaffold significantly promoted the differentiation and biomineralization of the human DPSCs. The alkaline phosphatase (ALP) activity and expressions of marker genes for odontogenic differentiation (Col I, ALP, OCN, DSPP and DMP-1) were all significantly higher in the NF-gelatin/SBG than in the NF-gelatin group. Those results were further confirmed by hematoxylin and eosin (H&E) and von Kossa staining, as evidenced by greater ECM secretion and mineral deposition in the hybrid scaffold. In summary, the biomimetic NF-gelatin/SBG hybrid scaffolds provide an excellent environment for the growth and differentiation of human DPSCs and are promising candidates for dentin/pulp tissue regeneration. PMID:24098854

  11. Human and Swine Dental Pulp Stem Cells Form a Vascularlike Network after Angiogenic Differentiation in Comparison with Endothelial Cells: A Quantitative Analysis.

    PubMed

    Aksel, Hacer; Huang, George T-J

    2017-04-01

    The aim of this study was to quantify vascular network formation capacity after angiogenic induction of human and swine dental pulp stem cells (DPSCs) in comparison with endothelial cells. Primary human DPSCs or swine DPSCs were induced in endothelial growth medium for 7 days. The expression of the endothelial marker von Willebrand factor was determined by immunostaining. Induced DPSCs (iDPSCs) and noninduced DPSCs (niDPSCs) were seeded at different cell numbers onto Matrigel (BD Biosciences, San Jose, CA) for vascular network formation assays and analyzed after 4, 8, 12, and 18 hours in comparison with human microvascular endothelial cells (hMECs). Quantitative analysis of vascular tubule formation was performed using ImageJ software (National Institutes of Health, Bethesda, MD). The vascular network formation was also conducted by coculturing of niDPSCs and iDPSCs. Von Willebrand factor was detected by immunofluorescence in both niDPSCs and iDPSCs (human and swine). Time-lapse microscopic observation showed that the vascular network was formed by iDPSCs but not niDPSCs. After 4 hours, iDPSCs showed vascular network formation, whereas niDPSCs started to aggregate and formed clusters. Human iDPSCs displayed a similar capacity to form vascular networks in Matrigel compared with hMECs based on quantitative analysis. Swine iDPSCs had a higher capacity compared with human iDPSCs or hMECs (P < .05) in forming the network structures including segments, nodes, and mesh. A coculture experiment showed that human niDPSCs colocalized on the angiogenic tubules and vascular networks that were formed by human iDPSCs. Our findings indicate that iDPSCs in combination with their noninduced counterparts may be used as a future clinical strategy for enhancing angiogenesis during the process of pulp-dentin regeneration. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  12. Biological characteristic effects of human dental pulp stem cells on poly-ε-caprolactone-biphasic calcium phosphate fabricated scaffolds using modified melt stretching and multilayer deposition.

    PubMed

    Wongsupa, Natkrita; Nuntanaranont, Thongchai; Kamolmattayakul, Suttatip; Thuaksuban, Nuttawut

    2017-02-01

    Craniofacial bone defects such as alveolar cleft affect the esthetics and functions that need bone reconstruction. The advanced techniques of biomaterials combined with stem cells have been a challenging role for maxillofacial surgeons and scientists. PCL-coated biphasic calcium phosphate (PCL-BCP) scaffolds were created with the modified melt stretching and multilayer deposition (mMSMD) technique and merged with human dental pulp stem cells (hDPSCs) to fulfill the component of tissue engineering for bone substitution. In the present study, the objective was to test the biocompatibility and biofunctionalities that included cell proliferation, cell viability, alkaline phosphatase activity, osteocalcin, alizarin red staining for mineralization, and histological analysis. The results showed that mMSMD PCL-BCP scaffolds were suitable for hDPSCs viability since the cells attached and spread onto the scaffold. Furthermore, the constructs of induced hDPSCs and scaffolds performed ALP activity and produced osteocalcin and mineralized nodules. The results indicated that mMSMD PCL-BCP scaffolds with hDPSCs showed promise in bone regeneration for treatment of osseous defects.

  13. Human dental pulp stem cells respond to cues from the rat retina and differentiate to express the retinal neuronal marker rhodopsin.

    PubMed

    Bray, A F; Cevallos, R R; Gazarian, K; Lamas, M

    2014-11-07

    Human adult dental pulp stem cells (DPSCs) are self-renewing stem cells that originate from the neural crest during development and remain within the dental pulp niche through adulthood. Due to their multi-lineage differentiation potential and their relative ease of access they represent an exciting alternative for autologous stem cell-based therapies in neurodegenerative diseases. In animal models, DPSCs transplanted into the brain differentiate into functional neurons or astrocytes in response to local environmental cues that appear to influence the fate of the surviving cells. Here we tested the hypothesis that DPSCs might be able to respond to factors present in the retina enabling the regenerative potential of these cells. We evaluated the response of DPSCs to conditioned media from organotypic explants from control and chemically damaged rat retinas. To evaluate cell differentiation, we analyzed the expression of glial fibrillary acidic protein (GFAP), early neuronal and retinal markers (polysialic acid-neural cell adhesion molecule (PSA-NCAM); Pax6; Ascl1; NeuroD1) and the late photoreceptor marker rhodopsin, by immunofluorescence and reverse transcription polymerase chain reaction (RT-PCR). Exposure of DPSC cultures to conditioned media from control retinas induced a 39% reduction on the number of DPSCs that expressed GFAP; the expression of Pax6, Ascl1, PSA-NCAM or NeuroD1 was undetectable or did not change significantly. Expression of rhodopsin was not detectable in control or after exposure of the cultures with retinal conditioned media. By contrast, 44% of DPSCs exposed to conditioned media from damaged retinas were immunopositive to this protein. This response could not be reproduced when conditioned media from Müller-enriched primary cultures was used. Finally, quantitative RT-PCR was performed to compare the relative expression of glial cell-derived neurotrophic factor (GDNF), nerve growth factor (NGF), ciliary neurotrophic factor (CNTF) and brain

  14. An In vivo Model for Short-Term Evaluation of the Implantation Effects of Biomolecules or Stem Cells in the Dental Pulp

    PubMed Central

    Lacerda-Pinheiro, Sally; Marchadier, Arnaud; Donãs, Patricio; Septier, Dominique; Benhamou, Laurent; Kellermann, Odile; Goldberg, Michel; Poliard, Anne

    2008-01-01

    The continuously growing rodent incisor is a widely used model to investigate odontogenesis and mineralized tissue formation. This study focused on evaluating the mouse mandibular incisor as an experimental biological tool for analyzing in vivo the capacity of odontoblast-like progenitors or bioactive molecules to contribute to reparative dentinogenesis. We describe here a surgical procedure allowing direct access to the forming part of the incisor dental pulp Amelogenin peptide A+4 adsorbed on agarose beads, or dental pulp progenitor cells were implanted in the pulp following this procedure. After 10 days A+4 induced the formation of an osteodentin occluding almost the totality of the pulp compartment. Implantation of progenitor cells leads to formation of islets of osteodentin-like structures located centrally in the pulp. These pilot studies validate the incisor as an experimental model to test the capacity of progenitor cells or bioactive molecules to induce the formation of reparative dentin. PMID:19088885

  15. Three-dimensional simulated microgravity culture improves the proliferation and odontogenic differentiation of dental pulp stem cell in PLGA scaffolds implanted in mice.

    PubMed

    Li, Yanping; He, Lina; Pan, Shuang; Zhang, Lin; Zhang, Weiwei; Yi, Hong; Niu, Yumei

    2017-02-01

    Tooth regeneration through stem cell-based therapy is a promising treatment for tooth decay and loss. Human dental pulp stem cells (hDPSCs) have been widely identified as the stem cells with the most potential for tooth tissue regeneration. However, the culture of hDPSCs in vitro for tissue engineering is challenging, as cells may proliferate slowly or/and differentiate poorly in vivo. Dynamic three‑dimensional (3D) simulated microgravity (SMG) created using the rotary cell culture system is considered to an effective tool, which contributes to several cell functions. Thus, the present study aimed to investigate the effect of dynamic 3D SMG culture on the proliferation and odontogenic differentiation abilities of hDPSCs in poly (lactic‑co‑glycolic acid) (PLGA) scaffolds in nude mice. The hDPSCs on PLGA scaffolds were maintained separately in the 3D SMG culture system and static 3D cultures with osteogenic medium for 7 days in vitro. Subsequently, the cell‑PLGA complexes were implanted subcutaneously on the backs of nude mice for 4 weeks. The results of histological and immunohistochemical examinations of Ki‑67, type I collagen, dentin sialoprotein and DMP‑1 indicated that the proliferation and odontogenic differentiation abilities of the hDPSCs prepared in the 3D SMG culture system were higher, compared with those prepared in the static culture system. These findings suggested that dynamic 3D SMG culture likely contributes to tissue engineering by improving the proliferation and odontogenic differentiation abilities of hDPSCs in vivo.

  16. Adipose-derived mesenchymal stromal (stem) cells differentiate to osteoblast and chondroblast lineages upon incubation with conditioned media from dental pulp stem cell-derived osteoblasts and auricle cartilage chondrocytes.

    PubMed

    Carbone, A; Valente, M; Annacontini, L; Castellani, S; Di Gioia, S; Parisi, D; Rucci, M; Belgiovine, G; Colombo, C; Di Benedetto, A; Mori, G; Lo Muzio, L; Maiorella, A; Portincasa, A; Conese, M

    2016-01-01

    The potential of adipose-derived mesenchymal stromal (stem) cells (ADSCs) to differentiate into either osteoblasts or chondrocytes is controversial. In this study we investigated the multicapacity potential of ADSCs to differentiate towards adipocyte, osteoblast, and chondrocyte lineages when cells are seeded onto plastic in comparison with incubation with conditioned media (CM) obtained from differentiated cell types.ADSCs, obtained from liposuctions, were characterized for mesenchymal and hematopoietic markers by cytofluorimetry. Their differentiation capacity towards adipocytes, osteoblasts, and chondrocytes was investigated by histochemistry methods (Oil-Red-O staining, Safranin O and Alizarin Red staining, respectively). Dental pulp stem cells (DPSCs) and dedifferentiated auricle derived-chondrocytes were differentiated towards osteoblastic and chondrocytic lineages respectively, and the CM obtained from these cultures was used to induce differentiation of ADSCs. ADSCs were positive for mesenchymal markers (CD29, CD105, CD73, CD44), but not for hematopoietic lineage markers (CD14, CD34, CD45) and this behavior was conserved from the isolation up to the fifth passage. While ADSCs were readily differentiated in adipocytes, they were not towards chondrocytes and osteoblastic lineages, a behavior different from that of bone marrow-derived MSCs that differentiated into the three lineages at two weeks post-induction. Only ADSCs treated with CM from cultured chondrocytes and DPSCs, produced glycosaminoglycans and mineralized matrix. These results indicate that ADSCs need growth/morphogenic factor supplementation from the tissue environment to be appropriately differentiated to mesodermic lineages.

  17. Characterization of Progenitor Cells in Pulps of Murine Incisors

    PubMed Central

    Balic, A.; Mina, M.

    2010-01-01

    The continuous growth of rodent incisors requires the presence of stem cells capable of generating ameloblasts and odontoblasts. While epithelial stem cells giving rise to ameloblasts have been well-characterized, cells giving rise to the odontoblasts in incisors have not been fully characterized. The goal of this study was to gain insight into the potential population in dental pulps of unerupted and erupted incisors that give rise to odontoblasts. We show that pulps from unerupted incisors contain a significant mesenchymal-stem-cell (MSC)-like population (cells expressing CD90+/CD45-, CD117+/CD45-, Sca-1+/CD45-) and few CD45+ cells. Our in vitro studies showed that these cells displayed extensive osteo-dentinogenic potential, but were unable to differentiate into chondrocytes and adipocytes. Dental pulps from erupted incisors displayed increased percentages of CD45+ and decreased percentages of cells expressing markers of an MSC-like population. Despite these differences, pulps from erupted incisors also displayed extensive osteo-dentinogenic potential and inability to differentiate into chondrocytes and adipocytes. These results provide evidence that continuous generation of odontoblasts and dentin on the labial and lingual sides of unerupted and erupted incisors is supported by a progenitor population and not multipotent MSCs in the dental pulp. PMID:20739699

  18. Odontoblastic differentiation of dental pulp stem cells from healthy and carious teeth on an original PCL-based 3D scaffold.

    PubMed

    Louvrier, A; Euvrard, E; Nicod, L; Rolin, G; Gindraux, F; Pazart, L; Houdayer, C; Risold, P Y; Meyer, F; Meyer, C

    2017-01-21

    To isolate and characterize dental pulp stem cells (DPSCs) obtained from carious and healthy mature teeth extracted when conservative treatment was not possible or for orthodontic reasons; to evaluate the ability of DPSCs to colonize, proliferate and differentiate into functional odontoblast-like cells when cultured onto a polycaprolactone cone made by jet-spraying and prototyped into a design similar to a gutta-percha cone. DPSCs were obtained from nine carious and 12 healthy mature teeth. Then cells were characterized by flow cytometry and submitted to multidifferentiation to confirm their multipotency. These DPSCs were then cultured on a polycaprolactone cone in an odontoblastic differentiation medium. Cell proliferation, colonization of the biomaterial and functional differentiation of cells were histologically assessed. For the characterization, a t-Student test was used to compare the two groups. In all cell cultures, characterization highlighted a mesenchymal stem cell phenotype (CD105+, CD90+, CD73+, CD11b-, CD34-, CD45-, HLA-DR-). No significant differences were found between cultures obtained from carious and healthy mature teeth. DPSCs from both origins were able to differentiate into osteocytes, adipocytes and chondrocytes. Cell colonization was observed both on the surface and in the thickness of polycaprolactone cones as well as a mineralized pericellular matrix deposit composed of type I collagen, alkaline phosphatase, osteocalcin and dentin sialophosphoprotein. DPSCs were isolated from both carious and healthy mature teeth. They were able to colonize and proliferate within a polycaprolactone cone and could be differentiated into functional odontoblast-like cells. © 2017 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  19. Odontogenic Differentiation of Human Dental Pulp Stem Cells on Hydrogel Scaffolds Derived from Decellularized Bone Extracellular Matrix and Collagen Type I.

    PubMed

    Paduano, Francesco; Marrelli, Massimo; White, Lisa J; Shakesheff, Kevin M; Tatullo, Marco

    2016-01-01

    The aim of this study was to evaluate the level of odontogenic differentiation of dental pulp stem cells (DPSCs) on hydrogel scaffolds derived from bone extracellular matrix (bECM) in comparison to those seeded on collagen I (Col-I), one of the main components of dental pulp ECM. DPSCs isolated from human third molars were characterized for surface marker expression and odontogenic potential prior to seeding into bECM or Col-I hydrogel scaffolds. The cells were then seeded onto bECM and Col-I hydrogel scaffolds and cultured under basal conditions or with odontogenic and growth factor (GF) supplements. DPSCs cultivated on tissue culture polystyrene (TCPS) with and without supplements were used as controls. Gene expression of dentin sialophosphoprotein (DSPP), dentin matrix protein 1 (DMP-1) and matrix extracellular phosphoglycoprotein (MEPE) was evaluated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and mineral deposition was observed by Von Kossa staining. When DPSCs were cultured on bECM hydrogels, the mRNA expression levels of DSPP, DMP-1 and MEPE genes were significantly upregulated with respect to those cultured on Col-I scaffolds or TCPS in the absence of extra odontogenic inducers. In addition, more mineral deposition was observed on bECM hydrogel scaffolds as demonstrated by Von Kossa staining. Moreover, DSPP, DMP-1 and MEPE mRNA expressions of DPSCs cultured on bECM hydrogels were further upregulated by the addition of GFs or osteo/odontogenic medium compared to Col-I treated cells in the same culture conditions. These results demonstrate the potential of the bECM hydrogel scaffolds to stimulate odontogenic differentiation of DPSCs.

  20. Odontogenic Differentiation of Human Dental Pulp Stem Cells on Hydrogel Scaffolds Derived from Decellularized Bone Extracellular Matrix and Collagen Type I

    PubMed Central

    White, Lisa J.; Shakesheff, Kevin M.; Tatullo, Marco

    2016-01-01

    Objectives The aim of this study was to evaluate the level of odontogenic differentiation of dental pulp stem cells (DPSCs) on hydrogel scaffolds derived from bone extracellular matrix (bECM) in comparison to those seeded on collagen I (Col-I), one of the main components of dental pulp ECM. Methods DPSCs isolated from human third molars were characterized for surface marker expression and odontogenic potential prior to seeding into bECM or Col-I hydrogel scaffolds. The cells were then seeded onto bECM and Col-I hydrogel scaffolds and cultured under basal conditions or with odontogenic and growth factor (GF) supplements. DPSCs cultivated on tissue culture polystyrene (TCPS) with and without supplements were used as controls. Gene expression of dentin sialophosphoprotein (DSPP), dentin matrix protein 1 (DMP-1) and matrix extracellular phosphoglycoprotein (MEPE) was evaluated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and mineral deposition was observed by Von Kossa staining. Results When DPSCs were cultured on bECM hydrogels, the mRNA expression levels of DSPP, DMP-1 and MEPE genes were significantly upregulated with respect to those cultured on Col-I scaffolds or TCPS in the absence of extra odontogenic inducers. In addition, more mineral deposition was observed on bECM hydrogel scaffolds as demonstrated by Von Kossa staining. Moreover, DSPP, DMP-1 and MEPE mRNA expressions of DPSCs cultured on bECM hydrogels were further upregulated by the addition of GFs or osteo/odontogenic medium compared to Col-I treated cells in the same culture conditions. Significance These results demonstrate the potential of the bECM hydrogel scaffolds to stimulate odontogenic differentiation of DPSCs. PMID:26882351

  1. Phenotype heterogeneity in dental pulp stem niches.

    PubMed

    Perlea, Paula; Rusu, Mugurel Constantin; Didilescu, Andreea Cristiana; Pătroi, Elvira Floriana; Leonardi, Rosalia Maria; Imre, Marina; Răducanu, Anca Maria

    2016-01-01

    Dental stem niches (DSNs) reside in different dental tissues, being of particular importance in tissue engineering and dental regeneration procedures. The present paper aims to review the DSNs from the view of niche inhabitants, either extrinsic, such as cells of the myeloid lineage, or intrinsic, such as endothelial cells, perivascular cells and spindle-shaped stromal cells, e.g., telocytes. DSNs harbor different dental stem÷progenitor cell morphologies, in different stages of differentiation and with various potentialities, the angiogenic potential with respect to regenerative endodontic procedures being emphasized here. It seems therefore important to consider the DSNs as being heterogeneous, for a better understanding of an accurate identification of niche team players in regenerative medicine.

  2. Connexin43-containing gap junctions potentiate extracellular Ca²⁺-induced odontoblastic differentiation of human dental pulp stem cells via Erk1/2.

    PubMed

    Li, Shiting; He, Haitao; Zhang, Gang; Wang, Fei; Zhang, Ping; Tan, Yinghui

    2015-10-15

    Extracellular Ca(2+) can promote dentin sialophosphoprotein (DSPP) expression and odontoblastic differentiation of dental pulp stem cells (DPSCs). Gap junctions mediated by connexin43 (Cx43) allow diffusion of small molecules (such as Ca(2+)) among cells to regulate cell-to-cell communications. However, it is unclear whether Cx43 is required for the Ca(2+)-induced cell differentiation. Here, we found that the influx of extracellular Ca(2+) through L-type Ca(2+) channels increases intracellular free Ca(2+) levels to promote DSPP expression. Cx43 overexpression potentiated the extracellular Ca(2+)-induced DSPP expression via Erk1/2. Flow cytometry analyses showed that Cx43 increased the percentage of p-Erk1/2 positive cells in response to Ca(2+), indicating that Cx43 in DPSCs possibly acts as a traditional gap junction channel, which permits the sharing of signals among coupled cells to make more DPSCs respond to Ca(2+). Furthermore, inhibition of Cx43 function and gap junction communication decreased Ca(2+)-induced the expression of DSPP, suggesting that cell-to-cell contacts are required for Cx43 to promote the Ca(2+)-induced cell differentiation. Similarly, the study performed on DPSCs cultured at low-density and high-density revealed that intercellular contacts are required to potentiate Erk1/2 activity and DSPP expression. In total, this study indicates that Cx43 increases Ca(2+)-induced DSPP expression and odontoblastic differentiation of DPSCs via Erk1/2 through gap junction-mediated cell-to-cell contacts. Copyright © 2015 Elsevier Inc. All rights reserved.

