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Células Madre Dentales ARTÍCULOS CIENTÍFICOS Junio 2011 Para solicitar los artículos completos en PDF:

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Presentación del tema: "Células Madre Dentales ARTÍCULOS CIENTÍFICOS Junio 2011 Para solicitar los artículos completos en PDF:"— Transcripción de la presentación:

1 Células Madre Dentales ARTÍCULOS CIENTÍFICOS Junio 2011 Para solicitar los artículos completos en PDF:

2 - A continuación se presentan algunos ARTÍCULOS CIENTÍFICOS sobre el potencial terapéutico de las células madre dentales. Los temas a encontrar en cada diapositiva, son: TÍTULO y AUTORES JOURNAL ABSTRACT RESUMEN TRATAMIENTO POTENCIAL NAVEGACIÓN

3 1. En el Menú de Temas, da Click en el tema de tu elección. Ejemplo: 2. Para seguir navegando utiliza los siguientes botones: Instrucciones: Regenaración de Piel ----Se recomienda visualizarlo en Pantalla Completa----

4 Células Madre Mesenquimales de Origen Dental Regeneración Dental Regeneración Huesos y Estructura Craneofacial Regeneración Cardiaca Problemas Neurológicos Regeneración de Córnea Regenaración de Piel Diabetes Regeneración Hepática Regeneración Muscular y Cirugía Plástica Enfermedades Auto-Inmunes y Lupus Otros Artículos

5 Células Madre Mesenquimales de Origen Dental Resumen del Tema: Los dientes primarios y algunos dientes permanentes son una fuente accesible de células madre multipotenciales que se pueden diferenciar en distintos tipos de células como células nerviosas, adipocitos, osteoblastos, condrocitos, miocitos y odontoblastos. Las células madre dentales se pueden multiplicar in-vitro varias generaciones y se pueden obtener trillones de células para su utlización terapéutica. De la misma forma éstas células se pueden crío-preservar sin perder su multipotencialidad. Debido a su accesibilidad y sus altas tasas de proliferación celular son candidatos idóneos para la ingeniería de tejidos y tratamientos regenerativos.

6 TÍTULO JOURNAL ABSTRACT RESUMEN Los dientes primarios son una fuente accesible de células madre multipotenciales que se pueden diferenciar en distintos tipos de células como células nerviosas, adipocitos, osteoblastos y odontoblastos. Debido a su accesibilidad y sus altas tasas de proliferación celular son candidatos idóneos para la ingeniería de tejidos y transplantes autólogos. Células Madre Mesenquimales de Origen Dental TRATAMIENTO POTENCIAL SHED: stem cells from human exfoliated deciduous teeth. Miura MMiura M, Gronthos S, Zhao M, Lu B, Fisher LW, Robey PG, Shi S.s S, Zhao her LWPG, Proc Natl Acad Sci U S A. 2003 May 13;100(10):5807-12. Epub 2003 Apr 25. To isolate high-quality human postnatal stem cells from accessible resources is an important goal for stem-cell research. In this study we found that exfoliated human deciduous tooth contains multipotent stem cells [stem cells from human exfoliated deciduous teeth (SHED)]. SHED were identified to be a population of highly proliferative, clonogenic cells capable of differentiating into a variety of cell types including neural cells, adipocytes, and odontoblasts. After in vivo transplantation, SHED were found to be able to induce bone formation, generate dentin, and survive in mouse brain along with expression of neural markers. Here we show that a naturally exfoliated human organ contains a population of stem cells that are completely different from previously identified stem cells. SHED are not only derived from a very accessible tissue resource but are also capable of providing enough cells for potential clinical application. Thus, exfoliated teeth may be an unexpected unique resource for stem-cell therapies including autologous stem-cell transplantation and tissue engineering.

7 TÍTULO ABSTRACT RESUMEN Las células madre de los dientes primarios son accesibles y fáciles de obtener, considerando que la dentición primaria se exfolia naturalmente. La crío-preservación de estás células madre es recomendable puesto que las mismas pueden ser utilizadas por el paciente, sin problemas de rechazo, en el tratamiento de varias enfermedades. Células Madre Mesenquimales de Origen Dental Banking stem cells from human exfoliated deciduous teeth (SHED): saving for the future. Arora VArora V, Arora P, Munshi AK., Munsh J Clin Pediatr Dent. 2009 Summer;33(4):289-94. Tooth derived cells are readily accessible and provide an easy and minimally invasive way to obtain and store stem cells for future use. Banking ones own tooth-derived stem cells is a reasonable and simple alternative to harvesting stem cells from other tissues. Obtaining stem cells from human exfoliated deciduous teeth (SHED) is simple and convenient, with little or no trauma. Every child loses primary teeth, which creates the perfect opportunity to recover and store this convenient source of stem cells--should they be needed to treat future injuries or ailments and presents a far better alternative to simply discarding the teeth or storing them as mementos from the past. Furthermore, using ones own stem cells poses few, if any, risks for developing immune reactions or rejection following transplantation and also eliminates the potential of contracting disease from donor cells. Stem cells can also be recovered from developing wisdom teeth and permanent teeth. Individuals have different opportunities at different stages of their life to bank these valuable cells. It is best to recover stem cells when a child is young and healthy and the cells are strong and proliferative. The purpose of this review is to discuss the present scenario as well as the technical details of tooth banking as related to SHED cells. JOURNAL TRATAMIENTO POTENCIAL

8 TÍTULO ABSTRACT RESUMEN Las células madre dentales se pueden diferenciar en tejido osteogénico, condrogénico, adipogénico, miogénico y neurogénico. También se comparan las células madre provenientes del tejido periodontal y de la papila apical, y del folículo dental. Mesenchymal stem cells derived from dental tissues vs. those from other sources: their biology and role in regenerative medicine. Huang GTHuang GT, Gronthos S, Shi S. S, Shi S. J Dent Res. 2009 Sep;88(9):792-806. To date, 5 different human dental stem/progenitor cells have been isolated and characterized: dental pulp stem cells (DPSCs), stem cells from exfoliated deciduous teeth (SHED), periodontal ligament stem cells (PDLSCs), stem cells from apical papilla (SCAP), and dental follicle progenitor cells (DFPCs). These postnatal populations have mesenchymal-stem-cell-like (MSC) qualities, including the capacity for self-renewal and multilineage differentiation potential. MSCs derived from bone marrow (BMMSCs) are capable of giving rise to various lineages of cells, such as osteogenic, chondrogenic, adipogenic, myogenic, and neurogenic cells. The dental-tissue-derived stem cells are isolated from specialized tissue with potent capacities to differentiate into odontogenic cells. However, they also have the ability to give rise to other cell lineages similar to, but different in potency from, that of BMMSCs. This article will review the isolation and characterization of the properties of different dental MSC-like populations in comparison with those of other MSCs, such as BMMSCs. Important issues in stem cell biology, such as stem cell niche, homing, and immunoregulation, will also be discussed. Células Madre Mesenquimales de Origen Dental TRATAMIENTO POTENCIAL JOURNAL

9 TÍTULO ABSTRACT RESUMEN Las células madre de las muelas del juicio se pueden reprogramar para convertirse en células madre pluripotentes obteniendo las mismas características de las células madre embrionarias. Células Madre Mesenquimales de Origen Dental The expression of four transcription factors (OCT3/4, SOX2, KLF4, and MYC) can reprogram mouse as well as human somatic cells to induced pluripotent stem (iPS) cells. We generated iPS cells from mesenchymal stromal cells (MSCs) derived from human third molars (wisdom teeth) by retroviral transduction of OCT3/4, SOX2, and KLF4 without MYC, which is considered as oncogene. Interestingly, some of the clonally expanded MSCs could be used for iPS cell generation with 30-100- fold higher efficiency when compared with that of other clonally expanded MSCs and human dermal fibroblasts. Global gene expression profiles demonstrated some up-regulated genes regarding DNA repair/histone conformational change in the efficient clones, suggesting that the processes of chromatin remodeling have important roles in the cascade of iPS cells generation. The generated iPS cells resembled human embryonic stem (ES) cells in many aspects, including morphology, ES marker expression, global gene expression, epigenetic states, and the ability to differentiate into the three germ layers in vitro and in vivo. Because human third molars are discarded as clinical waste, our data indicate that clonally expanded MSCs derived from human third molars are a valuable cell source for the generation of iPS cells. Induction of pluripotent stem cells from human third molar mesenchymal stromal cells. Oda YOda Y, Yoshimura Y, Ohnishi H, Tadokoro M, Katsube Y, Sasao M, Kubo Y, Hattori K, Saito S, Horimoto K, Yuba S, Ohgushi H.mura Y, Ohnoro M, Kasao M, Kub K, Saitoo K, Yuushi H J Biol Chem. 2010 Sep 17;285(38):29270-8. Epub 2010 Jul 1. TRATAMIENTO POTENCIAL JOURNAL

10 TÍTULO ABSTRACT RESUMEN Las células madre dentales pueden convertirse en odontoblastos, osteoblastos y condrocitos. Se analiza esta capacidad de multidiferenciación en distintos pases celulares. Células Madre Mesenquimales de Origen Dental Dental pulp stem cells (DPSCs) can be driven into odontoblast, osteoblast, and chondrocyte lineages in different inductive media. However, the differentiation potential of naive DPSCs after serial passaging in the routine culture system has not been fully elucidated. DPSCs were isolated from human/rat dental pulps by the magnetic activated cell sorting based on STRO-1 expression, cultured and passaged in the conventional culture media. The biological features of STRO-1+ DPSCs at the 1st and 9th passages were investigated. During the long-term passage, the proliferation ability of human STRO-1+ DPSCs was downregulated as indicated by the growth kinetics. When compared with STRO-1+ DPSCs at the 1st passage (DPSC- P1), the expression of mature osteoblast-specific genes/proteins (alkaline phosphatase, bone sialoprotein, osterix, and osteopontin), odontoblast-specific gene/protein (dentin sialophosphoprotein and dentin sialoprotein), and chondrocyte- specific gene/protein (type II collagen) was significantly upregulated in human STRO-1+ DPSCs at the 9th passage (DPSC-P9). Furthermore, human DPSC-P9 cells in the mineralization-inducing media presented higher levels of alkaline phosphatase at day 3 and day 7 respectively, and produced more mineralized matrix than DPSC-P9 cells at day 14. In vivo transplantation results showed that rat DPSC-P1 cell pellets developed into dentin, bone and cartilage structures respectively, while DPSC-P9 cells can only generate bone tissues. These findings suggest that STRO-1+ DPSCs consist of several interrelated subpopulations which can spontaneously differentiate into odontoblasts, osteoblasts, and chondrocytes. The differentiation capacity of these DPSCs changes during cell passaging, and DPSCs at the 9th passage restrict their differentiation potential to the osteoblast lineage in vivo. Differentiation potential of STRO-1+ dental pulp stem cells changes during cell passaging. Yu JYu J, He H, Tang C, Zhang G, Li Y, Wang R, Shi J, Jin Y., TaZhang Wang RJin BMC Cell Biol. 2010 May 8;11:32. TRATAMIENTO POTENCIAL JOURNAL

11 TÍTULO ABSTRACT RESUMEN Las células madre dentales pueden convertirse en dentina y en tejido óseo. Con la utilización de andamios en 3D se observa que éstas células pueden ser utilizadas exitosamente en la ingeniería de tejidos. Células Madre Mesenquimales de Origen Dental Human dental pulp stem cells: from biology to clinical applications. d'Aquino Rd'Aquino R, De Rosa A, Laino G, Caruso F, Guida L, Rullo R, Checchi V, Laino L, Tirino V, Papaccio G. Laino G,Guida L, Checch L, Tirpaccio J Exp Zool B Mol Dev Evol. 2009 Jul 15;312B(5):408-15. 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. TRATAMIENTO POTENCIAL JOURNAL

12 TÍTULO ABSTRACT RESUMEN Las células madre dentales pueden convertirse en distintos tipos de tejidos y pueden ser una fuente idónea de células para la ingeniería de tejidos. Los autores recomiendan mayor investigación en el tema. Células Madre Mesenquimales de Origen Dental In vivo evaluation of human dental pulp stem cells differentiated towards multiple lineages. Zhang WZhang W, Walboomers XF, Van Kuppevelt TH, Daamen WF, Van Damme PA, Bian Z, Jansen JA.ers XF, Van Kamen WF, Van Damsen JA. J Tissue Eng Regen Med. 2008 Mar-Apr;2(2-3):117-25. An increasing number of investigations supports that adult stem cells have the potential to differentiate into matured cell types beyond their origin, a property defined as plasticity. Previously, the plasticity of stem cells derived from dental pulp (DPSC) has been confirmed by culturing cells in lineage-specific media in vitro. In the current study, the in vivo differentiation or maturation potential of DPSC was further analysed, by transplanting human DPSC/collagen scaffold constructs into subcutaneous tissue of immunocompromised mice. Cells received odontogenic, adipogenic or myogenic pre-induction, whereas control samples received no stimulation. Also blank collagen scaffolds were implanted. The results indicated that seeded cells produced tissue within the implanted constructs after 3 weeks of implantation. According to morphological and phenotypical changes, the pre-induced DPSC showed the ability to further differentiate along odontogenic, myogenic and adipogenic pathways in vivo. Moreover, DPSC without pre-treatment were able to spontaneously differentiate along odontogenic and adipogenic directions in vivo. However, only limited mature morphological changes were detected in histology. In summary, stem cells derived from human dental pulp form a suitable source for tissue engineering and cell-mediated therapy, although additional analyses should be considered. TRATAMIENTO POTENCIAL JOURNAL

