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

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The reconstruction of the craniofacial skeleton in development between 2 and 10 years old, remains a major challenge for reconstructive plastic surgery. Local autogenous bone is practically unavailable, the distant bone graft has significant morbidity and use of alloplastic materials is incompatible with the growing facial skeleton. With the advent of bioengineered tissue, however, osteogenesis induced by the use of mesenchymal stem cells associated with biomaterials has become a potential solution to the shortage bone-related morbidity and donor bone in the region in pediatric patients.

The association of mesenchymal stem cells to biomaterials has provided new bone formation and a significant reduction of morbidity, for rehabilitation of the alveolar bone in patients with cleft lip palate.

To perform the rehabilitation of alveolar bone cleft, other donor regions of bone (iliac crest, ribs, skull) suffer morbidity for obtaining bone to be used in alveolar bone grafting. In order to eliminate the morbidity at the bone donor region for these patients and reduce costs of patient permanence in the operating room the aim of this study is to perform the bone tissue engineering to reconstruct the alveolar bone defect in cleft lip and palate patients using mesenchymal stem cells from deciduous dental pulp associated with a collagen and hydroxyapatite biomaterial (Geistlich Bio-Oss®) through prospective qualitative and quantitative analysis of bone neoformation.

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Researchers at the Nagoya University Graduate School of Medicine in Japan are exploiting the neural capabilities of dental pulp stem cells obtained from human wisdom teeth. In the study, researchers used stem cells recovered from wisdom teeth to treat a rat population with transected spinal cords. The rats were found to have recovered locomotive function in their hind legs and regenerated damaged nerve cells and oligodendrocytes. In a parallel study, stem cells from bone marrow and skin fibroblasts were utilized with statistically significantly lesser effect.

As the study demonstrates, teeth are a valuable source of powerful stem cells. Banking your own adult stem cells today will insure your future health as regenerative therapies, such as these, are developed to treat a wide array of disease, trauma and injury.

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Abstract

The differentiation of stem cells can be directed by the grade of stiffness of the developed tissue cells. For example a rigid extracellular matrix supports the osteogenic differentiation in bone marrow derived mesenchymal stem cells (MSCs). However, less is known about the relation of extracellular matrix stiffness and cell differentiation of ectomesenchymal dental precursor cells. Our study examined for the first time the influence of the surface stiffness on the proliferation and osteogenic differentiation of human dental follicle cells (DFCs). Cell proliferation of DFCs was only slightly decreased on cell culture surfaces with a bone-like stiffness. The osteogenic differentiation in DFCs could only be initiated with a dexamethasone based differentiation medium after using varying stiffness. Here, the softest surface improved the induction of osteogenic differentiation in comparison to that with the highest stiffness. In conclusion, different to bone marrow derived MSCs, soft ECMs have a superior capacity to support the osteogenic differentiation of DFCs.

J Biol Regul Homeost Agents. 2011 Jan-Mar;25(1):57-69.

D' Alimonte I, Nargi E, Mastrangelo F, Falco G, Lanuti P, Marchisio M, Miscia S, Robuffo I, Capogreco M, Buccella S, Caputi S, Caciagli F, Tetè S, Ciccarelli R.

Departments of Biomedical Sciences, University of Chieti, Italy

Cell Transplant. 2011 Mar 8.

Byeong GJ, Kang EJ, Kumar BM, Maeng GH, Ock SA, Kwack DO, Park BW, Rho GJ.

Eur Cell Mater. 2011 Mar 22;21:304-16.

d'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.

Department of Experimental Medicine, Section of Histology, TERM Laboratory, 2nd University of Naples, via L. Armanni 5, I-80138 Naples, [email protected].

Soria JM, Sancho-Tello M, Esparza MA, Mirabet V, Bagan JV, Monleón M, Carda C.

Facultad Ciencias de la Salud, Universidad CEU Cardenal Herrera, Avda Seminario sn. 46113 Moncada, Valencia, Spain. [email protected].

Odontology. 2011 Jan;99(1):1-7. Epub 2011 Jan 27.

Casagrande L, Cordeiro MM, Nör SA, Nör JE.

Department of Cariology, Restorative Sciences and Endodontics, University of Michigan School of Dentistry, 1011 N. University, Rm. 2309, Ann Arbor, MI, 48109-1078, USA.

Eur J Histochem. 2010 Nov 10;54(4):e46.

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

Dept. Anatomy and Histology, University of Modena and Reggio Emilia. [email protected].

J Biomed Mater Res A. 2011 Mar;96(3):528-34. doi: 10.1002/jbm.a.33002. Epub 2011 Jan 4.

Liu J, Jin TC, Chang S, Czajka-Jakubowska A, Clarkson BH.

Department of Cardiology, Restorative Sciences and Endodontics, Dental School, University of Michigan, Ann Arbor, Michigan. [email protected].

Int J Biomater. 2010;2010:915327. Epub 2010 Dec 28.

Lavenus S, Louarn G, Layrolle P.

Inserm U957, Bone Resorption Physiopathology and Primary Bone Tumors Therapy, Faculty of Medicine, University of Nantes - 1 rue Gaston Veil, 44035 Nantes cedex 1, France.

Arch Oral Biol. 2011 Jan 10.

Bakopoulou A, Leyhausen G, Volk J, Tsiftsoglou A, Garefis P, Koidis P, Geurtsen W.

Department of Fixed Prosthesis & Implant Prosthodontics, School of Dentistry, Aristotle University of Thessaloniki, Greece; Department of Conservative Dentistry, Periodontology and Preventive Dentistry, Hannover Medical School, Germany.

Abstract

OBJECTIVE: The aim of this study was to compare the in vitro osteo/odontogenic differentiation potential of mesenchymal stem cells (MSCs) derived from the dental pulp (dental pulp stem cells - DPSCs) or the apical papilla (stem cells from the apical papilla - SCAP) of permanent developing teeth.

DESIGN: DPSCs and SCAP cultures were established from impacted third molars of young healthy donors at the stage of root development. Cultures were analysed for stem cell markers, including STRO-1, CD146, CD34 and CD45 using flow cytometry. Cells were then induced for osteo/odontogenic differentiation by media containing dexamethasone, KH(2)PO(4) and ß-glycerophosphate. Cultures were analysed for morphology, growth characteristics, mineralization potential (Alizarin Red method) and differentiation markers (dentine sialophosphoprotein-DSPP, bone sialoprotein-BSP, osteocalcin-OCN, alkaline phosphatase-ALP), using immunocytochemistry and reverse transcriptase-polymerase chain reaction.

RESULTS: All DPSCs and SCAP cultures were positive for STRO-1, CD146 and CD34, in percentages varying according to cell type and donor, but negative for CD45. Both types of MSCs displayed an active potential for cellular migration, organization and mineralization, producing 3D mineralized structures. These structures progressively expressed differentiation markers, including DSPP, BSP, OCN, ALP, having the characteristics of osteodentin. SCAP, however, showed a significantly higher proliferation rate and mineralization potential, which might be of significance for their use in bone/dental tissue engineering.

CONCLUSIONS: This study provides evidence that different types of dental MSCs can be used in tissue engineering/regeneration protocols as an approachable stem cell source for osteo/odontogenic differentiation and biomineralization that could be further applied for stem cell-based clinical therapies.

Neurochem Int. 2011 Jan 8.

Király M, Kádár K, Horváthy DB, Nardai P, Rácz GZ, Lacza Z, Varga G, Gerber G.

Department of Oral Biology, Semmelweis University, Nagyvarad ter 4, 1089 Budapest, Hungary.

Abstract

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, NFneuronally 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.

Schweiz Monatsschr Zahnmed. 2010;120(10):860-72.

Ulmer FL, Winkel A, Kohorst P, Stiesch M.

Clinic for Dental Prosthodontics and Biomedical Materials Science Center for Dentistry and Oral Medicine Medical University of Hannover, Germany.

J Oral Sci. 2010;52(4):541-52.

Honda MJ, Imaizumi M, Tsuchiya S, Morsczeck C.

Department of Anatomy, Nihon University School of Dentistry, Chiyoda-ku, Tokyo, Japan. [email protected]

Regen Med. 2011 Jan;6(1):67-79.

Lee UL, Jeon SH, Park JY, Choung PH.

Department of Oral & Maxillofacial Surgery, School of Dentistry, Seoul National University, Seoul, South Korea.

Regen Med. 2011 Jan;6(1):111-24.

Galler KM, D'Souza RN.

Department of Operative Dentistry & Periodontology, University of Regensburg, Regensburg, Germany.

Latest Study Advances Muscular Dystrophy Research and Treatment.