  3. Expression of high mobility group box 1 in inflamed dental pulp and its chemotactic effect on dental pulp cells

    SciTech Connect

    Zhang, Xufang; Jiang, Hongwei; Gong, Qimei; Fan, Chen; Huang, Yihua; Ling, Junqi

    2014-08-08

    Highlights: • HMGB1 translocated from nucleus to cytoplasm during dental pulp inflammation. • HMGB1and its receptor RAGE were up-regulated in hDPCs under LPS stimulation. • HMGB1 enhanced hDPCs migration and induces cytoskeleton reorganization. • HMGB1 may play a critical role in dental pulp repair during inflamed state. - Abstract: High mobility group box 1 protein (HMGB1) is a chromatin protein which can be released extracellularly, eliciting a pro-inflammatory response and promoting tissue repair process. This study aimed to examine the expression and distribution of HMGB1 and its receptor RAGE in inflamed dental pulp tissues, and to assess its effects on proliferation, migration and cytoskeleton of cultured human dental pulp cells (DPCs). Our data demonstrated that cytoplasmic expression of HMGB1 was observed in inflamed pulp tissues, while HMGB1 expression was confined in the nuclei in healthy dental pulp. The mRNA expression of HMGB1 and RAGE were significantly increased in inflamed pulps. In in vitro cultured DPCs, expression of HMGB1 in both protein and mRNA level was up-regulated after treated with lipopolysaccharide (LPS). Exogenous HMGB1 enhanced DPCs migration in a dose-dependent manner and induced the reorganization of f-actin in DPCs. Our results suggests that HMGB1 are not only involved in the process of dental pulp inflammation, but also play an important role in the recruitment of dental pulp stem cells, promoting pulp repair and regeneration.

  4. Transplantation of stem cells obtained from murine dental pulp improves pancreatic damage, renal function, and painful diabetic neuropathy in diabetic type 1 mouse model.

    PubMed

    Guimarães, Elisalva Teixeira; Cruz, Gabriela da Silva; Almeida, Tiago Farias de; Souza, Bruno Solano de Freitas; Kaneto, Carla Martins; Vasconcelos, Juliana Fraga; Santos, Washington Luis Conrado dos; Santos, Ricardo Ribeiro-dos; Villarreal, Cristiane Flora; Soares, Milena Botelho Pereira

    2013-01-01

    Diabetes mellitus (DM) is one of the most common and serious chronic diseases in the world. Here, we investigated the effects of mouse dental pulp stem cell (mDPSC) transplantation in a streptozotocin (STZ)-induced diabetes type 1 model. C57BL/6 mice were treated intraperitoneally with 80 mg/kg of STZ and transplanted with 1 × 10(6) mDPSCs or injected with saline, by an endovenous route, after diabetes onset. Blood and urine glucose levels were reduced in hyperglycemic mice treated with mDPSCs when compared to saline-treated controls. This correlated with an increase in pancreatic islets and insulin production 30 days after mDPSC therapy. Moreover, urea and proteinuria levels normalized after mDPSC transplantation in diabetic mice, indicating an improvement of renal function. This was confirmed by a histopathological analysis of kidney sections. We observed the loss of the epithelial brush border and proximal tubule dilatation only in saline-treated diabetic mice, which is indicative of acute renal lesion. STZ-induced thermal hyperalgesia was also reduced after cell therapy. Three days after transplantation, mDPSC-treated diabetic mice exhibited nociceptive thresholds similar to that of nondiabetic mice, an effect maintained throughout the 90-day evaluation period. Immunofluorescence analyses of the pancreas revealed the presence of GFP(+) cells in, or surrounding, pancreatic islets. Our results demonstrate that mDPSCs may contribute to pancreatic β-cell renewal, prevent renal damage in diabetic animals, and produce a powerful and long-lasting antinociceptive effect on behavioral neuropathic pain. Our results suggest stem cell therapy as an option for the control of diabetes complications such as intractable diabetic neuropathic pain.

  5. Comparative analysis of the biological effects of the endodontic bioactive cements MTA Angelus, MTA Repair HP and NeoMTA Plus on human dental pulp stem cells.

    PubMed

    Tomás-Catalá, C J; Collado-González, M; García-Bernal, D; Oñate-Sánchez, R E; Forner, L; Llena, C; Lozano, A; Castelo-Baz, P; Moraleda, J M; Rodríguez-Lozano, F J

    2017-09-11

    To evaluate the biological effects in vitro of MTA Angelus (MTA-Ang; Angelus, Londrina, PR, Brazil), MTA Repair HP (MTA-HP; Angelus,) and NeoMTA Plus (NeoMTA-P; Avalon Biomed Inc, Bradenton, FL, USA) on human dental pulp stem cells (hDPSCs). Cell viability and cell migration assays were performed using eluates of each material. To evaluate cell morphology and cell attachment to the different materials, hDPSCs were directly seeded onto the material surfaces and analyzed by immunocytofluorescence and scanning electron microscopy, respectively. The chemical composition of the materials was determined by energy dispersive X-ray (EDX) and eluates were analyzed by inductively coupled plasma-mass spectrometry (ICP-MS). Statistical analysis was performed with analysis of variance and Bonferroni or Tukey post-test (α <0.05). Undiluted MTA-Ang, MTA-HP and NeoMTA-P displayed a significant increase in cell viability greater than that obtained using complete medium alone (control) (*p<0.05; **p<0.01; ***p<0.001). Moreover, a cell migration assay revealed cell migration rates after incubation with extracts of MTA-Ang, MTA-HP and NeoMTA-P, that were similar to levels obtained in the control group. In addition, stretched cytoskeletal F-actin fibres were detected in the cells treated with the three material extracts. SEM studies revealed a high degree of cell proliferation and attachment on all three materials. EDX analysis demonstrated similar weight percentages of C, O and Ca in all three materials, while other elements such as Al, Si and S were also found. MTA-Ang, MTA-HP, and NeoMTA-P were associated with biological effects on hDPSCs in terms of cell proliferation, morphology, migration and attachment. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  6. Early transplantation of human immature dental pulp stem cells from baby teeth to golden retriever muscular dystrophy (GRMD) dogs: Local or systemic?

    PubMed Central

    Kerkis, Irina; Ambrosio, Carlos E; Kerkis, Alexandre; Martins, Daniele S; Zucconi, Eder; Fonseca, Simone AS; Cabral, Rosa M; Maranduba, Carlos MC; Gaiad, Thais P; Morini, Adriana C; Vieira, Natassia M; Brolio, Marina P; Sant'Anna, Osvaldo A; Miglino, Maria A; Zatz, Mayana

    2008-01-01

    Background The golden retriever muscular dystrophy (GRMD) dogs represent the best available animal model for therapeutic trials aiming at the future treatment of human Duchenne muscular dystrophy (DMD). We have obtained a rare litter of six GRMD dogs (3 males and 3 females) born from an affected male and a carrier female which were submitted to a therapeutic trial with adult human stem cells to investigate their capacity to engraft into dogs muscles by local as compared to systemic injection without any immunosuppression. Methods Human Immature Dental Pulp Stem Cells (hIDPSC) were transplanted into 4 littermate dogs aged 28 to 40 days by either arterial or muscular injections. Two non-injected dogs were kept as controls. Clinical translation effects were analyzed since immune reactions by blood exams and physical scores capacity of each dog. Samples from biopsies were checked by immunohistochemistry (dystrophin markers) and FISH for human probes. Results and Discussion We analyzed the cells' ability in respect to migrate, engraftment, and myogenic potential, and the expression of human dystrophin in affected muscles. Additionally, the efficiency of single and consecutive early transplantation was compared. Chimeric muscle fibers were detected by immunofluorescence and fluorescent in situ hybridisation (FISH) using human antibodies and X and Y DNA probes. No signs of immune rejection were observed and these results suggested that hIDPSC cell transplantation may be done without immunosuppression. We showed that hIDPSC presented significant engraftment in GRMD dog muscles, although human dystrophin expression was modest and limited to several muscle fibers. Better clinical condition was also observed in the dog, which received monthly arterial injections and is still clinically stable at 25 months of age. Conclusion Our data suggested that systemic multiple deliveries seemed more effective than local injections. These findings open important avenues for further

  7. Mineral trioxide aggregate enhances the odonto/osteogenic capacity of stem cells from inflammatory dental pulps via NF-κB pathway.

    PubMed

    Wang, Y; Yan, M; Fan, Z; Ma, L; Yu, Y; Yu, J

    2014-10-01

    This study was designed to investigate the effects of mineral trioxide aggregate (MTA) on the osteo/odontogenic differentiation of inflammatory dental pulp stem cells (iDPSCs). inflammatory DPSCs were isolated from the inflammatory pulps of rat incisors and cocultured with MTA-conditioned medium. MTT assay and flow cytometry were performed to evaluate the proliferation of iDPSCs. Alkaline phosphatase (ALP) activity, alizarin red staining, real-time RT-PCR, and Western blot assay were used to investigate the differentiation capacity as well as the involvement of NF-κB pathway in iDPSCs. Mineral trioxide aggregate-treated iDPSCs demonstrated the higher ALP activity and formed more mineralized nodules than the untreated group. The odonto/osteoblastic markers (Alp, Runx2/RUNX2, Osx/OSX, Ocn/OCN, and Dspp/DSP, respectively) in MTA-treated iDPSCs were significantly upregulated as compared with untreated iDPSCs. Mechanistically, cytoplastic phos-P65 and nuclear P65 in MTA-treated iDPSCs were significantly increased in a time-dependent manner. Moreover, the inhibition of NF-κB pathway suppressed the MTA-induced odonto/osteoblastic differentiation of iDPSCs, as indicated by decreased ALP levels, weakened mineralization capacity and downregulated levels of odonto/osteoblastic genes (Osx, Ocn, and Dspp). Mineral trioxide aggregate enhances the odonto/osteogenic capacity of DPSCs from inflammatory sites via activating the NF-κB pathway. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  8. MEPE-Derived ASARM Peptide Inhibits Odontogenic Differentiation of Dental Pulp Stem Cells and Impairs Mineralization in Tooth Models of X-Linked Hypophosphatemia

    PubMed Central

    Khaddam, Mayssam; Naji, Jiar; Coyac, Benjamin R.; Baroukh, Brigitte; Letourneur, Franck; Lesieur, Julie; Decup, Franck; Le Denmat, Dominique; Nicoletti, Antonino; Poliard, Anne; Rowe, Peter S.; Huet, Eric; Vital, Sibylle Opsahl; Linglart, Agnès; McKee, Marc D.; Chaussain, Catherine

    2013-01-01

    Mutations in PHEX (phosphate-regulating gene with homologies to endopeptidases on the X-chromosome) cause X-linked familial hypophosphatemic rickets (XLH), a disorder having severe bone and tooth dentin mineralization defects. The absence of functional PHEX leads to abnormal accumulation of ASARM (acidic serine- and aspartate-rich motif) peptide − a substrate for PHEX and a strong inhibitor of mineralization − derived from MEPE (matrix extracellular phosphoglycoprotein) and other matrix proteins. MEPE-derived ASARM peptide accumulates in tooth dentin of XLH patients where it may impair dentinogenesis. Here, we investigated the effects of ASARM peptides in vitro and in vivo on odontoblast differentiation and matrix mineralization. Dental pulp stem cells from human exfoliated deciduous teeth (SHEDs) were seeded into a 3D collagen scaffold, and induced towards odontogenic differentiation. Cultures were treated with synthetic ASARM peptides (phosphorylated and nonphosphorylated) derived from the human MEPE sequence. Phosphorylated ASARM peptide inhibited SHED differentiation in vitro, with no mineralized nodule formation, decreased odontoblast marker expression, and upregulated MEPE expression. Phosphorylated ASARM peptide implanted in a rat molar pulp injury model impaired reparative dentin formation and mineralization, with increased MEPE immunohistochemical staining. In conclusion, using complementary models to study tooth dentin defects observed in XLH, we demonstrate that the MEPE-derived ASARM peptide inhibits both odontogenic differentiation and matrix mineralization, while increasing MEPE expression. These results contribute to a partial mechanistic explanation of XLH pathogenesis: direct inhibition of mineralization by ASARM peptide leads to the mineralization defects in XLH teeth. This process appears to be positively reinforced by the increased MEPE expression induced by ASARM. The MEPE-ASARM system can therefore be considered as a potential therapeutic

  9. MEPE-derived ASARM peptide inhibits odontogenic differentiation of dental pulp stem cells and impairs mineralization in tooth models of X-linked hypophosphatemia.

    PubMed

    Salmon, Benjamin; Bardet, Claire; Khaddam, Mayssam; Naji, Jiar; Coyac, Benjamin R; Baroukh, Brigitte; Letourneur, Franck; Lesieur, Julie; Decup, Franck; Le Denmat, Dominique; Nicoletti, Antonino; Poliard, Anne; Rowe, Peter S; Huet, Eric; Vital, Sibylle Opsahl; Linglart, Agnès; McKee, Marc D; Chaussain, Catherine

    2013-01-01

    Mutations in PHEX (phosphate-regulating gene with homologies to endopeptidases on the X-chromosome) cause X-linked familial hypophosphatemic rickets (XLH), a disorder having severe bone and tooth dentin mineralization defects. The absence of functional PHEX leads to abnormal accumulation of ASARM (acidic serine- and aspartate-rich motif) peptide - a substrate for PHEX and a strong inhibitor of mineralization - derived from MEPE (matrix extracellular phosphoglycoprotein) and other matrix proteins. MEPE-derived ASARM peptide accumulates in tooth dentin of XLH patients where it may impair dentinogenesis. Here, we investigated the effects of ASARM peptides in vitro and in vivo on odontoblast differentiation and matrix mineralization. Dental pulp stem cells from human exfoliated deciduous teeth (SHEDs) were seeded into a 3D collagen scaffold, and induced towards odontogenic differentiation. Cultures were treated with synthetic ASARM peptides (phosphorylated and nonphosphorylated) derived from the human MEPE sequence. Phosphorylated ASARM peptide inhibited SHED differentiation in vitro, with no mineralized nodule formation, decreased odontoblast marker expression, and upregulated MEPE expression. Phosphorylated ASARM peptide implanted in a rat molar pulp injury model impaired reparative dentin formation and mineralization, with increased MEPE immunohistochemical staining. In conclusion, using complementary models to study tooth dentin defects observed in XLH, we demonstrate that the MEPE-derived ASARM peptide inhibits both odontogenic differentiation and matrix mineralization, while increasing MEPE expression. These results contribute to a partial mechanistic explanation of XLH pathogenesis: direct inhibition of mineralization by ASARM peptide leads to the mineralization defects in XLH teeth. This process appears to be positively reinforced by the increased MEPE expression induced by ASARM. The MEPE-ASARM system can therefore be considered as a potential therapeutic

  10. Use Electrospinning to Introduce Graphene into Poly(4-Vinylpridine) (P4VP) Polymer Fibers and Their Biocompatibility with Dental Pulp Stem Cells (DPSCs)

    NASA Astrophysics Data System (ADS)

    Zhang, Linxi; Chang, Chung-Chueh; Rafailovich, Miriam

    Graphene-polymer composite materials have been popularized in tissue engineering due to the outstanding thermal, electrical and mechanical properties of graphene. Most of the current studies, however, focus on 2-D structured films which hardly represent the real conditions of scaffolds in vivo environment and dispersion of graphene in polymer matrix has always been challenging since the graphene tends to aggregate. In our study, we have successfully introduced graphene nanoplatelets (GNPs) into poly(4-vinphylpridine) (P4VP) matrix and fabricated nano- and micro-scale size fibers by using electrospinning technique. SEM and TEM reveal uniform defect-free fiber structures and good dispersion of graphene; DSC and AFM indicate the enhancement of physical properties. The biocompatibility of the electrospun 3-D scaffolds with dental pulp stem cells (DPSCs) has been examined. Our results show that the cells can accelerate proliferation to respond to the existence of GNPs. SEM with EDAX reveals a deposition of mineralized calcium matrix on the fibers after 35-day incubation, which has possibly been caused by cell differentiation induced by fibrous scaffolds. Electrospinning Nano- and Mirco-scale size Poly(4-vinphylpridine) Fibers Loaded with Graphene Nano Platelets (GNPs).

  11. Differentiation of dental pulp stem cells into chondrocytes upon culture on porous chitosan-xanthan scaffolds in the presence of kartogenin.

    PubMed

    Westin, Cecília B; Trinca, Rafael B; Zuliani, Carolina; Coimbra, Ibsen B; Moraes, Ângela M

    2017-11-01

    Adhesion, proliferation and differentiation of dental pulp stem cells (DPSCs) into chondrocytes were investigated in this work with the purpose of broadening the array of cell alternatives to the therapy of cartilage lesions related to tissue engineering approaches. A porous chitosan-xanthan (C-X) matrix was used as scaffold and kartogenin was used as a selective chondrogenic differentiation promoter. The scaffold was characterized regarding aspect and surface morphology, absorption and stability in culture medium, thickness, porosity, thermogravimetric behavior, X-ray diffraction, mechanical properties and indirect cytocompatibility. The behavior of DPSCs cultured on the scaffold was evaluated by scanning electron microscopy and cell differentiation, by histological analysis. A sufficiently stable amorphous scaffold with mean thickness of 0.89±0.01mm and high culture medium absorption capacity (13.20±1.88g/g) was obtained, and kartogenin concentrations as low as 100nmol/L were sufficient to efficiently induce DPSCs differentiation into chondrocytes, showing that the strategy proposed may be a straightforward and effective approach for tissue engineering aiming at the therapy of cartilage lesions. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Factors secreted from dental pulp stem cells show multifaceted benefits for treating acute lung injury in mice.

    PubMed

    Wakayama, Hirotaka; Hashimoto, Naozumi; Matsushita, Yoshihiro; Matsubara, Kohki; Yamamoto, Noriyuki; Hasegawa, Yoshinori; Ueda, Minoru; Yamamoto, Akihito

    2015-08-01

    Acute respiratory distress syndrome (ARDS) is a severe inflammatory disorder characterized by acute respiratory failure, resulting from severe, destructive lung inflammation and irreversible lung fibrosis. We evaluated the use of stem cells derived from human exfoliated deciduous teeth (SHEDs) or SHED-derived serum-free conditioned medium (SHED-CM) as treatments for bleomycin (BLM)-induced mice acute lung injury (ALI), exhibiting several pathogenic features associated with the human disease ARDS. Mice with BLM-induced ALI with or without SHED or SHED-CM treatment were examined for weight loss and survival. The lung tissue was characterized by histological and real-time quantitative polymerase chain reaction analysis. The effects of SHED-CM on macrophage differentiation in vitro were also assessed. A single intravenous administration of either SHEDs or SHED-CM attenuated the lung injury and weight loss in BLM-treated mice and improved their survival rate. Similar recovery levels were seen in the SHEDs and SHED-CM treatment groups, suggesting that SHED improves ALI by paracrine mechanisms. SHED-CM contained multiple therapeutic factors involved in lung-regenerative mechanisms. Importantly, SHED-CM attenuated the BLM-induced pro-inflammatory response and generated an anti-inflammatory/tissue-regenerating environment, accompanied by the induction of anti-inflammatory M2-like lung macrophages. Furthermore, SHED-CM promoted the in vitro differentiation of bone marrow-derived macrophages into M2-like cells, which expressed high levels of Arginase1, CD206 and Ym-1. Our results suggest that SHED-secreted factors provide multifaceted therapeutic effects, including a strong M2-inducing activity, for treating BLM-induced ALI. This work may open new avenues for research on stem cell-based ARDS therapies. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  13. Stem cells.

    PubMed

    Behr, Björn; Ko, Sae Hee; Wong, Victor W; Gurtner, Geoffrey C; Longaker, Michael T

    2010-10-01

    Stem cells are self-renewing cells capable of differentiating into multiple cell lines and are classified according to their origin and their ability to differentiate. Enormous potential exists in use of stem cells for regenerative medicine. To produce effective stem cell-based treatments for a range of diseases, an improved understanding of stem cell biology and better control over stem cell fate are necessary. In addition, the barriers to clinical translation, such as potential oncologic properties of stem cells, need to be addressed. With renewed government support and continued refinement of current stem cell methodologies, the future of stem cell research is exciting and promises to provide novel reconstructive options for patients and surgeons limited by traditional paradigms.

  14. Chitosan-Intercalated Montmorillonite/Poly(vinyl alcohol) Nanofibers as a Platform to Guide Neuronlike Differentiation of Human Dental Pulp Stem Cells.