13 TÍTULO ABSTRACT RESUMEN Las células madre dentales están en investigación para tratar el Parkinson, degeneración neural, problemas cardiovasculares y enfermedades auto-inmunes. Éstas Células podrán ser utilizadas en odontología para reemplazar tejido craneofacial, tratar glándulas salivales defectuosas entre otras. En general se repasan los beneficios de la preservación de las células madre dentales y el papel que juegan los dentistas en este proceso. Stem Cells: Sources, Therapies and The Dental Professional Células Madre Mesenquimales de Origen Dental TRATAMIENTO POTENCIAL JOURNAL

14 TÍTULO ABSTRACT RESUMEN Las células madre dentales pueden diferenciarse en osteoblastos, condrocitos y adipositos. La manera de obtenerlas es muy sencilla y además son capaces de reproducirse para proveer las cantidades celulares necesarias para aplicaciones clínicas. Células Madre Mesenquimales de Origen Dental Evaluation of pluripotency in human dental pulp cells. Koyama NKoyama N, Okubo Y, Nakao K, Bessho K. Nakao K. J Oral Maxillofac Surg. 2009 Mar;67(3):501-6. Postnatal stem cells have been isolated from various tissues, including bone marrow, neural tissue, skin, retina, and dental epithelium. Recently, adult stem cells have been isolated from human dental pulp. Postnatal stem cells have been isolated from a variety of tissues. Previously, it was generally accepted that the differentiation potential of postnatal stem cells was lineage restricted. Normal impacted third molars were collected from adults and normal exfoliated deciduous teeth (SHED; stem cells from human exfoliated deciduous teeth) by single-colony selection and magnetic activated cell sorting. BMP-2 treatment groups produced alkaline phosphatase in the cells and also produced and secreted osteocalcin in the culture medium, and were capable of inducing an upregulated expression of Osteocalcin or Sox9, Col 2, and Col X by reverse transcriptase polymerase chain reaction (RT-PCR). For adipogenic differentiation, there is potential for SHED and dental pulp stem cells (DPSC) to express 2 adipocyte- specific transcripts, PPARgamma2 and LPL, in vitro, as do bone marrow mesenchymal stem cells by RT- PCR. This study demonstrated that pluripotential cells isolated from the pulp of human teeth expanded in vitro and differentiated into osteoblasts, chondrocytes, and adipocytes. DPSC and SHED are not only derived from a very accessible tissue resource but also capable of providing enough cells for potential clinical applications. TRATAMIENTO POTENCIAL JOURNAL

15 TÍTULO ABSTRACT RESUMEN Las células madre dentales fueron crío-preservadas y después se les hicieron pruebas de multipotencialidad para verificar si seguían teniendo la posibilidad de diferenciarse en distintos tipos de tejidos. El experimento demostró que aún despues de la crío-preservación las células madre dentales seguían manteniendo su multi-potencialidad. Células Madre Mesenquimales de Origen Dental Multilineage differentiation potential of stem cells derived from human dental pulp after cryopreservation. Zhang WZhang W, Walboomers XF, Shi S, Fan M, Jansen JA.ers XF, Shi S JA. Tissue Eng. 2006 Oct;12(10):2813-23. The current study aimed to prove that human dental pulp stem cells (hDPSCs) isolated from the pulp of third molars can show multilineage differentiation after cryopreservation. First, hDPSC were isolated via enzymatic procedures, and frozen in liquid nitrogen until use. After defrosting, cells were analyzed for proliferative potential and the expression of the stem cell marker STRO-1. Subsequently, cells were cultured in neurogenic, osteogenic/odontogenic, adipogenic, myogenic, and chondrogenic inductive media, and analyzed on basis of morphology, immunohistochemistry, and reverse transcriptase-polymerase chain reaction (RT-PCR) for specific marker genes. All data were replicated, and the results of the primary cells were compared to similar tests with an additional primary dental pulp stem cell strain, obtained from the National Institutes of Health (NIH). Results showed that our cell population could be maintained for at least 25 passages. The existence of stem/ progenitor cells in both cell strains was proven by the STRO-1 staining. Under the influence of the 5 different media, both cell strains were capable to advance into all 5 differentiation pathways. Still differences between both strains were found. In general, our primary culture performed better in myogenic differentiation, while the externally obtained cells were superior in the odontogenic/osteogenic and chondrogenic differentiation pathways. In conclusion, the pulp tissue of the third molar may serve as a suitable source of multipotent stem cells for future tissue engineering strategies and cell-based therapies, even after cryopreservation. TRATAMIENTO POTENCIAL JOURNAL

16 TÍTULO ABSTRACT RESUMEN Se cultivaron células madre dentales y se lograron más de 40 dupliaciones de población celular sin afectar la viabilidad celular ni las propiedades biológicas y fenotípicas de las células madre. Células Madre Mesenquimales de Origen Dental TRATAMIENTO POTENCIAL JOURNAL

17 TÍTULO ABSTRACT RESUMEN Optimized cryopreservation method for human dental pulp- derived stem cells and their tissues of origin for banking and clinical use. Woods EJWoods EJ, Perry BC, Hockema JJ, Larson L, Zhou D, Goebel WS., Hockemon L, ZhouS. 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 120h 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 Me(2)SO 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 2x10(6) cells/mL. It was further established that DPSC can be stored at -85 degrees C or -196 degrees 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. Se muestra que las células madre dentales pueden ser expandidas in-vitro para despues crío-preservarlas. Las células madre no pierden sus propiedades despúes de ser descongeladas. Células Madre Mesenquimales de Origen Dental TRATAMIENTO POTENCIAL JOURNAL

18 TÍTULO ABSTRACT RESUMEN Isolation and characterization of stem cells derived from human third molar tooth germs of young adults: implications in neo-vascularization, osteo-, adipo- and neurogenesis. Yalvac MEYalvac ME, Ramazanoglu M, Rizvanov AA, Sahin F, Bayrak OF, Salli U, Palotás A, Kose M, RizvanoBayrak OF, tás A, Pharmacogenomics J. 2010 Apr;10(2):105-13. Epub 2009 Sep 1. A number of studies have reported in the last decade that human tooth germs contain multipotent cells that give rise to dental and peri-odontal structures. The dental pulp, third molars in particular, have been shown to be a significant stem cell source. In this study, we isolated and characterized human tooth germ stem cells (hTGSCs) from third molars and assessed the expression of developmentally important transcription factors, such as oct4, sox2, klf4, nanog and c-myc, to determine their pluri-potency. Flow- cytometry analysis revealed that hTGSCs were positive for CD73, CD90, CD105 and CD166, but negative for CD34, CD45 and CD133, suggesting that these cells are mesenchymal-like stem cells. Under specific culture conditions, hTGSCs differentiated into osteogenic, adipogenic and neurogenic cells, as well as formed tube-like structures in Matrigel assay. hTGSCs showed significant levels of expression of sox2 and c-myc messenger RNA (mRNA), and a very high level of expression of klf4 mRNA when compared with human embryonic stem cells. This study reports for the first time that hTGSCs express developmentally important transcription factors that could render hTGSCs an attractive candidate for future somatic cell re-programming studies to differentiate germs into various tissue types, such as neurons and vascular structures. In addition, these multipotential hTGSCs could be important stem cell sources for autologous transplantation. Se prueba que las células de la pulpa dental de los terceros molares son células madre mesenquimales que bajo condiciones controladas se pueden diferenciar en tejido osteogénico, adipogénico y neurogénico. Los terceros molares pueden ser una fuente ideal de células madre para transplantes autólogos. Células Madre Mesenquimales de Origen Dental TRATAMIENTO POTENCIAL JOURNAL

19 TÍTULO ABSTRACT RESUMEN Se prueba que las células madre dentales se pueden multplicar varias generaciones sin perder su multipotencialidad. Gracias a esto, los dientes son considerados como una fuente ideal para la obtención de células madre adultas con fines terapéuticos. Human dental pulp stem cells--isolation and long term cultivation. Suchánek JSuchánek J, Soukup T, Ivancaková R, Karbanová J, Hubková V, Pytlík R, Kucerová L.Ivancakoanová J, Hub R, Kucerov Cell Prolif. 2009 Oct;42(5):587-94. Epub 2009 Jul 14. Human adult mesenchymal stem cells (MSCs) are rare elements living in various organs (e.g. bone marrow, skeletal muscle), with capability to differentiate in various cell types (e.g. chondrocytes, adipocytes and osteoblasts). In the year 2000, Gronthos and co-workers isolated stem cells from the human dental pulp (DPSCs). Later on, stem cells from exfoliated tooth were also obtained. The aims of our study were to establish protocol of DPSCs isolation and to cultivate DPSCs either from adult or exfoliated tooth, and to compare these cells with mesenchymal progenitor cell (MPCs) cultures. MPCs were isolated from the human bone marrow of proximal femur. DPSCs were isolated from deciduous and permanent teeth. Both cell types were cultivated under the same conditions in the media with 2% of FCS supplemented with PDGF and EGF growth factors. We have cultivated undifferentiated DPSCs for long time, over 60 population doublings in cultivation media designed for bone marrow MPCs. After reaching Hayflick's limit, they still have normal karyotype. Initial doubling time of our cultures was from 12 to 50 hours for first 40 population doublings, after reaching 50 population doublings, doubling time had increased to 60-90 hours. Regression analysis of uncumulated population doublings proved tight dependence of population doublings on passage number and slow decrease of proliferation potential. In comparison with bone marrow MPCs, DPSCs share similar biological characteristics and stem cell properties. The results of our experiments proved that the DPSCs and MPCs are highly proliferative, clonogenic cells that can be expanded beyond Hayflick's limit and remain cytogenetically stable. Moreover we have probably isolated two different populations of DPSCs. These DPSCs lines differed one from another in morphology. Because of their high proliferative and differentiation potential, DPSCs can become more attractive, easily accessible source of adult stem cells for therapeutic purposes. Células Madre Mesenquimales de Origen Dental TRATAMIENTO POTENCIAL JOURNAL

20 TÍTULO ABSTRACT RESUMEN Las células madre obtenidas de los terceros molares se pueden multiplicar en grandes cantidades y pueden formar tejido osteoblástico y odontobástico. Se comparan distintos niveles de proliferación celular. Characterization of dental pulp stem cells of human tooth germs. Takeda TTakeda T, Tezuka Y, Horiuchi M, Hosono K, Iida K, Hatakeyama D, Miyaki S, Kunisada T, Shibata T, Tezuka K., Horiucno K, Iidama D, Minisadaata T, Tezuk J Dent Res. 2008 Jul;87(7):676-81. In previous studies, human dental pulp stem cells (hDPSCs) were mainly isolated from adults. In this present study, we characterized hDPSCs isolated from an earlier developmental stage to evaluate the potential usage of these cells for tissue-regenerative therapy. hDPSCs isolated at the crown-completed stage showed a higher proliferation rate than those isolated at a later stage. When the cells from either group were cultured in medium promoting differentiation toward cells of the osteo/odontoblastic lineage, both became alkaline-phosphatase-positive, produced calcified matrix, and were also capable of forming dentin-like matrix on scaffolds in vivo. However, during long-term passage, these cells underwent a change in morphology and lost their differentiation ability. The results of a DNA array experiment showed that the expression of several genes, such as WNT16, was markedly changed with an increasing number of passages, which might have caused the loss of their characteristics as hDPSCs. Células Madre Mesenquimales de Origen Dental TRATAMIENTO POTENCIAL JOURNAL

21 TÍTULO ABSTRACT RESUMEN Las células madre obtenidas de los gérmenes dentales comparten varias características con las células madre embrionarias, como la pluripotencia. Ya que estas células son fácilmente accesibles y se multiplican en grandes cantidades son candidatos idóneos para la terapia regenerativa. Células Madre Mesenquimales de Origen Dental Human neural crest-derived postnatal cells exhibit remarkable embryonic attributes either in vitro or in vivo. d'Aquino Rd'Aquino R, Tirino V, Desiderio V, Studer M, De Angelis GC, Laino L, De Rosa A, Di Nucci D, Martino S, Paino F, Sampaolesi M, Papaccio G.Desiderir M, De Ango L, De Nucci D, Mar, SampaPapaccio Eur Cell Mater. 2011 Mar 22;21:304-16. During human embryonic development, odontogenic tissues, deriving from the neural crest, remain undifferentiated until the adult age. This study was aimed at characterising the cells of the follicle enveloping the dental germ, due to its direct origin from neural crests. Sixty dental follicles were collected from patients aged 18 to 45 years. This research has clarified that dental follicles, if extracted in a very early stage, when dental roots did not start to be formed, contain a lineage of cells, characterised by a high degree of plasticity in comparison with other adult stem cell populations. In particular, we found that these cells share the following features with ES: (i) high levels of embryonic stem cell markers (CD90, TRA1-60, TRA1-81, OCT-4, CD133, and SSEA-4); (ii) mRNA transcripts for Nanog and Rex-1; (iii) broader potency, being able to differentiate in cell types of all three germ layer, including smooth and skeletal muscle, osteoblasts, neurons, glial cells, and adipocytes; (iv) high levels of telomerase activity; (v) ability to form embryoid bodies; (vi) ability, after injection in murine blastocysts, to be localised within the inner cell mass; (vii) no teratoma formation after injection; (viii) in vivo tissue formation after transplantation. Our results demonstrate that these cells represent a very easy accessible and extraordinary source of pluripotent cells and point out the fact that they own the cardinal feature of embryonic stem cells. TRATAMIENTO POTENCIAL JOURNAL