Dental Stem Cells, in the latest example of their plasticity, were differentiated into dystrophin producing multi-nucleated muscle cells.   The research, published recently in PLoS One was led by Dr. Jeremy Mao, Professor and Director of the Tissue Engineering and Regenerative Medicine Laboratory (TERML) of Columbia University Medical Center and Chief Scientific Advisor to StemSave.

Dr. Mao’s utilization of myogenic progenitor cells derived from dental stem cells demonstrated significantly higher numbers of dystrophin producing cells than the parent heterogeneous stem cells from which they were derived.  The latest research suggests therapeutic potential for muscle regeneration and has implications for disorders such as Muscular Dystrophy where the inability of the body to produce dystrophin results in health complications. 

The latest research, along with recently published research demonstrating the ability of dental stem cells to differentiate into bone, myocardiocytes (heart muscle) and insulin producing pancreatic beta cells, supports the wisdom of banking stem cells from teeth. 

The ability of dental stem cells to differentiate into various types of cells (often referred to as their ‘plasticity’) makes teeth a source of valuable stem cells to utilize in the treatment of a potentially wide range of trauma and disease.   These valuable stem cells can be recovered during routine dental procedures thus making it both convenient and affordable to bank your own stem cells for use in emerging regenerative therapies.

by Dr. Dean Vafiadis, DDS

Cells Tissues Organs. 2010 Nov 11. 

Karbanová J, Soukup T, Suchánek J, Pytlík R, Corbeil D, Mokrý J.

Department of Histology and Embryology, Charles University in Prague, Faculty of Medicine, Prague, Czech Republic.

We isolated and expanded stem cells from dental pulp from extracted third molars using an innovative culture method consisting of low serum-containing medium supplemented with epidermal growth factor and platelet-derived growth factor BB. We evaluated the differentiation potential of these cells when they were growing either adherently or as micromass/spheroid cultures in various media. Undifferentiated and differentiated cells were analyzed by flow cytometry, immunocytochemistry and immunoblotting. The flow cytometry results showed that the dental pulp stem cells (DPSCs) were positive for mesenchymal stromal cell markers, but negative for hematopoietic markers. Immunocytochemical and/or immunoblotting analyses revealed the expression of numerous stem cell markers, including nanog, Sox2, nestin, Musashi-1 and nucleostemin, whereas they were negative for markers associated with differentiated neural, vascular and hepatic cells. Surprisingly, the cells were only slightly positive for a-smooth muscle actin, and a heterogeneous expression of CD146 was observed. When cultured in osteogenic media, they expressed osteonectin, osteopontin and procollagen I, and in micromass cultures, they produced collagen I. DPSCs cultured in TGF-ß1/3-supplemented media produced extracellular matrix typical of cartilaginous tissue. The addition of vascular endothelial growth factor to serum-free media resulted in the expression of endothelial markers. Interestingly, when cultured in neurogenic media, DPSCs exhibited de novo or upregulated markers of undifferentiated and differentiated neural cells. Collectively, our data show that DPSCs are self-renewing and able to express markers of bone, cartilage, vascular and neural tissues, suggesting their multipotential capacity. Their easy accessibility makes these cells a suitable source of somatic stem cells for tissue engineering.

Cell Transplant. 2010 Nov 5.

Yamada Y, Ito K, Nakamura S, Ueda M, Nagasaka T.

J Biomed Biotechnol. 2010;2010:673513. Epub 2010 Oct 4.

Mokry J, Soukup T, Micuda S, Karbanova J, Visek B, Brcakova E, Suchanek J, Bouchal J, Vokurkova D, Ivancakova R.

Department of Histology and Embryology, Faculty of Medicine in Hradec Kralove, Charles University in Prague, Simkova 870, 50038 Hradec Kralove, Czech Republic. [email protected]

Eur Cell Mater. 2010 Oct 7;20:295-305.

Paino F, Ricci G, De Rosa A, D'Aquino R, Laino L, Pirozzi G, Tirino V, Papaccio G.

Department of Experimental Medicine, Histology and Embryology, Tissue Engineering and Regenerative Medicine (TERM) Division, School of Medicine, Second University of Naples, 5 via L. Armanni, I-80138 Napoli, [email protected].

Shanghai Kou Qiang Yi Xue. 2010 Aug;19(4):398-402.

Zhang XY, Li XT, Zeng XL.

Department of Orthodontics, Peking University School and Hospital of Stomatology. Beijing 100081, China.

Brain Res. 2010 Sep 17.

Nesti C, Pardini C, Barachini S, D'Alessandro D, Siciliano G, Murri L, Petrini M, Vaglini F.

RRMR/CUCCS (Rete Regionale di Medicina Rigenerativa/Center for the Clinical Use of Stem Cells), Italy; Stella Maris Scientific Institute, Pisa, Italy.

PMID: 20854799 [PubMed - as supplied by publisher]

Acta Medica (Hradec Kralove). 2010;53(2):93-9.

Suchánek J, Visek B, Soukup T, El-Din Mohamed SK, Ivancaková R, Mokr? J, Aboul-Ezz EH, Omran A.

Charles University in Prague, Faculty of Medicine, Department of Dentistry and University Hospital Hradec Králové, Czech Republic. [email protected]

J Endod. 2010 Sep;36(9):1504-1515.

Govindasamy V, Abdullah AN, Sainik Ronald V, Musa S, Che Ab Aziz ZA, Zain RB, Totey S, Bhonde RR, Abu Kasim NH.

Stempeutics Research Malaysia Sdn Bhd, Kuala Lumpur, Malaysia

Stem Cells Dev. 2010 Aug 23

Tomic S, Djokic J, Vasilijic S, Vucevic D, Todorovic V, Supic G, Colic M.

Institute for Medical Research, Military Medical Academy, Belgrade, Serbia; [email protected].

In Vitro Cell Dev Biol Anim. 2010 Aug 20.

Govindasamy V, Ronald VS, Totey S, Binti Din S, Mustafa WM, Totey S, Zakaria Z, Bhonde RR.

Stempeutics Research Malaysia, Sdn Bhd (773817-K) Lot 3-i-7, Enterprise 4, Technology Park Malaysia, Bukit Jalil, 57000, Kuala Lumpur, Malaysia

Stem Cells Dev. 2010 Feb 4.

Wang J, Wang X, Sun Z, Wang X, Yang H, Shi S, Wang S.

1 Salivary Gland Disease Center and the Molecular Laboratory for Gene Therapy and Tooth Regeneration, Capital Medical University School of Stomatology , Beijing, People's Republic of China .

Med Hypotheses. 2010 Aug 25.

Zuolin J, Hong Q, Jiali T.

Department of Orthodontics, School of Stomatology, The Fourth Military Medical University, No. 145 West Changle Road, Xi'an 710032, China.

J Cell Physiol. 2010 Jul 12.

S A T, S S, D K.

Department of Periodontics and Oral Medicine, University of Michigan; Ann Arbor, MI USA;

J Endod. 2010 Jul;36(7):1139-44. Epub 2010 Apr 10.

Hirata TM, Ishkitiev N, Yaegaki K, Calenic B, Ishikawa H, Nakahara T, Mitev V, Tanaka T, Haapasalo M.

Department of Oral Health, Nippon Dental University, School of Life Dentistry at Tokyo, Tokyo, Japan.

Arch Oral Biol. 2010 Jul 12.

Iida K, Takeda-Kawaguchi T, Tezuka Y, Kunisada T, Shibata T, Tezuka KI.

Department of Oral and Maxillofacial Science, Gifu University Graduate School of Medicine, 1-1 Yanagido, Gifu City, Gifu 501-1194, Japan.

J Endod. 2010 Aug;36(8):1336-1340. Epub 2010 Jun 23.

Lee SY, Chiang PC, Tsai YH, Tsai SY, Jeng JH, Kawata T, Huang HM.

School of Dentistry, Taipei Medical University, Taipei, Taiwan.

Stem Cell Res Ther. 2010 Mar 15;1(1):5.

Yamaza T, Kentaro A, Chen C, Liu Y, Shi Y, Gronthos S, Wang S, Shi S.

Center for Craniofacial Molecular Biology, University of Southern California School of Dentistry, 2250 Alcazar Street, CSA 103, Los Angeles, CA 90033, USA. [email protected].

Tamaoki N, Takahashi K, Tanaka T, Ichisaka T, Aoki H, Takeda-Kawaguchi T, Iida K, Kunisada T, Shibata T, Yamanaka S, Tezuka K.

Miyoshi K, Tsuji D, Kudoh K, Satomura K, Muto T, Itoh K, Noma T.

Department of Molecular Biology, Institute of Health Biosciences, The University of Tokushima Graduate School, 3-18-15 Kuramoto, Tokushima 770-8504, Japan.