    PubMed

    Ghasemi Hamidabadi, Hatef; Rezvani, Zahra; Nazm Bojnordi, Maryam; Shirinzadeh, Haji; Seifalian, Alexander M; Joghataei, Mohammad Taghi; Razaghpour, Mojgan; Alibakhshi, Abbas; Yazdanpanah, Abolfazl; Salimi, Maryam; Mozafari, Masoud; Urbanska, Aleksandra M; Reis, Rui L; Kundu, Subhas C; Gholipourmalekabadi, Mazaher

    2017-03-27

    In this study, we present a novel chitosan-intercalated montmorillonite/poly(vinyl alcohol) (OMMT/PVA) nanofibrous mesh as a microenvironment for guiding differentiation of human dental pulp stem cells (hDPSCs) toward neuronlike cells. The OMMT was prepared through ion exchange reaction between the montmorillonite (MMT) and chitosan. The PVA solutions containing various concentrations of OMMT were electrospun to form 3D OMMT-PVA nanofibrous meshes. The biomechanical and biological characteristics of the nanofibrous meshes were evaluated by ATR-FTIR, XRD, SEM, MTT, and LDH specific activity, contact angle, and DAPI staining. They were carried out for mechanical properties, overall viability, and toxicity of the cells. The hDPSCs were seeded on the prepared scaffolds and induced with neuronal specific differentiation media at two differentiation stages (2 days at preinduction stage and 6 days at induction stage). The neural differentiation of the cells cultured on the meshes was evaluated by determining the expression of Oct-4, Nestin, NF-M, NF-H, MAP2, and βIII-tubulin in the cells after preinduction, at induction stages by real-time PCR (RT-PCR) and immunostaining. All the synthesized nanofibers exhibited a homogeneous morphology with a favorable mechanical behavior. The population of the cells differentiated into neuronlike cells in all the experimental groups was significantly higher than that in control group. The expression level of the neuronal specific markers in the cells cultured on 5% OMMT/PVA meshes was significantly higher than the other groups. This study demonstrates the feasibility of the OMMT/PVA artificial nerve graft cultured with hDPSCs for regeneration of damaged neural tissues. These fabricated matrices may have a potential in neural tissue engineering applications.

  15. Effects of total flavonoids from Drynaria fortunei on the proliferation and osteogenic differentiation of rat dental pulp stem cells.

    PubMed

    Huang, Xiao-Fei; Yuan, Su-Jian; Yang, Cheng

    2012-09-01

    Dental pulp stem cells (DPSCs) have the potential to form bone, nerve and fat, and are a candidate for use in regenerative medicine. Previous studies indicated that total flavonoids from Drynaria fortunei show a stimulative effect on the proliferation and osteogenic differentiation of osteoblastic MC3T3-E1 cells in vitro. This study aimed to investigate the effect of total flavonoids from Drynaria fortunei on the proliferation and osteogenic differentiation of rat DPSCs, and to further clarify the mechanisms involved. DPSCs were isolated by enzymatic digestion and identified using the CD44, CD29 and CD34 markers by immunohistochemistry, and exposed to 0.01, 0.05 and 0.1 g/l total flavonoids from Drynaria fortunei media. Total flavonoids from Drynaria fortunei promoted the proliferation of DPSCs in a dose-dependent manner and this effect may depend on the shortening of the G0/G1 phase and promotion of the S phase. Compared with the control group, the levels of alkaline phosphatase (ALP) and the expression of osteogenic genes increased with the concentrations of total flavonoids from Drynaria fortunei, and the volume and number of calcified nodules in the Drynaria groups was bigger compared to the control group. These results suggest that total flavonoid from Drynaria fortunei directly stimulates DPSC proliferation and osteogenic differentiation, and may serve as a new promising candidate drug for dental tissue engineering and bone regeneration.

  16. Effects of a discoloration-resistant calcium aluminosilicate cement on the viability and proliferation of undifferentiated human dental pulp stem cells.

    PubMed

    Niu, Li-na; Watson, Devon; Thames, Kyle; Primus, Carolyn M; Bergeron, Brian E; Jiao, Kai; Bortoluzzi, Eduardo A; Cutler, Christopher W; Chen, Ji-hua; Pashley, David H; Tay, Franklin R

    2015-11-30

    Discoloration-resistant calcium aluminosilicate cement has been formulated to overcome the timely problem of tooth discoloration reported in the clinical application of bismuth oxide-containing hydraulic cements. The present study examined the effects of this experimental cement (Quick-Set2) on the viability and proliferation of human dental pulp stem cells (hDPSCs) by comparing the cellular responses with commercially available calcium silicate cement (white mineral trioxide aggregate; WMTA) after different aging periods. Cell viability and proliferation were examined using assays that examined plasma membrane integrity, leakage of cytosolic enzyme, caspase-3 activity for early apoptosis, oxidative stress, mitochondrial metabolic activity and intracellular DNA content. Results of the six assays indicated that both Quick-Set2 and WMTA were initially cytotoxic to hDPSCs after setting for 24 h, with Quick-Set2 being comparatively less cytotoxic than WMTA at this stage. After two aging cycles, the cytotoxicity profiles of the two hydraulic cements were not significantly different and were much less cytotoxic than the positive control (zinc oxide-eugenol cement). Based on these results, it is envisaged that any potential beneficial effect of the discoloration-resistant calcium aluminosilicate cement on osteogenesis by differentiated hDPSCs is more likely to be revealed after outward diffusion and removal of its cytotoxic components.

  17. Effects of a discoloration-resistant calcium aluminosilicate cement on the viability and proliferation of undifferentiated human dental pulp stem cells

    PubMed Central

    Niu, Li-na; Watson, Devon; Thames, Kyle; Primus, Carolyn M.; Bergeron, Brian E.; Jiao, Kai; Bortoluzzi, Eduardo A.; Cutler, Christopher W.; Chen, Ji-hua; Pashley, David H.; Tay, Franklin R.

    2015-01-01

    Discoloration-resistant calcium aluminosilicate cement has been formulated to overcome the timely problem of tooth discoloration reported in the clinical application of bismuth oxide-containing hydraulic cements. The present study examined the effects of this experimental cement (Quick-Set2) on the viability and proliferation of human dental pulp stem cells (hDPSCs) by comparing the cellular responses with commercially available calcium silicate cement (white mineral trioxide aggregate; WMTA) after different aging periods. Cell viability and proliferation were examined using assays that examined plasma membrane integrity, leakage of cytosolic enzyme, caspase-3 activity for early apoptosis, oxidative stress, mitochondrial metabolic activity and intracellular DNA content. Results of the six assays indicated that both Quick-Set2 and WMTA were initially cytotoxic to hDPSCs after setting for 24 h, with Quick-Set2 being comparatively less cytotoxic than WMTA at this stage. After two aging cycles, the cytotoxicity profiles of the two hydraulic cements were not significantly different and were much less cytotoxic than the positive control (zinc oxide–eugenol cement). Based on these results, it is envisaged that any potential beneficial effect of the discoloration-resistant calcium aluminosilicate cement on osteogenesis by differentiated hDPSCs is more likely to be revealed after outward diffusion and removal of its cytotoxic components. PMID:26617338

  18. Types of Stem Cells

    MedlinePlus

    ... Stem Cell Glossary Search Toggle Nav Types of Stem Cells Stem cells are the foundation from which all ... Learn About Stem Cells > Types of Stem Cells Stem cells Stem cells are the foundation for every organ ...

  19. Effects of Recombinant Overexpression of Bcl2 on the Proliferation, Apoptosis, and Osteogenic/Odontogenic Differentiation Potential of Dental Pulp Stem Cells.

    PubMed

    Heng, Boon Chin; Ye, Xin; Liu, Yuan; Dissanayaka, Waruna Lakmal; Cheung, Gary Shun Pan; Zhang, Chengfei

    2016-04-01

    The therapeutic usefulness of dental pulp stem cells (DPSCs) is severely limited by low survivability upon transplantation in situ because of the presence of various proapoptotic factors within damaged/diseased tissues (ie, hypoxia and inflammation). One strategy to enhance the survivability of grafted DPSCs could be recombinant overexpression of antiapoptotic genes, such as the B-cell lymphoma 2 gene (Bcl2). DPSCs were transfected with the Bcl2 and/or GFP gene. Cell density and mitotic activity of transfected DPSCs within in vitro culture were evaluated with the water soluble tetrazolium salt-8 (WST-8) and bromodeoxyuridine assay, respectively, whereas apoptosis was evaluated through the detection of cytoplasmic histone-associated DNA fragments. The osteogenic/odontogenic differentiation potential of these cells was evaluated with quantitative real-time polymerase chain reaction, alkaline phosphatase, and alizarin red staining. Bcl2-transfected DPSCs exhibited consistently higher cell densities than the GFP-transfected control within in vitro culture, and this was not because of the higher mitotic rate but was instead attributed to enhanced cell survivability because of the inhibition of apoptosis by Bcl2. Recombinant overexpression of Bcl2 inhibited the osteogenic/odontogenic potential of DPSCs, as indicated by lower levels of alkaline phosphatase activity and mineralized calcium deposition, together with the down-regulated expression of several key osteogenic/odontogenic gene markers including collagen I, osteocalcin, dentin matrix protein-1, bone sialoprotein, and alkaline phosphatase. The results place a "caveat" or limitation on the use of recombinant Bcl2 overexpression as a therapeutic strategy for improving the survivability of grafted DPSCs in that the osteogenic/odontogenic potential of these cells may be compromised despite enhanced survival within the host. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights

  20. Human dental pulp stem cells transplantation combined with treadmill training in rats after traumatic spinal cord injury

    PubMed Central

    Nicola, F.C.; Rodrigues, L.P.; Crestani, T.; Quintiliano, K.; Sanches, E.F.; Willborn, S.; Aristimunha, D.; Boisserand, L.; Pranke, P.; Netto, C.A.

    2016-01-01

    Spinal cord injury (SCI) is a disabling condition resulting in deficits of sensory and motor functions, and has no effective treatment. Considering that protocols with stem cell transplantation and treadmill training have shown promising results, the present study evaluated the effectiveness of stem cells from human exfoliated deciduous teeth (SHEDs) transplantation combined with treadmill training in rats with experimental spinal cord injury. Fifty-four Wistar rats were spinalized using NYU impactor. The rats were randomly distributed into 5 groups: Sham (laminectomy with no SCI, n=10); SCI (laminectomy followed by SCI, n=12); SHEDs (SCI treated with SHEDs, n=11); TT (SCI treated with treadmill training, n=11); SHEDs+TT (SCI treated with SHEDs and treadmill training; n=10). Treatment with SHEDs alone or in combination with treadmill training promoted functional recovery, reaching scores of 15 and 14, respectively, in the BBB scale, being different from the SCI group, which reached 11. SHEDs treatment was able to reduce the cystic cavity area and glial scar, increase neurofilament. Treadmill training alone had no functional effectiveness or tissue effects. In a second experiment, the SHEDs transplantation reduced the TNF-α levels in the cord tissue measured 6 h after the injury. Contrary to our hypothesis, treadmill training either alone or in combination, caused no functional improvement. However, SHEDs showed to be neuroprotective, by the reduction of TNF-α levels, the cystic cavity and the glial scar associated with the improvement of motor function after SCI. These results provide evidence that grafted SHEDs might be an effective therapy to spinal cord lesions, with possible anti-inflammatory action. PMID:27509306

  1. Human dental pulp stem cells transplantation combined with treadmill training in rats after traumatic spinal cord injury.

    PubMed

    Nicola, F C; Rodrigues, L P; Crestani, T; Quintiliano, K; Sanches, E F; Willborn, S; Aristimunha, D; Boisserand, L; Pranke, P; Netto, C A

    2016-08-08

    Spinal cord injury (SCI) is a disabling condition resulting in deficits of sensory and motor functions, and has no effective treatment. Considering that protocols with stem cell transplantation and treadmill training have shown promising results, the present study evaluated the effectiveness of stem cells from human exfoliated deciduous teeth (SHEDs) transplantation combined with treadmill training in rats with experimental spinal cord injury. Fifty-four Wistar rats were spinalized using NYU impactor. The rats were randomly distributed into 5 groups: Sham (laminectomy with no SCI, n=10); SCI (laminectomy followed by SCI, n=12); SHEDs (SCI treated with SHEDs, n=11); TT (SCI treated with treadmill training, n=11); SHEDs+TT (SCI treated with SHEDs and treadmill training; n=10). Treatment with SHEDs alone or in combination with treadmill training promoted functional recovery, reaching scores of 15 and 14, respectively, in the BBB scale, being different from the SCI group, which reached 11. SHEDs treatment was able to reduce the cystic cavity area and glial scar, increase neurofilament. Treadmill training alone had no functional effectiveness or tissue effects. In a second experiment, the SHEDs transplantation reduced the TNF-α levels in the cord tissue measured 6 h after the injury. Contrary to our hypothesis, treadmill training either alone or in combination, caused no functional improvement. However, SHEDs showed to be neuroprotective, by the reduction of TNF-α levels, the cystic cavity and the glial scar associated with the improvement of motor function after SCI. These results provide evidence that grafted SHEDs might be an effective therapy to spinal cord lesions, with possible anti-inflammatory action.

  2. Differential Neuronal Plasticity of Dental Pulp Stem Cells From Exfoliated Deciduous and Permanent Teeth Towards Dopaminergic Neurons.

    PubMed

    Majumdar, Debanjana; Kanafi, Mohammad; Bhonde, Ramesh; Gupta, Pawan; Datta, Indrani

    2016-09-01

    Based on early occurrence in chronological age, stem-cells from human exfoliated deciduous teeth (SHED) has been reported to possess better differentiation-potential toward certain cell-lineage in comparison to stem-cells from adult teeth (DPSCs). Whether this same property between them extends for the yield of functional central nervous system neurons is still not evaluated. Hence, we aim to assess the neuronal plasticity of SHED in comparison to DPSCs toward dopaminergic-neurons and further, if the difference is reflected in a differential expression of sonic-hedgehog (SHH)-receptors and basal-expressions of tyrosine-hydroxylase [TH; through cAMP levels]. Human SHED and DPSCs were exposed to midbrain-cues [SHH, fibroblast growth-factor8, and basic fibroblast growth-factor], and their molecular, immunophenotypical, and functional characterization was performed at different time-points of induction. Though SHED and DPSCs spontaneously expressed early-neuronal and neural-crest marker in their naïve state, only SHED expressed a high basal-expression of TH. The upregulation of dopaminergic transcription-factors Nurr1, Engrailed1, and Pitx3 was more pronounced in DPSCs. The yield of TH-expressing cells decreased from 49.8% to 32.16% in SHED while it increased from 8.09% to 77.47% in DPSCs. Dopamine release and intracellular-Ca(2+) influx upon stimulation (KCl and ATP) was higher in induced DPSCs. Significantly lower-expression of SHH-receptors was noted in naïve SHED than DPSCs, which may explain the differential neuronal plasticity. In addition, unlike DPSCs, SHED showed a down-regulation of cyclic adenosine-monophosphate (cAMP) upon exposure to SHH; possibly another contributor to the lesser differentiation-potential. Our data clearly demonstrates for the first time that DPSCs possess superior neuronal plasticity toward dopaminergic-neurons than SHED; influenced by higher SHH-receptor and lower basal TH expression. J. Cell. Physiol. 231: 2048-2063, 2016. © 2016

  3. Pulp cell tracking by radionuclide imaging for dental tissue engineering.

    PubMed

    Souron, Jean-Baptiste; Petiet, Anne; Decup, Franck; Tran, Xuan Vinh; Lesieur, Julie; Poliard, Anne; Le Guludec, Dominique; Letourneur, Didier; Chaussain, Catherine; Rouzet, Francois; Opsahl Vital, Sibylle

    2014-03-01

    Pulp engineering with dental mesenchymal stem cells is a promising therapy for injured teeth. An important point is to determine the fate of implanted cells in the pulp over time and particularly during the early phase following implantation. Indeed, the potential engraftment of the implanted cells in other organs has to be assessed, in particular, to evaluate the risk of inducing ectopic mineralization. In this study, our aim was to follow by nuclear imaging the radiolabeled pulp cells after implantation in the rat emptied pulp chamber. For that purpose, indium-111-oxine (¹¹¹In-oxine)-labeled rat pulp cells were added to polymerizing type I collagen hydrogel to obtain a pulp equivalent. This scaffold was implanted in the emptied pulp chamber space in the upper first rat molar. Labeled cells were then tracked during 3 weeks by helical single-photon emission computed tomography (SPECT)/computed tomography performed on a dual modality dedicated small animal camera. Negative controls were performed using lysed radiolabeled cells obtained in a hypotonic solution. In vitro data indicated that ¹¹¹In-oxine labeling did not affect cell viability and proliferation. In vivo experiments allowed a noninvasive longitudinal follow-up of implanted living cells for at least 3 weeks and indicated that SPECT signal intensity was related to implanted cell integrity. Notably, there was no detectable systemic release of implanted cells from the tooth. In addition, histological analysis of the samples showed mitotically active fibroblastic cells as well as neoangiogenesis and nervous fibers in pulp equivalents seeded with entire cells, whereas pulp equivalents prepared from lysed cells were devoid of cell colonization. In conclusion, our study demonstrates that efficient labeling of pulp cells can be achieved and, for the first time, that these cells can be followed up after implantation in the tooth by nuclear imaging. Furthermore, it appears that grafted cells retained the label

  4. Pulp Cell Tracking by Radionuclide Imaging for Dental Tissue Engineering

    PubMed Central

    Souron, Jean-Baptiste; Petiet, Anne; Decup, Franck; Tran, Xuan Vinh; Lesieur, Julie; Poliard, Anne; Le Guludec, Dominique; Letourneur, Didier; Chaussain, Catherine; Rouzet, Francois

    2014-01-01

    Pulp engineering with dental mesenchymal stem cells is a promising therapy for injured teeth. An important point is to determine the fate of implanted cells in the pulp over time and particularly during the early phase following implantation. Indeed, the potential engraftment of the implanted cells in other organs has to be assessed, in particular, to evaluate the risk of inducing ectopic mineralization. In this study, our aim was to follow by nuclear imaging the radiolabeled pulp cells after implantation in the rat emptied pulp chamber. For that purpose, indium-111-oxine (111In-oxine)-labeled rat pulp cells were added to polymerizing type I collagen hydrogel to obtain a pulp equivalent. This scaffold was implanted in the emptied pulp chamber space in the upper first rat molar. Labeled cells were then tracked during 3 weeks by helical single-photon emission computed tomography (SPECT)/computed tomography performed on a dual modality dedicated small animal camera. Negative controls were performed using lysed radiolabeled cells obtained in a hypotonic solution. In vitro data indicated that 111In-oxine labeling did not affect cell viability and proliferation. In vivo experiments allowed a noninvasive longitudinal follow-up of implanted living cells for at least 3 weeks and indicated that SPECT signal intensity was related to implanted cell integrity. Notably, there was no detectable systemic release of implanted cells from the tooth. In addition, histological analysis of the samples showed mitotically active fibroblastic cells as well as neoangiogenesis and nervous fibers in pulp equivalents seeded with entire cells, whereas pulp equivalents prepared from lysed cells were devoid of cell colonization. In conclusion, our study demonstrates that efficient labeling of pulp cells can be achieved and, for the first time, that these cells can be followed up after implantation in the tooth by nuclear imaging. Furthermore, it appears that grafted cells retained the label and

  5. Influence of Partial O₂ Pressure on the Adhesion, Proliferation, and Osteogenic Differentiation of Human Dental Pulp Stem Cells on β-Tricalcium Phosphate Scaffold.

    PubMed

    Viña-Almunia, Jose; Mas-Bargues, Cristina; Borras, Consuelo; Gambini, Juan; El Alami, Marya; Sanz-Ros, Jorge; Peñarrocha, Miguel; Vina, Jose

    2017-09-22

    To analyze, in vitro, the influence of O₂ pressure on the adhesion, proliferation, and osteogenic differentiation of human dental pulp stem cells (DPSC) on β-tricalcium phosphate (β-TCP) scaffold. DPSC, positive for the molecular markers CD133, Oct4, Nestin, Stro-1, and CD34, and negative for CD45, were isolated from extracted third molars. Experiments were started by seeding 200,000 cells on β-TCP cultured under 3% or 21% O₂ pressure. No osteogenic medium was used. Eight different cultures were performed at each time point under each O₂ pressure condition. Cell adhesion, proliferation, and differentiation over the biomaterial were evaluated at 7, 13, 18, and 23 days of culture. Cell adhesion was determined by light microscopy, proliferation by DNA quantification, and osteogenic differentiation by alkaline phosphatase (ALP) activity analysis. DPSC adhered to β-TCP with both O₂ conditions. Cell proliferation was found from day 7 of culture. Higher values were recorded at 3% O₂ in each time point. Statistically significant differences were recorded at 23 days of culture (P = .033). ALP activity was not detectable at 7 days. There was, however, an increase in ALP activity over time in both groups. At 13, 18, and 23 days of culture, higher ALP activity was recorded under 3% O₂ pressure. Statistical differences were found at day 23 (P = .014). DPSC display capacity of adhering to β-TCP under 3% or 21% O₂ pressure conditions. Cell proliferation on β-TCP phosphate is significantly higher at 3% than at 21% O₂ pressure, the most frequently used O₂ tension. β-TCP can itself promote osteogenic differentiation of DPSC and is enhanced under 3% O₂ compared with 21%.