22 TÍTULO ABSTRACT RESUMEN Las células madre dentales se pueden aislar de los dientes de leche y de algunos dientes permanentes. Estas células puedes ser extremadamente útiles para su preservación y para futuras aplicaciones terapéuticas. Stem cells from human exfoliated deciduous teeth-- isolation, long term cultivation and phenotypical analysis. Suchánek JSuchánek J, Visek B, Soukup T, El-Din Mohamed SK, Ivancaková R, Mokr J, Aboul-Ezz EH, Omran A.oukup TMohamed ková R, Mokr J, A. Acta Medica (Hradec Kralove). 2010;53(2):93-9. Aims: Our aims were to isolate stem cells from human exfoliated deciduous teeth (SHED), to cultivate them in vitro and to investigate their basic biological properties, phenotype and to compare our findings with dental pulp stem cells (DPSC) isolated from permanent teeth. Methods: Dental pulp was gently evacuated from exfoliated teeth. After enzymatic dissociation of dental pulp, SHED were cultivated in modified cultivation media for mesenchymal adult proge- nitor cells containing 2 % FCS and supplemented with growth factors and insulin, transferrin, sodium (ITS) supplement. Cell viability and other biological properties were examined using a Vi-Cell analyzer and a Z2-Counter. DNA analyses and phenotyping were performed with flow cytometry. Results: We were able to cultivate SHED over 45 population doublings. Our results showed that SHED cultivated under same conditions as DPSC had longer average population doubling time (41.3 hrs for SHED vs. 24.5 hrs for DPSC). Phenotypic comparison of cultivated SHED to that of cultivated DPSC showed differential expression CD29, CD44, CD71, CD117, CD166. During long-term cultivation, SHED did not showed any signs of degeneration or spontaneous differentiation. Conclusions: We isolated stem cells from exfoliated teeth. In comparison to DPSC, SHED proliferation rate was about 50% slower, and SHED showed slightly different phenotype. These cells may be extremely useful for stem cell tissue banking, further stem cell research and future therapeutic appli- cations. Células Madre Mesenquimales de Origen Dental TRATAMIENTO POTENCIAL JOURNAL

23 TÍTULO ABSTRACT RESUMEN Se crío-preservaron células madre dentales para evaluar que su potencialidad siga efectiva después de ser descongeladas. Después de 2 años de crío- preservación los científicos demostraron que las células madre dentales se podían seguir diferenciando en osteoblastos dando la posibilidad de preservar las células para un futuro uso clínico. Células Madre Mesenquimales de Origen Dental Long-term cryopreservation of dental pulp stem cells (SBP- DPSCs) and their differentiated osteoblasts: a cell source for tissue repair. Papaccio GPapaccio G, Graziano A, d'Aquino R, Graziano MF, Pirozzi G, Menditti D, De Rosa A, Carinci F, Laino G., d'Aquino MF, Pirozti D, De Ro F, Laino J Cell Physiol. 2006 Aug;208(2):319-25. 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. TRATAMIENTO POTENCIAL JOURNAL

24 TÍTULO ABSTRACT RESUMEN Se obtienen células madre de los dientes natales y se demuestra su capacidad de convertirse en tejido adipogénico, osteogénico, condrogénico, miogénico y neurogénico. Isolation and in vitro characterisation of dental pulp stem cells from natal teeth Karaöz EKaraöz E, Doğan BN, Aksoy A, Gacar G, Akyüz S, Ayhan S, Genç ZS, Yürüker S, Duruksu G, Demircan PC, Sariboyaci AE., Aksoy, Akyüz S, Genüker S,G, DemiSariboy Histochem Cell Biol (2010) 133:95–112 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. Células Madre Mesenquimales de Origen Dental TRATAMIENTO POTENCIAL JOURNAL

25 Regeneración Dental Resumen del Tema: Las células madre dentales se pueden diferenciar en tejidos dentales como la dentina y la pulpa abriendo la posibilidad de generar piezas dentales completas para reemplazar dientes perdidos y utilizar dientes biológicos en lugar de implantes artificiales. Asimismo, las células madre dentales se pueden utilizar para tratar problemas con tejidos dentales específicos como problemas relacionados a la dentina.

26 TÍTULO ABSTRACT RESUMEN Bioengineered dental tissues grown in the rat jaw. Duailibi SEDuailibi SE, Duailibi MT, Zhang W, Asrican R, Vacanti JP, Yelick PC., Zhang W, canti J PC. J Dent Res. 2008 Aug;87(8):745-50. Our long-term objective is to develop methods to form, in the jaw, bioengineered replacement teeth that exhibit physical properties and functions similar to those of natural teeth. Our results show that cultured rat tooth bud cells, seeded onto biodegradable scaffolds, implanted into the jaws of adult rat hosts and grown for 12 weeks, formed small, organized, bioengineered tooth crowns, containing dentin, enamel, pulp, and periodontal ligament tissues, similar to identical cell- seeded scaffolds implanted and grown in the omentum. Radiographic, histological, and immunohistochemical analyses showed that bioengineered teeth consisted of organized dentin, enamel, and pulp tissues. This study advances practical applications for dental tissue engineering by demonstrating that bioengineered tooth tissues can be regenerated at the site of previously lost teeth, and supports the use of tissue engineering strategies in humans, to regenerate previously lost and/or missing teeth. The results presented in this report support the feasibility of bioengineered replacement tooth formation in the jaw. Utilizando las células madre dentales de una rata, se demuestra la viabilidad de formar dientes creados mediante la bioingeniería. Regeneración Dental TRATAMIENTO POTENCIAL JOURNAL

27 TÍTULO ABSTRACT RESUMEN Utilizando las células madre dentales porcinas se realiza la creación de un diente que contiene dentina y esmalte. Esto abre la posibilidad de crear dientes de bioingeniería para personas que requieren remplazar dientes perdidos. Regeneración Dental Tissue engineering of complex tooth structures on biodegradable polymer scaffolds. Young CSYoung CS, Terada S, Vacanti JP, Honda M, Bartlett JD, Yelick PC., Vacanta M, Bartlck PC. J Dent Res. 2002 Oct;81(10):695-700. Tooth loss due to periodontal disease, dental caries, trauma, or a variety of genetic disorders continues to affect most adults adversely at some time in their lives. A biological tooth substitute that could replace lost teeth would provide a vital alternative to currently available clinical treatments. To pursue this goal, we dissociated porcine third molar tooth buds into single-cell suspensions and seeded them onto biodegradable polymers. After growing in rat hosts for 20 to 30 weeks, recognizable tooth structures formed that contained dentin, odontoblasts, a well-defined pulp chamber, putative Hertwig's root sheath epithelia, putative cementoblasts, and a morphologically correct enamel organ containing fully formed enamel. Our results demonstrate the first successful generation of tooth crowns from dissociated tooth tissues that contain both dentin and enamel, and suggest the presence of epithelial and mesenchymal dental stem cells in porcine third molar tissues. TRATAMIENTO POTENCIAL JOURNAL

28 TÍTULO ABSTRACT RESUMEN Las células madre dentales se pueden diferenciar en tejidos dentales como la dentina y la pulpa. Al lograr una reproducción rápida de las células, se sugiere la posibilidad de llegar a tratar problemas de la dentina en un futuro no muy distante. Regeneración Dental Postnatal human dental pulp stem cells (DPSCs) in vitro and in vivo. Gronthos SGronthos S, Mankani M, Brahim J, Robey PG, Shi S. Brahim J Shi S. Proc Natl Acad Sci U S A. 2000 Dec 5;97(25):13625-30. Dentinal repair in the postnatal organism occurs through the activity of specialized cells, odontoblasts, that are thought to be maintained by an as yet undefined precursor population associated with pulp tissue. In this study, we isolated a clonogenic, rapidly proliferative population of cells from adult human dental pulp. These DPSCs were then compared with human bone marrow stromal cells (BMSCs), known precursors of osteoblasts. Although they share a similar immunophenotype in vitro, functional studies showed that DPSCs produced only sporadic, but densely calcified nodules, and did not form adipocytes, whereas BMSCs routinely calcified throughout the adherent cell layer with clusters of lipid- laden adipocytes. When DPSCs were transplanted into immunocompromised mice, they generated a dentin-like structure lined with human odontoblast-like cells that surrounded a pulp-like interstitial tissue. In contrast, BMSCs formed lamellar bone containing osteocytes and surface-lining osteoblasts, surrounding a fibrous vascular tissue with active hematopoiesis and adipocytes. This study isolates postnatal human DPSCs that have the ability to form a dentin/pulp-like complex. TRATAMIENTO POTENCIAL JOURNAL

29 TÍTULO ABSTRACT RESUMEN Las células madre dentales pueden ser utilizadas para regenerar tejidos dentales in-vivo. Regeneración Dental The efficacy of mesenchymal stem cells to regenerate and repair dental structures. Shi SShi S, Bartold PM, Miura M, Seo BM, Robey PG, Gronthos S.ld PM, Miu, Robeythos S Orthod Craniofac Res. 2005 Aug;8(3):191-9. OBJECTIVES: Identification, characterization, and potential application of mesenchymal stem cells (MSC) derived from human dental tissues. RESULTS: MSC were identified in adult human dental pulp (dental pulp stem cells, DPSC), human primary teeth (stem cells from human exfoliated deciduous teeth, SHED), and periodontal ligament (periodontal ligament stem cells, PDLSC) by their capacity to generate clongenic cell clusters in culture. Ex vivo expanded DPSC, SHED, and PDLSC populations expressed a heterogeneous assortment of makers associated with MSC, dentin, bone, smooth muscle, neural tissue, and endothelium. PDLSC were also found to express the tendon specific marker, Scleraxis. Xenogeneic transplants containing HA/TCP with either DPSC or SHED generated donor-derived dentin-pulp-like tissues with distinct odontoblast layers lining the mineralized dentin-matrix. In parallel studies, PDLSC generated cementum-like structures associated with PDL-like connective tissue when transplanted with HA/TCP into immunocompromised mice. CONCLUSION: Collectively, these data revealed the presence of distinct MSC populations associated with dental structures with the potential of stem cells to regenerate living human dental tissues in vivo. TRATAMIENTO POTENCIAL JOURNAL

30 TÍTULO ABSTRACT RESUMEN Los avances en las investigaciones con células madre dentales y los andamios óptimos soportan la viabilidad de regenerar dientes enteros mediante la ingeniería de tejidos. Regeneración Dental Dental tissue regeneration - a mini-review. Yen AHYen AH, Yelick PC. PC. Gerontology. 2011;57(1):85-94. Epub 2010 May 6. OBJECTIVE: in this review, we summarize the emerging concepts of whole-tooth replacement strategies, using postnatal dental stem cells (DSCs) and dental tissue engineering approaches. RESULTS: current approaches to achieve clinically relevant biological replacement tooth therapies rely on the cultivation of DSCs capable of relaying odontogenic induction signals, through dental epithelial- mesenchymal cell interactions. DSC expansion and differentiation can be achieved by programming progenitor stem cells to adopt dental lineages, using instructive, bioengineered scaffold materials. Periodontal ligament regeneration in particular has demonstrated significant progress recently, despite the somewhat unpredictable clinical outcomes, with regard to its capacity to augment conventional metallic dental implants and as an important component for whole-tooth tissue engineering. Following recent advances made in DSC and tissue engineering research, various research groups are in the midst of performing 'proof of principle' experiments for whole-tooth regeneration, with associated functional periodontal tissues. This mini-review focuses on recent and promising developments in the fields of pulp and periodontal tissue DSCs that are of particular relevance for dental tissue and whole- tooth regeneration. CONCLUSION: continued advances in the derivation of useable DSC populations and optimally designed scaffold materials unequivocally support the feasibility of dental tissue and whole-tooth tissue engineering. TRATAMIENTO POTENCIAL JOURNAL

31 TÍTULO ABSTRACT RESUMEN Las células madre dentales son la fuente más accesible de células madre. Éstas tienen las características de mulipotencialidad y se reproducen en grandes cantidades por lo que son una fuente ideal para la regeneración de tejidos. Regeneración Dental Dental pulp stem cells in regenerative dentistry. Casagrande LCasagrande L, Cordeiro MM, Nör SA, Nör JE. Nör SA, Nö Odontology. 2011 Jan;99(1):1-7. Epub 2011 Jan 27. 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. TRATAMIENTO POTENCIAL JOURNAL

32 TÍTULO ABSTRACT RESUMEN Después de una exhaustiva revisión de literatura se determina que la regeneración de estructuras complejas como los dientes ya no es un fin inalcanzable. Regeneración Dental Dental pulp tissue engineering. Demarco FFDemarco FF, Conde MC, Cavalcanti BN, Casagrande L, Sakai VT, Nör JE.Cavalcanagrande L, Sa Braz Dent J. 2011;22(1):3-13. Dental pulp is a highly specialized mesenchymal tissue that has a limited regeneration capacity due to anatomical arrangement and post-mitotic nature of odontoblastic cells. Entire pulp amputation followed by pulp space disinfection and filling with an artificial material cause loss of a significant amount of dentin leaving as life-lasting sequelae a non-vital and weakened tooth. However, regenerative endodontics is an emerging field of modern tissue engineering that has demonstrated promising results using stem cells associated with scaffolds and responsive molecules. Thereby, this article reviews the most recent endeavors to regenerate pulp tissue based on tissue engineering principles and provides insightful information to readers about the different aspects involved in tissue engineering. Here, we speculate that the search for the ideal combination of cells, scaffolds, and morphogenic factors for dental pulp tissue engineering may be extended over future years and result in significant advances in other areas of dental and craniofacial research. The findings collected in this literature review show that we are now at a stage in which engineering a complex tissue, such as the dental pulp, is no longer an unachievable goal and the next decade will certainly be an exciting time for dental and craniofacial research. TRATAMIENTO POTENCIAL JOURNAL

33 Regeneración Huesos y Estructura Craneofacial Resumen del Tema: Las células madre dentales pueden ser diferenciadas en osteblastos (células óseas) y pueden ser utilizadas para corregir defectos craneofaciales. Al combinar las células madre con matrices de colágena se ha demostrado que las heridas y los defectos craneofaciales pueden ser corregidos de forma más rápida y con estructuras óseas mejor formadas que esperando la regeneración natural de los tejidos.