Cell Prolif. 2010 Jun;43(3):219-28.

Pan K, Sun Q, Zhang J, Ge S, Li S, Zhao Y, Yang P.

Department of Periodontology and Institute of Oral Biomedicine, School of Dentistry, Shandong University, Jinan, China.

J Biol Regul Homeost Agents. 2010 Apr-Jun;24(2):167-75.

Mori G, Centonze M, Brunetti G, Ballini A, Oranger A, Mori C, Lo Muzio L, Tetè S, Ciccolella F, Colucci S, Grano M, Grassi FR.

Department of Biomedical Science, University of Foggia Medical School, Foggia, Italy. [email protected]

Regen Med. 2010 May 14.

Alongi DJ, Yamaza T, Song Y, Fouad AF, Romberg EE, Shi S, Tuan RS, Huang GT.

University of Maryland, College of Dental Surgery, MD, USA; and Boston University School of Dental Medicine, Department of Endodontics, Boston, MA 02118, USA.

Stem Cells. 2010 May;28(5):984-95.

Marynka-Kalmani K, Treves S, Yafee M, Rachima H, Gafni Y, Cohen MA, Pitaru S.

Department of Oral Biology, School of Dental Medicine Faculty of Medicine, Tel Aviv University, Tel Aviv, Sheba Medical Center, Tel Aviv University, Tel Aviv 69978, Israel.

Biomaterials. 2010 May;31(13):3543-51. Epub 2010 Feb 1.

Mangano C, De Rosa A, Desiderio V, d'Aquino R, Piattelli A, De Francesco F, Tirino V, Mangano F, Papaccio G.

Dipartimento di Scienze dei Biomateriali, Università dell'Insubria-Varese, Italy.

Cell Tissue Res. 2010 Mar 23.

Chadipiralla K, Yochim JM, Bahuleyan B, Huang CY, Garcia-Godoy F, Murray PE, Stelnicki EJ.

Craniofacial Research Laboratory, College of Dental Medicine, Nova Southeastern University, Fort Lauderdale, Fla., USA.

Sakai VT, Zhang Z, Dong Z, Neiva KG, Machado MA, Shi S, Santos CF, Nör JE.

Curr Neurovasc Res. 2010 Feb 1;7(1):49-58.

Yalvaç ME, Ramazanoglu M, Tekguc M, Bayrak OF, Shafigullina AK, Salafutdinov II, Blatt NL, Kiyasov AP, Sahin F, Palotás A, Rizvanov AA.

Department of Genetics and BioEngineering, College of Engineering and Architecture, Yeditepe University, Campus, Kayisdagi cad., Kayisdagi, TR-34755 Istanbul, Turkey.

J Cell Physiol. 2010 Jan 15.

Ding G, Wang W, Liu Y, An Y, Zhang C, Shi S, Wang S.

Salivary Gland Disease Center and the Molecular Laboratory for Gene Therapy & Tooth Regeneration, Capital Medical University School of Stomatology, Beijing 100050, China.

Yamada Y, Nakamura S, Ito K, Sugito T, Yoshimi R, Nagasaka T, Ueda M.

Nagoya University School of Medicine, Center for Genetic and Regenerative Medicine, 65 Tsuruma-cho, Showa-ku, Nagoya, Aichi, Japan, 466-8550, +81-52-744-2348, +81-52-744-2348; [email protected].

Saber Sel-D.

Department of Endodontics, Faculty of Dentistry, Ain Shams University, Cairo, Egypt. [email protected]

Tsuchiya S, Ohshima S, Yamakoshi Y, Simmer JP, Honda MJ.

Department of Anatomy, Nihon University School of Dentistry, Division of Stem Cell Engineering, Institute of Medical Science, University of Tokyo, Tokyo, Japan; Department of Biologic and Materials Sciences, University of Michigan School of Dentistry, Michigan, USA.

Ji YM, Jeon SH, Park JY, Chung JH, Choung YH, Choung PH.

Department of Oral and Maxillofacial Surgery, Dental Research Institute, Seoul, Korea, Republic of; [email protected].

Huang CY, Pelaez D, Bendala JD, Garcia-Godoy F, Cheung HS.

Department of Biomedical Engineering, University of Miami, Coral Gables, FL, USA.

Yalvac ME, Rizvanov AA, Kilic E, Sahin F, Mukhamedyarov MA, Islamov RR, Palotás A.

Department of Genetics and BioEngineering, College of Engineering and Architecture, Yeditepe University, Istanbul, Turkey.

Yagyuu T, Ikeda E, Ohgushi H, Tadokoro M, Hirose M, Maeda M, Inagake K, Kirita T.

Department of Oral and Maxillofacial Surgery, Nara Medical University, 840 Shijo-cho, Kashihara, Nara 634-8521, Japan; Tissue Engineering Research Group, Research Institute for Cell Engineering, National Institute of Advanced Industrial Science and Technology, 3-11-46 Nakoji, Amagasaki, Hyogo 661-0974, Japan.

Zou D, Zhao J, Ding W, Xia L, Jang X, Huang Y.

School of Stomatology, Tongji University, Shanghai 200072, China; Department of Stomatology, Shanghai East Hospital Affiliated to Tongji University, No.150 JiMO Road, Shanghai City 200011, China.

Kim SH, Kim KH, Seo BM, Koo KT, Kim TI, Seol YJ, Ku Y, Rhyu IC, Chung CP, Lee YM.

Department of Periodontology and Dental Research Institute, School of Dentistry, Seoul National University, Seoul, Korea.

J Endod. 2009 Nov;35(11):1536-42. Epub 2009 Sep 20.

Nakamura S, Yamada Y, Katagiri W, Sugito T, Ito K, Ueda M.

Department of Oral and Maxillofacial Surgery, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan.

Zhang W, Walboomers XF, Shi S, Fan M, Jansen JA.

Radboud University Nijmegen Medical Centre, Periodontology & Biomaterials, Nijmegen, The Netherlands.

Kádár K, Porcsalmy B, Király M, Molnár B, Jobbágy-Ovári G, Somogyi E, Hermann P, Gera I, Varga G.

Semmelweis Egyetem Orálbiológiai Tanszék, Budapest.

Völlner F, Ernst W, Driemel O, Morsczeck C.

Institute of Human Genetics, Franz-Josef Strauss Allee 11, University of Regensburg, 93053 Regensburg, Germany.

Nakashima M, Iohara K, Sugiyama M.

Department of Oral Disease Research, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, Obu, Aichi 474-8522, Japan.

Rimondini L, Mele S.

Department of Medical Science, Piemonte Orientale University, "A. Avogadro", Novara, Italy - :[email protected].

Columbia University, Dentistry, New York, New York, United States; [email protected].

Generation of induced pluripotent stem (iPS) cells holds a great promise for regenerative medicine and other aspects of clinical applications. Many types of cells have been successfully reprogrammed into iPS cells in the mouse system, however, reprogramming human cells have been more difficult. To date, human dermal fibroblasts are the most accessible and feasible cell source for iPS generation. Dental tissues derived from ectomesenchyme harbor mesenchymal-like stem/progenitor cells and some of the tissues have been treated as biomedical wastes, e.g., exfoliated primary teeth and extracted third molars. We asked whether stem/progenitor cells from discarded dental tissues can be reprogrammed into iPS cells. The four factors Lin28/Nanog/Oct4/Sox2 or c-Myc/Klf4/Oct4/Sox2 carried by viral vectors were used to reprogram three different dental stem/progenitor cells: stem cells from exfoliated deciduous teeth (SHED), stem cells from apical papilla (SCAP) and dental pulp stem cells (DPSCs). We showed that all three can be reprogrammed into iPS cells and appeared to be at a higher rate than fibroblasts. They exhibited a morphology indistinguishable from human embryonic stem (hES) cells in cultures and expressed hES cell markers SSEA-4, TRA-1-60, TRA-1-80, TRA-2-49, Nanog, Oct4 and Sox2. They formed embryoid bodies in vitro and teratomas in vivo containing tissues of all three germ layers. We conclude that cells of ectomesenchymal origin serve as an excellent alternative source for generating iPS cells.

Petrovic V, Stefanovic V.

University of Nis School of Medicine, Nis, Serbia. [email protected]

Karaöz E, Dogan BN, Aksoy A, Gacar G, Akyüz S, Ayhan S, Genç ZS, Yürüker S, Duruksu G, Demircan PC, Sariboyaci AE.

Stem Cell and Gene Therapy Research and Applied Center, Kocaeli University, 41380, Kocaeli, Turkey, [email protected].

Huang AH, Chen YK, Lin LM, Shieh TY, Chan AW.