  6. Cytotoxic Effects of Various Mineral Trioxide Aggregate Formulations, Calcium-Enriched Mixture and a New Cement on Human Pulp Stem Cells

    PubMed Central

    Jaberiansari, Zahra; Naderi, Seddigheh; Tabatabaei, Fahimeh Sadat

    2014-01-01

    Introduction: This in vitro study compared the cytotoxic effects of three commercially available MTA formulations naming ProRoot MTA (PMTA), Angelus MTA (AMTA), and Root MTA (RMTA), with calcium-enriched mixture (CEM) cement and a new nanohybrid MTA (NMTA) on human dental pulp stem cells (DPSC). Methods and Materials: Four disc-shaped specimens of each material were prepared. After completion of setting, 2 different (neat and 1/2) elutes of the test materials were made. Then in each cavity of a 96-well plate, 3000 cells were seeded and incubated in a humidified incubator with 5% CO2 and 95% air at 37°C for 24 h. After this period, the culture medium of each well was replaced with 200 μL of test material elutes. Plain culture medium was used as the negative control and distilled water as the positive control group. Cell viability was assessed using 2, 5-diphenyl-SH-tetrazelium bromide colorimetric assay, aka Mosmann’s tetrazolium toxicity (MTT) assay, at three time intervals (24, 48, and 72 h after mixing). Data were analyzed using the ANOVA and Tukey’s post hoc test (P=0.05). Results: After 24 h, the viability of cells in neat concentration had no significant differences (P>0.05) except for the NMTA. However, CEM and AMTA, at 1/2 concentration exerted significant proliferative effects on cells. At 48 and 72-h intervals, significant proliferation of DPSCs was seen in all samples, except for the NMTA which exerted toxic effects on cells. Conclusion: All of the three commercial MTAs and CEM cement showed comparative biocompatibility. However, NMTA had cytotoxic effects on DPSCs at all the time intervals. PMID:25386208

  7. An in vivo swine study for xeno-grafts of calcium sulfate-based bone grafts with human dental pulp stem cells (hDPSCs).

    PubMed

    Kuo, Tzong-fu; Lee, Sheng-Yang; Wu, Hong-Da; Poma, Malosi; Wu, Yu-Wei; Yang, Jen-Chang

    2015-05-01

    The purpose of this in vivo study was to evaluate the effect of human dental pulp stem cells (hDPSCs) on various resorbable calcium sulfate/calcium phosphate bone grafts in bone regeneration. Granular particles of calcium sulfate dehydrate (CSD), α-calcium sulfate hemihydrate/amorphous calcium phosphate (α-CSH/ACP), and CSD/β-tricalcium phosphates (β-TCP) were prepared for in vitro dissolution and implantation test. The chemical compositions of specimen residues after dissolution test were characterized by XRD. The ratios of new bone formation for implanted grafts/hDPSCs were evaluated using mandible bony defect model of Lanyu pig. All the graft systems exhibited a similar two-stage dissolution behavior and phase transformation of poor crystalline HAp. Eight weeks post-operation, the addition of hDPSCs to various graft systems showed statistically significant increasing in the ratio of new bone formation (p<0.05). Null hypothesis of hDPSCs showing no scaffold dependence in bone regeneration was rejected. The results suggest that the addition of hDPSCs to calcium sulfate based xenografts could enhance the bone regeneration in the bony defect. Copyright © 2015 Elsevier B.V. All rights reserved.

  8. Human plasma and human platelet-rich plasma as a substitute for fetal calf serum during long-term cultivation of mesenchymal dental pulp stem cells.

    PubMed

    Suchánková Kleplová, Tereza; Soukup, Tomáš; Řeháček, Vít; Suchánek, Jakub

    2014-01-01

    Our aims were to isolate and cultivate mesenchymal dental pulp stem cells (DPSC) in various media enriched with human blood components, and subsequently to investigate their basic biological properties. DPSC were cultivated in five different media based on α MEM containing different concentrations of human plasma (HP), platelet-rich plasma (PRP), or fetal calf serum (FCS). The DPSC biological properties were examined periodically. We cultivated DPSC in the various cultivation media over 15 population doublings except for the medium supplemented with 10% HP. Our results showed that DPSC cultivated in medium supplemented with 10% PRP showed the shortest average population doubling time (DT) (28.6 ± 4.6 hours), in contrast to DPSC cultivated in 10% HP which indicated the longest DT (156.2 ± 17.8 hours); hence this part of the experiment had been cancelled in the 6th passage. DPSC cultivated in media with 2% FCS+ITS (DT 47.3 ± 10.4 hours), 2% PRP (DT 40.1 ± 5.7 hours) and 2% HP (DT 49.0 ± 15.2 hours) showed almost the same proliferative activity. DPSC's viability in the 9th passage was over 90% except for the DPSC cultivated in the 10% HP media. We proved that human blood components are suitable substitution for FCS in cultivation media for long-term DPSC cultivation.

  9. Purmorphamine increased adhesion, proliferation and expression of osteoblast phenotype markers of human dental pulp stem cells cultured on beta-tricalcium phosphate.

    PubMed

    Rezia Rad, Maryam; Khojaste, Moein; Hasan Shahriari, Mehrnoosh; Asgary, Saeed; Khojasteh, Arash

    2016-08-01

    Growth factors play a significant role in cell proliferation and differentiation during different stages of the bone repair. However, several limitations have been brought researchers attention to an osteoinductive small molecule including Purmorphamine. In this study, we aimed to evaluate the effect of Purmorphamine on adhesion, proliferation and differentiation of human dental pulp stem cells (hDPSCs) seaded on beta-tricalcium phosphate (β-TCP) granules. hDPSCs were established from extracted wisdom teeth of healthy volenteers. Cells at passage 3 were seeded on β-TCP in the presence or absence of Purmorphamine. Cell adhesion and proliferation were assessed using scanning electeron microscopy (SEM) and DNA counting assay, respectively, after 1, 3 and 5days. Then, hDPSCs seeded on β-TCP were subjected to osteogenic medium with or without Purmorphamine. After 7 and 14days osteogenic diffrentiation capability of hDPSCs were determined using real-time RT-PCR and alkaline phosphatase (ALP) activity assay. The significant increase in amount of DNA was observed at day 3 and 5 in the presence of Purmorphamine. SEM imaging also was confirmed the DNA counting assay; in all given time points, hDPSC attachment and growth was significantly higher in the presence of Purmorphamine. ALP activity was increased by Purmorphamine at both 7 and 14days of induction. Purmorphamine showed to effect on osteopontin expression at earlier stage of osteogenic differentiation, whereas for osteocalcin expression, this effect was more evident at later stage of differentiation. Purmorphamine had a promotive effect on adhesion, proliferation and osteogenic differentiation of hDPSCs cultured on β-TCP. The outcome of the current study would help in development of in vitro culture conditions for better osteogenic differentiation of hDPSCs prior to transplantation. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  10. Conditioned medium from the stem cells of human dental pulp improves cognitive function in a mouse model of Alzheimer's disease.

    PubMed

    Mita, Tsuneyuki; Furukawa-Hibi, Yoko; Takeuchi, Hideyuki; Hattori, Hisashi; Yamada, Kiyofumi; Hibi, Hideharu; Ueda, Minoru; Yamamoto, Akihito

    2015-10-15

    Alzheimer's disease (AD) is a progressive, neurodegenerative disease characterized by a decline in cognitive abilities and the appearance of β-amyloid plaques in the brain. Although the pathogenic mechanisms associated with AD are not fully understood, activated microglia releasing various neurotoxic factors, including pro-inflammatory cytokines and oxidative stress mediators, appear to play major roles. Here, we investigated the therapeutic benefits of a serum-free conditioned medium (CM) derived from the stem cells of human exfoliated deciduous teeth (SHEDs) in a mouse model of AD. The intranasal administration of SHEDs in these mice resulted in substantially improved cognitive function. SHED-CM contained factors involved in multiple neuroregenerative mechanisms, such as neuroprotection, axonal elongation, neurotransmission, the suppression of inflammation, and microglial regulation. Notably, SHED-CM attenuated the pro-inflammatory responses induced by β-amyloid plaques, and generated an anti-inflammatory/tissue-regenerating environment, which was accompanied by the induction of anti-inflammatory M2-like microglia. Our data suggest that SHED-CM may provide significant therapeutic benefits for AD.

  11. Biodentine induces human dental pulp stem cell differentiation through mitogen-activated protein kinase and calcium-/calmodulin-dependent protein kinase II pathways.

    PubMed

    Luo, Zhirong; Kohli, Meetu R; Yu, Qing; Kim, Syngcuk; Qu, Tiejun; He, Wen-xi

    2014-07-01

    Biodentine (Septodont, Saint-Maur-des-Fossès, France), a new tricalcium silicate cement formulation, has been introduced as a bioactive dentine substitute to be used in direct contact with pulp tissue. The aim of this study was to investigate the response of human dental pulp stem cells (hDPSCs) to the material and whether mitogen-activated protein kinase (MAPK), nuclear factor-kappa B (NF-κB), and calcium-/calmodulin-dependent protein kinase II (CaMKII) signal pathways played a regulatory role in Biodentine-induced odontoblast differentiation. hDPCs obtained from impacted third molars were incubated with Biodentine. Odontoblastic differentiation was evaluated by alkaline phosphatase activity, alizarin red staining, and quantitative real-time reverse-transcriptase polymerase chain reaction for the analysis of messenger RNA expression of the following differentiation gene markers: osteocalcin (OCN), dentin sialophosprotein (DSPP), dentin matrix protein 1 (DMP1), and bone sialoprotein (BSP). Cell cultures in the presence of Biodentine were exposed to specific inhibitors of MAPK (U0126, SB203580, and SP600125), NF-κB (pyrrolidine dithiocarbamate), and CaMKII (KN-93) pathways to evaluate the regulatory effect on the expression of these markers and mineralization assay. Biodentine significantly increased alkaline phosphatase activity and mineralized nodule formation and the expression of OCN, DSPP, DMP1, and BSP. The MAPK inhibitor for extracellular signal-regulated kinase 1/2 (U0126) and Jun N-terminal kinase (SP600125) significantly decreased the Biodentine-induced mineralized differentiation of hDPSCs and OCN, DSPP, DMP1, and BSP messenger RNA expression, whereas p38 MAPK inhibitors (SB203580) had no effect. The CaMKII inhibitor KN-93 significantly attenuated and the NF-κB inhibitor pyrrolidine dithiocarbamate further enhanced the up-regulation of Biodentine-induced gene expression and mineralization. Biodentine is a bioactive and biocompatible material capable

  12. Dental stem cells and their sources.

    PubMed

    Sedgley, Christine M; Botero, Tatiana M

    2012-07-01

    The search for more accessible mesenchymal stem cells than those found in bone marrow has propelled interest in dental tissues. Human dental stem/progenitor cells (collectively termed dental stem cells [DSCs]) that have been isolated and characterized include dental pulp stem cells, stem cells from exfoliated deciduous teeth, stem cells from apical papilla, periodontal ligament stem cells, and dental follicle progenitor cells. Common characteristics of these cell populations are the capacity for self-renewal and the ability to differentiate into multiple lineages. In vitro and animal studies have shown that DSCs can differentiate into osseous, odontogenic, adipose, endothelial, and neural-like tissues.

  13. Odontogenic differentiation and biomineralization potential of dental pulp stem cells inside Mg-based bioceramic scaffolds under low-level laser treatment.

    PubMed

    Theocharidou, Anna; Bakopoulou, Athina; Kontonasaki, Eleana; Papachristou, Eleni; Hadjichristou, Christina; Bousnaki, Maria; Theodorou, George; Papadopoulou, Lambrini; Kantiranis, Nikolaos; Paraskevopoulos, Konstantinos; Koidis, Petros

    2017-01-01

    This study aimed to investigate the potential of low-level laser irradiation (LLLI) to promote odontogenic differentiation and biomineralization by dental pulp stem cells (DPSCs) seeded inside bioceramic scaffolds. Mg-based, Zn-doped bioceramic scaffolds, synthesized by the sol-gel technique, were spotted with DPSCs and exposed to LLLI at 660 nm with maximum output power of 140 mw at fluencies (a) 2 and 4 J/cm(2) to evaluate cell viability/proliferation by the MTT assay and (b) 4 J/cm(2) to evaluate cell differentiation, using real-time PCR (expression of odontogenic markers) and a p-nitrophenylphosphate (pNPP)-based assay for alkaline phosphatase (ALP) activity measurement. Scanning electron microscopy (SEM) and X-ray diffraction (XRD) analysis were used for structural/chemical characterization of the regenerated tissues. Exposure of the DPSCs/scaffold complexes to the proposed LLLI scheme was associated with statistically significant increase of odontogenesis-related markers (bone morphogenetic protein 2 (BMP-2): 22.4-fold, dentin sialophosphoprotein (DSPP): 28.4-fold, Osterix: 18.5-fold, and Runt-related transcription factor 2 (Runx2): 3.4-fold). ALP activity was significantly increased at 3 and 7 days inside the irradiated compared to that in the non-irradiated SC/DPSC complexes, but gradually decreased until 14 days. Newly formed Ca-P tissue was formed on the SC/DPSC complexes after 28 days of culture that attained the characteristics of bioapatite. Overall, LLLI treatment proved to be beneficial for odontogenic differentiation and biomineralization of DPSCs inside the bioceramic scaffolds, making this therapeutic modality promising for targeted dentin engineering.

  14. Influence of the mechanical environment on the engineering of mineralised tissues using human dental pulp stem cells and silk fibroin scaffolds.

    PubMed

    Woloszyk, Anna; Holsten Dircksen, Sabrina; Bostanci, Nagihan; Müller, Ralph; Hofmann, Sandra; Mitsiadis, Thimios A

    2014-01-01

    Teeth constitute a promising source of stem cells that can be used for tissue engineering and regenerative medicine purposes. Bone loss in the craniofacial complex due to pathological conditions and severe injuries could be treated with new materials combined with human dental pulp stem cells (hDPSCs) that have the same embryonic origin as craniofacial bones. Optimising combinations of scaffolds, cells, growth factors and culture conditions still remains a great challenge. In the present study, we evaluate the mineralisation potential of hDPSCs seeded on porous silk fibroin scaffolds in a mechanically dynamic environment provided by spinner flask bioreactors. Cell-seeded scaffolds were cultured in either standard or osteogenic media in both static and dynamic conditions for 47 days. Histological analysis and micro-computed tomography of the samples showed low levels of mineralisation when samples were cultured in static conditions (0.16±0.1 BV/TV%), while their culture in a dynamic environment with osteogenic medium and weekly µCT scans (4.9±1.6 BV/TV%) significantly increased the formation of homogeneously mineralised structures, which was also confirmed by the elevated calcium levels (4.5±1.0 vs. 8.8±1.7 mg/mL). Molecular analysis of the samples showed that the expression of tooth correlated genes such as Dentin Sialophosphoprotein and Nestin were downregulated by a factor of 6.7 and 7.4, respectively, in hDPSCs when cultured in presence of osteogenic medium. This finding indicates that hDPSCs are able to adopt a non-dental identity by changing the culture conditions only. Also an increased expression of Osteocalcin (1.4x) and Collagen type I (1.7x) was found after culture under mechanically dynamic conditions in control medium. In conclusion, the combination of hDPSCs and silk scaffolds cultured under mechanical loading in spinner flask bioreactors could offer a novel and promising approach for bone tissue engineering where appropriate and rapid bone

  15. Influence of the Mechanical Environment on the Engineering of Mineralised Tissues Using Human Dental Pulp Stem Cells and Silk Fibroin Scaffolds

    PubMed Central

    Woloszyk, Anna; Holsten Dircksen, Sabrina; Bostanci, Nagihan; Müller, Ralph; Hofmann, Sandra; Mitsiadis, Thimios A.

    2014-01-01

    Teeth constitute a promising source of stem cells that can be used for tissue engineering and regenerative medicine purposes. Bone loss in the craniofacial complex due to pathological conditions and severe injuries could be treated with new materials combined with human dental pulp stem cells (hDPSCs) that have the same embryonic origin as craniofacial bones. Optimising combinations of scaffolds, cells, growth factors and culture conditions still remains a great challenge. In the present study, we evaluate the mineralisation potential of hDPSCs seeded on porous silk fibroin scaffolds in a mechanically dynamic environment provided by spinner flask bioreactors. Cell-seeded scaffolds were cultured in either standard or osteogenic media in both static and dynamic conditions for 47 days. Histological analysis and micro-computed tomography of the samples showed low levels of mineralisation when samples were cultured in static conditions (0.16±0.1 BV/TV%), while their culture in a dynamic environment with osteogenic medium and weekly µCT scans (4.9±1.6 BV/TV%) significantly increased the formation of homogeneously mineralised structures, which was also confirmed by the elevated calcium levels (4.5±1.0 vs. 8.8±1.7 mg/mL). Molecular analysis of the samples showed that the expression of tooth correlated genes such as Dentin Sialophosphoprotein and Nestin were downregulated by a factor of 6.7 and 7.4, respectively, in hDPSCs when cultured in presence of osteogenic medium. This finding indicates that hDPSCs are able to adopt a non-dental identity by changing the culture conditions only. Also an increased expression of Osteocalcin (1.4x) and Collagen type I (1.7x) was found after culture under mechanically dynamic conditions in control medium. In conclusion, the combination of hDPSCs and silk scaffolds cultured under mechanical loading in spinner flask bioreactors could offer a novel and promising approach for bone tissue engineering where appropriate and rapid bone

  16. Isolation and characterization of lymphoid enhancer factor-1-positive deciduous dental pulp stem-like cells after transfection with a piggyBac vector containing LEF1 promoter-driven selection markers.

    PubMed

    Murakami, Tomoya; Saitoh, Issei; Sato, Masahiro; Inada, Emi; Soda, Miki; Oda, Masataka; Domon, Hisanori; Iwase, Yoko; Sawami, Tadashi; Matsueda, Kazunari; Terao, Yutaka; Ohshima, Hayato; Noguchi, Hirofumi; Hayasaki, Haruaki

    2017-09-01

    Lymphoid enhancer-binding factor-1 (LEF1) is a 48-kD nuclear protein that is expressed in pre-B and T cells. LEF1 is also an important member of the Wnt/β-catenin signaling pathway that plays important roles in the self-renewal and differentiation of embryonic stem cells. We speculated that LEF1 might function in the stem cells from human exfoliated deciduous teeth (SHED). In this study, we attempted to isolate such LEF1-positive cells from human deciduous dental pulp cells (HDDPCs) by genetic engineering technology, using the human LEF1 promoter. A piggyBac transposon plasmid (pTA-LEN) was introduced into HDDPCs, using the Neon(®) transfection system. After G418 selection, the emerging colonies were assessed for EGFP-derived fluorescence by fluorescence microscopy. Reverse transcription polymerase chain reaction (RT-PCR) analysis was performed using RNA isolated from these colonies to examine stem cell-specific transcript expression. Osteoblastic or neuronal differentiation was induced by cultivating the LEF1-positive cells with differentiation-inducing medium. RT-PCR analysis confirmed the expression of several stem cell markers, including OCT3/4, SOX2, REX1, and NANOG, in LEF1-positive HDDPCs, which could be differentiated into osteoblasts and neuronal cells. The isolated LEF1-positive HDDPCs exhibited the properties of stem cells, suggesting that LEF1 might serve as a marker for SHED. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  17. Functionalization of Polycaprolactone Scaffolds with Hyaluronic Acid and β-TCP Facilitates Migration and Osteogenic Differentiation of Human Dental Pulp Stem Cells In Vitro

    PubMed Central

    Kraft, David Christian Evar; Lysdahl, Helle; Foldager, Casper Bindzus; Chen, Muwan; Kristiansen, Asger Albæk; Rölfing, Jan Hendrik Duedal; Bünger, Cody Eric

    2015-01-01

    In this study, we sought to assess the osteogenic potential of human dental pulp stem cells (DPSCs) on three different polycaprolactone (PCL) scaffolds. The backbone structure of the scaffolds was manufactured by fused deposition modeling (PCL scaffold). The composition and morphology was functionalized in two of the scaffolds. The first underwent thermal induced phase separation of PCL infused into the pores of the PCL scaffold. This procedure resulted in a highly variable micro- and nanostructured porous (NSP), interconnected, and isotropic tubular morphology (NSP-PCL scaffold). The second scaffold type was functionalized by dip-coating the PCL scaffold with a mixture of hyaluronic acid and β-TCP (HT-PCL scaffold). The scaffolds were cylindrical and measured 5 mm in height and 10 mm in diameter. They were seeded with 1×106 human DPSCs, a cell type known to express bone-related markers, differentiate into osteoblasts-like cells, and to produce a mineralized bone-like extracellular matrix. DPSCs were phenotypically characterized by flow cytometry for CD90+, CD73+, CD105+, and CD14−. DNA, ALP, and Ca2+ assays and real-time quantitative polymerase chain reaction for genes involved in osteogenic differentiation were analyzed on day 1, 7, 14, and 21. Cell viability and distribution were assessed on day 1, 7, 14, and 21 by fluorescent-, scanning electron-, and confocal microscopy. The results revealed that the DPSCs expressed relevant gene expression consistent with osteogenic differentiation. The NSP-PCL and HT-PCL scaffolds promoted osteogenic differentiation and Ca2+ deposition after 21 days of cultivation. Different gene expressions associated with mature osteoblasts were upregulated in these two scaffold types, suggesting that the methods in which the scaffolds promote osteogenic differentiation, depends on functionalization approaches. However, only the HT-PCL scaffold was also able to support cell proliferation and cell migration resulting in even cell

  18. Ginsenoside Rg1 of Panax ginseng stimulates the proliferation, odontogenic/osteogenic differentiation and gene expression profiles of human dental pulp stem cells.