34 TÍTULO ABSTRACT RESUMEN Todas las estructuras craneofaciales son derivadas del tejido mesenquimal. Las células madre dentales pueden regenerar varios tejidos craneofaciales por lo que pueden representar un significativo avance en la odonotología regenerativa. Regeneración Huesos y Estructura Craneofacial Craniofacial tissue engineering by stem cells. Mao JJMao JJ, Giannobile WV, Helms JA, Hollister SJ, Krebsbach PH, Longaker MT, Shi S.bile WV, Helm SJ, Kre Longaker MT J Dent Res. 2006 Nov;85(11):966-79. Craniofacial tissue engineering promises the regeneration or de novo formation of dental, oral, and craniofacial structures lost to congenital anomalies, trauma, and diseases. Virtually all craniofacial structures are derivatives of mesenchymal cells. Mesenchymal stem cells are the offspring of mesenchymal cells following asymmetrical division, and reside in various craniofacial structures in the adult. Cells with characteristics of adult stem cells have been isolated from the dental pulp, the deciduous tooth, and the periodontium. Several craniofacial structures--such as the mandibular condyle, calvarial bone, cranial suture, and subcutaneous adipose tissue--have been engineered from mesenchymal stem cells, growth factor, and/or gene therapy approaches. As a departure from the reliance of current clinical practice on durable materials such as amalgam, composites, and metallic alloys, biological therapies utilize mesenchymal stem cells, delivered or internally recruited, to generate craniofacial structures in temporary scaffolding biomaterials. Craniofacial tissue engineering is likely to be realized in the foreseeable future, and represents an opportunity that dentistry cannot afford to miss. TRATAMIENTO POTENCIAL JOURNAL

35 TÍTULO ABSTRACT RESUMEN Las células madre dentales se pueden utilizar para la regeneración de hueso por lo que presentan un gran potencial para revolucionar la implantología dental. Regeneración Huesos y Estructura Craneofacial The osteoblastic differentiation of dental pulp stem cells and bone formation on different titanium surface textures. Mangano CMangano C, De Rosa A, Desiderio V, d'Aquino R, Piattelli A, De Francesco F, Tirino V, Mangano F, Papaccio G., Desiderino R, Piatrancesco FMangano F, Biomaterials. 2010 May;31(13):3543-51. Epub 2010 Feb 1. Bone Tissue Engineering (BTE) and Dental Implantology (DI) require the integration of implanted structures, with well characterized surfaces, in bone. In this work we have challenged acid-etched titanium (AET) and Laser Sintered Titanium (LST) surfaces with either human osteoblasts or stem cells from human dental pulps (DPSCs), to understand their osteointegration and clinical use capability of derived implants. DPSCs and human osteoblasts were challenged with the two titanium surfaces, either in plane cultures or in a roller apparatus within a culture chamber, for hours up to a month. During the cultures cells on the titanium surfaces were examined for histology, protein secretion and gene expression. Results show that a complete osteointegration using human DPSCs has been obtained: these cells were capable to quickly differentiate into osteoblasts and endotheliocytes and, then, able to produce bone tissue along the implant surfaces. Osteoblast differentiation of DPSCs and bone morphogenetic protein production was obtained in a better and quicker way, when challenging stem cells with the LST surfaces. This successful BTE in a comparatively short time gives interesting data suggesting that LST is a promising alternative for clinical use in DI. TRATAMIENTO POTENCIAL JOURNAL

36 TÍTULO ABSTRACT RESUMEN Las células madre dentales son una herramienta prometedora para la regeneración de huesos y la facilidad de utilizarlas las hace candidatas idóneas para protocolos experimentales en humanos. Regeneración Huesos y Estructura Craneofacial Dental pulp stem cells: a promising tool for bone regeneration. Graziano AGraziano A, d'Aquino R, Laino G, Papaccio G., Laino G, Stem Cell Rev. 2008 Spring;4(1):21-6. Human tissues are different in term of regenerative properties. Stem cells are a promising tool for tissue regeneration, thanks to their particular characteristics of proliferation, differentiation and plasticity. Several "loci" or "niches" within the adult human body are colonized by a significant number of stem cells. However, access to these potential collection sites often is a limiting point. The interaction with biomaterials is a further point that needs to be considered for the therapeutic use of stem cells. Dental pulp stem cells (DPSCs) have been demonstrated to answer all of these issues: access to the collection site of these cells is easy and produces very low morbidity; extraction of stem cells from pulp tissue is highly efficiency; they have an extensive differentiation ability; and the demonstrated interactivity with biomaterials makes them ideal for tissue reconstruction. SBP-DPSCs are a multipotent stem cell subpopulation of DPSCs which are able to differentiate into osteoblasts, synthesizing 3D woven bone tissue chips in vitro and that are capable to synergically differentiate into osteoblasts and endotheliocytes. Several studied have been performed on DPSCs and they mainly found that these cells are multipotent stromal cells that can be safety cryopreserved, used with several scaffolds, that can extensively proliferate, have a long lifespan and build in vivo an adult bone with Havers channels and an appropriate vascularization. A definitive proof of their ability to produce dentin has not been yet done. Interestingly, they seem to possess immunoprivileges as they can be grafted into allogenic tissues and seem to exert anti-inflammatory abilities, like many other mesenchymal stem cells. The easy management of dental pulp stem cells make them feasible for use in clinical trials on human patients. TRATAMIENTO POTENCIAL JOURNAL

37 TÍTULO ABSTRACT RESUMEN Las células madre dentales pueden ser utilizadas para corregir grandes defectos craneofaciales en ratas por lo que constituyen un modelo prometedor para la reconstrucción de problemas craneofaciales en humanos. Regeneración Huesos y Estructura Craneofacial Reconstruction of large cranial defects in nonimmunosuppressed experimental design with human dental pulp stem cells. de Mendonça Costa Ade Mendonça Costa A, Bueno DF, Martins MT, Kerkis I, Kerkis A, Fanganiello RD, Cerruti H, Alonso N, Passos- Bueno MRT, Kerkis A, Fangaerruti H, Passos J Craniofac Surg. 2008 Jan;19(1):204-10. The main aim of this study is to evaluate the capacity of human dental pulp stem cells (hDPSC), isolated from deciduous teeth, to reconstruct large-sized cranial bone defects in nonimmunosuppressed (NIS) rats. To our knowledge, these cells were not used before in similar experiments. We performed two symmetric full-thickness cranial defects (5 x 8 mm) on each parietal region of eight NIS rats. In six of them, the left side was supplied with collagen membrane only and the right side (RS) with collagen membrane and hDPSC. In two rats, the RS had collagen membrane only and nothing was added at the left side (controls). Cells were used after in vitro characterization as mesenchymal cells. Animals were euthanized at 7, 20, 30, 60, and 120 days postoperatively and cranial tissue samples were taken from the defects for histologic analysis. Analysis of the presence of human cells in the new bone was confirmed by molecular analysis. The hDPSC lineage was positive for the four mesenchymal cell markers tested and showed osteogenic, adipogenic, and myogenic in vitro differentiation. We observed bone formation 1 month after surgery in both sides, but a more mature bone was present in the RS. Human DNA was polymerase chain reaction-amplified only at the RS, indicating that this new bone had human cells. The use of hDPSC in NIS rats did not cause any graft rejection. Our findings suggest that hDPSC is an additional cell resource for correcting large cranial defects in rats and constitutes a promising model for reconstruction of human large cranial defects in craniofacial surgery. TRATAMIENTO POTENCIAL JOURNAL

38 TÍTULO ABSTRACT RESUMEN Este estudio demuestra que las células madre dentales y las matrices de colágena pueden corregir completamente los defectos de la mandíbula así como de otros tejidos y órganos. Regeneración Huesos y Estructura Craneofacial Human mandible bone defect repair by the grafting of dental pulp stem/progenitor cells and collagen sponge biocomplexes. d'Aquino Rd'Aquino R, De Rosa A, Lanza V, Tirino V, Laino L, Graziano A, Desiderio V, Laino G, Papaccio G. Lanza V,Laino Lo A, DesLaino Go G. In this study we used a biocomplex constructed from dental pulp stem/progenitor cells (DPCs) and a collagen sponge scaffold for oro-maxillo-facial (OMF) bone tissue repair in patients requiring extraction of their third molars. The experiments were carried out according to our Internal Ethical Committee Guidelines and written informed consent was obtained from the patients. The patients presented with bilateral bone reabsorption of the alveolar ridge distal to the second molar secondary to impaction of the third molar on the cortical alveolar lamina, producing a defect without walls, of at least 1.5 cm in height. This clinical condition does not permit spontaneous bone repair after extraction of the third molar, and eventually leads to loss also of the adjacent second molar. Maxillary third molars were extracted first for DPC isolation and expansion. The cells were then seeded onto a collagen sponge scaffold and the obtained biocomplex was used to fill in the injury site left by extraction of the mandibular third molars. Three months after autologous DPC grafting, alveolar bone of patients had optimal vertical repair and complete restoration of periodontal tissue back to the second molars, as assessed by clinical probing and X-rays. Histological observations clearly demonstrated the complete regeneration of bone at the injury site. Optimal bone regeneration was evident one year after grafting. This clinical study demonstrates that a DPC/collagen sponge biocomplex can completely restore human mandible bone defects and indicates that this cell population could be used for the repair and/or regeneration of tissues and organs. Eur Cell Mater. 2009 Nov 12;18:75-83. TRATAMIENTO POTENCIAL JOURNAL

39 Regeneración Cardiaca Resumen del Tema: Las células madre de origen dental tienen la posibilidad de diferenciarse en células cardiacas (cardiomiocitos). Esta cualidad sugiere que las células madre dentales se pueden usar en tratamientos regenerativos de corazón después de un infarto. De la misma forma, las células mesenquimales de los dientes pueden jugar un papel clave en la generación de válvulas cardiacas para uso humano.

40 TÍTULO ABSTRACT RESUMEN Se utilizaron células madre dentales para tratar un infarto cardiaco en ratas. Al revisar los resultados, se sugiere que las células madre dentales podrían ser una buena alternativa para la regeneración de corazones que han sufrido infartos. Regeneración Cardiaca Human dental pulp stem cells improve left ventricular function, induce angiogenesis, and reduce infarct size in rats with acute myocardial infarction. Gandia CGandia C, Armiñan A, García-Verdugo JM, Lledó E, Ruiz A, Miñana MD, Sanchez-Torrijos J, Payá R, Mirabet V, Carbonell-Uberos F, Llop M, Montero JA, Sepúlveda P.A, García, Lledó E, Ruiz Az-Torriyá R, V, CarbonF, Llop M, Montero Stem Cells. 2008 Mar;26(3):638-45. Epub 2007 Dec 13. Human dental pulp contains precursor cells termed dental pulp stem cells (DPSC) that show self-renewal and multilineage differentiation and also secrete multiple proangiogenic and antiapoptotic factors. To examine whether these cells could have therapeutic potential in the repair of myocardial infarction (MI), DPSC were infected with a retrovirus encoding the green fluorescent protein (GFP) and expanded ex vivo. Seven days after induction of myocardial infarction by coronary artery ligation, 1.5 x 10(6) GFP- DPSC were injected intramyocardially in nude rats. At 4 weeks, cell-treated animals showed an improvement in cardiac function, observed by percentage changes in anterior wall thickening left ventricular fractional area change, in parallel with a reduction in infarct size. No histologic evidence was seen of GFP+ endothelial cells, smooth muscle cells, or cardiac muscle cells within the infarct. However, angiogenesis was increased relative to control-treated animals. Taken together, these data suggest that DPSC could provide a novel alternative cell population for cardiac repair, at least in the setting of acute MI. TRATAMIENTO POTENCIAL JOURNAL

41 Problemas Neurológicos Resumen del Tema: Las células madre dentales producen factores neurotróficos y pueden reparar lesiones de médula espinal. Las células madre dentales se pueden diferenciar en el laboratorio y formar neuronas activas y funcionales. Las células dentales son capaces de migrar a las zonas afectadas del cerebro, inducir la vascularización y mejorar la isquemia cerebral. Las células madre dentales fueron diferenciadas en tejidos de lineaje neural y mostraron propiedades de protección a las neuronas dopaminérgicos por lo que presentan una posibilidad para tratar enfermedades neurodegenerativas como el Mal de Parkinson. Las células madre dentales mantienen su potencial de neuro-protección después de la crío-preservación de largo plazo.