Grace Dental Clinic, School of Dentistry, Kaosiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan. [email protected]

Mesenchymal Stem Cell Group, CSCR University of Adelaide, Adelaide, South Australia, Australia.

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. STEM CELLS 2009;27:2229-2237.

Department of Genetics and BioEngineering, College of Engineering and Architecture, Yeditepe University, Kayisdagi, Istanbul, Turkey.

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.The Pharmacogenomics Journal advance online publication, 1 September 2009; doi:10.1038/tpj.2009.40.

Department of Conservative Dentistry and Endodontics, K.D. Dental College and Hospital, Mathura, India. [email protected]

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.

Department of Oral and Maxillofacial Surgery, Graduate School of Medicine, Kyoto University, Shogoin, Sakyo-ku, Kyoto, Japan.

PURPOSE: 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. MATERIALS AND METHODS: 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. RESULTS: 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. CONCLUSION: 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.

University of Maryland, College of Dental Surgery, Dental School, Department of Endodontics, Prosthodontics and Operative Dentistry, 650 West Baltimore St., Baltimore, MD 21201, USA. [email protected]

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.

Department of Oral and Maxillofacial Surgery, Institute of Health Sciences, Gyeongsang National University School of Medicine, Gyeongsangnam-do, South Korea.

PURPOSE: This study examined the osteogenic phenotypes and mineralization of cultured human dental papilla-derived cells. MATERIALS AND METHODS: Dental papillae were harvested from mandibles during surgical extraction of lower impacted third molars from 3 patients aged 13 to 15 years. The dental papilla-derived cells were introduced into the cell culture. After passage 3, the dental papilla-derived cells were further cultured for 42 days in an osteogenic inductive culture medium containing dexamethasone, ascorbic acid, and beta-glycerophosphate. We examined the histochemical detection of alkaline phosphatase (ALP), the reverse transcriptase-polymerase chain reaction (RT-PCR) analysis for ALP and osteocalcin, and von Kossa staining in the dental papilla-derived cells. RESULTS: It was observed that ALP was strongly expressed in the earlier stage of osteoblastic differentiation, whereas osteocalcin was mainly expressed and secreted into the medium at the later stage. Von Kossa-positive mineralization nodules were first observed on day 14, which increased in number during the entire culture period. CONCLUSIONS: These results suggest that dental papilla-derived cell have osteogenic potential and could be used as an additional source of cells for bone tissue engineering.

Methods: Cell cultures were isolated from milk tooth and third molar pulp and were grown in DMEM supplemented with 10 % FBS. Cells were characterized for expressing stem cell markers CD117, CD44H, Oct3/4 by immunofluorescency and flow-cytometry. After 3 to 5 passages we added to the media 20 ng/ml hepatocyte growth factor (HGF) for 5 days for hepatic commitment. For hepatic differentiation the cells were cultured in DMEM, 20 ng/ml HGF, 10 nM dexamethasone, insulin-transferrin-selenium X, 10 ng/ml oncostatin and 2% FBS for 15 days.

Results: Both mesenchymal cell lines were proven to be positive for pluripotent cell markers CD117, CD44H, Oct3/4. After hepatic induction both cell types changed from spindle shaped, fibroblast like to polygonal, parenchimal-like morphology. The alpha feto-protein and albumin expression were found during the differentiating process by immunofluorescency and ELISA. Mesenchymal cells were expanded in vitro and maintained in an undifferentiated state for more than 50 population doublings. Thus the cells differentiated into cells with morphological, phenotypic, and functional characteristics of hepatocytes.

Yalvac ME, Ramazanoglu M, Rizvanov AA, Sahin F, Bayrak OF, Salli U, Palotás A, Kose GT.

Department of Genetics and BioEngineering, College of Engineering and Architecture, Yeditepe University, Kayisdagi, Istanbul, Turkey.

Morsczeck C, Völlner F, Saugspier M, Brandl C, Reichert TE, Driemel O, Schmalz G.

Department of Conservative Dentistry and Periodontology, University of Regensburg, Franz-Josef-Strauss-Allee 11, 93053, Regensburg, Germany, [email protected].

Department of Oral Pathology, College of Dental Medicine, Kaohsiung Medical University, Taiwan.

INTRODUCTION: Human dental pulp stem/stromal cells (hDPSCs) in adults are primarily derived from the pulp tissues of permanent third molar teeth in existing literatures, whereas no reports exist, to our knowledge, on deriving hDPSCs from a tooth without the need for surgical procedure. The aim of this study was to raise a novel idea to source hDPSCs from complicated crown-fractured teeth requiring root canal therapy. METHODS: hDPSCs were harvested from the pulp tissues for two complicated crown-fractured teeth requiring root canal therapy, retaining the teeth for subsequent prosthodontic rehabilitation, in a 41-year-old woman who had suffered a motorcycle accident. Pulp tissue from the left lower deciduous canine of a healthy 10-year-old boy (the positive control) was also removed because of high mobility and cultured for hDPSCs. RESULTS: The hDPSCs derived from the two complicated crown-fractured teeth and the deciduous tooth were able to differentiate into adipogenic, chondrogenic, and osteogenic lineages and also expressed stem cells markers and differentiation markers, which indicated their stem cell origin and differentiation capability. In addition, hDPSCs from both the complicated crown-fractured teeth and the deciduous tooth showed high expression for bone marrow stem cell markers including CD29, CD90, and CD105 and exhibited very low expression of markers specific for hematopoietic cells such as CD14, CD34, and CD45. CONCLUSIONS: This report describes the successful isolation and characterization of hDPSCs from the pulp tissue of complicated crown-fractured teeth without tooth extraction. Therefore, pulp exposed in complicated crown-fractured teeth might represent a valuable source of personal hDPSCs.

Institute of Human Genetics, University of Regensburg, Regensburg, Germany. [email protected]

Complex human tissues harbour stem cells and/or precursor cells, which are responsible for tissue development or repair. Recently, dental tissues such as periodontal ligament (PDL), dental papilla or dental follicle have been identified as easily accessible sources of undifferentiated cells. The dental stem cell biology might provide meaningful insights into the development of dental tissues and cellular differentiation processes. Dental stem cells could also be feasible tools for dental tissue engineering. Constructing complex structures like a periodontium, which provides the functional connection between a tooth or an implant and the surrounding jaw, could effectively improve modern dentistry. Dental precursor cells are attractive for novel approaches to treat diseases like periodontitis, dental caries or to improve dental pulp healing and the regeneration of craniofacial bone and teeth. These cells are easily accessible and, in contrast to bone-marrow-derived mesenchymal stem cells, are more closely related to dental tissues. This review gives a short overview of stem cells of dental origin.

School of Dentistry, The University of Adelaide, South Australia.

Periodontitis is an inflammatory disease which manifests clinically as loss of supporting periodontal tissues including periodontal ligament and alveolar bone. For decades periodontists have sought ways to repair the damage which occurs during periodontitis. This has included the use of a range of surgical procedures, the use of a variety of grafting materials and growth factors, and the use of barrier membranes. To date periodontal regeneration is considered to be biologically possible but clinically unpredictable. Recently, reports have begun to emerge demonstrating that populations of adult stem cells reside in the periodontal ligament of humans and other animals. This opens the way for new cell-based therapies for periodontal regeneration. For this to become a reality a thorough understanding of adult human stem cells is needed. This review provides an overview of adult human stem cells and their potential use in periodontal regeneration.

School of Dentistry, University of Adelaide, Adelaide, SA, Australia.

BACKGROUND AND OBJECTIVE: Human postnatal stem cells have been identified in periodontal ligament, with the potential to regenerate the periodontium in vivo. However, it is unclear if periodontal ligament stem cells are present in regenerating periodontal tissues. The aim of this study was to identify and localize putative stem cells in block biopsies and explant cultures of regenerating human periodontal tissues. MATERIAL AND METHODS: Guided tissue regeneration was carried out on the molars of three human volunteers. After 6 wk, the teeth with the surrounding regenerating tissues and bone were surgically removed and processed for immunohistochemistry. The mesenchymal stem cell-associated markers STRO-1, CD146 and CD44 were used to identify putative stem cells. Cell cultures established from regenerating tissue explants were analysed by flow cytometry to assess the expression of these markers. Mineralization, calcium concentration and adipogenic potential of regenerating tissue cells were assessed and compared with periodontal ligament stem cells, bone marrow stromal stem cells and gingival fibroblasts. RESULTS: STRO-1(+), CD44(+) and CD146(+) cells were identified in the regenerating tissues. They were found mainly in the paravascular and extravascular regions. Flow cytometry revealed that cultured regenerating tissue cells expressed all three mesenchymal stem cell associated markers. The regenerating tissue cells were able to form mineral deposits and lipid-containing adipocytes. However, the level of mineralization in these cells was lower than that of periodontal ligament stem cells and bone marrow stromal stem cells. CONCLUSION: Cells with characteristics of putative mesenchymal stem cells were found in regenerating periodontal tissues, implying their involvement in periodontal regeneration.