    PubMed

    Wang, Ping; Wei, Xi; Zhang, Fujun; Yang, Kai; Qu, Chen; Luo, Huiqiong; He, Longzhu

    2014-01-15

    Ginsenoside Rg1 is one of the major active components of Panax ginseng C. A. Mey. Human dental pulp stem cells (hDPSCs) play an important role in the dentin formation, reparation and tooth tissue engineering. This study investigated the effects of ginsenoside Rg1 on the proliferation, odontogenic differentiation of hDPSCs and revealed the underlying molecular mechanisms. [³H]-thymidine incorporation assay and cell cycle analysis were applied to investigate the proliferation of hDPSCs after the treatment of ginsenoside Rg1. Immunocytochemistry analysis and fluorescent quantitative reverse transcriptase-polymerase chain reaction (FQ-PCR) were performed to evaluate the odontogenic differentiation of hDPSCs. Gene and protein expressions of bone morphogenetic protein-2 (BMP-2) and fibroblast growth factor 2 (FGF2) were detected by FQ-PCR and enzyme-linked immunosorbent assay. The Roche Nimblegen Whole Human Genome Expression profile microarray was used to detected representative gene expression profiles of hDPSCs by ginsenoside Rg1. The results indicated that ginsenoside Rg1 significantly increased hDPSCs proliferation (p<0.05). Gene expressions of DSPP, ALP, OCN, BMP-2, FGF2 and protein expressions of BMP-2 and FGF2 were increased compared with the untreated group (p<0.05). Gene expression profile analysis revealed that 2059 differentially expressed genes were detected by ginsenoside Rg1. Ginsenoside Rg1 promoted the proliferation and differentiation of hDPSCs through alteration of gene expression profiles. Copyright © 2013 Elsevier GmbH. All rights reserved.

  19. Insulin-Like Growth Factor 1 Receptor and p38 Mitogen-Activated Protein Kinase Signals Inversely Regulate Signal Transducer and Activator of Transcription 3 Activity to Control Human Dental Pulp Stem Cell Quiescence, Propagation, and Differentiation

    PubMed Central

    Vandomme, Jerome; Touil, Yasmine; Ostyn, Pauline; Olejnik, Cecile; Flamenco, Pilar; El Machhour, Raja; Segard, Pascaline; Masselot, Bernadette; Bailliez, Yves; Formstecher, Pierre

    2014-01-01

    Dental pulp stem cells (DPSCs) remain quiescent until activated in response to severe dental pulp damage. Once activated, they exit quiescence and enter regenerative odontogenesis, producing reparative dentin. The factors and signaling molecules that control the quiescence/activation and commitment to differentiation of human DPSCs are not known. In this study, we determined that the inhibition of insulin-like growth factor 1 receptor (IGF-1R) and p38 mitogen-activated protein kinase (p38 MAPK) signaling commonly activates DPSCs and promotes their exit from the G0 phase of the cell cycle as well as from the pyronin Ylow stem cell compartment. The inhibition of these two pathways, however, inversely determines DPSC fate. In contrast to p38 MAPK inhibitors, IGF-1R inhibitors enhance dental pulp cell sphere-forming capacity and reduce the cells' colony-forming capacity without inducing cell death. The inverse cellular changes initiated by IGF-1R and p38 MAPK inhibitors were accompanied by inverse changes in the levels of active signal transducer and activator of transcription 3 (STAT3) factor, inactive glycogen synthase kinase 3, and matrix extracellular phosphoglycoprotein, a marker of early odontoblast differentiation. Our data suggest that there is cross talk between the IGF-1R and p38 MAPK signaling pathways in DPSCs and that the signals provided by these pathways converge at STAT3 and inversely regulate its activity to maintain quiescence or to promote self-renewal and differentiation of the cells. We propose a working model that explains the possible interactions between IGF-1R and p38 MAPK at the molecular level and describes the cellular consequences of these interactions. This model may inspire further fundamental study and stimulate research on the clinical applications of DPSC in cellular therapy and tissue regeneration. PMID:24266654

  20. Insulin-like growth factor 1 receptor and p38 mitogen-activated protein kinase signals inversely regulate signal transducer and activator of transcription 3 activity to control human dental pulp stem cell quiescence, propagation, and differentiation.

    PubMed

    Vandomme, Jerome; Touil, Yasmine; Ostyn, Pauline; Olejnik, Cecile; Flamenco, Pilar; El Machhour, Raja; Segard, Pascaline; Masselot, Bernadette; Bailliez, Yves; Formstecher, Pierre; Polakowska, Renata

    2014-04-15

    Dental pulp stem cells (DPSCs) remain quiescent until activated in response to severe dental pulp damage. Once activated, they exit quiescence and enter regenerative odontogenesis, producing reparative dentin. The factors and signaling molecules that control the quiescence/activation and commitment to differentiation of human DPSCs are not known. In this study, we determined that the inhibition of insulin-like growth factor 1 receptor (IGF-1R) and p38 mitogen-activated protein kinase (p38 MAPK) signaling commonly activates DPSCs and promotes their exit from the G0 phase of the cell cycle as well as from the pyronin Y(low) stem cell compartment. The inhibition of these two pathways, however, inversely determines DPSC fate. In contrast to p38 MAPK inhibitors, IGF-1R inhibitors enhance dental pulp cell sphere-forming capacity and reduce the cells' colony-forming capacity without inducing cell death. The inverse cellular changes initiated by IGF-1R and p38 MAPK inhibitors were accompanied by inverse changes in the levels of active signal transducer and activator of transcription 3 (STAT3) factor, inactive glycogen synthase kinase 3, and matrix extracellular phosphoglycoprotein, a marker of early odontoblast differentiation. Our data suggest that there is cross talk between the IGF-1R and p38 MAPK signaling pathways in DPSCs and that the signals provided by these pathways converge at STAT3 and inversely regulate its activity to maintain quiescence or to promote self-renewal and differentiation of the cells. We propose a working model that explains the possible interactions between IGF-1R and p38 MAPK at the molecular level and describes the cellular consequences of these interactions. This model may inspire further fundamental study and stimulate research on the clinical applications of DPSC in cellular therapy and tissue regeneration.

  1. Perspectives for Cell-homing Approaches to Engineer Dental Pulp.

    PubMed

    Galler, Kerstin M; Widbiller, Matthias

    2017-09-01

    Sufficient proof is available today to demonstrate that dental pulp tissue engineering is possible. The body of evidence was generated mainly on cell transplantation; however, because of several severe problems afflicted with this approach, it might not be feasible for a clinical setting in the near future. More recently, cell homing has been proposed as a viable alternative. We suggest a modification of the tissue engineering paradigm, where resident cells are attracted by endogenous, dentin-derived growth factors that further induce cell proliferation and differentiation and a bioactive scaffold material laden with these growth factors that serves as a template for tissue formation. This article highlights the latest developments regarding scaffold materials, stem cells, and dentin-derived growth factors specifically for a cell-homing approach to engineer dental pulp and summarizes new ideas. Copyright © 2017 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  2. Stem/Progenitor Cell–Mediated De Novo Regeneration of Dental Pulp with Newly Deposited Continuous Layer of Dentin in an In Vivo Model

    PubMed Central

    Yamaza, Takayoshi; Shea, Lonnie D.; Djouad, Farida; Kuhn, Nastaran Z.; Tuan, Rocky S.; Shi, Songtao

    2010-01-01

    The ultimate goal of this study is to regenerate lost dental pulp and dentin via stem/progenitor cell–based approaches and tissue engineering technologies. In this study, we tested the possibility of regenerating vascularized human dental pulp in emptied root canal space and producing new dentin on existing dentinal walls using a stem/progenitor cell–mediated approach with a human root fragment and an immunocompromised mouse model. Stem/progenitor cells from apical papilla and dental pulp stem cells were isolated, characterized, seeded onto synthetic scaffolds consisting of poly-D,L-lactide/glycolide, inserted into the tooth fragments, and transplanted into mice. Our results showed that the root canal space was filled entirely by a pulp-like tissue with well-established vascularity. In addition, a continuous layer of dentin-like tissue was deposited onto the canal dentinal wall. This dentin-like structure appeared to be produced by a layer of newly formed odontoblast-like cells expressing dentin sialophosphoprotein, bone sialoprotein, alkaline phosphatase, and CD105. The cells in regenerated pulp-like tissue reacted positively to anti-human mitochondria antibodies, indicating their human origin. This study provides the first evidence showing that pulp-like tissue can be regenerated de novo in emptied root canal space by stem cells from apical papilla and dental pulp stem cells that give rise to odontoblast-like cells producing dentin-like tissue on existing dentinal walls. PMID:19737072

  3. Interaction of Substrate Mechanics with Dental Pulp Stem Cells (DPSCs) differentiation to generate a scaffold for Bone regeneration

    NASA Astrophysics Data System (ADS)

    Rafailovich, Miriam; Bhatnagar, Divya; Bherwani, Aneel; Simon, Marcia

    2012-02-01

    This work investigates the interaction of the substrate mechanics with the differentiation in the absence of chemical induction and only resulting from the stimuli of the substrate mechanics and chemistry. We chose enzymatically cross-linked gelatin hydrogels substrates of different stiffness varying from 8KPa to 100Pa. DPSCs were cultured and differentiated on the substrates for 7, 14 and 21 days with and without dexamethasone induction media. SEM and EDX analysis after 21 days indicate that cells produced a sheet of biomineralized deposits, several tenths of mm thick on the hard substrate irrespective of chemical induction. Modulli of the cells was independent of the induction and stiffness of the hydrogels. RT-PCR assays indicated that cells expressed more osteocalcin when cultured in non-induction media and harder substrate. The shape of the deposits was more uniform and in close packing on the harder substrate with a higher Ca:P ratio. On soft substrate the deposits were more flat with less Ca:P ratio. Further experiments indicated that conformational change due to the crosslinking of gelatin could be the reason for biomineralization.

  4. Stem cells, tissue engineering and periodontal regeneration.

    PubMed

    Han, J; Menicanin, D; Gronthos, S; Bartold, P M

    2014-06-01

    The aim of this review is to discuss the clinical utility of stem cells in periodontal regeneration by reviewing relevant literature that assesses the periodontal-regenerative potential of stem cells. We consider and describe the main stem cell populations that have been utilized with regard to periodontal regeneration, including bone marrow-derived mesenchymal stem cells and the main dental-derived mesenchymal stem cell populations: periodontal ligament stem cells, dental pulp stem cells, stem cells from human exfoliated deciduous teeth, stem cells from apical papilla and dental follicle precursor cells. Research into the use of stem cells for tissue regeneration has the potential to significantly influence periodontal treatment strategies in the future.

  5. Dental Stem Cell in Tooth Development and Advances of Adult Dental Stem Cell in Regenerative Therapies.

    PubMed

    Tan, Jiali; Xu, Xin; Lin, Jiong; Fan, Li; Zheng, Yuting; Kuang, Wei

    2015-01-01

    Stem cell-based therapies are considered as a promising treatment for many clinical usage such as tooth regeneration, bone repairation, spinal cord injury, and so on. However, the ideal stem cell for stem cell-based therapy still remains to be elucidated. In the past decades, several types of stem cells have been isolated from teeth, including dental pulp stem cells (DPSCs), stem cells from human exfoliated deciduous teeth (SHED), periodontal ligament stem cells (PDLSCs), dental follicle progenitor stem cells (DFPCs) and stem cells from apical papilla (SCAP), which may be a good source for stem cell-based therapy in certain disease, especially when they origin from neural crest is considered. In this review, the specific characteristics and advantages of the adult dental stem cell population will be summarized and the molecular mechanisms of the differentiation of dental stem cell during tooth development will be also discussed.

  6. Adult stem cell-based apexogenesis

    PubMed Central

    Li, Yao; Shu, Li-Hong; Yan, Ming; Dai, Wen-Yong; Li, Jun-Jun; Zhang, Guang-Dong; Yu, Jin-Hua

    2014-01-01

    Generally, the dental pulp needs to be removed when it is infected, and root canal therapy (RCT) is usually required in which infected dental pulp is replaced with inorganic materials (paste and gutta percha). This treatment approach ultimately brings about a dead tooth. However, pulp vitality is extremely important to the tooth itself, since it provides nutrition and acts as a biosensor to detect the potential pathogenic stimuli. Despite the reported clinical success rate, RCT-treated teeth are destined to be devitalized, brittle and susceptible to postoperative fracture. Recently, the advances and achievements in the field of stem cell biology and regenerative medicine have inspired novel biological approaches to apexogenesis in young patients suffering from pulpitis or periapical periodontitis. This review mainly focuses on the benchtop and clinical regeneration of root apex mediated by adult stem cells. Moreover, current strategies for infected pulp therapy are also discussed here. PMID:25332909

  7. A monoclonal antibody recognizes undifferentiation-specific carbohydrate moieties expressed on cell surface of the human dental pulp cells.

    PubMed

    Kang, Kyung-Jung; Ko, Seon-Yle; Ryu, Chun-Jeih; Jang, Young-Joo

    2017-05-01

    Human dental pulp cells are obtained from dental pulp tissue, and have the ability to form dentin and a pulp-like complex. Although adult stem cells have been identified from the primary culture by using specific cell surface markers, the identity of surface markers for the purification of stem cells within the dental pulp population are still unclear. Previously, we had constructed monoclonal antibodies against the undifferentiated cell-specific surface markers of human dental pulp cells (hDPCs) by performing decoy immunization. Among them, a monoclonal antibody against the cell surface antigen of the undifferentiated hDPCs (named UPSA-1) was purified and its heavy and light chain consensus regions were analyzed. The cell surface binding affinity of UPSA-1 mAb on the undifferentiated hDPCs was stronger than that on the differentiated cells. When tunicamycin was applied to hDPSCs during culture, the cell surface binding affinity of the antibody was dramatically decreased, and dentinogenic differentiation was reduced. The purified UPSA-1 antigen band resulting from immunoprecipitation disappeared or shifted down on the SDS-PAGE by deglycosylation. These data suggested that glycosylation on the cell surface might be a marker of an undifferentiated state, and that UPSA-1 mAb might be useful for identifying the carbohydrate moiety on the cell surface of undifferentiated pulp cells. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  8. Regenerative endodontics in light of the stem cell paradigm.

    PubMed

    Rosa, Vinicius; Botero, Tatiana M; Nör, Jacques E

    2011-08-01

    Stem cells play a critical role in development and in tissue regeneration. The dental pulp contains a small sub-population of stem cells that are involved in the response of the pulp to caries progression. Specifically, stem cells replace odontoblasts that have undergone cell death as a consequence of the cariogenic challenge. Stem cells also secrete factors that have the potential to enhance pulp vascularisation and provide the oxygen and nutrients required for the dentinogenic response that is typically observed in teeth with deep caries. However, the same angiogenic factors that are required for dentine regeneration may ultimately contribute to the demise of the pulp by enhancing vascular permeability and interstitial pressure. Recent studies focused on the biology of dental pulp stem cells revealed that the multipotency and angiogenic capacity of these cells could be exploited therapeutically in dental pulp tissue engineering. Collectively, these findings suggest new treatment paradigms in the field of endodontics. The goal of this review is to discuss the potential impact of dental pulp stem cells to regenerative endodontics. © 2011 FDI World Dental Federation.

  9. Regenerative Endodontics in light of the stem cell paradigm

    PubMed Central

    Rosa, Vinicius; Botero, Tatiana M.; Nör, Jacques E.

    2013-01-01

    Stem cells play a critical role in development and in tissue regeneration. The dental pulp contains a small sub-population of stem cells that are involved in the response of the pulp to caries progression. Specifically, stem cells replace odontoblasts that have undergone cell death as a consequence of the cariogenic challenge. Stem cells also secrete factors that have the potential to enhance pulp vascularization and provide the oxygen and nutrients required for the dentinogenic response that is typically observed in teeth with deep caries. However, the same angiogenic factors that are required for dentin regeneration may ultimately contribute to the demise of the pulp by enhancing vascular permeability and interstitial pressure. Recent studies focused on the biology of dental pulp stem cells revealed that the multipotency and angiogenic capacity of these cells could be exploited therapeutically in dental pulp tissue engineering. Collectively, these findings suggest new treatment paradigms in the field of Endodontics. The goal of this review is to discuss the potential impact of dental pulp stem cells to Regenerative Endodontics. PMID:21726222

  10. Static Magnetic Field Attenuates Lipopolysaccharide-Induced Inflammation in Pulp Cells by Affecting Cell Membrane Stability

    PubMed Central

    Tsao, Jeng-Ting; Lee, Lin-Wen; Lin, Che-Tong

    2015-01-01

    One of the causes of dental pulpitis is lipopolysaccharide- (LPS-) induced inflammatory response. Following pulp tissue inflammation, odontoblasts, dental pulp cells (DPCs), and dental pulp stem cells (DPSCs) will activate and repair damaged tissue to maintain homeostasis. However, when LPS infection is too serious, dental repair is impossible and disease may progress to irreversible pulpitis. Therefore, the aim of this study was to examine whether static magnetic field (SMF) can attenuate inflammatory response of dental pulp cells challenged with LPS. In methodology, dental pulp cells were isolated from extracted teeth. The population of DPSCs in the cultured DPCs was identified by phenotypes and multilineage differentiation. The effects of 0.4 T SMF on DPCs were observed through MTT assay and fluorescent anisotropy assay. Our results showed that the SMF exposure had no effect on surface markers or multilineage differentiation capability. However, SMF exposure increases cell viability by 15%. In addition, SMF increased cell membrane rigidity which is directly related to higher fluorescent anisotropy. In the LPS-challenged condition, DPCs treated with SMF demonstrated a higher tolerance to LPS-induced inflammatory response when compared to untreated controls. According to these results, we suggest that 0.4 T SMF attenuates LPS-induced inflammatory response to DPCs by changing cell membrane stability. PMID:25884030

  11. Investigation of modified platelet-rich plasma (mPRP) in promoting the proliferation and differentiation of dental pulp stem cells from deciduous teeth

    PubMed Central

    Wen, J.; Li, H.T.; Li, S.H.; Li, X.; Duan, J.M.

    2016-01-01

    Stem cells from human exfoliated deciduous teeth (SHEDs) have great potential to treat various dental-related diseases in regenerative medicine. They are usually maintained with 10% fetal bovine serum (FBS) in vitro. Modified platelet-rich plasma (mPRP) would be a safe alternative to 10% FBS during SHEDs culture. Therefore, our study aimed to compare the proliferation and differentiation of SHEDs cultured in mPRP and FBS medium to explore an optimal concentration of mPRP for SHEDs maintenance. Platelets were harvested by automatic blood cell analyzer and activated by repeated liquid nitrogen freezing and thawing. The platelet-related cytokines were examined and analyzed by ELISA. SHEDs were extracted and cultured with different concentrations of mPRP or 10% FBS medium. Alkaline phosphatase (ALP) activity was measured. Mineralization factors, RUNX2 and OCN, were measured by real-time PCR. SHEDs were characterized with mesenchymal stem cells (MSCs) markers including vimentin, CD44, and CD105. mPRP at different concentrations (2, 5, 10, and 20%) enhanced the growth of SHEDs. Moreover, mPRP significantly stimulated ALP activity and promoted expression of RUNX2 and OCN compared with 10% FBS. mPRP could efficiently facilitate proliferation and differentiation of SHEDs, and 2% mPRP would be an optimal substitute for 10% FBS during SHEDs expansion and differentiation in clinical scale manufacturing. PMID:27599200

  12. Stem Cell Sciences plc.

    PubMed

    Daniels, Sebnem

    2006-09-01

    Stem Cell Sciences' core objective is to develop safe and effective stem cell-based therapies for currently incurable diseases. In order to achieve this goal, Stem Cell Sciences recognizes the need for multiple technologies and a globally integrated stem cell initiative. The key challenges for the successful application of stem cells in the clinic is the need for a reproducible supply of pure, fully characterized stem cells that have been grown in suitable conditions for use in the clinic.