42 TÍTULO ABSTRACT RESUMEN Las células madre dentales producen factores neurotróficos y pueden reparar lesiones de médula espinal. Problemas Neurológicos Dental pulp cells produce neurotrophic factors, interact with trigeminal neurons in vitro, and rescue motoneurons after spinal cord injury. Nosrat IVNosrat IV, Widenfalk J, Olson L, Nosrat CA. J, Olson L Dev Biol. 2001 Oct 1;238(1):120-32. Interactions between ingrowing nerve fibers and their target tissues form the basis for functional connectivity with the central nervous system. Studies of the developing dental pulp innervation by nerve fibers from the trigeminal ganglion is an excellent example of nerve-target tissue interactions and will allow specific questions regarding development of the dental pulp nerve system to be addressed. Dental pulp cells (DPC) produce an array of neurotrophic factors during development, suggesting that these proteins might be involved in supporting trigeminal nerve fibers that innervate the dental pulp. We have established an in vitro culture system to study the interactions between the dental pulp cells and trigeminal neurons. We show that dental pulp cells produce several neurotrophic factors in culture. When DPC are cocultured with trigeminal neurons, they promote survival and a specific and elaborate neurite outgrowth pattern from trigeminal neurons, whereas skin fibroblasts do not provide a similar support. In addition, we show that dental pulp tissue becomes innervated when transplanted ectopically into the anterior chamber of the eye in rats, and upregulates the catecholaminergic nerve fiber density of the irises. Interestingly, grafting the dental pulp tissue into hemisected spinal cord increases the number of surviving motoneurons, indicating a functional bioactivity of the dental pulp-derived neurotrophic factors in vivo by rescuing motoneurons. Based on these findings, we propose that dental pulp-derived neurotrophic factors play an important role in orchestrating the dental pulp innervation. TRATAMIENTO POTENCIAL JOURNAL

43 TÍTULO ABSTRACT RESUMEN Las células madre dentales se pueden diferenciar en el laboratorio y formar neuronas activas y funcionales. Problemas Neurológicos Simultaneous PKC and cAMP activation induces differentiation of human dental pulp stem cells into functionally active neurons. Király MKirály M, Porcsalmy B, Pataki A, Kádár K, Jelitai M, Molnár B, Hermann P, Gera I, Grimm WD, Ganss B, Zsembery A, Varga G.y B, PatakiJelitai B, Hermra I, Gris B, Zsearga G. Neurochem Int. 2009 Sep;55(5):323-32. Epub 2009 Apr 5. The plasticity of dental pulp stem cells (DPSCs) has been demonstrated by several studies showing that they appear to self-maintain through several passages, giving rise to a variety of cells. The aim of the present study was to differentiate DPSCs to mature neuronal cells showing functional evidence of voltage gated ion channel activities in vitro. First, DPSC cultures were seeded on poly-l-lysine coated surfaces and pretreated for 48h with a medium containing basic fibroblast growth factor and the demethylating agent 5-azacytidine. Then neural induction was performed by the simultaneous activation of protein kinase C and the cyclic adenosine monophosphate pathway. Finally, maturation of the induced cells was achieved by continuous treatment with neurotrophin-3, dibutyryl cyclic AMP, and other supplementary components. Non-induced DPSCs already expressed vimentin, nestin, N-tubulin, neurogenin-2 and neurofilament-M. The inductive treatment resulted in decreased vimentin, nestin, N-tubulin and increased neurogenin-2, neuron-specific enolase, neurofilament-M and glial fibrillary acidic protein expression. By the end of the maturation period, all investigated genes were expressed at higher levels than in undifferentiated controls except vimentin and nestin. Patch clamp analysis revealed the functional activity of both voltage-dependent sodium and potassium channels in the differentiated cells. Our results demonstrate that although most surviving cells show neuronal morphology and express neuronal markers, there is a functional heterogeneity among the differentiated cells obtained by the in vitro differentiation protocol described herein. Nevertheless, this study clearly indicates that the dental pulp contains a cell population that is capable of neural commitment by our three step neuroinductive protocol. TRATAMIENTO POTENCIAL JOURNAL

44 TÍTULO ABSTRACT RESUMEN Las células madre dentales mantienen su potencial de neuro-protección después de la crío-preservación de largo plazo. Esto sugiere que sería recomendable preservar las células madre de los niños para un posible uso futuro en enfermedades degenerativas relacionadas con la edad. Problemas Neurológicos Human tooth germ stem cells preserve neuro-protective effects after long-term cryo-preservation. Yalvaç MEYalvaç ME, Ramazanoglu M, Tekguc M, Bayrak OF, Shafigullina AK, Salafutdinov II, Blatt NL, Kiyasov AP, Sahin F, Palotás A, Rizvanov M, Tekguc hafigulllafutdino NL, Kiyasov APs A, Rizvanov A Curr Neurovasc Res. 2010 Feb 1;7(1):49-58. The use of mesenchymal stem cells (MSCs) has been shown to be promising in chronic disorders such as diabetes, Alzheimer's dementia, Parkinson's disease, spinal cord injury and brain ischemia. Recent studies revealed that human tooth germs (hTG) contain MSCs which can be easily isolated, expanded and cryo- preserved. In this report, we isolated human tooth germ stem cells (hTGSCs) with MSC characteristics from third molar tooth germs, cryo-preserved them at -80( degrees )C for 6 months, and evaluated for their surface antigens, expression of pluri-potency associated genes, differentiation capacity, karyotype, and proliferation rate. These characteristics were compared to their non-frozen counterparts. In addition, neuro- protective effects of cryo-preserved cells on neuro-blastoma SH-SY5Y cells were also assessed after exposure to stress conditions induced by hydrogen-peroxide (oxidative stress) and paclitaxel (microtubule stabilizing mitotic inhibitor). After long term cryo-preservation hTGSCs expressed surface antigens CD29, CD73, CD90, CD105, and CD166, but not CD34, CD45 or CD133, which was typical for non-frozen hTGSCs. Cryo-preserved hTGSCs were able to differentiate into osteo-, adipo- and neuro-genic cells. They also showed normal karyotype after high number of population doublings and unchanged proliferation rate. On the other hand, cryo-preserved cells demonstrated a tendency for lower level of pluri-potency associated gene expression (nanog, oct4, sox2, klf4, c-myc) than non-frozen hTGSCs. hTGSCs conditioned media increased survival of SH-SY5Y cells exposed to oxidative stress or paclitaxel. These findings confirm that hTGSCs preserve their major characteristics and exert neuro-protection after long- term cryo-preservation, suggesting that hTGSCs, harvested from young individuals and stored for possible use later as they grow old, might be employed in cellular therapy of age-related degenerative disorders. TRATAMIENTO POTENCIAL JOURNAL

45 TÍTULO ABSTRACT RESUMEN Células madre dentales fueron transplantas en ratas con isquemia cerebral. Las células fueron capaces de migrar a las zonas afectadas del cerebro, inducir la vascularización y mejoraron la isquemia cerebral. Problemas Neurológicos Dental Pulp-Derived CD31(-)/CD146(-) Side Population Stem/Progenitor Cells Enhance Recovery of Focal Cerebral Ischemia in Rats. Sugiyama MSugiyama M, Iohara K, Wakita H, Hattori H, Ueda M, Matsushita K, Nakashima M.Wakita HH, Ueda ita K, Na Tissue Eng Part A. 2011 May;17(9-10):1303-11. Epub 2011 Feb 25. Regenerative therapy using stem cells is a promising approach for the treatment of stroke. Recently, we reported that CD31(-)/CD146(-) side population (SP) cells from porcine dental pulp exhibit highly vasculogenic potential in hindlimb ischemia. In this study, we investigated the influence of CD31(- )/CD146(-) SP cells after transient middle cerebral artery occlusion (TMCAO). Adult male Sprague- Dawley rats were subjected to 2 h of TMCAO. Twenty-four hours after TMCAO, CD31(-)/CD146(-) SP cells were transplanted into the brain. Motor function and infarct volume were evaluated. Neurogenesis and vasculogenesis were determined with immunochemical markers, and the levels of neurotrophic factors were assayed with real-time reverse transcription-polymerase chain reaction. In the cell transplantation group, the number of doublecortin-positive cells increased twofold, and the number of NeuN-positive cells increased eightfold, as compared with the control phosphate-buffered saline group. The vascular endothelial growth factor level in the ischemic brain with transplanted cells was 28 times higher than that in the normal brain. In conclusion, CD31(-)/CD146(-) SP cells promoted migration and differentiation of the endogenous neuronal progenitor cells and induced vasculogenesis, and ameliorated ischemic brain injury after TMCAO. TRATAMIENTO POTENCIAL JOURNAL

46 TÍTULO ABSTRACT RESUMEN Las células madre dentales tienen el potencial de generar neuronas activas y pueden inducir la neuroplasticidad en el sistema nervioso. Problemas Neurológicos Implanted adult human dental pulp stem cells induce endogenous axon guidance. Arthur AArthur A, Shi S, Zannettino AC, Fujii N, Gronthos S, Koblar SA.annet, Fujii N, Grr SA. Stem Cells. 2009 Sep;27(9):2229-37. The human central nervous system has limited capacity for regeneration. Stem cell-based therapies may overcome this through cellular mechanisms of neural replacement and/or through molecular mechanisms, whereby secreted factors induce change in the host tissue. To investigate these mechanisms, we used a readily accessible human cell population, dental pulp progenitor/stem cells (DPSCs) that can differentiate into functionally active neurons given the appropriate environmental cues. We hypothesized that implanted DPSCs secrete factors that coordinate axon guidance within a receptive host nervous system. An avian embryonic model system was adapted to investigate axon guidance in vivo after transplantation of adult human DPSCs. Chemoattraction of avian trigeminal ganglion axons toward implanted DPSCs was mediated via the chemokine, CXCL12, also known as stromal cell-derived factor-1, and its receptor, CXCR4. These findings provide the first direct evidence that DPSCs may induce neuroplasticity within a receptive host nervous system. TRATAMIENTO POTENCIAL JOURNAL

47 TÍTULO ABSTRACT RESUMEN Las células madre dentales promueven la proliferación y diferenciación de células neurales y son buenas candidatas para la terapia personalizada ya que evitan los problemas de rechazo. Problemas Neurológicos Putative dental pulp-derived stem/stromal cells promote proliferation and differentiation of endogenous neural cells in the hippocampus of mice. Huang AHHuang AH, Snyder BR, Cheng PH, Chan AW.R, Cheng. Stem Cells. 2008 Oct;26(10):2654-63. Epub 2008 Aug 7. Until now, interest in dental pulp stem/stromal cell (DPSC) research has centered on mineralization and tooth repair. Beginning a new paradigm in DPSC research, we grafted undifferentiated, untreated DPSCs into the hippocampus of immune-suppressed mice. The rhesus DPSC (rDPSC) line used was established from the dental pulp of rhesus macaques and found to be similar to human bone marrow/mesenchymal stem cells, which express Nanog, Rex-1, Oct-4, and various cell surface antigens, and have multipotent differentiation capability. Implantation of rDPSCs into the hippocampus of mice stimulated proliferation of endogenous neural cells and resulted in the recruitment of pre-existing Nestin(+) neural progenitor cells (NPCs) and beta-tubulin-III(+) mature neurons to the site of the graft. Additionally, many cells born during the first 7 days after implantation proliferated, forming NPCs and neurons, and, to a lesser extent, underwent astrogliosis, forming astrocytes and microglia, by 30 days after implantation. Although the DPSC graft itself was short term, it had long-term effects by promoting growth factor signaling. Implantation of DPSCs enhanced the expression of ciliary neurotrophic factor, vascular endothelial growth factor, and fibroblast growth factor for up to 30 days after implantation. In conclusion, grafting rDPSCs promotes proliferation, cell recruitment, and maturation of endogenous stem/progenitor cells by modulating the local microenvironment. Our results suggest that DPSCs have a valuable, unique therapeutic potential, specifically as a stimulator and modulator of the local repair response in the central nervous system. DPSCs would be a preferable cell source for therapy due to the possibility of a "personalized" stem cell, avoiding the problems associated with host immune rejection. Disclosure of potential conflicts of interest is found at the end of this article. TRATAMIENTO POTENCIAL JOURNAL

48 TÍTULO ABSTRACT RESUMEN Los resultados del estudio muestran que las células madre dentales transplantadas In-Vivo tienen propiedades neuronales importantes que pueden reparar cerebros que estan lastimados. Problemas Neurológicos Integration of neuronally predifferentiated human dental pulp stem cells into rat brain in vivo. Király MKirály M, Kádár K, Horváthy DB, Nardai P, Rácz GZ, Lacza Z, Varga G, Gerber G. Horvátrdai P, Rác, Varga G. Neurochem Int. 2011 Jan 8. Pluripotency and their neural crest origin make dental pulp stem cells (DPSCs) an attractive donor source for neuronal cell replacement. Despite recent encouraging results in this field, little is known about the integration of transplanted DPSC derived neuronal pecursors into the central nervous system. To address this issue, neuronally predifferentiated DPSCs, labeled with a vital cell dye Vybrant DiD were introduced into postnatal rat brain. DPSCs were transplanted into the cerebrospinal fluid of 3-day-old male Wistar rats. Cortical lesion was induced by touching a cold (-60°C) metal stamp to the calvaria over the forelimb motor cortex. Four weeks later cell localization was detected by fluorescent microscopy and neuronal cell markers were studied by immunohistochemistry. To investigate electrophysiological properties of engrafted, fluorescently labeled DPSCs, 300μm-thick horizontal brain slices were prepared and the presence of voltage-dependent sodium and potassium channels were recorded by patch clamping. Predifferentiated donor DPSCs injected into the cerebrospinal fluid of newborn rats migrated as single cells into a variety of brain regions. Most of the cells were localized in the normal neural progenitor zones of the brain, the subventricular zone (SVZ), subgranular zone (SGZ) and subcallosal zone (SCZ). Immunohistochemical analysis revealed that transplanted DPSCs expressed the early neuronal marker N-tubulin, the neuronal specific intermediate filament protein NF-M, the postmitotic neuronal marker NeuN, and glial GFAP. Moreover, the cells displayed TTX sensitive voltage dependent (VD) sodium currents (I(Na)) and TEA sensitive delayed rectifier potassium currents (K(DR)). Four weeks after injury, fluorescently labeled cells were detected in the lesioned cortex. Neurospecific marker expression was increased in DPSCs found in the area of the cortical lesions compared to that in fluorescent cells of uninjured brain. TTX sensitive VD sodium currents and TEA sensitive K(DR) significantly increased in labeled cells of the cortically injured area. In conclusion, our data demonstrate that engrafted DPSC-derived cells integrate into the host brain and show neuronal properties not only by expressing neuron-specific markers but also by exhibiting voltage dependent sodium and potassium channels. This proof of concept study reveals that predifferentiated hDPSCs may serve as useful sources of neuro- and gliogenesis in vivo, especially when the brain is injured. TRATAMIENTO POTENCIAL JOURNAL