Oral Biology, The School of Dentistry, The University of Birmingham, St. Chads Queensway, Birmingham B4 6NN, UK.

OBJECTIVE: Primary pulp cell cultures are frequently used to study cellular responses, odontogenic potential and stem cell responses. Their isolation and expansion via a range of technical approaches are widely reported. The purpose of this study was to investigate the influence of isolation approach and extended expansion on cell phenotype and behaviour. DESIGN: To determine viable cell isolation, enzymatic dissociation was performed on rodent incisor pulps using collagenase, trypsin, hyaluronidase and ficin. Extended expansion culture of released cells was performed in DMEM and alpha-MEM media. Cultures were subsequently analysed for gene expression, cell proliferation, cell morphology and differentiation capacity up to passage 20. RESULTS: Data indicated that incubation of extirpated and mechanically minced rodent pulpal tissue with 0.25% Trypsin:EDTA and subsequent culture in alpha-MEM medium provided optimal conditions for maximal cell growth and expansion. Under these conditions, extended culture decreased cellular proliferative capacity up to passage 7, whilst higher passages demonstrated recovered growth rates. In general gene expression analysis of osteogenic and dentinogenic associated markers decreased with increasing passage number. Notably expression of TGFbetas-1, -2 and -3 increased up to passage 10 as did the stem cell and pericyte/myofibroblast markers, CD74, Neuroserpin and alpha-SMA. Analysis of molecular phenotypes indicated little difference in lineage differentiation capacity between earlier and later passages. CONCLUSIONS: The present study characterizes conditions for primary pulp cell isolation and expansion and indicates that both earlier and later passages maintain differentiation capacity. Continued passage however may result in selection for cells with a pericyte/myofibroblast phenotype.

Department of Histology and Medical Embryology, University of Rome "La Sapienza", Rome, Italy.

Abstract Numerous stem cell niches are present in the different tissues and organs of the adult human body. Among these tissues, dental pulp, entrapped within the 'sealed niche' of the pulp chamber, is an extremely rich site for collecting stem cells. In this study, we demonstrate that the isolation of human dental pulp stem cells by the explants culture method (hD-DPSCs) allows the recovery of a population of dental mesenchymal stem cells that exhibit an elevated proliferation potential. Moreover, we highlight that hD-DPSCs are not only capable of differentiating into osteoblasts and chondrocytes but are also able to switch their genetic programme when co-cultured with murine myoblasts. High levels of MyoD expression were detected, indicating that muscle-specific genes in dental pulp cells can be turned on through myogenic fusion, confirming thus their multipotency. A perivascular niche may be the potential source of hD-DPSCs, as suggested by the consistent Ca(2+) release from these cells in response to endothelin-1 (ET-1) treatment, which is also able to significantly increase cell proliferation. Moreover, response to ET-1 has been found to be superior in hD-DPSCs than in DPSCs, probably due to the isolation method that promotes release of stem/progenitor cells from perivascular structures. The ability to isolate, expand and direct the differentiation of hD-DPSCs into several lineages, mainly towards myogenesis, offers an opportunity for the study of events associated with cell commitment and differentiation. Therefore, hD-DPSCs display enhanced differentiation abilities when compared to DPSCs, and this might be of relevance for their use in therapy.

Department of Operative Dentistry and Periodontology, University of Regensburg, Franz-Josef-Strauss-Allee 11, 93053, Regensburg, Germany, [email protected].

Recently, osteogenic precursor cells were isolated from human dental follicles, which differentiate into cementoblast- or osteoblast-like cells under in vitro conditions after the induction with dexamethasone or insulin. However, mechanisms for osteogenic differentiation are not understood in detail. In a previous study, real-time RT-PCR results demonstrated molecular mechanisms in dental follicle cells (DFCs) during osteogenic differentiation that are different from those in bone-marrow-derived mesenchymal stem cells. We analysed gene expression profiles in DFCs before and after osteogenic differentiation with the Affymetrix GeneChip(R) Human Gene 1.0 ST Array. Transcripts of 98 genes were up-regulated after differentiation. These genes could be clustered into subcategories such as cell differentiation, cell morphogenesis, and skeletal development. Osteoblast-specific transcription factors like osterix and runx2 were constitutively expressed in differentiated DFCs. In contrast, the transcription factor ZBTB16, which promotes the osteoblastic differentiation of mesenchymal stem cells as an up-stream regulator of runx2, was differentially expressed after differentiation. Transcription factors NR4A3, KLF9 and TSC22D3, involved in the regulation of cellular development, were up-regulated as well. In conclusion, we present the first transcriptome of human DFCs before and after osteogenic differentiation. This study sheds new light on the complex mechanism of osteogenic differentiation in DFCs.

Department of Biological Science, College of Natural Sciences, Wonkwang University, Iksan, Jeonbuk 570-749, Republic of Korea.

Human dental pulp-derived stem cells (hDPSCs) have been considered alternative sources of adult stem cells because of their potential to differentiate into multiple cell lineages. This study investigated the possible role of gangliosides in the neural differentiation of hDPSCs. When hDPSCs were cultured under neural differentiation conditions, expression of neural cell maker genes such as Nestin, MAP-2, and NeuN was detected. Immunostaining and high-performance thin-layer chromatography analysis showed that an increase in ganglioside biosynthesis was associated with neural differentiation of hDPSCs. Specifically, a significant increase in GD3 and GD1a expression was observed during neural differentiation. To confirm the role of gangliosides in neural differentiation, ganglioside biosynthesis was inhibited in hDPSCs by knockdown of UDP-glucose ceramide glucosyltransferase (Ugcg), which prevented differentiation into neural cells. These results suggest that gangliosides may play a role in the neural differentiation process of hDPSCs.

Tzu Chi Stem Cells Centre, Tzu Chi General Hospital, Hualien, Taiwan.

Background aims We have isolated human neuronal stem cells from exfoliated third molars (wisdom teeth) using a simple and efficient method. The cultured neuronal stem cells (designated tNSC) expressed embryonic and adult stem cell markers, markers for chemotatic factor and its corresponding ligand, as well as neuron proteins. The tNSC expressed genes of Nurr1, NF-M and nestin. They were used to treat middle cerebral artery occlusion (MCAO) surgery-inflicted Sprague-Dawley (SD) rats to assess their therapeutic potential for stroke therapy. Methods. For each tNSC cell line, a normal human impacted wisdom tooth was collected from a donor with consent. The tooth was cleaned thoroughly with normal saline. The molar was vigorously shaken or vortexed for 30 min in a 50-mL conical tube with 15-20 mL normal saline. The mixture of dental pulp was collected by centrifugation and cultured in a 25-cm(2) tissue culture flask with 4-5 mL Medium 199 supplemented with 5-10% fetal calf serum. The tNSC harvested from tissue culture, at a concentration of 1-2x10(5), were suspended in 3 microL saline solution and injected into the right dorsolateral striatum of experimental animals inflicted with MCAO. Results. Behavioral measurements of the tNSC-treated SD rats showed a significant recovery from neurologic dysfunction after MCAO treatment. In contrast, a sham group of SD rats failed to recover from the surgery. Immunohistochemistry analysis of brain sections of the tNSC-treated SD rats showed survival of the transplanted cells. Conclusions. These results suggest that adult neuronal stem cells may be procured from third molars, and tNSC thus cultivated have potential for treatment of stroke-inflicted rats.

Department of Oral Biology, Semmelweis University, Budapest, Hungary.

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.

School of Dentistry, Federal University of Pelotas, Pelotas, RS, Brazil. [email protected]

AIM: The aim of this paper is to present a review and discussion of the current status of stem cell research with regard to tooth generation. BACKGROUND: Stem cells have been isolated from the pulp tissue of both deciduous and permanent teeth as well as from the periodontal ligament. Dental pulp stem cells demonstrate the capacity to form a dentin pulp-like complex in immunocompromised mice. A tooth-like structure was successfully formed, using a heterogeneous mixture of dental enamel epithelium, pulp mesenchymal cells, and scaffolds. CONCLUSION: The scientific community understands the need for more investigations to completely understand the conditions that would best favor the creation of a tooth substitute. Recent gains in the understanding of the molecular regulation of tooth morphogenesis, stem cell biology, and biotechnology offers the opportunity to realize this goal. CLINICAL SIGNIFICANCE: These findings, combined with the recent progress in stem cell research and tissue engineering, might allow the development of alternatives for current materials and therapies used to treat tooth tissue loss (e.g., enamel, dentin, pulp), reconstruct dentoalveolar and craniofacial bone defects, and eventually replace an entire tooth.