  13. Regenerative capability of dental pulp cells after crown fracture.

    PubMed

    Shima, Hidehito; Matsuzaka, Kenichi; Kokubu, Eitoyo; Inoue, Takashi

    2013-02-01

    The purpose of this study was to evaluate the characteristics of dental pulp cells for tissue engineering derived from the fractured incisal portion of tooth crowns. Thirty Sprague-Dawley rats were used for histological and immunohistochemical analysis of nestin protein expression and to measure levels of mRNAs encoding osteocalcin, osteopontin, bone sialoprotein (BSP), dentin sialoprotein (DSP), heat shock protein (HSP) 27, vascular endothelial growth factor (VEGF), ATP-binding cassette transporter G2 (ABCG2), nestin, and p57(Kip2) . Odontoblasts at the incisal portion in the control group were oriented in a regular pattern, but those in the experimental group were randomly stratified. Immunohistochemically, only a few odontoblasts were positive for nestin at the incisal portion in the experimental group at 2 days. Some cells in the inner area in the control group were positive for nestin, but nestin-positive cells in the experimental group at the incisal portion were not observed. The mRNA expression for osteogenic or odontogenic markers in the experimental group was higher than in the control group. HSP27 mRNA expression in the experimental group at 2 days was higher than in the control group and in the experimental group at 7 days. mRNA expression of stem cell markers, such as ABCG2 and nestin, in the experimental group tended to decrease compared with the control. In conclusion, this study demonstrates that dental pulp stem cells derived from fractured teeth differentiate to osteogenic or odontogenic cells. © 2012 John Wiley & Sons A/S.

  14. Quality evaluation of dissolving pulp fabricated from banana plant stem and its potential for biorefinery.

    PubMed

    Das, Atanu Kumar; Nakagawa-Izumi, Akiko; Ohi, Hiroshi

    2016-08-20

    The study was conducted to evaluate the quality of dissolving pulp of Musa sapientum L. (banana) plant stem and its potential for biorefinery. Introduction of pre-hydrolysis prior to any alkaline pulping process helps to reduce the content of hemicellulose and consequently produce acceptably high content of cellulose pulp. Water pre-hydrolysis was done at 150°C for 90min. The amount of lignin, xylan and glucan in the extracted pre-hydrolysis liquor (PHL) was 1.6, 4.9 and 1.6%, respectively. Pulping of pre-extracted chips was done following soda-AQ, alkaline sulfite and kraft process. The ratio of chip to liquor was 1:7 for both pre-hydrolysis and pulping. The kraft pulping process with 20% active alkali and 25% sulfidity at 150°C for 90min showed the best result. The lowest kappa number was 26.2 with a considerable pulp yield of 32.7%. The pulp was bleached by acidic NaClO2 and the consistency was 10% based on air-dried pulp. The lowest amount of 7% NaClO2 was used for the bleaching sequence of D0ED1ED2. After D0ED1ED2 bleaching, the pulp showed that α-cellulose, brightness and ash were 91.9, 77.9 and 1.6% respectively. The viscosity was 19.9cP. Hence, there is a possibility to use banana plant stem as a raw material for dissolving grade pulp and other bioproducts. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Stem cells from oral niches: a review

    PubMed Central

    Giordano, Guido; La Monaca, Gerardo; Annibali, Susanna; Cicconetti, Andrea; Ottolenghi, Livia

    2011-01-01

    Summary Aim Stem cell research in recent years have been considered the most advanced sort of medical-scientific research and early results have aroused great expectations. Also in dentistry many studies were performed with the final aim of obtaining new bone and new teeth. In this work we describe the state of the art in dental science stem cell research. Methods We have performed a web-based research on MEDLINE within (www.pubmed.gov). We have used “stem cells from human exfoliated deciduous teeth” (24 paper found), “periodontal ligament stem cells” (32 paper found), “stem cell apical papilla” (16 paper found), “dental pulp stem cells” (136 paper found) as keywords for research. For each keyword we have performed a complete review focusing on knowledge upgrade. Results For each topic was created a selection of papers in chronological order of publication date so to give a timetable of the development of the research for each niche. Conclusion Research about stem cell from oral niches began in 2000 and every year papers publicated were more than the precedent. This review analysed about 180 articles most of which in the last 5 years. Dentla pulp from adult as from deciduous teeth seems to be the most valuable font of stem cells due to the pluripotential type of cells. PMID:22238715

  16. Stem Cell Basics

    MedlinePlus

    ... healthy cells replace damaged cells in adult organisms. Stem cell research is one of the most fascinating areas of ... as with many expanding fields of scientific inquiry, research on stem cells raises scientific questions as rapidly as it generates ...

  17. Stem cell-based biological tooth repair and regeneration

    PubMed Central

    Volponi, Ana Angelova; Pang, Yvonne; Sharpe, Paul T.

    2010-01-01

    Teeth exhibit limited repair in response to damage, and dental pulp stem cells probably provide a source of cells to replace those damaged and to facilitate repair. Stem cells in other parts of the tooth, such as the periodontal ligament and growing roots, play more dynamic roles in tooth function and development. Dental stem cells can be obtained with ease, making them an attractive source of autologous stem cells for use in restoring vital pulp tissue removed because of infection, in regeneration of periodontal ligament lost in periodontal disease, and for generation of complete or partial tooth structures to form biological implants. As dental stem cells share properties with mesenchymal stem cells, there is also considerable interest in their wider potential to treat disorders involving mesenchymal (or indeed non-mesenchymal) cell derivatives, such as in Parkinson's disease. PMID:21035344

  18. [Pancreatic cancer stem cell].

    PubMed

    Hamada, Shin; Masamune, Atsushi; Shimosegawa, Tooru

    2015-05-01

    Prognosis of pancreatic cancer remains dismal due to the resistance against conventional therapies. Metastasis and massive invasion toward surrounding organs hamper radical resection. Small part of entire cancer cells reveal resistance against chemotherapy or radiotherapy, increased tumorigenicity and migratory phenotype. These cells are called as cancer stem cells, as a counter part of normal stem cells. In pancreatic cancer, several cancer stem cell markers have been identified, which enabled detailed characterization of pancreatic cancer stem cells. Recent researches clarified that conventional chemotherapy itself could increase cancer cells with stem cell-phenotype, suggesting the necessity of cancer stem cell-targeting therapy. Based on these observations, pancreatic cancer stem cell-targeting therapies have been tested, which effectively eliminated cancer stem cell fraction and attenuated cancer progression in experimental models. Clinical efficacy of these therapies need to be evaluated, and cancer stem cell-targeting therapy will contribute to improve the prognosis of pancreatic cancer.

  19. Application of Stem Cell Technology in Dental Regenerative Medicine.

    PubMed

    Feng, Ruoxue; Lengner, Chistopher

    2013-07-01

    In this review, we summarize the current literature regarding the isolation and characterization of dental tissue-derived stem cells and address the potential of these cell types for use in regenerative cell transplantation therapy. Looking forward, platforms for the delivery of stem cells via scaffolds and the use of growth factors and cytokines for enhancing dental stem cell self-renewal and differentiation are discussed. We aim to understand the developmental origins of dental tissues in an effort to elucidate the molecular pathways governing the genesis of somatic dental stem cells. The advantages and disadvantages of several dental stem cells are discussed, including the developmental stage and specific locations from which these cells can be purified. In particular, stem cells from human exfoliated deciduous teeth may act as a very practical and easily accessibly reservoir for autologous stem cells and hold the most value in stem cell therapy. Dental pulp stem cells and periodontal ligament stem cells should also be considered for their triple lineage differentiation ability and relative ease of isolation. Further, we address the potentials and limitations of induced pluripotent stem cells as a cell source in dental regenerative. From an economical and a practical standpoint, dental stem cell therapy would be most easily applied in the prevention of periodontal ligament detachment and bone atrophy, as well as in the regeneration of dentin-pulp complex. In contrast, cell-based tooth replacement due to decay or other oral pathology seems, at the current time, an untenable approach.

  20. Therapeutic potential of dental stem cells

    PubMed Central

    Chalisserry, Elna Paul; Nam, Seung Yun; Park, Sang Hyug; Anil, Sukumaran

    2017-01-01

    Stem cell biology has become an important field in regenerative medicine and tissue engineering therapy since the discovery and characterization of mesenchymal stem cells. Stem cell populations have also been isolated from human dental tissues, including dental pulp stem cells, stem cells from human exfoliated deciduous teeth, stem cells from apical papilla, dental follicle progenitor cells, and periodontal ligament stem cells. Dental stem cells are relatively easily obtainable and exhibit high plasticity and multipotential capabilities. The dental stem cells represent a gold standard for neural-crest-derived bone reconstruction in humans and can be used for the repair of body defects in low-risk autologous therapeutic strategies. The bioengineering technologies developed for tooth regeneration will make substantial contributions to understand the developmental process and will encourage future organ replacement by regenerative therapies in a wide variety of organs such as the liver, kidney, and heart. The concept of developing tooth banking and preservation of dental stem cells is promising. Further research in the area has the potential to herald a new dawn in effective treatment of notoriously difficult diseases which could prove highly beneficial to mankind in the long run. PMID:28616151

  1. Therapeutic potential of dental stem cells.

    PubMed

    Chalisserry, Elna Paul; Nam, Seung Yun; Park, Sang Hyug; Anil, Sukumaran

    2017-01-01

    Stem cell biology has become an important field in regenerative medicine and tissue engineering therapy since the discovery and characterization of mesenchymal stem cells. Stem cell populations have also been isolated from human dental tissues, including dental pulp stem cells, stem cells from human exfoliated deciduous teeth, stem cells from apical papilla, dental follicle progenitor cells, and periodontal ligament stem cells. Dental stem cells are relatively easily obtainable and exhibit high plasticity and multipotential capabilities. The dental stem cells represent a gold standard for neural-crest-derived bone reconstruction in humans and can be used for the repair of body defects in low-risk autologous therapeutic strategies. The bioengineering technologies developed for tooth regeneration will make substantial contributions to understand the developmental process and will encourage future organ replacement by regenerative therapies in a wide variety of organs such as the liver, kidney, and heart. The concept of developing tooth banking and preservation of dental stem cells is promising. Further research in the area has the potential to herald a new dawn in effective treatment of notoriously difficult diseases which could prove highly beneficial to mankind in the long run.

  2. Effect of EDTA on TGF-β1 released from the dentin matrix and its influence on dental pulp stem cell migration.

    PubMed

    Gonçalves, Lidiany Freitas; Fernandes, Ana Paula; Cosme-Silva, Leopoldo; Colombo, Fabio Antonio; Martins, Natália Silva; Oliveira, Thais Marchini; Araujo, Tomaz Henrique; Sakai, Vivien Thiemy

    2016-12-22

    Bioactive molecules stored in dentin, such as transforming growth factor beta1 (TGF-b1), may be involved in the signaling events related to dental tissue repair. The authors conducted an in vitro evaluation of the amount of TGF-b1 released from dentin slices after treatment with 10% ethylenediaminetetraacetic acid (EDTA), 2.5% sodium hypochlorite (NaOCl) or phosphate-buffered saline (PBS), and the effect of this growth factor on stem cell migration from human exfoliated deciduous teeth (SHED). Sixty 1-mm-thick tooth slices were prepared with or without the predentin layer, and treated with either 10% EDTA for 1 minute, 2.5% NaOCl for 5 days or kept in PBS. Tooth slice conditioned media were prepared and used for TGF-b1 ELISA and migration assays. Culture medium with different concentrations of recombinant human TGF-b1 (0.5, 1.0, 5.0 or 10.0 ng/mL) was also tested by migration assay. The data were evaluated by ANOVA and Tukey's test. Optical density values corresponding to media conditioned by tooth slices either containing or not containing the predentin layer and treated with 10% EDTA were statistically greater than the other groups and close to 1 ng/mL. Increased rates of migration toward media conditioned by tooth slices containing the predentin layer and treated with PBS, 10% EDTA or 2.5% NaOCl were observed. Recombinant human TGF-b1 also stimulated migration of SHED, irrespective of the concentration used. EDTA may be considered an effective extractant of TGF-b1 from the dentin matrix. However, it does not impact SHED migration, suggesting that other components may account for the cell migration.

  3. Mesenchymal dental stem cells in regenerative dentistry.

    PubMed

    Rodríguez-Lozano, Francisco-Javier; Insausti, Carmen-Luisa; Iniesta, Francisca; Blanquer, Miguel; Ramírez, María-del-Carmen; Meseguer, Luis; Meseguer-Henarejos, Ana-Belén; Marín, Noemí; Martínez, Salvador; Moraleda, José-María

    2012-11-01

    In the last decade, tissue engineering is a field that has been suffering an enormous expansion in the regenerative medicine and dentistry. The use of cells as mesenchymal dental stem cells of easy access for dentist and oral surgeon, immunosuppressive properties, high proliferation and capacity to differentiate into odontoblasts, cementoblasts, osteoblasts and other cells implicated in the teeth, suppose a good perspective of future in the clinical dentistry. However, is necessary advance in the known of growth factors and signalling molecules implicated in tooth development and regeneration of different structures of teeth. Furthermore, these cells need a fabulous scaffold that facility their integration, differentiation, matrix synthesis and promote multiple specific interactions between cells. In this review, we give a brief description of tooth development and anatomy, definition and classification of stem cells, with special attention of mesenchymal stem cells, commonly used in the cellular therapy for their trasdifferentiation ability, non ethical problems and acceptable results in preliminary clinical trials. In terms of tissue engineering, we provide an overview of different types of mesenchymal stem cells that have been isolated from teeth, including dental pulp stem cells (DPSCs), stem cells from human exfoliated deciduous teeth (SHEDs), periodontal ligament stem cells (PDLSCs), dental follicle progenitor stem cells (DFPCs), and stem cells from apical papilla (SCAPs), growth factors implicated in regeneration teeth and types of scaffolds for dental tissue regeneration.

  4. Mapping of BrdU label-retaining dental pulp cells in growing teeth and their regenerative capacity after injuries.

    PubMed

    Ishikawa, Yuko; Ida-Yonemochi, Hiroko; Suzuki, Hironobu; Nakakura-Ohshima, Kuniko; Jung, Han-Sung; Honda, Masaki J; Ishii, Yumiko; Watanabe, Nobukazu; Ohshima, Hayato

    2010-09-01

    Recent studies have demonstrated that human dental pulp contains adult stem cells. A pulse of the thymidine analog BrdU given to young animals at the optimal time could clarify where slow-cycling long-term label-retaining cells (LRCs), putative adult stem cells, reside in the pulp tissue. This study focuses on the mapping of LRCs in growing teeth and their regenerative capacity after tooth injuries. Two to seven peritoneal injections of BrdU into pregnant Wistar rats revealed slow-cycling long-term dense LRCs in the mature tissues of born animals. Numerous dense LRCs were postnatally decreased in number and reached a plateau at 4 weeks after birth when they mainly resided in the center of the dental pulp, associating with blood vessels. Mature dental pulp cells were stained with Hoechst 33342 and sorted into (<0.76%) side population cells using FACS, which included dense LRCs. Some dense LRCs co-expressed mesenchymal stem cell markers such as STRO-1 or CD146. Tooth injuries caused degeneration of the odontoblast layer, and newly differentiated odontoblast-like cells contained LRCs. Thus, dense LRCs in mature pulp tissues were supposed to be dental pulp stem cells possessing regenerative capacity for forming newly differentiated odontoblast-like cells. The present study proposes the new hypothesis that both granular and dense LRCs are equipped in the dental pulp and that the dense LRCs with proliferative capacity play crucial roles in the pulpal healing process following exogenous stimuli in cooperation with the granular LRCs.

  5. Human dental pulp cell culture and cell transplantation with an alginate scaffold.

    PubMed

    Kumabe, Shunji; Nakatsuka, Michiko; Kim, Gi-Seup; Jue, Seong-Suk; Aikawa, Fumiko; Shin, Je-Won; Iwai, Yasutomo

    2006-02-01

    Many studies on tissue stem cells have been conducted in the field of regenerative medicine, and some studies have indicated that cultured dental pulp mesenchymal cells secrete dentin matrix. In the present study we used alginate as a scaffold to transplant subcultured human dental pulp cells subcutaneously into the backs of nude mice. We found that when beta-glycerophosphate was added to the culture medium, dentin sialophosphoprotein mRNA coding dentin sialoprotein (DSP) was expressed. An increase in alkaline phosphatase, which is an early marker for odontoblast differentiation, was also demonstrated. At 6 weeks after implantation the subcutaneous formation of radio-opaque calcified bodies was observed in situ. Immunohistochemical and fine structure studies identified expression of type I collagen, type III collagen, and DSP in the mineralizing transplants. Isolated odontoblast-like cells initiated dentin-like hard tissue formation and scattered autolyzing apoptotic cells were also observed in the transplants. The study showed that subcultured dental pulp cells actively differentiate into odontoblast-like cells and induce calcification in an alginate scaffold.

  6. Liver cancer stem cells.

    PubMed

    Sell, Stewart; Leffert, Hyam L

    2008-06-10

    In an effort to review the evidence that liver cancer stem cells exist, two fundamental questions must be addressed. First, do hepatocellular carcinomas (HCC) arise from liver stem cells? Second, do HCCs contain cells that possess properties of cancer stem cells? For many years the finding of preneoplastic nodules in the liver during experimental induction of HCCs by chemicals was interpreted to support the hypothesis that HCC arose by dedifferentiation of mature liver cells. More recently, recognition of the role of small oval cells in the carcinogenic process led to a new hypothesis that HCC arises by maturation arrest of liver stem cells. Analysis of the cells in HCC supports the presence of cells with stem-cell properties (ie, immortality, transplantability, and resistance to therapy). However, definitive markers for these putative cancer stem cells have not yet been found and a liver cancer stem cell has not been isolated.

  7. Considerations on the ultrastructural particularities of the dental pulp cells.

    PubMed

    Manolea, H; Deva, V; Bogdan, Fl; Moraru, Iren; Pancă, Oana-Adina; Caraivan, O

    2008-01-01

    We realized an ultrastructural study of the cells of the dental pulp, having in view their particularities relative to other types of conjunctive tissue. For this purpose, we selected five cases represented by teeth without subjective or objective symptomatology. Within the paper there are exposed the morphological aspects observed by means of electron microscopy. The results are then discussed in relation with a series of observations made by other researchers regarding the particularities of the pulp cells structures.

  8. Stem Cell Transplant

    MedlinePlus

    ... transplant is a procedure that infuses healthy blood stem cells into your body to replace your damaged or ... A bone marrow transplant is also called a stem cell transplant. A bone marrow transplant may be necessary ...

  9. Tooth slice/scaffold model of dental pulp tissue engineering.

    PubMed

    Sakai, V T; Cordeiro, M M; Dong, Z; Zhang, Z; Zeitlin, B D; Nör, J E

    2011-07-01

    Multipotency is a defining characteristic of post-natal stem cells. The human dental pulp contains a small subpopulation of stem cells that exhibit multipotency, as demonstrated by their ability to differentiate into odontoblasts, neural cells, and vascular endothelial cells. These discoveries highlight the fundamental role of stem cells in the biology of the dental pulp and suggest that these cells are uniquely suited for dental pulp tissue-engineering purposes. The availability of experimental approaches specifically designed for studies of the differentiation potential of dental pulp stem cells has played an important role in these discoveries. The objective of this review is to describe the development and characterization of the Tooth Slice/Scaffold Model of Dental Pulp Tissue Engineering. In addition, we discuss the multipotency of dental pulp stem cells, focusing on the differentiation of these cells into functional odontoblasts and into vascular endothelial cells.