49 TÍTULO ABSTRACT RESUMEN Las células madre dentales fueron diferenciadas en tejidos de lineaje neural por lo que pueden ayudar en tratamientos de medicina regenerativa. Problemas Neurológicos Differentiation of dental pulp stem cells into a neural lineage Masaharu Takeyasu Masaharu Takeyasu 1), Tadashige Nozaki 2) and Michiharu Daito 1)) and Michiharu Pediatric Dental Journal - Vol. 16 (2006), No. 2 pp.154-162 We previously investigated whether dental pulp-derived cells possess similar pluripotency to bone marrow cells, and reported their capacity to differentiate into osteoblasts, as well as the characteristics of the stem cells present in dental pulp. In the present study, we hypothesized that neural stem cells would also exist in rat dental pulp, similar to bone marrow and the brain, and attempted to induce their differentiation into a neural lineage by applying an in vitro study design previously reported to induce differentiation of human bone marrow cells. Before inducing differentiation, we detected cells expressing nestin (Nes), which is known to be a marker for neural stem cells, within primary cultures of rat dental pulp-derived cells, suggesting the existence of neural stem cells in dental pulp. Quantitative analyses of the mRNA and protein expression levels revealed downregulation of both the Nes mRNA and protein levels to about 68.1% and 12.4%, respectively, after the induction of differentiation compared to the corresponding levels before induction. Conversely, the glial fibrillary acidic protein (Gfap) mRNA level was elevated by 1.3-fold after the induction of differentiation compared with the level before induction. The reduced number of Nes-positive cells and decreased Nes mRNA and protein levels after the induction of differentiation may be attributed to differentiation of neural stem cells into a neural lineage. Moreover, the increased number of Gfap- positive cells and increased Gfap mRNA level after the induction of differentiation most likely support their progressive differentiation into a glial cell lineage, since Gfap is a marker that is upregulated in glial cells. Our present data demonstrate the existence of neural stem cells in tissues other than the central nervous system, and may represent a significant step toward providing more diverse and multiple sources of stem cells for future regenerative medicine. TRATAMIENTO POTENCIAL JOURNAL

50 TÍTULO ABSTRACT RESUMEN Las células madre dentales fueron diferenciadas en tejidos de lineaje neural y mostraron propiedades de protección a las neuronas dopaminérgicos por lo que presentan una posibilidad para tratar enfermedades neurodegenerativas como el Mal de Parkinson. Problemas Neurológicos Human dental pulp stem cells protect mouse dopaminergic neurons against MPP+ or rotenone. Nesti CNesti C, Pardini C, Barachini S, D'Alessandro D, Siciliano G, Murri L, Petrini M, Vaglini F. C, Baraclessandro D, Murri L, PetF. Brain Res. 2011 Jan 7;1367:94-102. Epub 2010 Sep 18. Parkinson's disease (PD) is a neurodegenerative disorder characterized by the progressive death of substantia nigra dopaminergic neurons that results in a regional loss of striatal dopamine (DA) levels. Dental pulp contains ex vivo-expandable cells called dental pulp stem cells (DPSCs), with the capacity to differentiate into multiple cell lineages. More interestingly, due to their embryonic origin, DPSCs express neurotrophic factors such as brain- derived neurotrophic factor, nerve growth factor and glial cell-derived neurotrophic factor. The aim of the present study was to investigate the neuroprotective effects of DPSCs against MPP+ (2.5, 5, and 10 μM) and rotenone (0.25, 0.5 and 1 μM) in an in vitro model of PD, using an indirect co-culture system with mesencephalic cell cultures. When mesencephalic cultures were challenged with MPP+ or rotenone, in the presence of DPSCs a statistically significant protective effect was observed at all the tested doses in terms of DA uptake. DPSCs protective effect on DA neurons was also confirmed by immunocytochemistry: an increased number of spared tyrosine hydroxylase (TH)+ cells was observed in co-culture conditions compared to controls, and neurons showed longer processes in comparison with mesencephalic cells grown without DPSCs. In conclusion, the co- culture with DPSCs significantly attenuated MPP+ or rotenone-induced toxicity in primary cultures of mesencephalic neurons. Considering that the direct contact between the two cell types was prevented, it can be speculated that neuroprotection could be due to soluble factors such as BDNF and NGF, released by DPSCs. Blocking BDNF and NGF with neutralizing antibodies, the neuroprotecting effect of DPSCs was completely abolished. Therefore DPSCs can be viewed as possible candidates for studies on cell-based therapy in neurodegenerative disorders. TRATAMIENTO POTENCIAL JOURNAL

51 TÍTULO ABSTRACT RESUMEN Bajo condiciones controladas, las células madre dentales se pueden convertir en neuronas activas con el potencial de utilizarlas en terapias regenerativas. Problemas Neurológicos Adult human dental pulp stem cells differentiate toward functionally active neurons under appropriate environmental cues. Arthur AArthur A, Rychkov G, Shi S, Koblar SA, Gronthos S.G, Shi S, Gron Stem Cells. 2008 Jul;26(7):1787-95. Epub 2008 May 22. Human adult dental pulp stem cells (DPSCs) reside within the perivascular niche of dental pulp and are thought to originate from migrating cranial neural crest (CNC) cells. During embryonic development, CNC cells differentiate into a wide variety of cell types, including neurons of the peripheral nervous system. Previously, we have demonstrated that DPSCs derived from adult human third molar teeth differentiate into cell types reminiscent of CNC embryonic ontology. We hypothesized that DPSCs exposed to the appropriate environmental cues would differentiate into functionally active neurons. The data demonstrated that ex vivo-expanded human adult DPSCs responded to neuronal inductive conditions both in vitro and in vivo. Human adult DPSCs, but not human foreskin fibroblasts (HFFs), acquired a neuronal morphology, and expressed neuronal-specific markers at both the gene and protein levels. Culture-expanded DPSCs also exhibited the capacity to produce a sodium current consistent with functional neuronal cells when exposed to neuronal inductive media. Furthermore, the response of human DPSCs and HFFs to endogenous neuronal environmental cues was determined in vivo using an avian xenotransplantation assay. DPSCs expressed neuronal markers and acquired a neuronal morphology following transplantation into the mesencephalon of embryonic day-2 chicken embryo, whereas HFFs maintained a thin spindle fibroblastic morphology. We propose that adult human DPSCs provide a readily accessible source of exogenous stem/precursor cells that have the potential for use in cell-therapeutic paradigms to treat neurological disease. TRATAMIENTO POTENCIAL JOURNAL

52 TÍTULO ABSTRACT RESUMEN Las células madre dentales fueron utilizadas terapéuticamente para tratar ratas con Mal de Parkinson logrando mejoras en el sistema nervioso de los animales. Esto abre la posibilidad de tratar el Parkinson en humanos. Problemas Neurológicos Stem cells from human-exfoliated deciduous teeth can differentiate into dopaminergic neuron-like cells. Wang JWang J, Wang X, Sun Z, Wang X, Yang H, Shi S, Wang S., Sun X, Yahi S, Stem Cells Dev. 2010 Sep;19(9):1375-83. Stem cells from human exfoliated deciduous teeth (SHED) have been identified as a novel population of postnatal stem cells capable of differentiating into neural cells, odontogenic cells, and adipocytes. SHED were reported to differentiate into neural cells based on cellular morphology and the expression of early neuronal markers when cultured under neural inductive conditions. This study therefore investigated the therapeutic efficacy of SHED in alleviating Parkinson's disease (PD) in a rat model. We found that SHED could be induced to form neural-like spheres in a medium optimized for neural stem cells in vitro. After incubation with a cocktail of cytokines including sonic hedgehog, fibroblast growth factor 8, glial cell line-derived neurotrophic factor, and forskolin, these SHED-derived spheres further differentiated into a cell population that contained specific dopaminergic neurons. Moreover, transplantation of SHED spheres into the striatum of parkinsonian rats partially improved the apomorphine-evoked rotation of behavorial disorders compared to transplantation of control SHED. Our data indicate that SHED, potentially derived from neural crest cells, may be an optimal source of postnatal stem cells for PD treatment. TRATAMIENTO POTENCIAL JOURNAL

53 TÍTULO ABSTRACT RESUMEN Este artículo repasa la potencialidad que tienen las células madre dentales en generar tejido nervioso y actuar como inmunomoduladores y realizar tareas de neuro-protección y neurotróficas. En ese sentido las Células Madre Dentales se pueden utilizar para tratar la Isquemia Cerebral. Problemas Neurológicos Potential role of dental stem cells in the cellular therapy of cerebral ischemia. Yalvac MEYalvac ME, Rizvanov AA, Kilic E, Sahin F, Mukhamedyarov MA, Islamov RR, Palotás A.AA, Kilic EkhamedyIslamovtás A. Curr Pharm Des. 2009;15(33):3908-16. Stem cell based therapies for cerebral ischemia (CI) utilize different cell sources including embryonic stem cells (ESCs), neural stem cells (NSCs), umbilical cord blood cells (UCBCs), mesenchymal stem cells (MSCs), and some immortalized cell lines. To date, experimental studies showed that all of these cell sources have been successful to some extent in attenuating the ischemic damage and improving functional recovery after brain injury. Bone marrow derived MSCs seem to be the most widely used and well characterized cell source, which can be also employed for autologous transplantation. Currently, there are two main theories behind the therapeutic effect of stem cell transplantation for treating CIs. The first concept is cell replacement theory in which transplanted stem cells differentiate into progenitor and specialized somatic cells to supersede dying cells. The other hypothesis is based on immuno-modulatory, neuro-protective and neuro-trophic abilities of stem cells which help reducing stroke size and increasing the recovery of behavioral functions. Recent studies focusing on alternative stem cell sources have revealed that dental stem cells (DSCs), including dental pulp stem cells (DPSCs) and dental follicle cells (DFCs) possess properties of MSCs and NSCs. They differentiate into neural linage cells and some other cell types such as osteocytes, adipocytes, chondrocytes, muscle cells and hepatocytes. This review is intended to examine stem cell therapy approaches for CI and emphasize potential use of DSCs as an alternative cell source for the treatment of brain ischemia. TRATAMIENTO POTENCIAL JOURNAL

54 Regeneración de Córnea Resumen del Tema: Las Células madre dentales tiene la potencialidad de diferenciarse en células límbicas (del ojo) y se están utilizando en la regenración de córneas.

55 TÍTULO ABSTRACT RESUMEN Células madre dentales humanas fueron utilizadas terapéuticamente para regenerar córneas en un modelo animal. Regeneración de Córnea Corneal reconstruction with tissue-engineered cell sheets composed of human immature dental pulp stem cells. Gomes JAGomes JA, Geraldes Monteiro B, Melo GB, Smith RL, Cavenaghi Pereira da Silva M, Lizier NF, Kerkis A, Cerruti H, Kerkis I. Monteiro B, Melo Gi Pereiva M, Lierkis A, Cerruti H, Kerkis I Invest Ophthalmol Vis Sci. 2010 Mar;51(3):1408-14. Epub 2009 Nov 5. OBJECTIVES: To determine the outcome of the use of a tissue-engineered cell sheet composed of human undifferentiated immature dental pulp stem cells (hIDPSC) for ocular surface reconstruction in an animal model of total limbal stem cell deficiency (LSCD). RESULTS: Corneal transparency of the rabbit eyes that underwent hIDPSC transplantation was improved throughout the follow-up, while the control corneas developed total conjunctivalization and opacification. Rabbits from the MCB group showed clearer corneas with less neovascularization. The clinical data were confirmed by histologic analysis that showed healthy uniform corneal epithelium, especially in the MCB group. The presence of hIDPSC was detected using an anti-hIDPSC antibody. The corneal tissue also showed positive immunostaining with anti-human antibodies. In the control corneas, none of these antigens were detected. CONCLUSIONS: Overall, these data showed that transplantation of a tissue-engineered hIDPSC sheet was successful for the reconstruction of corneal epithelium in an animal model of LSCD. TRATAMIENTO POTENCIAL JOURNAL

56 TÍTULO ABSTRACT RESUMEN En este estudio, se tomaron Células madre dentales humanas y fueron transformadas en células límbicas que sirven para la reconstrucción de córneas. Regeneración de Córnea Human immature dental pulp stem cells share key characteristic features with limbal stem cells. Monteiro BGMonteiro BG, Serafim RC, Melo GB, Silva MC, Lizier NF, Maranduba CM, Smith RL, Kerkis A, Cerruti H, Gomes JA, Kerkis I. Melo GB, zier NFba CM, Serkis A, Gomes JA, Ke Cell Prolif. 2009 Oct;42(5):587-94. Epub 2009 Jul 14. Limbal stem cells (LSC) are self-renewing, highly proliferative cells in vitro, which express a set of specific markers and in vivo have the capacity to reconstruct the entire corneal epithelium in cases of ocular surface injury. Currently, LSC transplantation is a commonly used procedure in patients with either uni- or bilateral total limbal stem cells deficiency (TLSCD). Although LSC transplantation holds great promise for patients, several problems need to be overcome. In order to find an alternative source of cells that can partially substitute LSC in cornea epithelium reconstruction, we aimed at investigating whether human immature dental pulp stem cells (hIDPSC) would present similar key characteristics as LSC and whether they could be used for corneal surface reconstruction in a rabbit TLSCD model. We have demonstrated, using immunohistochemistry and reverse transcription-polymerase chain reaction, that hIDPSCs express markers in common with LSC, such as ABCG2, integrin beta1, vimentin, p63, connexin 43 and cytokeratins 3/12. They were also capable of reconstructing the eye surface after induction of unilateral TLSCD in rabbits, as shown by morphological and immunohistochemical analysis using human- specific antibodies against limbal and corneal epithelium. Our data suggest that hIDPSCs share similar characteristics with LSC and might be used as a potential alternative source of cells for corneal reconstruction. TRATAMIENTO POTENCIAL JOURNAL

57 Regeneración de Piel Resumen del Tema: Las Células madre dentales humanas tienen la posibilidad de diferenciarse en células formadoras de piel por lo que se están utilizando para tratar problemas de quemaduras y heridas epiteliales.