Mesenchymal Stem Cell Group, Division of Haematology, Institute of Medical and Veterinary Science/Hanson Institute, CSCR University of Adelaide, Adelaide 5000, SA, Australia.

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. A readily accessible human cell population to investigate these mechanisms are dental pulp progenitor/stem cells (DPSC) that can differentiate into functionally active neurons given the appropriate environmental cues. We hypothesized that implanted DPSC 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 following transplantation of adult human DPSC. Chemo-attraction of avian trigeminal ganglion axons towards implanted DPSC 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 DPSC may induce neuroplasticity within a receptive host nervous system.

General BioTechnology, LLC, Indianapolis, IN, USA; Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN, USA.

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.

Laboratory of Molecular Genetics and Stem Cell Differentiation, Dental Research Institute, School of Dentistry, Seoul National University, Seoul 110-749, Republic of Korea.

It is well known that interactions between epithelial components and mesenchymal components are essential for tooth development. Therefore, it has been postulated that both types of stem cells might be involved in the regeneration of dental hard tissues. Recently, mesenchymal dental pulp stem cells that have odontogenic potential were identified from human dental pulp. However, the existence of epithelial cells has never been reported in human dental pulp. In the present study, we isolated and characterized epithelial cell-like cells from human deciduous dental pulp. They had characteristic epithelial morphology and expressed epithelial markers. Moreover, they expressed epithelial stem cell-related genes such as ABCG2, Bmi-1, DeltaNp63, and p75. Taken together, our findings suggest that epithelial stem cell-like cells might exist in human deciduous dental pulp and might play a role as an epithelial component for the repair or regeneration of teeth.

Department of Restorative Dentistry, Harvard School of Medicine, Boston, MA 02115, USA. [email protected]

INTRODUCTION: Postnatal human dental pulp is a potentially promising source of progenitor cells. Sustaining and amplifying progenitor cell populations would be beneficial for basic science research with application in pulpal regeneration. Hypoxia has been observed to promote the undifferentiated cell state in various stem cell populations. The purpose of this study was to examine human dental pulp cells (DPCs) proliferation in normoxia and hypoxia. METHODS: Dental pulp cells were obtained from third molars of adult patients and cultured in alpha modification of Eagle's medium culture medium with 10% fetal bovine serum. For cell proliferation, DPCs were divided into two groups: (1) DPCs incubated in normoxic conditions (20% oxygen tension) and (2) DPC incubated in hypoxic conditions (3% oxygen tension). Cell proliferation assays were performed every 2 to 3 days from day 3 to day 14 by trypsinization and quantification of cells with a hemacytometer. Fluorescence-activated cell sorting analysis was completed to investigate stem cell markers, CD133, and STRO-1. RESULTS: DPCs proliferated significantly more in hypoxia than in normoxia (ie, two-fold throughout the experiment, p < 0.0001). The primitive stem cell marker, CD133, decreased in hypoxia, whereas the osteoprogenitor marker, STRO-1, increased by 8.5-fold. CONCLUSIONS: This study suggested that hypoxia is an effective treatment to amplify numbers of progenitor cells from human dental pulp.

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

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

ABSTRACT

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

Read More:  http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2757034/

Tissue Eng Part C Methods. 2009 Dec;15(4):659-67.

De Rosa A, De Francesco F, Tirino V, Ferraro GA, Desiderio V, Paino F, Pirozzi G, D'Andrea F, Papaccio G.

Dipartimento di Discipline Odontostomatologiche, Ortodontiche e Chirurgiche, Seconda Università degli Studi di Napoli , Naples, Italy.

Mineralised Tissue Research Group, Tissue Engineering and Regenerative Dentistry, School of Dentistry Cardiff University, Cardiff, UK. [email protected]

INTRODUCTION: It is now accepted that progenitor/stem cells reside within the post-natal dental pulp. Studies have identified several niches of multipotent mesenchymal progenitor cells, known as dental pulp stem cells, which have a high proliferative potential for self-renewal. These progenitor stem cells are now recognized as being vital to the dentine regeneration process following injury. Understanding the nature of these progenitor/stem cell populations in the pulp is important in determining their potentialities and development of isolation or recruitment strategies for use in regeneration and tissue engineering. Characterization of these cells, and determination of their potentialities in terms of specificity of regenerative response, may help direct new clinical treatment modalities. Such novel treatments may involve controlled direct recruitment of the cells in situ and possible seeding of stem cells at sites of injury for regeneration or use of the stem cells with appropriate scaffolds for tissue engineering solutions. Such approaches may provide an innovative and novel biologically based new generation of clinical materials and/or treatments for dental disease. AIM: This study aimed to review the body of knowledge relating to stem cells and to consider the possibility of these cell populations, and related technology, in future clinical applications.

Department of Endodontics, College of Dental Medicine, Nova Southeastern University, Fort Lauderdale, FL 33328-2018, USA.

The purpose of this study was to measure cell survival and degradation within tissue-engineered dental constructs. Dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PLSCs) were seeded on three types of tissue engineering scaffolds: a synthetic open-cell D,D-L,L-polylactic acid (polymer) scaffold, a bovine collagen scaffold (collagen), and a calcium phosphate bioceramic (calcium phosphate) scaffold. The dental pulp and periodontal constructs (n = 144) were maintained in cell culture for between 3 and 14 days. The cell survival and degradation within the constructs were measured using histologic criteria. The DPSC and PLSC survival was optimal in the polymer and collagen constructs but not the calcium phosphate constructs, especially over longer time periods. These in vitro results suggest that both the polymer and collagen scaffolds and the DPSCs and PLSCs can be combined to create pulp and periodontal constructs for use in future regenerative dental treatments.

Tissue Engineering and Reparative Dentistry, School of Dentistry, Cardiff University, Cardiff, UK. [email protected]

The present study compared the cellular characteristics of progenitor stem cell populations present in adult dental pulp, isolated by different methods utilizing 2 different features of stem cell biology. One population expressing high levels of beta1 integrin was isolated by preferential selection of adherent cells to fibronectin over 20 min. In an alternative approach, cells expressing the embryonic neural crest cell marker, low-affinity nerve growth factor receptor (LANGFR), were selected by magnetic-activated cell sorting. For each method, clonal cell lines were established and expanded in culture. One clone derived via the respective methods was examined for embryonic/progenitor cell markers by immunocytochemistry and RT-PCR. Both clonal populations demonstrated the expression of stro-1 and stained positive for vimentin, demonstrating mesenchymal lineage. Of note, cells selected for LANGFR cells demonstrated the additional expression of CD105 and Notch 2. For both clonal populations, expanded cultures demonstrated the ability to differentiate into osteoblasts, adipocytes and chondrocytes. These results would suggest the potential isolation of 2 progenitor cell populations exhibiting different cellular characteristics in terms of their embryonic nature. The potential for both cell populations to derive from a common origin is discussed. Copyright 2008 S. Karger AG, Basel.

Department of Oral and Maxillofacial Pathology, Division of Craniofacial and Molecular Genetics, Tufts University, Boston, Massachusetts, USA.

Based on the successful use of silk scaffolds in bone tissue engineering, we examined their utility for mineralized dental tissue engineering. Four types of hexafluoroisopropanol (HFIP) silk scaffolds-(250 and 550 microm diameter pores, with or without arginine-glycine-aspartic acid (RGD) peptide) were seeded with cultured 4-day postnatal rat tooth bud cells and grown in the rat omentum for 20 weeks. Analyses of harvested implants revealed the formation of bioengineered mineralized tissue that was most robust in 550 microm pore RGD-containing scaffolds and least robust in 250 microm pore sized scaffolds without RGD. The size and shape of the silk scaffold pores appeared to guide mineralized tissue formation, as revealed using polarized light imaging of collagen fiber alignment along the scaffold surfaces. This study is the first to characterize bioengineered tissues generated from tooth bud cells seeded onto silk scaffolds and indicates that silk scaffolds may be useful in forming mineralized osteodentin of specified sizes and shapes.

Institute of Human Genetics, University of Regensburg, Regensburg, Germany. [email protected]

Complex human tissues harbour stem cells and/or precursor cells, which are responsible for tissue development or repair. Recently, dental tissues such as periodontal ligament (PDL), dental papilla or dental follicle have been identified as easily accessible sources of undifferentiated cells. The dental stem cell biology might provide meaningful insights into the development of dental tissues and cellular differentiation processes. Dental stem cells could also be feasible tools for dental tissue engineering. Constructing complex structures like a periodontium, which provides the functional connection between a tooth or an implant and the surrounding jaw, could effectively improve modern dentistry. Dental precursor cells are attractive for novel approaches to treat diseases like periodontitis, dental caries or to improve dental pulp healing and the regeneration of craniofacial bone and teeth. These cells are easily accessible and, in contrast to bone-marrow-derived mesenchymal stem cells, are more closely related to dental tissues. This review gives a short overview of stem cells of dental origin.