  10. Plant stem cell niches.

    PubMed

    Aichinger, Ernst; Kornet, Noortje; Friedrich, Thomas; Laux, Thomas

    2012-01-01

    Multicellular organisms possess pluripotent stem cells to form new organs, replenish the daily loss of cells, or regenerate organs after injury. Stem cells are maintained in specific environments, the stem cell niches, that provide signals to block differentiation. In plants, stem cell niches are situated in the shoot, root, and vascular meristems-self-perpetuating units of organ formation. Plants' lifelong activity-which, as in the case of trees, can extend over more than a thousand years-requires that a robust regulatory network keep the balance between pluripotent stem cells and differentiating descendants. In this review, we focus on current models in plant stem cell research elaborated during the past two decades, mainly in the model plant Arabidopsis thaliana. We address the roles of mobile signals on transcriptional modules involved in balancing cell fates. In addition, we discuss shared features of and differences between the distinct stem cell niches of Arabidopsis.

  11. Schwann Cell Phenotype Changes in Aging Human Dental Pulp.

    PubMed

    Couve, E; Lovera, M; Suzuki, K; Schmachtenberg, O

    2017-10-01

    Schwann cells are glial cells that support axonal development, maintenance, defense, and regeneration in the peripheral nervous system. There is limited knowledge regarding the organization, plasticity, and aging of Schwann cells within the dental pulp in adult permanent teeth. The present study sought to relate changes in the pattern of Schwann cell phenotypes between young and old adult teeth with neuronal, immune, and vascular components of the dental pulp. Schwann cells are shown to form a prominent glial network at the dentin-pulp interface, consisting of nonmyelinating and myelinating phenotypes, forming a multicellular neuroimmune interface in association with nerve fibers and dendritic cells. Schwann cell phenotypes are recognized by the expression of S100, glial fibrillary acidic protein (GFAP), myelin basic protein (MBP), Sox10, GAP43, and p75NTR markers. In young adult teeth, a dense population of nonmyelinating Schwann cells projects processes in close association with sensory nerve terminals through the odontoblast layer, reaching the adjacent predentin/dentin domain. While GAP43 and p75NTR are highly expressed in nonmyelinating Schwann cells from young adult teeth, the presence of these markers declines significantly in old adult teeth. Myelinated axons, identified by MBP expression, are mainly present at the Raschkow plexus and within nerve bundles in the dental pulp, but their density is significantly reduced in old adult versus young adult teeth. These data reveal age-related changes within the glial network of the dental pulp, in association with a reduction of coronal dental pulp innervation in old adult versus young adult teeth. The prominence of Schwann cells as a cellular component at the dentin-pulp interface supports the notion that their association with sensory nerve terminals and immune system components forms part of an integrated multicellular barrier for defense against pathogens and dentin repair.

  12. Chitosan-collagen biomembrane embedded with calcium-aluminate enhances dentinogenic potential of pulp cells.

    PubMed

    Soares, Diana Gabriela; Rosseto, Hebert Luís; Basso, Fernanda Gonçalves; Scheffel, Débora Salles; Hebling, Josimeri; Costa, Carlos Alberto de Souza

    2016-01-01

    The development of biomaterials capable of driving dental pulp stem cell differentiation into odontoblast-like cells able to secrete reparative dentin is the goal of current conservative dentistry. In the present investigation, a biomembrane (BM) composed of a chitosan/collagen matrix embedded with calcium-aluminate microparticles was tested. The BM was produced by mixing collagen gel with a chitosan solution (2:1), and then adding bioactive calcium-aluminate cement as the mineral phase. An inert material (polystyrene) was used as the negative control. Human dental pulp cells were seeded onto the surface of certain materials, and the cytocompatibility was evaluated by cell proliferation and cell morphology, assessed after 1, 7, 14 and 28 days in culture. The odontoblastic differentiation was evaluated by measuring alkaline phosphatase (ALP) activity, total protein production, gene expression of DMP-1/DSPP and mineralized nodule deposition. The pulp cells were able to attach onto the BM surface and spread, displaying a faster proliferative rate at initial periods than that of the control cells. The BM also acted on the cells to induce more intense ALP activity, protein production at 14 days, and higher gene expression of DSPP and DMP-1 at 28 days, leading to the deposition of about five times more mineralized matrix than the cells in the control group. Therefore, the experimental biomembrane induced the differentiation of pulp cells into odontoblast-like cells featuring a highly secretory phenotype. This innovative bioactive material can drive other protocols for dental pulp exposure treatment by inducing the regeneration of dentin tissue mediated by resident cells.

  13. Nail stem cells.

    PubMed

    Sellheyer, Klaus

    2013-03-01

    Our knowledge on stem cells of the hair follicle has increased exponentially after the bulge was characterized as the stem cell niche two decades ago. In contrast, little is known about stem cells in the nail unit. Whereas hair follicles are plentiful and easy to access, the human body has only twenty nails and they are rarely biopsied. Therefore, examining fetal material offers unique advantages. In the following mini-review, our current knowledge on nail stem cells is summarized and analogies to the hair follicle stem cells are drawn.

  14. Stem cells remember their grade.

    PubMed

    Lo Celso, Cristina; Scadden, David T

    2007-08-16

    The stem cell state is understood based on what cells do in performance assays, crude measures of a highly refined state. In this issue of Cell Stem Cell, Dykstra et al. (2007) reveal stem cell gradation and the extent to which that gradation is retained in stem cell daughters of hematopoietic stem cells.

  15. Angiogenic signaling triggered by cariogenic bacteria in pulp cells.

    PubMed

    Soden, R I; Botero, T M; Hanks, C T; Nör, J E

    2009-09-01

    The inflammation observed in the dental pulp of teeth with deep caries lesions is characterized by a significant increase in blood vessel density. It is known that lipoteichoic acid (LTA) from Gram-positive cariogenic bacteria induces expression of vascular endothelial growth factor (VEGF) in dental pulp cells. The hypothesis underlying this study was that LTA induces VEGF expression in dental pulp cells through TLR2 and PI3k/Akt signaling. Odontoblast-like cells (MDPC-23) and undifferentiated pulp cells (OD-21) were exposed to LTA from Streptococcus sanguis, and the role of TLR2, PI3K/Akt, and IKK signaling in LTA-induced VEGF expression was evaluated. These studies demonstrated that TLR2 signaling through the PI3K-Akt pathway is necessary for LTA-induced VEGF expression in pulp cells. In contrast, inhibition of IKK signaling did not prevent VEGF up-regulation in response to LTA. Understanding signaling pathways triggered by cariogenic bacteria may reveal novel therapeutic targets for the clinical management of pulpitis.

  16. Stem Cells and Aging.

    PubMed

    Koliakos, George

    2017-02-01

    The article is a presentation at the 4th Conference of ESAAM, which took place on October 30-31, 2015, in Athens, Greece. Its purpose was not to cover all aspects of cellular aging but to share with the audience of the Conference, in a 15-minute presentation, current knowledge about the rejuvenating and repairing somatic stem cells that are distinct from other stem cell types (such as embryonic or induced pluripotent stem cells), emphasize that our body in old age cannot take advantage of these rejuvenating cells, and provide some examples of novel experimental stem cell applications in the field of rejuvenation and antiaging biomedical research.

  17. Stress and stem cells.

    PubMed

    Tower, John

    2012-01-01

    The unique properties and functions of stem cells make them particularly susceptible to stresses and also lead to their regulation by stress. Stem cell division must respond to the demand to replenish cells during normal tissue turnover as well as in response to damage. Oxidative stress, mechanical stress, growth factors, and cytokines signal stem cell division and differentiation. Many of the conserved pathways regulating stem cell self-renewal and differentiation are also stress-response pathways. The long life span and division potential of stem cells create a propensity for transformation (cancer) and specific stress responses such as apoptosis and senescence act as antitumor mechanisms. Quiescence regulated by CDK inhibitors and a hypoxic niche regulated by FOXO transcription factor function to reduce stress for several types of stem cells to facilitate long-term maintenance. Aging is a particularly relevant stress for stem cells, because repeated demands on stem cell function over the life span can have cumulative cell-autonomous effects including epigenetic dysregulation, mutations, and telomere erosion. In addition, aging of the organism impairs function of the stem cell niche and systemic signals, including chronic inflammation and oxidative stress.

  18. Stress and stem cells

    PubMed Central

    Tower, John

    2013-01-01

    The unique properties and functions of stem cells make them particularly susceptible to stresses and also lead to their regulation by stress. Stem cell division must respond to the demand to replenish cells during normal tissue turnover as well as in response to damage. Oxidative stress, mechanical stress, growth factors, and cytokines signal stem cell division and differentiation. Many of the conserved pathways regulating stem cell self-renewal and differentiation are also stress-response pathways. The long life span and division potential of stem cells create a propensity for transformation (cancer) and specific stress responses such as apoptosis and senescence act as antitumor mechanisms. Quiescence regulated by CDK inhibitors and a hypoxic niche regulated by FOXO transcription factor function to reduce stress for several types of stem cells to facilitate long-term maintenance. Aging is a particularly relevant stress for stem cells, because repeated demands on stem cell function over the life span can have cumulative cell-autonomous effects including epigenetic dysregulation, mutations, and telomere erosion. In addition, aging of the organism impairs function of the stem cell niche and systemic signals, including chronic inflammation and oxidative stress. PMID:23799624

  19. Stem Cell Transplantation for Pulpal Regeneration: A Systematic Review

    PubMed Central

    Jakusz, Kimberley; Jochens, Arne; Dörfer, Christof; Schwendicke, Falk

    2015-01-01

    For treating pulpal pathological conditions, pulpal regeneration through transplanted stem/progenitor cells might be an alternative to conventional root canal treatment. A number of animal studies demonstrated beneficial effects of stem/progenitor cell transplantation for pulp–dentin complex regeneration, that is, pulpal tissue, neural, vascular, and dentinal regeneration. We systematically reviewed animal studies investigating stem/progenitor cell-mediated pulp–dentin complex regeneration. Studies quantitatively comparing pulp–dentin complex regeneration after transplantation of stem/progenitor cells versus no stem/progenitor cell transplantation controls in intraoral in vivo teeth animal models were analyzed. The following outcomes were investigated: regenerated pulp area per root canal total area, capillaries per total surface, regenerated dentinal area per total defect area, and nerves per total surface. PubMed and EMBASE were screened for studies published until July 2014. Cross-referencing and hand searching were used to identify further articles. Standardized mean differences (SMD) and 95% confidence intervals (95% CI) were calculated using random-effects meta-analysis. To assess possible bias, SYRCLE's risk of bias tool for animal studies was used. From 1364 screened articles, five studies (representing 64 animals) were included in the quantitative analysis. Risk of bias of all studies was high. Stem/progenitor cell-transplanted pulps showed significantly larger regenerated pulp area per root canal total area (SMD [95% CI]: 2.28 [0.35–4.21]) and regenerated dentin area per root canal total area (SMD: 6.91 [5.39–8.43]) compared with no stem/progenitor cell transplantation controls. Only one study reported on capillaries per or nerves per total surface and found both significantly increased in stem/progenitor cell-transplanted pulps compared with controls. Stem/progenitor cell transplantation seems to enhance pulp–dentin complex regeneration in

  20. Colorectal cancer stem cells.

    PubMed

    Salama, Paul; Platell, Cameron

    2009-10-01

    Somatic stem cells reside at the base of the crypts throughout the colonic mucosa. These cells are essential for the normal regeneration of the colonic epithelium. The stem cells reside within a special 'niche' comprised of intestinal sub-epithelial myofibroblasts that tightly control their function. It has been postulated that mutations within these adult colonic stem cells may induce neoplastic changes. Such cells can then dissociate from the epithelium and travel into the mesenchyme and thus form invasive cancers. This theory is based on the observation that within a colon cancer, less than 1% of the neoplastic cells have the ability to regenerate the tumour. It is this group of cells that exhibits characteristics of colonic stem cells. Although anti-neoplastic agents can induce remissions by inhibiting cell division, the stem cells appear to be remarkably resistant to both standard chemotherapy and radiotherapy. These stem cells may therefore persist after treatment and form the nucleus for cancer recurrence. Hence, future treatment modalities should focus specifically on controlling the cancer stem cells. In this review, we discuss the biology of normal and malignant colonic stem cells.

  1. Immunological characteristics of mesenchymal stem cells

    PubMed Central

    Machado, Cíntia de Vasconcellos; Telles, Paloma Dias da Silva; Nascimento, Ivana Lucia Oliveira

    2013-01-01

    Although bone marrow is the main source, mesenchymal stem cells have already been isolated from various other tissues, such as the liver, pancreas, adipose tissue, peripheral blood and dental pulp. These plastic adherent cells are morphologically similar to fibroblasts and have a high proliferative potential. This special group of cells possesses two essential characteristics: self-renewal and differentiation, with appropriate stimuli, into various cell types. Mesenchymal stem cells are considered immunologically privileged, since they do not express costimulatory molecules, required for complete T cell activation, on their surface. Several studies have shown that these cells exert an immunosuppressive effect on cells from both innate and acquired immunity systems. Mesenchymal stem cells can regulate the immune response in vitro by inhibiting the maturation of dendritic cells, as well as by suppressing the proliferation and function of T and B lymphocytes and natural killer cells. These special properties of mesenchymal stem cells make them a promising strategy in the treatment of immune mediated disorders, such as graft-versus-host disease and autoimmune diseases, as well as in regenerative medicine. The understanding of immune regulation mechanisms of mesenchymal stem cells, and also those involved in the differentiation of these cells in various lineages is primordial for their successful and safe application in different areas of medicine. PMID:23580887

  2. Establishment of in vitro culture system for evaluating dentin-pulp complex regeneration with special reference to the differentiation capacity of BrdU label-retaining dental pulp cells.

    PubMed

    Ida-Yonemochi, Hiroko; Nakatomi, Mitsushiro; Ohshima, Hayato

    2014-09-01

    We have proposed the new hypothesis that dental pulp stem cells play crucial roles in the pulpal healing process following exogenous stimuli in cooperation with progenitors. This study aimed to establish an in vitro culture system for evaluating dentin-pulp complex regeneration with special reference to the differentiation capacity of slow-cycling long-term label-retaining cells (LRCs). Three intraperitoneal injections of BrdU were given to pregnant ICR mice to map LRCs in the mature tissues of born animals. The upper bilateral first molars of 3-week-old mice were extracted and divided into two pieces and cultured for 0, 1, 3, 5 and 7 days using the Trowel's method. We succeeded in establishing an in vitro culture system for evaluating dentin-pulp complex regeneration, where most odontoblasts were occasionally degenerated and lost nestin immunoreactivity because of the separation of cell bodies from cellular processes in the dentin matrix by the beginning of in vitro culture. Numerous dense LRCs mainly resided in the center of the dental pulp associating with blood vessels throughout the experimental periods. On postoperative days 1-3, the periphery of the pulp tissue including the odontoblast layer showed degenerative features. By Day 7, nestin-positive odontoblast-like cells were arranged along the pulp-dentin border and dense LRCs were committed in the odontoblast-like cells. These results suggest that dense LRCs in the center of the dental pulp associating with blood vessels were supposed to be dental pulp stem/progenitor cells possessing regenerative capacity for forming newly differentiated odontoblast-like cells.

  3. Phenotypic and growth characterization of human mesenchymal stem cells cultured from permanent and deciduous teeth.

    PubMed

    Shekar, Revathi; Ranganathan, Kannan

    2012-01-01

    A step-by-step approach to harvesting stem cells from the pulp of permanent and deciduous teeth, the problems faced during culture, and the differences in the growth properties and morphology of cells obtained from both the sources has been discussed as a precursor to the use of these cells in therapy. To isolate, culture, and study the morphology and growth characteristics of mesenchymal stem cells from the dental pulp of permanent teeth (dental pulp stem cells; DPSC) and exfoliated deciduous teeth (stem cells from human exfoliated deciduous; SHED). Cell culture study carried out at the Department of Oral and Maxillofacial Pathology, Ragas Dental College and Hospital, Chennai. Fifteen permanent teeth and thirteen deciduous teeth from ten subjects were used. The growth characteristics and phenotype of cultured DPSCs and SHED were studied from the fourth passage on 24-well plates. Data analysis was done using SPSS ® version 10.05. Linear regression analysis was performed to derive the slope from growth curves and Mann-Whitney U test was performed to compare the fibroblastoid: epithelioid cell ratio between permanent and deciduous tooth pulp groups. Protocol for the culture of DPSC and SHED was standardized. DPSC and SHED populations were morphologically distinct. The cells from permanent tooth pulp showed a higher proportion of spindle-shaped fibroblastoid cells, whereas deciduous pulp culture showed a higher proportion of epithelioid cells. The seeding efficiency of DPSC - 88.9% (14 th permanent tooth pulp) and 91.7% (15 th permanent tooth pulp) was higher as compared to SHED - 84.25% (10 th deciduous tooth pulp). Permanent and deciduous teeth are both viable sources of stem cells. The permanent teeth were easier to culture because of a lower chance of contamination with oral microflora. The growth characteristics of the cells obtained from both these sources were similar. However, there was a difference in the ratio of fibroblastoid cells to epithelioid cells

  4. Intraoperative Stem Cell Therapy

    PubMed Central

    Coelho, Mónica Beato; Cabral, Joaquim M.S.; Karp, Jeffrey M.

    2013-01-01

    Stem cells hold significant promise for regeneration of tissue defects and disease-modifying therapies. Although numerous promising stem cell approaches are advancing in clinical trials, intraoperative stem cell therapies offer more immediate hope by integrating an autologous cell source with a well-established surgical intervention in a single procedure. Herein, the major developments in intraoperative stem cell approaches, from in vivo models to clinical studies, are reviewed, and the potential regenerative mechanisms and the roles of different cell populations in the regeneration process are discussed. Although intraoperative stem cell therapies have been shown to be safe and effective for several indications, there are still critical challenges to be tackled prior to adoption into the standard surgical armamentarium. PMID:22809140

  5. Isolated rat dental pulp cell culture and transplantation with an alginate scaffold.

    PubMed

    Fujiwara, Shiro; Kumabe, Shunji; Iwai, Yasutomo

    2006-05-01

    Many studies have been conducted on tissue stem cells in the field of regenerative medicine, and cultured dental pulp mesenchymal cells have been reported to secrete dentin matrix. In the present study we used alginate as a scaffold to transplant subcultured rat dental-pulp-derived cells subcutaneously into the back of nude mice. We found that when beta-glycerophosphate was added to the culture medium, the mRNA of the dentin sialophosphoprotein (DSPP) gene coding dentin sialoprotein (DSP) and dentin phosphoprotein (DPP) was expressed, and an increase in alkaline phosphatase, an early marker of odontoblast differentiation, was also demonstrated. Six weeks after implantation, subcutaneous formation of radiopaque calcified bodies was observed in situ. Immunohistochemical and fine structure studies identified expression of type I collagen, type III collagen, and DSP in the mineralizing transplants, and isolated odontoblast-like cells began to form dentin-like hard tissue formation. Scattered autolyzing apoptotic cells were also observed in the transplants. The study showed that subcultured rat dental-pulp-derived cells actively differentiate into odontoblast-like cells and induce calcification in an alginate scaffold.

  6. Myeloproliferative neoplasm stem cells.

    PubMed

    Mead, Adam J; Mullally, Ann

    2017-03-23

    Myeloproliferative neoplasms (MPNs) arise in the hematopoietic stem cell (HSC) compartment as a result of the acquisition of somatic mutations in a single HSC that provides a selective advantage to mutant HSC over normal HSC and promotes myeloid differentiation to engender a myeloproliferative phenotype. This population of somatically mutated HSC, which initiates and sustains MPNs, is termed MPN stem cells. In >95% of cases, mutations that drive the development of an MPN phenotype occur in a mutually exclusive manner in 1 of 3 genes: JAK2, CALR, or MPL The thrombopoietin receptor, MPL, is the key cytokine receptor in MPN development, and these mutations all activate MPL-JAK-STAT signaling in MPN stem cells. Despite common biological features, MPNs display diverse disease phenotypes as a result of both constitutional and acquired factors that influence MPN stem cells, and likely also as a result of heterogeneity in the HSC in which MPN-initiating mutations arise. As the MPN clone expands, it exerts cell-extrinsic effects on components of the bone marrow niche that can favor the survival and expansion of MPN stem cells over normal HSC, further sustaining and driving malignant hematopoiesis. Although developed as targeted therapies for MPNs, current JAK2 inhibitors do not preferentially target MPN stem cells, and as a result, rarely induce molecular remissions in MPN patients. As the understanding of the molecular mechanisms underlying the clonal dominance of MPN stem cells advances, this will help facilitate the development of therapies that preferentially target MPN stem cells over normal HSC.