58 TÍTULO ABSTRACT RESUMEN Células madre dentales humanas fueron capaces de regenerar heridas en un modelo animal lo que abre la posibilidad de tratar heridas y problemas de la piel con células madre en humanos. Regeneración de Piel Stem cells from human exfoliated deciduous teeth (SHED) enhance wound healing and the possibility of novel cell therapy. Nishino YNishino Y, Yamada Y, Ebisawa K, Nakamura S, Okabe K, Umemura E, Hara K, Ueda M. Ebisawara S, Okara E, Hara Cytotherapy. 2011 May;13(5):598-605. Epub 2011 Feb 22 In recent years, stem cells from human exfoliated deciduous teeth (SHED) have received attention as a novel stem cell source with multipotent potential. We examined the effect on wound-healing promotion with unique stem cells from deciduous teeth as a medical waste. Methods. An excisional wound-splinting mouse model was used and the effect of wound healing among SHED, human mesenchymal stromal cells (hMSCs), human fibroblasts (hFibro) and a control (phosphate-buffered saline; PBS) was evaluated by macroscopy, histology and enzyme-linked immunosorbent assay (ELISA), and the expression of hyaluronan (HA), which is related to wound healing, investigated. Results. SHED and hMSCs accelerated wound healing compared with hFibro and the control. There was a statistically significant difference in wound healing area among hFibro, hMSCs and SHED compared with the control after day 5. At days 7 and 14 after cell transplantation, the histologic observation showed that transplanted PKH26-positive cells were surrounded by human HA binding protein, especially in hMSCs and SHED. HA expression volume values were 1558.41 ± 60.33 (control), 2092.75 ± 42.56 (hFibro), 2342.07 ± 188.10 (hMSCs) and 2314.85 ± 164.91 (SHED) ng/mg, respectively, and significantly higher in hMSCs and SHED compared with hFibro and control at days 7 and 14 (P < 0.05). Conclusions. Our results show that SHED hMSCs have similar effects of wound-healing promotion as hFibro and controls. This implies that SHED might offer a unique stem cell resource and the possibility of novel cell therapies for wound healing in the future. TRATAMIENTO POTENCIAL JOURNAL

59 Diabetes Resumen del Tema: Las Células madre dentales pueden ser diferenciadas en células pancreáticas productoras de insulina, lo que abre la posibilidad de tratar la diabetes (tipo 1 y 2).

60 TÍTULO ABSTRACT RESUMEN Células madre dentales fueron diferenciadas en células pancreáticas productoras de insulina, lo que abre la posibilidad de tratar la diabetes con células madre de los dientes. Diabetes Differentiation of Dental Pulp Stem Cells into Islet-like Aggregates. Govindasamy VGovindasamy V, Ronald VS, Abdullah AN, Nathan KR, Ab Aziz ZA, Abdullah M, Musa S, Kasim NH, Bhonde RR.dullah AN, Ab Aziz Z, Musa S, Bhonde RR. J Dent Res. 2011 May;90(5):646-52. Epub 2011 Feb 18. The post-natal dental pulp tissue contains a population of multipotent mesenchymal progenitor cells known as dental pulp stromal/stem cells (DPSCs), with high proliferative potential for self- renewal. In this investigation, we explored the potential of DPSCs to differentiate into pancreatic cell lineage resembling islet-like cell aggregates (ICAs). We isolated, propagated, and characterized DPSCs and demonstrated that these could be differentiated into adipogenic, chondrogenic, and osteogenic lineage upon exposure to an appropriate cocktail of differentiating agents. Using a three-step protocol reported previously by our group, we succeeded in obtaining ICAs from DPSCs. The identity of ICAs was confirmed as islets by dithiozone-positive staining, as well as by expression of C-peptide, Pdx-1, Pax4, Pax6, Ngn3, and Isl-1. There were several-fold up-regulations of these transcription factors proportional to days of differentiation as compared with undifferentiated DPSCs. Day 10 ICAs released insulin and C-peptide in a glucose-dependent manner, exhibiting in vitro functionality. Our results demonstrated for the first time that DPSCs could be differentiated into pancreatic cell lineage and offer an unconventional and non- controversial source of human tissue that could be used for autologous stem cell therapy in diabetes. TRATAMIENTO POTENCIAL JOURNAL

61 Regeneración Hepática Resumen del Tema: Las Células madre dentales tienen la potencialidad de diferenciarse en células hepáticas (misma morfología y funcionalidad), abriendo la posibilidad de tratar enfermedades del hígado.

62 TÍTULO ABSTRACT RESUMEN Las Células madre dentales fueron diferenciadas en células hepáticas abriendo la posibilidad de tratar enfermedades del hígado. Regeneración Hepática Multipotent cells from the human third molar: feasibility of cell- based therapy for liver disease. Ikeda EIkeda E, Yagi K, Kojima M, Yagyuu T, Ohshima A, Sobajima S, Tadokoro M, Katsube Y, Isoda K, Kondoh M, Kawase M, Go MJ, Adachi H, Yokota Y, Kirita T, Ohgushi H. Kojimyuu T, OSobajimaro M, Katda K, Kond M, Go MJ,okota Y, hgushi Differentiation. 2008 May;76(5):495-505. Epub 2007 Dec 17. Adult stem cells have been reported to exist in various tissues. The isolation of high-quality human stem cells that can be used for regeneration of fatal deseases from accessible resources is an important advance in stem cell research. In the present study, we identified a novel stem cell, which we named tooth germ progenitor cells (TGPCs), from discarded third molar, commonly called as wisdom teeth. We demonstrated the characterization and distinctiveness of the TGPCs, and found that TGPCs showed high proliferation activity and capability to differentiate in vitro into cells of three germ layers including osteoblasts, neural cells, and hepatocytes. TGPCs were examined by the transplantation into a carbon tetrachloride (CCl4)-treated liver injured rat to determine whether this novel cell source might be useful for cell-based therapy to treat liver diseases. The successful engraftment of the TGPCs was demonstrated by PKH26 fluorescence in the recipient's rat as to liver at 4 weeks after transplantation. The TGPCs prevented the progression of liver fibrosis in the liver of CCl4- treated rats and contributed to the restoration of liver function, as assessed by the measurement of hepatic serum markers aspartate aminotransferase and alanine aminotransferase. Furthermore, the liver functions, observed by the levels of serum bilirubin and albumin, appeared to be improved following transplantation of TGPCs. These findings suggest that multipotent TGPCs are one of the candidates for cell-based therapy to treat liver diseases and offer unprecedented opportunities for developing therapies in treating tissue repair and regeneration. TRATAMIENTO POTENCIAL JOURNAL

63 TÍTULO ABSTRACT RESUMEN Las Células madre dentales fueron diferenciadas en células con la misma funcionalidad y la misma morfología que las células hepáticas. Regeneración Hepática Deciduous and permanent dental pulp mesenchymal cells acquire hepatic morphologic and functional features in vitro. Ishkitiev NIshkitiev N, Yaegaki K, Calenic B, Nakahara T, Ishikawa H, Mitiev V, Haapasalo M.Calenic BT, Ishikav V, Haapa J Endod. 2010 Mar;36(3):469-74. Mesenchymal stem cells display extensive proliferative capacity of multilineage differentiation. The stromal compartment of mesenchymal tissues is considered to harbor stem cells. We assessed the endodermal differentiation of mesenchymal cells from deciduous and wisdom tooth pulp. METHODS: Dental mesenchymal cells were isolated and expanded in vitro. After cell cultures had been established, cells were characterized using known stem cell markers. For hepatic differentiation the media was supplemented with hepatic growth factor, dexamethasone, Insulin-Transferrin- Selenium-X, and oncostatin. RESULTS: Both cultures showed a number of cells positive for specific hepatic markers including alpha-fetoprotein, albumin, and hepatic nuclear factor 4alpha after differentiation. Also, small clusters of cells positive for insulin-like growth factor 1 were found. The concentration of urea increased significantly in the media. Moreover, a significant amount of glycogen was found in the cells. CONCLUSION: Because the cells proved to produce specific hepatic proteins and to start functions specific for hepatocytes, such as storing glycogen and urea production, we may state that the mesenchymal cell cultures from wisdom and deciduous tooth pulp acquired morphologic and functional characteristics of hepatocytes. TRATAMIENTO POTENCIAL JOURNAL

64 Regeneración Muscular y Cirugía Plástica Resumen del Tema: Las Células madre dentales tienen la posibilidad de ser diferenciadas en miocitos, que son las células musculares y pueden llegar a tratar problemas de este tipo. Por otro lado, las células madre de origen dental tienen la posibilidad de ayudar en la cirugía plástica y en cirugías reconstructivas.

65 TÍTULO ABSTRACT RESUMEN Las Células madre del tejido periodontal pueden ser utilizadas en cirugías reconstructivas y cirugías plásticas para reparar y regenerar el tejido dañado. Transplantation of mesenchymal stem cells is an optimal approach for plastic surgery. Fang DFang D, Seo BM, Liu Y, Sonoyama W, Yamaza T, Zhang C, Wang S, Shi S., Liu ama Wa T, ZhangShi S. Stem Cells. 2007 Apr;25(4):1021-8. Epub 2006 Dec 14. Mesenchymal stem cells (MSCs) are able to differentiate into a variety of cell types, offering promising approaches for stem cell-mediated tissue regeneration. Here, we explored the potential of utilizing MSCs to reconstruct orofacial tissue, thereby altering the orofacial appearance. We demonstrated that bone marrow MSCs were capable of generating bone structures and bone-associated marrow elements on the surfaces of the orofacial bone. This resulted in significant recontouring of the facial appearance in mouse and swine. Notably, the newly formed bone and associated marrow tissues integrated with the surfaces of the recipient bones and re-established a functional bone marrow organ- like system. These data suggested that MSC-mediated tissue regeneration led to a body structure extension, with the re-establishment of all functional components necessary for maintaining the bone and associated marrow organ. In addition, we found that the subcutaneous transplantation of another population of MSCs, the human periodontal ligament stem cells (PDLSCs), could form substantial amounts of collagen fibers and improve facial wrinkles in mouse. By contrast, bone marrow MSCs failed to survive at 8 weeks post-transplantation under the conditions used for the PDLSC transplantation. This study suggested that the mutual interactions between donor MSCs and recipient microenvironment determine long-term outcome of the functional tissue regeneration. Regeneración Muscular y Cirugía Plástica TRATAMIENTO POTENCIAL JOURNAL

66 TÍTULO ABSTRACT RESUMEN Las Células madre dentales fueron utilizadas para tratar distrofia muscular en un modelo canino. Esto abre la posibilidad para tratar problemas musculares y distróficos en humanos. Early transplantation of human immature dental pulp stem cells from baby teeth to golden retriever muscular dystrophy (GRMD) dogs: Local or systemic? Kerkis IKerkis I, Ambrosio CE, Kerkis A, Martins DS, Zucconi E, Fonseca SA, Cabral RM, Maranduba CM, Gaiad TP, Morini AC, Vieira NM, Brolio MP, Sant'Anna OA, Miglino MA, Zatz M. CE, KerkisS, Zuccoeca SA, Caanduba CM Morini AC Brolio Ma OA, Miglin J Transl Med. 2008 Jul 3;6:35. 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. 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 researches. Regeneración Muscular y Cirugía Plástica TRATAMIENTO POTENCIAL JOURNAL

67 Enfermedades Auto-Inmunes y Lupus Resumen del Tema: Las Células madre dentales pueden ser utilizadas para tratar enfermedades auto-inmunes como el Lupus.

68 TÍTULO ABSTRACT RESUMEN Las Células madre dentales fueron utilizadas para tratar Lupus Erythematosus en un modelo animal. Los resultados muestran que las células madre de dientes temporales pueden ser utilizadas para tratar enfermedades auto-inmunes como el Lupus. Enfermedades Auto-Inmunes y Lupus Immunomodulatory properties of stem cells from human exfoliated deciduous teeth. Yamaza TYamaza T, Kentaro A, Chen C, Liu Y, Shi Y, Gronthos S, Wang S, Shi S.A, Chen Ci Y, GS, Wahi S. Stem Cell Res Ther. 2010 Mar 15;1(1):5. INTRODUCTION : Stem cells from human exfoliated deciduous teeth (SHED) have been identified as a population of postnatal stem cells capable of differentiating into osteogenic and odontogenic cells, adipogenic cells, and neural cells. Herein we have characterized mesenchymal stem cell properties of SHED in comparison to human bone marrow mesenchymal stem cells (BMMSCs). METHODS : We used in vitro stem cell analysis approaches, including flow cytometry, inductive differentiation, telomerase activity, and Western blot analysis to assess multipotent differentiation of SHED and in vivo implantation to assess tissue regeneration of SHED. In addition, we utilized systemic SHED transplantation to treat systemic lupus erythematosus (SLE)-like MRL/lpr mice. RESULTS : We found that SHED are capable of differentiating into osteogenic and adipogenic cells, expressing mesenchymal surface molecules (STRO-1, CD146, SSEA4, CD73, CD105, and CD166), and activating multiple signaling pathways, including TGFbeta, ERK, Akt, Wnt, and PDGF. Recently, BMMSCs were shown to possess an immunomodulatory function that leads to successful therapies for immune diseases. We examined the immunomodulatory properties of SHED in comparison to BMMSCs and found that SHED had significant effects on inhibiting T helper 17 (Th17) cells in vitro. Moreover, we found that SHED transplantation is capable of effectively reversing SLE-associated disorders in MRL/lpr mice. At the cellular level, SHED transplantation elevated the ratio of regulatory T cells (Tregs) via Th17 cells. CONCLUSIONS : These data suggest that SHED are an accessible and feasible mesenchymal stem cell source for treating immune disorders like SLE. TRATAMIENTO POTENCIAL JOURNAL