Department of Genetics and BioEngineering, Yeditepe University, 26 Agustos Campus, Kayisdagi cad., Kayisdagi, 34755, Istanbul, Turkey.

Introduction Human dental follicle cells (HDFCs) derived from human impacted third molars (wisdom teeth) have been shown to be a significant source of adult stem cells. Generation of mesenchymal stem cell-like cells from dental follicles causes minimal surgical stress. In vitro and in vivo reports showed that HDFCs can be utilized in gene and cell therapy applications which make them an attractive alternative source for different gene-cell therapy applications. However, there are currently no systematic comparative studies on transfection potential of HDFC cells using different chemical and electro-poration techniques. Methods Stem cells from impacted third tooth molars were isolated, and analyzed for expression of surface markers. Transfection efficiencies of four commercially available transfection reagents (Transfast, Escort V, Superfect and FuGene HD) and electro-poration on isolated stem cells were compared. Results Isolated HDFCs were stained positive for CD105, CD90, CD73, CD166, and negative for CD34, CD45, and CD133. Among the chemical transfection reagents used in this study, FuGene HD was the most efficient in transfecting HDFCs, even in the presence of 10% serum. Conclusion Electro-poration of HDFCs yield relatively high transfection rates and cell viability when compared to chemical transfection techniques. Our observations might be useful for developing gene and cell therapy applications using dental follicle stem cells.

Dipartimento di Medicina Sperimentale, Sezione di Istologia ed Embriologia, TESLab, Secondo Ateneo di Napoli, Napoli, Italy.

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. J. Exp. Zool. (Mol. Dev. Evol.) 310B, 2008. (c) 2008 Wiley-Liss, Inc.

Tissue Engineering and Reparative Dentistry, School of Dentistry, Cardiff University, Cardiff, UK. [email protected]

The present study compared the cellular characteristics of progenitor stem cell populations present in adult dental pulp, isolated by different methods utilizing 2 different features of stem cell biology. One population expressing high levels of beta1 integrin was isolated by preferential selection of adherent cells to fibronectin over 20 min. In an alternative approach, cells expressing the embryonic neural crest cell marker, low-affinity nerve growth factor receptor (LANGFR), were selected by magnetic-activated cell sorting. For each method, clonal cell lines were established and expanded in culture. One clone derived via the respective methods was examined for embryonic/progenitor cell markers by immunocytochemistry and RT-PCR. Both clonal populations demonstrated the expression of stro-1 and stained positive for vimentin, demonstrating mesenchymal lineage. Of note, cells selected for LANGFR cells demonstrated the additional expression of CD105 and Notch 2. For both clonal populations, expanded cultures demonstrated the ability to differentiate into osteoblasts, adipocytes and chondrocytes. These results would suggest the potential isolation of 2 progenitor cell populations exhibiting different cellular characteristics in terms of their embryonic nature. The potential for both cell populations to derive from a common origin is discussed. Copyright 2008 S. Karger AG, Basel.

J Cell Physiol. 2006 Aug;208(2):319-25.

Papaccio G, Graziano A, d'Aquino R, Graziano MF, Pirozzi G, Menditti D, De Rosa A, Carinci F, Laino G.

Dipartimento di Medicina Sperimentale, Sezione di Istologia ed Embriologia, Secondo Ateneo di Napoli, Napoli, Italy. [email protected]

Grace Dental Clinic, Kaohsiung City, Taiwan

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.

Departamento de Estomatologia, Centro de Ciências da Saúde, Universidade Federal de Santa Catarina, Brazil. [email protected]

BACKGROUND AND OBJECTIVE: The periodontal ligament is a specialized connective tissue, derived from dental follicle and originated from neural crest cells. Recently it has been suggested, based on animal models, that periodontal ligament could be a niche for neural crest stem cells. However, there is still little knowledge on this subject. The identification of neural crest adult stem cells has received much attention based on its potential in tissue regeneration. The objective of the present work was to verify the human periodontal ligament as a niche for neural crest stem cells. MATERIAL AND METHODS: Cells from human periodontal ligament were isolated from 10 teeth of seven individuals (periodontal ligament pool group) and also from four teeth of one individual (periodontal ligament single group), after enzymatic digestion. The cells were cultured in specific inductive medium. Analyses of protein and gene expression were performed through immunocytochemistry and reverse transcription-polymerase chain reaction techniques, respectively. RESULTS: Mesodermal phenotypes (adipogeneic, osteogenic and myofibroblastic) were identified after culture in inductive medium. Immunocytochemistry analyses showed the presence of the nestin marker of neural stem cells and also markers of undifferentiated neural crest cells (HNK1, p75). When cultured in inductive medium that allowed neural differentiation, the cells showed markers for beta-tubulin III, neurofilament M, peripherin, microtubule-associated protein 2 and protein zero. The results were similar between the two study groups (the periodontal ligament pool group and the periodontal ligament single group). CONCLUSION: This research provides evidence that human periodontal ligament, in addition to its mesodermal derivatives, produces neural crest-like cells. Such features suggest a recapitulation of their embryonic state. The human periodontal ligament revealed itself as a viable alternative source for possible primitive precursors to be used in stem-cell therapies.

Institut für Humangenetik, Universität Regensburg, Franz-Josef-Strauss-Allee 11, Regensburg, Germany. [email protected]

BACKGROUND: Undifferentiated human dental cells and especially human dental follicle cells are interesting for potential dental treatments. These somatic stem cells are cultured usually in cell culture medium containing bovine serum. In the age of bovine spongiform encephalopathy (BSE), a serum-free cell culture system for dental follicle cells are recommended, if these cells will be applied in dentistry. PURPOSE: However, less is known about the cultivation of dental follicle cells in serum-replacement medium. In this study, we cultivated dental follicle cells in serum-free cell culture medium, which is normally applied for neuronal stem/progenitor cells. MATERIALS AND METHODS: Dental follicle cells were cultivated in both serum-free and serum-containing cell culture media, and gene expression profiles were recorded for connective tissue markers collagen type I and type III and for the human dental follicle cell marker nestin. RESULTS: It is interesting to note that the gene expressions of collagens and nestin were similar after applying both cell culture conditions. CONCLUSION: Although the gene expression of dental follicle cell markers was unchanged, a more appropriate serum-free cell culture medium is recommended for cell proliferation of dental follicle cells.

Department of Endodontics, College of Stomatology, Fourth Military Medical University, Xi'an, Shaanxi, China.

Stem cell-mediated root regeneration offers opportunities to regenerate a bio-root and its associated periodontal tissues to restore tooth loss. Periodontal ligament (PDL) and cementum complex and dentin pulp complex have been tissue engineered using human dental pulp stem cells and PDL stem cells, respectively. The aim of this study was to explore whether dentin formation could be induced using an inductive substrate and whether bioengineered dentin could induce cementum and PDL formation. First, dentin was bioengineered from tooth papillae of Sprague-Dawley (SD) rats with an inductive substrate, and its phenotype was characterized; then primarily cultured human PDL cells were seeded on the surface of dentin and transplanted under the skin of immunocompromised mice. Histological, immunohistochemical, and scanning electronic microscopy examinations results showed that bioengineered dentin could induce cementogenesis and PDL formation, and condense PDL arranged perpendicularly on the dentin surface via a layer of cementum-like tissue. The results indicated that tissue-engineered dentin could be induced using an inductive substrate and could be used as a further substrate for cementum and PDL tissue engineering.

Institute of Stomatology, Nanjing Medical University, Nanjing, Jiangsu, P.R. China., Department of Endodontics, School of Stomatology, Fourth Military Medical University, Xi'an, Shaanxi, P.R. China., Research and Development Center for Tissue Engineering, Fourth Military Medical University, Xi'an, Shaanxi, P.R. China.

Tooth loss adversely affects not only mouth functions but also the esthetics of one's face. To repair these defects, current treatment methods mainly depend on nonbiological materials or artificial implants that also can, sometimes, reduce the quality of life because of their limited physiological function, or elicit an immunological rejection. Theoretically, a biological tooth (bio-tooth) that is made from the patient's own cells and grows in its intended location should be the best choice for treating tooth loss, although such bioengineered teeth have been nothing more than a dream for many centuries. Recently, significant advances in the fields of tissue engineering, stem cell biology, developmental biology, molecular genetics, and bionics have brought us close to the realization of a bio-tooth. However, issues involving in the reconstruction of a bio-tooth regarding the shape determination, size control, availability of dental epithelium, directional growth and eruption, and graft rejection in the jaws remain to be resolved. Here, this review outlines the current approaches toward the tooth regeneration, and focuses on several key challenges that must be met in the making of a bio-tooth.