  7. Fish Stem Cell Cultures

    PubMed Central

    Hong, Ni; Li, Zhendong; Hong, Yunhan

    2011-01-01

    Stem cells have the potential for self-renewal and differentiation. First stem cell cultures were derived 30 years ago from early developing mouse embryos. These are pluripotent embryonic stem (ES) cells. Efforts towards ES cell derivation have been attempted in other mammalian and non-mammalian species. Work with stem cell culture in fish started 20 years ago. Laboratory fish species, in particular zebrafish and medaka, have been the focus of research towards stem cell cultures. Medaka is the second organism that generated ES cells and the first that gave rise to a spermatogonial stem cell line capable of test-tube sperm production. Most recently, the first haploid stem cells capable of producing whole animals have also been generated from medaka. ES-like cells have been reported also in zebrafish and several marine species. Attempts for germline transmission of ES cell cultures and gene targeting have been reported in zebrafish. Recent years have witnessed the progress in markers and procedures for ES cell characterization. These include the identification of fish homologs/paralogs of mammalian pluripotency genes and parameters for optimal chimera formation. In addition, fish germ cell cultures and transplantation have attracted considerable interest for germline transmission and surrogate production. Haploid ES cell nuclear transfer has proven in medaka the feasibility of semi-cloning as a novel assisted reproductive technology. In this special issue on “Fish Stem Cells and Nuclear Transfer”, we will focus our review on medaka to illustrate the current status and perspective of fish stem cells in research and application. We will also mention semi-cloning as a new development to conventional nuclear transfer. PMID:21547056

  8. Analysis of side population cells derived from dental pulp tissue.

    PubMed

    Kenmotsu, M; Matsuzaka, K; Kokubu, E; Azuma, T; Inoue, T

    2010-12-01

    To investigate the characteristics of side population (SP) cells derived from the dental pulp of young and aged rats. Maxillary and mandibular incisors were extracted from 5-week-old (young) rats and 60- to 80-week-old (aged) rats. Coronal pulp tissue was removed mechanically, and single-cell suspensions were prepared using collagenase and dispase. Cells were stained with Hoechst 33342 and sorted with an fluorescence-activated cell sorter (FACS). Isolated SP and main population (MP) cells were analysed by real-time reverse transcription polymerase chain reaction, immunohistochemical localization and cell cycle determination. Two-way analysis of variance and the multiple comparison Scheffè test were used for statistical analysis (P<0.05). Approximately 0.40% of pulp cells in young rats and 0.11% in aged rats comprised SP cells. SP cells expressed a higher mRNA level of ATP-binding cassette transporter G2 (ABCG2), but lower mRNA levels of nestin, alkaline phosphatase, p16 and p57 than MP cells in both age groups. Immunohistochemical observation revealed ABCG2-positive cells localized in the cell-rich zone and nestin in the odontoblastic layer in both groups. Furthermore, the majority of both young and aged SP and MP cells were in growth arrest of the G(0) /G(1) phase. The FACS analysis revealed a decrease in the proportion of SP cells with age, whilst p16 mRNA expression indicated an increase in cell senescence. The cell cycles of SP and MP cells from both young and aged dental pulp were generally in the G0/G1 phase. © 2010 International Endodontic Journal.

  9. Application of Stem Cell Technology in Dental Regenerative Medicine

    PubMed Central

    Feng, Ruoxue; Lengner, Chistopher

    2013-01-01

    Significance In this review, we summarize the current literature regarding the isolation and characterization of dental tissue-derived stem cells and address the potential of these cell types for use in regenerative cell transplantation therapy. Recent Advances Looking forward, platforms for the delivery of stem cells via scaffolds and the use of growth factors and cytokines for enhancing dental stem cell self-renewal and differentiation are discussed. Critical Issues We aim to understand the developmental origins of dental tissues in an effort to elucidate the molecular pathways governing the genesis of somatic dental stem cells. The advantages and disadvantages of several dental stem cells are discussed, including the developmental stage and specific locations from which these cells can be purified. In particular, stem cells from human exfoliated deciduous teeth may act as a very practical and easily accessibly reservoir for autologous stem cells and hold the most value in stem cell therapy. Dental pulp stem cells and periodontal ligament stem cells should also be considered for their triple lineage differentiation ability and relative ease of isolation. Further, we address the potentials and limitations of induced pluripotent stem cells as a cell source in dental regenerative. Future Directions From an economical and a practical standpoint, dental stem cell therapy would be most easily applied in the prevention of periodontal ligament detachment and bone atrophy, as well as in the regeneration of dentin-pulp complex. In contrast, cell-based tooth replacement due to decay or other oral pathology seems, at the current time, an untenable approach. PMID:24527351

  10. Regenerative Endodontic Procedures: A Perspective from Stem Cell Niche Biology.

    PubMed

    Marí-Beffa, Manuel; Segura-Egea, Juan José; Díaz-Cuenca, Aránzazu

    2017-01-01

    Endodontics uses cell therapy strategies to treat pulpal and periapical diseases. During these therapies, surgeons aim to reconstruct the natural microenvironments that regulate the activity of dental stem cells. We searched for more than 400 articles in PubMed using key words from regenerative endodontics and dental stem cell biology. In 268 articles, we reviewed what factors may influence histologic results after preclinical dental treatments that use regenerative endodontic procedures after pulpectomy. Several factors, such as the origin of stem cells, the biomimicry of scaffolds used, and the size of lesions, are considered to influence the histologic appearance of the regenerated pulp-dentin complex after treatments. Information is accumulating on transcription factors that generate the pulp-dentin complex and survival/trophic factors that would benefit niche recovery and histologic results. In this article, we discuss the noninterchangeability of stem cells, the influence of dentin-entrapped molecule release on pulp regeneration and survival of stem cells, and the need of positional markers to assess treatments histologically. The ex vivo amplification of appropriate dental stem cells, the search for scaffolds storing the molecular diversity entrapped in the dentin, and the use of positional transcription factors as histologic markers are necessary to improve future preclinical experiments. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  11. Stem Cell Transplants (For Teens)

    MedlinePlus

    ... Loss Surgery? A Week of Healthy Breakfasts Shyness Stem Cell Transplants KidsHealth > For Teens > Stem Cell Transplants Print ... Does it Take to Recover? Coping What Are Stem Cells? As you probably remember from biology class, every ...

  12. Information on Stem Cell Research

    MedlinePlus

    ... Home » Current Research » Focus on Research Focus on Stem Cell Research Stem cells possess the unique ability to differentiate ... virus infection. To search the complete list of stem cell research projects funded by NIH please go to NIH ...

  13. Stem Cell Transplants (For Teens)

    MedlinePlus

    ... Loss Surgery? A Week of Healthy Breakfasts Shyness Stem Cell Transplants KidsHealth > For Teens > Stem Cell Transplants A ... Does it Take to Recover? Coping What Are Stem Cells? As you probably remember from biology class, every ...

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

    PubMed

    Hayashi, Y; Murakami, M; Kawamura, R; Ishizaka, R; Fukuta, O; Nakashima, M

    2015-05-29

    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. 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. 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

  15. Hematopoietic Stem Cells Therapies.

    PubMed

    Chivu-Economescu, Mihaela; Rubach, Martin

    2017-01-01

    Stem cell-based therapies are recognized as a new way to treat various diseases and injuries, with a wide range of health benefits. The goal is to heal or replace diseased or destroyed organs or body parts with healthy new cells provided by stem cell transplantation. The current practical form of stem cell therapy is the hematopoietic stem cells transplant applied for the treatment of hematological disorders. There are over 2100 clinical studies in progress concerning hematopoietic stem cell therapies. All of them are using hematopoietic stem cells to treat various diseases like: cancers, leukemia, lymphoma, cardiac failure, neural disorders, auto-immune diseases, immunodeficiency, metabolic or genetic disorders. Several challenges are to be addressed prior to developing and applying large scale cell therapies: 1) to explain and control the mechanisms of differentiation and development toward a specific cell type needed to treat the disease, 2) to obtain a sufficient number of desired cell type for transplantation, 3) to overcome the immune rejection and 4) to show that transplanted cells fulfill their normal functions in vivo after transplants.

  16. Stem cells-the hidden treasure: A strategic review.

    PubMed

    Chopra, Hitesh; Hans, Manoj Kumar; Shetty, Shashit

    2013-07-01

    In today's scenario, medical and dental professionals face a mammoth task while treating perplexing medical situations like organ failure or tissue loss. Though, different strategies exist to replace them, but ideal one is the same natural tissue or organ. In this aspect, stem cells have emerged in a promising way to provide an ideal replacement. There are different types of stem cells starting from the embryonic stage referred to as human embryonic stem cells to adult stem cells. Though in dentistry stem cell research is lagging as compared to the medical field but still a lot progress has been achieved in recent years. The stem cells have been isolated from dental pulp, human exfoliated deciduous teeth, and apical papilla and so on. These stem cells have provided exciting results like dentin-pulp regeneration, periodontal regeneration but ambiguity still prevails. As a result, much has to be further researched before its clinical application becomes a reality. Hence, these stem cells opened a new avenue in the field of regenerative dentistry.

  17. Stem Cell Organoid Engineering

    PubMed Central

    Yin, Xiaolei; Mead, Benjamin E.; Safaee, Helia; Langer, Robert; Karp, Jeffrey M.; Levy, Oren

    2016-01-01

    Organoid systems leverage the self-organizing properties of stem cells to create diverse multi-cellular tissue proxies. Most organoid models only represent single or partial components of a tissue, and it is often difficult to control the cell type, organization, and cell-cell/cell-matrix interactions within these systems. Herein, we discuss basic approaches to generate stem cell-based organoids, their advantages and limitations, and how bioengineering strategies can be used to steer the cell composition and their 3D organization within organoids to further enhance their utility in research and therapies. PMID:26748754

  18. Engineering Stem Cell Organoids.

    PubMed

    Yin, Xiaolei; Mead, Benjamin E; Safaee, Helia; Langer, Robert; Karp, Jeffrey M; Levy, Oren

    2016-01-07

    Organoid systems leverage the self-organizing properties of stem cells to create diverse multi-cellular tissue proxies. Most organoid models only represent single or partial components of a tissue, and it is often difficult to control the cell type, organization, and cell-cell/cell-matrix interactions within these systems. Herein, we discuss basic approaches to generate stem cell-based organoids, their advantages and limitations, and how bioengineering strategies can be used to steer the cell composition and their 3D organization within organoids to further enhance their utility in research and therapies.

  19. [On plant stem cells and animal stem cells].

    PubMed

    You, Yun; Jiang, Chao; Huang, Lu-Qi

    2014-01-01

    A comparison of plant and animal stem cells can highlight core aspects of stem-cell biology. In both kingdoms, stem cells are defined by their clonogenic properties and are maintained by intercellular signals. The signaling molecules are different in plants and animals stem cell niches, but the roles of argonaute and polycomb group proteins suggest that there are some molecular similarities.

  20. Manufacturing of dental pulp cell-based products from human third molars: current strategies and future investigations.

    PubMed

    Ducret, Maxime; Fabre, Hugo; Degoul, Olivier; Atzeni, Gianluigi; McGuckin, Colin; Forraz, Nico; Alliot-Licht, Brigitte; Mallein-Gerin, Frédéric; Perrier-Groult, Emeline; Farges, Jean-Christophe

    2015-01-01

    In recent years, mesenchymal cell-based products have been developed to improve surgical therapies aimed at repairing human tissues. In this context, the tooth has recently emerged as a valuable source of stem/progenitor cells for regenerating orofacial tissues, with easy access to pulp tissue and high differentiation potential of dental pulp mesenchymal cells. International guidelines now recommend the use of standardized procedures for cell isolation, storage and expansion in culture to ensure optimal reproducibility, efficacy and safety when cells are used for clinical application. However, most dental pulp cell-based medicinal products manufacturing procedures may not be fully satisfactory since they could alter the cells biological properties and the quality of derived products. Cell isolation, enrichment and cryopreservation procedures combined to long-term expansion in culture media containing xeno- and allogeneic components are known to affect cell phenotype, viability, proliferation and differentiation capacities. This article focuses on current manufacturing strategies of dental pulp cell-based medicinal products and proposes a new protocol to improve efficiency, reproducibility and safety of these strategies.

  1. Manufacturing of dental pulp cell-based products from human third molars: current strategies and future investigations

    PubMed Central

    Ducret, Maxime; Fabre, Hugo; Degoul, Olivier; Atzeni, Gianluigi; McGuckin, Colin; Forraz, Nico; Alliot-Licht, Brigitte; Mallein-Gerin, Frédéric; Perrier-Groult, Emeline; Farges, Jean-Christophe

    2015-01-01

    In recent years, mesenchymal cell-based products have been developed to improve surgical therapies aimed at repairing human tissues. In this context, the tooth has recently emerged as a valuable source of stem/progenitor cells for regenerating orofacial tissues, with easy access to pulp tissue and high differentiation potential of dental pulp mesenchymal cells. International guidelines now recommend the use of standardized procedures for cell isolation, storage and expansion in culture to ensure optimal reproducibility, efficacy and safety when cells are used for clinical application. However, most dental pulp cell-based medicinal products manufacturing procedures may not be fully satisfactory since they could alter the cells biological properties and the quality of derived products. Cell isolation, enrichment and cryopreservation procedures combined to long-term expansion in culture media containing xeno- and allogeneic components are known to affect cell phenotype, viability, proliferation and differentiation capacities. This article focuses on current manufacturing strategies of dental pulp cell-based medicinal products and proposes a new protocol to improve efficiency, reproducibility and safety of these strategies. PMID:26300779

  2. Fibroblast growth factor-2 regulates the cell function of human dental pulp cells.

    PubMed

    Shimabukuro, Yoshio; Ueda, Maki; Ozasa, Masao; Anzai, Jun; Takedachi, Masahide; Yanagita, Manabu; Ito, Masako; Hashikawa, Tomoko; Yamada, Satoru; Murakami, Shinya

    2009-11-01

    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). 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. 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. These results provide new evidence for the possible involvement of FGF-2 not only in homeostasis but also in regeneration of dentin-pulp complex.

  3. Mesenchymal dental pulp cells attenuate dentin resorption in homeostasis.

    PubMed

    Zheng, Y; Chen, M; He, L; Marão, H F; Sun, D M; Zhou, J; Kim, S G; Song, S; Wang, S L; Mao, J J

    2015-06-01

    Dentin in permanent teeth rarely undergoes resorption in development, homeostasis, or aging, in contrast to bone that undergoes periodic resorption/remodeling. The authors hypothesized that cells in the mesenchymal compartment of dental pulp attenuate osteoclastogenesis. Mononucleated and adherent cells from donor-matched rat dental pulp (dental pulp cells [DPCs]) and alveolar bone (alveolar bone cells [ABCs]) were isolated and separately cocultured with primary rat splenocytes. Primary splenocytes readily aggregated and formed osteoclast-like cells in chemically defined osteoclastogenesis medium with 20 ng/mL of macrophage colony-stimulating factor (M-CSF) and 50 ng/mL of receptor activator of nuclear factor κB ligand (RANKL). Strikingly, DPCs attenuated osteoclastogenesis when cocultured with primary splenocytes, whereas ABCs slightly but significantly promoted osteoclastogenesis. DPCs yielded ~20-fold lower RANKL expression but >2-fold higher osteoprotegerin (OPG) expression than donor-matched ABCs, yielding a RANKL/OPG ratio of 41:1 (ABCs:DPCs). Vitamin D3 significantly promoted RANKL expression in ABCs and OPG in DPCs. In vivo, rat maxillary incisors were atraumatically extracted (without any tooth fractures), followed by retrograde pulpectomy to remove DPCs and immediate replantation into the extraction sockets to allow repopulation of the surgically treated root canal with periodontal and alveolar bone-derived cells. After 8 wk, multiple dentin/root resorption lacunae were present in root dentin with robust RANKL and OPG expression. There were areas of dentin resoprtion alternating with areas of osteodentin formation in root dentin surface in the observed 8 wk. These findings suggest that DPCs of the mesenchymal compartment have an innate ability to attenuate osteoclastogenesis and that this innate ability may be responsible for the absence of dentin resorption in homeostasis. Mesenchymal attenuation of dentin resorption may have implications in internal

  4. Mesenchymal Dental Pulp Cells Attenuate Dentin Resorption in Homeostasis

    PubMed Central

    Zheng, Y.; Chen, M.; He, L.; Marão, H.F.; Sun, D.M.; Zhou, J.; Kim, S.G.; Song, S.; Wang, S.L.

    2015-01-01

    Dentin in permanent teeth rarely undergoes resorption in development, homeostasis, or aging, in contrast to bone that undergoes periodic resorption/remodeling. The authors hypothesized that cells in the mesenchymal compartment of dental pulp attenuate osteoclastogenesis. Mononucleated and adherent cells from donor-matched rat dental pulp (dental pulp cells [DPCs]) and alveolar bone (alveolar bone cells [ABCs]) were isolated and separately cocultured with primary rat splenocytes. Primary splenocytes readily aggregated and formed osteoclast-like cells in chemically defined osteoclastogenesis medium with 20 ng/mL of macrophage colony-stimulating factor (M-CSF) and 50 ng/mL of receptor activator of nuclear factor κB ligand (RANKL). Strikingly, DPCs attenuated osteoclastogenesis when cocultured with primary splenocytes, whereas ABCs slightly but significantly promoted osteoclastogenesis. DPCs yielded ~20-fold lower RANKL expression but >2-fold higher osteoprotegerin (OPG) expression than donor-matched ABCs, yielding a RANKL/OPG ratio of 41:1 (ABCs:DPCs). Vitamin D3 significantly promoted RANKL expression in ABCs and OPG in DPCs. In vivo, rat maxillary incisors were atraumatically extracted (without any tooth fractures), followed by retrograde pulpectomy to remove DPCs and immediate replantation into the extraction sockets to allow repopulation of the surgically treated root canal with periodontal and alveolar bone–derived cells. After 8 wk, multiple dentin/root resorption lacunae were present in root dentin with robust RANKL and OPG expression. There were areas of dentin resoprtion alternating with areas of osteodentin formation in root dentin surface in the observed 8 wk. These findings suggest that DPCs of the mesenchymal compartment have an innate ability to attenuate osteoclastogenesis and that this innate ability may be responsible for the absence of dentin resorption in homeostasis. Mesenchymal attenuation of dentin resorption may have implications in internal

  5. Melanoma stem cells.

    PubMed

    Roesch, Alexander

    2015-02-01

    The cancer stem cell concept significantly broadens our understanding of melanoma biology. However, this concept should be regarded as an integral part of a holistic cancer model that also includes the genetic evolution of tumor cells and the variability of cell phenotypes within a dynamic tumor microenvironment. The biologic complexity and methodological difficulties in identifying cancer stem cells and their biomarkers are currently impeding the direct translation of experimental findings into clinical practice. Nevertheless, it is these methodological shortcomings that provide a new perspective on the phenotypic heterogeneity and plasticity of melanoma with important consequences for future therapies. The development of new combination treatment strategies, particularly with regard to overcoming treatment resistance, could significantly benefit from targeted elimination of cell subpopulations with cancer stem cell properties. © 2015 Deutsche Dermatologische Gesellschaft (DDG). Published by John Wiley & Sons Ltd.

  6. Odontoblastic inductive potential of epithelial cells derived from human deciduous dental pulp.

    PubMed

    Lee, Hye-Kyung; Park, Ji-Won; Seo, You-Mi; Kim, Ha Hoon; Lee, Gene; Bae, Hyun-Sook; Park, Joo-Cheol

    2016-06-01

    For the dentin regeneration, dental epithelial cells are indispensible and must possess odontoblastic induction capability. Epithelial cell-like stem cells were recently identified in human deciduous dental pulp (DPESCs). However, their cellular characteristics remain poorly defined. The purpose of this study was to characterize DPESCs compared to HAT-7 ameloblastic cells. Expression levels of ameloblast-specific markers [odontogenic ameloblast-associated protein (Odam), matrix metalloproteinase (Mmp)-20, amelogenin, and ameloblastin] were detected in DPESCs. Co-culturing odontoblastic MDPC-23 cells with DPESCs increased expression of odontoblast differentiation markers (Dmp1 and Dspp) from days 4 to 10, while the expression of bone sialoprotein rapidly decreased. MDPC-23 cells cultured in DPESC-conditioned medium (CM) showed increased Dspp promoter activity compared with control MDPC-23 cultures. Mineralization was first observed in the CM groups from day 4 and proceeded rapidly until day 14, whereas mineralized nodules were found from day 7 in control media-cultured cells. In conclusion, DPESCs in human deciduous pulp possess ameloblast-like characteristics and differentiation properties, and substances derived from DPESCs promote odontoblastic differentiation. Thus, our results indicate that DPESCs can be a realistic epithelial source for