69 Otros Artículos

70 Biological approaches toward dental pulp regeneration by tissue engineering. Sun HH, Jin T, Yu Q, Chen FM. 2011 April 5 Comparative Analysis of Telomere Length, Telomerase and Reverse Transcriptase Activity in Human Dental Stem Cells. Jeon BG, Kang EJ, Mohana Kumar B, Maeng GH, Ock SA, Kwack DO, Park BW, Rho GJ. 2011 March 8 Expression Pattern of Oct-4, Sox2, and c-Myc in the Primary Culture of Human Dental Pulp Derived Cells. Liu L, Wei X, Ling J, Wu L, Xiao Y. 2011 Feb 25 Stem cell-based biological tooth repair and regeneration Ana Angelova Volponi, Yvonne Pang and Paul T. Sharpe Advanced Centre for Biochemical Engineering, University College London, London, UK - Web 2011 Jan 05 Human dental pulp stem cells produce mineralized matrix in 2D and 3D cultures. M. Riccio, E. Resca, T. Maraldi, A. Pisciotta, A. Ferrari, G. Bruzzesi, A. De Po1 European Journal of Histochemistry - 2010 Oct 10 Suppression of T Cell Proliferation By Root Apical Papilla Stem Cells in Vitro. G Ding, Y Liu, Y An, C Zhang, S Shi, W Wang, and S Wang. Cells Tissues Organs - 2010 191(5):357-364. Effect of Cryopreservation on Biological and Immunological Properties of Stem Cells From Apical Papilla. G Ding, Y Liu, Y An, C Zhang, S Shi, W Wang, and S Wang. J Cell Physiol - 2010 223(2):415-422. Tgf-Beta Stimulates Glial-Like Differentiation in Murine Dental Follicle Precursor Cells (Mdfpcs). O Felthaus, W Ernst, O Driemel, TE Reichert, G Schmalz, and C Morsczeck. Neurosci Lett - 2010 471(3):179-184. Inherent Differential Propensity of Dental Pulp Stem Cells Derived From Human Deciduous and Permanent Teeth. V Govindasamy, AN Abdullah, VS Ronald, S Musa, ZA Ab Aziz, RB Zain, S Totey, RR Bhonde, and NH Abu Kasim. J Endod - 2010 36(9):1504-1515. Gene-Modified Stem Cells Combined With Rapid Prototyping Techniques: A Novel Strategy for Periodontal Regeneration. H He, J Cao, D Wang, B Gu, H Guo, and H Liu. Stem Cell Rev - 2010 6(1):137-141. Expression of Multiple Stem Cell Markers in Dental Pulp Cells Cultured in Serum-Free Media. TM Hirata, N Ishkitiev, K Yaegaki, B Calenic, H Ishikawa, T Nakahara, V Mitev, T Tanaka, and M Haapasalo. J Endod - 2010 36(7):1139-1144. Osteogenic Differentiation of Human Dental Pulp-Derived Stem Cells Under Various Ex-Vivo Culture Conditions. J Karbanodva, T Soukup, J Suchanek, and J Mokry. Acta Medica (Hradec Kralove) - 2010 53(2):79-84. The Osteoblastic Differentiation of Dental Pulp Stem Cells and Bone Formation on Different Titanium Surface Textures. C Mangano, A De Rosa, V Desiderio, R d'Aquino, A Piattelli, F De Francesco, V Tirino, F Mangano, and G Papaccio. Biomaterials - 2010 31(13):3543-3551. Otros Artículos

71 Osteogenic Properties of Human Dental Pulp Stem Cells. G Mori, M Centonze, G Brunetti, A Ballini, A Oranger, C Mori, L Lo Muzio, S Tete, F Ciccolella, S Colucci, M Grano, and FR Grassi. J Biol Regul Homeost Agents - 2010 24(2):167-175. Proteomic Characterization of Mesenchymal Stem Cell-Like Populations Derived From Ovine Periodontal Ligament, Dental Pulp and Bone Marrow: Analysis of Differentially Expressed Proteins. KM Mrozik, PS Zilm, C Bagley, S Hack, P Hoffmann, S Gronthos, and PM Bartold. Stem Cells Dev - 2010 19(10):1485-1499. Explant-Derived Human Dental Pulp Stem Cells Enhance Differentiation and Proliferation Potentials. L Spath, V Rotilio, M Alessandrini, G Gambara, L De Angelis, M Mancini, TA Mitsiadis, E Vivarelli, F Naro, A Filippini, and G Papaccio. J Cell Mol Med - 2010 14(6B):1635-1644. Stem Cells From Human Exfoliated Deciduous Teeth--Isolation, Long Term Cultivation and Phenotypical Analysis. J Suchanek, B Visek, T Soukup, SK El-Din Mohamed, R Ivancakova, J Mokry, EH Aboul-Ezz, and A Omran. Acta Medica (Hradec Kralove) - 2010 53(2):93-99. Isolation and Characterization of Stem Cells Derived From Human Third Molar Tooth Germs of Young Adults: Implications in Neo-Vascularization, Osteo-, Adipo- and Neurogenesis. ME Yalvac, M Ramazanoglu, AA Rizvanov, F Sahin, OF Bayrak, U Salli, A Palotas, and GT Kose. Pharmacogenomics J - 2010 10(2):105-113. A Feasibility of Useful Cell-Based Therapy By Bone Regeneration With Deciduous Tooth Stem Cells, Dental Pulp Stem Cells, Or Bone Marrow-Derived Mesenchymal Stem Cells for Clinical Study Using Tissue Engineering Technology. Y Yamada, S Nakamura, K Ito, T Sugito, R Yoshimi, T Nagasaka, and M Ueda. Tissue Eng Part A - 2010 16(6):1891-1900. A Journey From Dental Pulp Stem Cells to a Bio-Tooth. M Yan, Y Yu, G Zhang, C Tang, and J Yu. Stem Cell Rev - 2010; Epub ahead of print Dental Tissue Regeneration - a Mini-Review. AH Yen, and PC Yelick. Gerontology - 2010 May 6; Epub ahead of print Stem Cell-Based Dental Tissue Engineering. P Zivkovic, V Petrovic, S Najman, and V Stefanovic. ScientificWorldJournal - 2010 10:901-916. Dental tissue -- new source for stem cells. Petrovic V, Stefanovic V. Scientific World Journal - 2009 Oct 14;9:1167-77 Mesenchymal stem cells derived from dental tissues vs. those from other sources: their biology and role in regenerative medicine. Huang GT, Gronthos S, Shi S. J Dent Res - 2009 Sep;88(9):792-806 Hypoxia affects the proliferation and migration abilities of dental pulp J. Quan, Q. Gong, H. Jiang, and J. Ling IADR Pan Asian Pacific Federation Conference - 2009 Sept Hepatic lineage differentiation of milk and third molar pulp cells N. Ishkitiew, T. Nakahara, T. Sato, V. Mitev, and K. Yaegaki IADR/AADR/CADR 87th General Session Conference - 2009 April

72 A simple and efficient method for generating Nurr1-positive neuronal stem cells from human wisdom teeth (tNSC) and the potential of tNSC for stroke therapy. Yang KL, Chen MF, Liao CH, Pang CY, Lin PY. Cytotherapy - 2009;11(5):606-17 Simultaneous PKC and cAMP activation induces differentiation of human dental pulp stem cells into functionally active neurons. Király M, Porcsalmy B, Pataki A, Kádár K, Jelitai M, Molnár B, Hermann P, Gera I, Grimm WD, Ganss B, Zsembery A, Varga G. Neurochem Int - 2009 Sep;55(5):323-32 Optimized cryopreservation method for human dental pulp-derived stem cells and their tissues of origin for banking and clinical use. Király M, Porcsalmy B, Pataki A, Kádár K, Jelitai M, Molnár B, Hermann P, Gera I, Grimm WD, Ganss B, Zsembery A, Varga G. Cryobiology - 2009 Oct;59(2):150-7 Identification of novel epithelial stem cell-like cells in human deciduous dental pulp. Nam H, Lee G. Biochem Biophys Res Commun - 2009 Aug 14;386(1):135-9 Hypoxia-amplified proliferation of human dental pulp cells. Sakdee JB, White RR, Pagonis TC, Hauschka PV. J Endod - 2009 Jun;35(6):818-23 Dental pulp stem cells and their characterization. Suchanek J, Soukup T, Visek B, Ivancakova R, Kucerova L, Mokry J. Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub - 2009 Mar;153(1):31-5 Differentiation and regenerative capacities of human odontoma-derived mesenchymal cells. Song JS, Stefanik D, Damek-Poprawa M, Alawi F, Akintoye SO. Differentiation - 2009 Jan;77(1):29-37 Evaluation of pluripotency in human dental pulp cells. Koyama N, Okubo Y, Nakao K, Bessho K. J Oral Maxillofac Surg - 2009 Mar;67(3):501-6 Dental pulp stem cells: what, where, how? Sloan AJ, Waddington RJ. Int J Paediatr Dent - 2009 Jan;19(1):61-70 Cardiac differentiation is driven by NKX2.5 and GATA4 nuclear translocation in tissue-specific mesenchymal stem cells. Armiñán A, Gandía C, Bartual M, García-Verdugo JM, Lledó E, Mirabet V, Llop M, Barea J, Montero JA, Sepúlveda P. Stem Cells Dev - 2009 Jul-Aug;18(6):907-18 Regenerating dentistry: the new realm of stem cells. Murray P, Pocock N. Todays FDA - 2008 Oct;20(10):35-6 Whole-tooth regeneration: it takes a village of scientists, clinicians, and patients. Snead ML. J Dent Educ - 2008 Aug;72(8):903-11 Self-assembling peptide amphiphile nanofibers as a scaffold for dental stem cells. Galler KM, Cavender A, Yuwono V, Dong H, Shi S, Schmalz G, Hartgerink JD, D'Souza RN. Tissue Eng Part A - 2008 Dec;14(12):2051-8 Dental pulp tissue engineering with stem cells from exfoliated deciduous teeth. Cordeiro MM, Dong Z, Kaneko T, Zhang Z, Miyazawa M, Shi S, Smith AJ, Nör JE. J Endod - 2008 Aug;34(8):962-9

73 Gene expression of nestin, collagen type I and type III in human dental follicle cells after cultivation in serum-free medium. Morsczeck C, Ernst W, Florian C, Reichert TE, Proff P, Bauer R, Müller-Richter U, Driemel O. Oral Maxillofac Surg - 2008 Jul;12(2):89-92 Inflammatory and immunological aspects of dental pulp repair. Goldberg M, Farges JC, Lacerda-Pinheiro S, Six N, Jegat N, Decup F, Septier D, Carrouel F, Durand S, Chaussain-Miller C, Denbesten P, Veis A, Poliard A. Pharmacol Res - 2008 Aug;58(2):137-47 A novel stem cell source for vasculogenesis in ischemia: subfraction of side population cells from dental pulp. Iohara K, Zheng L, Wake H, Ito M, Nabekura J, Wakita H, Nakamura H, Into T, Matsushita K, Nakashima M. Stem Cells - 2008 Sep;26(9):2408-18 Comparison between genetic portraits of osteoblasts derived from primary cultures and osteoblasts obtained from human pulpar stem cells. Carinci F, Papaccio G, Laino G, Palmieri A, Brunelli G, D'Aquino R, Graziano A, Lanza V, Scapoli L, Martinelli M, Pezzetti F. J Craniofac Surg - 2008 May;19(3):616-25; discussion 626-7 Adult human dental pulp stem cells differentiate toward functionally active neurons under appropriate environmental cues. Arthur A, Rychkov G, Shi S, Koblar SA, Gronthos S. Stem Cells - 2008 Jul;26(7):1787-95 The hidden treasure in apical papilla: the potential role in pulp/dentin regeneration and bioroot engineering. Huang GT, Sonoyama W, Liu Y, Liu H, Wang S, Shi S. J Endod - 2008 Jun;34(6):645-51 Stem cells and the future of dental care. Mao JJ. N Y State Dent J - 2008 Mar;74(2):20-4 Postnatal stem/progenitor cells derived from the dental pulp of adult chimpanzee. Cheng PH, Snyder B, Fillos D, Ibegbu CC, Huang AH, Chan AW. BMC Cell Biol - 2008 Apr 22;9:20 A paradigm shift in endodontic management of immature teeth: conservation of stem cells for regeneration. Huang GT. J Dent - 2008 Jun;36(6):379-86 Human dental pulp stem cells differentiate into neural crest-derived melanocytes and have label-retaining and sphere-forming abilities. Stevens A, Zuliani T, Olejnik C, LeRoy H, Obriot H, Kerr-Conte J, Formstecher P, Bailliez Y, Polakowska RR. Stem Cells Dev - 2008 Dec;17(6):1175-84 An ultrastructural investigation of tissue-engineered pulp constructs implanted within endodontically treated teeth. Gotlieb EL, Murray PE, Namerow KN, Kuttler S, Garcia-Godoy F. J Am Dent Assoc - 2008 Apr;139(4):457-65 Defining properties of neural crest-derived progenitor cells from the apex of human developing tooth. Degistirici O, Jaquiery C, Schönebeck B, Siemonsmeier J, Götz W, Martin I, Thie M. Tissue Eng Part A - 2008 Feb;14(2):317-30 Isolation and characterization of dental pulp stem cells from a supernumerary tooth. Huang AH, Chen YK, Lin LM, Shieh TY, Chan AW. J Oral Pathol Med - 2008 Oct;37(9):571-4 Characterisation of human dental stem cells and buccal mucosa fibroblasts. Lindroos B, Mäenpää K, Ylikomi T, Oja H, Suuronen R, Miettinen S. Biochem Biophys Res Commun- 2008 Apr 4;368(2):329-35

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