Department of Comparative Biomedical Sciences, Louisiana State University, School of Veterinary Medicine, Baton Rouge, LA 70803, USA.

The dental follicle (DF) differentiates into the periodontal ligament. In addition, it may be the precursor of other cells of the periodontium, including osteoblasts and cementoblasts. We hypothesized that stem cells may be present in the DF and be capable of differentiating into cells of the periodontium. Stem cells were identified in the DF of the rat first mandibular molar by Hoechst staining, alkaline phosphatase staining, and expression of side-population stem cell markers. These cells were shown to be able to differentiate into osteoblasts/cementoblasts, adipocytes, and neurons. Treating the DF cell population with doxorubicin, followed by incubation in an adipogenesis medium, suggested that the adipocytes originated from stem cells. Thus, a possibly puripotent stem cell population is present in the rat DF.

Dental Faculty, Tehran University of Medical Sciences, Tehran, Iran.

OBJECTIVE: Implant placement in the posterior maxilla may often be contraindicated because of insufficient bone volume and presence of the maxillary sinus. In these situations, sinus floor augmentation frequently has been proposed as the best treatment. This clinical study was based on the hypothesis that the clinical effectiveness of adult mesenchymal stem cells (MSCs) loaded to the biphasic scaffold. METHODS: In this report, the clinical and radiographic results are presented on 6 consecutively treated patients using MSCs in combination with biphasic hydroxyl apatite/ beta-tricalcium phosphate (HA/TCP) for sinus elevation. All the patients in the study had less than 3 mm initial bone height in the posterior maxillary area (IBH). MSCs were cultured and expanded from bone marrow aspirate for each patient. Three months after sinus elevation, radiographic evaluation was performed for the patients and the secondary bone height was measured (SBH(1)). In the second stage surgery, 30 implants were placed. Trephine bur was used as a pilot drill and a core biopsy was obtained from each implant site. Prosthetic rehabilitation of the patients was performed after 4 months. Secondary bone height was measured 9 months after implant placement (SBH(2)). RESULTS: Of 30 implants, 28 (93%) were considered clinically successful. Two implants were removed due to mobility at the time of surgical exposure. Histologic evaluation of the biopsy specimens revealed numerous areas of osteoid and bone formation HA/TCP, with no evidence of inflammatory cell infiltrate. Mean bone regenerate was 41.34%. Clinically, no complications were observed, and all implants were considered clinically osseointegrated after 4 months. Mean bone height was measured 3 and 12 months after sinus grafting (mean of SBH(1)= 12.08 mm and mean of SBH(2)= 10.08 mm). CONCLUSIONS: These clinical and histological findings suggest that sinus grafting with HA/TCP in combination with MSCs provide a viable therapeutic alternative for implant placement. The findings suggest that the addition of MSCs to bone derivative/substitute materials may enhance bone formation in the maxillary sinus area. Of course more studies with the control groups are needed for the evaluation of this method as a clinical solution for the patients.

Researchers from the University of Michigan School of Dentistry, Michigan, United States, have successfully used stem cells from human exfoliated deciduous teeth (SHED) to produce tissues that closely resemble physiologic dental pulp tissue. The findings of the study are published in the Journal of Endodontics.

Stem cells from human exfoliated deciduous teeth are multipotent cells. The team of researchers led by Mabel Mariela Rodriguez Cordeiro sought to assess morphological characteristics of the tissues produced from SHED. Biodegradable scaffolds within the human tooth slices were prepared, which were seeded with SHED and introduced into immunodeficient mice. The tissue formed by this technique closely resembled physiological dental pulp, both in terms of architecture and cellularity. SHED differentiated into odontoblast-like cells and endothelial-like cells. The researchers thereby concluded that the exfoliated primary teeth could be a potential source for multipotent stem cells.

A study by Dr Masako Miura and colleagues (Proceedings of the National Academy of Sciences of the United States of America, 2003) revealed that the primary dental pulp could be a viable source for stem cells as they remain vital even after exfoliation of the tooth. The researchers isolated multipotent stem cells from exfoliated deciduous teeth that were highly proliferative, and could be differentiated into various other types of cells such as neural cells, adipocytes and odontoblasts. Following in vivo transplantation, these stem cells showed the potential to form bone and generate dentin. In another study, Seo BM (Lancet, 2004) isolated stem cells from periodontal ligaments and used them for successful production of cementum and periodontal ligament-like tissue, in vivo.

Craniofacial tissue engineering aims at the formation of craniofacial structures absent due to congenital abnormalities, along with regeneration of dental and craniofacial structures lost due to trauma or diseases. Dental caries and periodontitis are the major cause for the loss of teeth and related structures. The National Institute of Dental and Craniofacial Research (NIDCR, 2004), estimates that about 42% of children, between 2 to 11 years of age, experience dental caries in primary teeth, while 59% of adolescents, between the age group of 12 and 19, have caries in their permanent dentition.

Stem cell therapies and dental pulp tissue engineering can be used to decrease the incidence of various dental disease conditions. The easy availability and accessibility of SHED, along with their capability to differentiate into various dental structures, such as bone, dentin and pulp, make them a potential boon for various stem cell therapies and dental pulp tissue engineering.

Department of Oral Disease Research, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, Obu, Aichi 474-8522, Japan.

Center of Advanced European Studies and Research, Bonn, Germany. [email protected]

The connective tissue of the human tooth arises from cells that are derived from the cranial neural crest and, thus, are termed as "ectomesenchymal cells." Here, cells being located in a pad-like tissue adjacent to the apex of the developing tooth, which we designated the third molar pad, were separated by the microexplant technique. When outgrowing from the explant, dental neural crest-derived progenitor cells (dNC-PCs) adhered to plastic, proliferated steadily, and displayed a fibroblast-like morphology. At the mRNA level, dNC-PCs expressed neural crest marker genes like Sox9, Snail1, Snail2, Twist1, Msx2, and Dlx6. Cytofluorometric analysis indicated that cells were positive for CD49d (alpha4 integrin), CD56 (NCAM), and PDGFRalpha, while negative for CD31, CD34, CD45, and STRO-1. dNC-PCs could be differentiated into neurogenic, chondrogenic, and osteogenic lineages and were shown to produce bone matrix in athymic mice. These results demonstrate that human third molar pad possesses neural crest-derived cells that represent multipotent stem/progenitor cells. As a rather large amount of dNC-PCs could be obtained from each single third molar, cells may be used to regenerate a wide range of tissues within the craniofacial region of humans.

Maxillofacial Surgery, Maxillofacial Reconstruction and Function, Division of Maxillofacial and Neck Reconstruction, Graduate School, Tokyo Medical and Dental University, 1-5-45, Yushima, Bunkyo-ku, Tokyo 113-8549, Japan.

Recent studies indicate that dental pulp is a new source of adult stem cells. The human tooth with an immature apex is a developing organ, and the apical pulp of this tooth may contain a variety of progenitor/stem cells, which participate in root formation. We investigated the hard tissue regeneration potential of apical pulp derived cells (APDCs) from human tooth with an immature apex. APDCs cultured with a mineralization-promoting medium showed alkaline phosphatase activity in porous hydroxyapatite (HA) scaffolds. The composites of APDCs and HA were implanted subcutaneously in immunocompromised rats and harvested at 12 weeks after implantation. In histological analysis, the APDCs/HA composites exhibited bone- and dentine-like mineralized tissues in the pore areas of HA. This study suggests that the human tooth with an immature apex is an effective source of cells for hard tissue regeneration.

Institute of Human Genetics, University of Regensburg, Regensburg, Germany, [email protected].

Department of Stomatology, Chinese PLA General Hospital and Postgraduate Military Medical School, Beijing 100853, China; Research and Development Center for Tissue Engineering, Fourth Military Medical University, Xi'an, Shaanxi 710032, China.

Two crucial growth factors, FGF2 and TGFbeta(1), were investigated in this study to determine their inductive effects on the odontoblastic differentiation of human dental pulp stem cells (DPSCs) in vitro. DPSCs were isolated by immunomagnetic bead selection using the STRO-1 antibody, and then co-cultured respectively with FGF2, TGFbeta(1) and FGF2+TGFbeta(1). The results showed that FGF2 can exert a significant effect on the cell proliferation, while TGFbeta(1) or FGF2+TGFbeta(1) can initiate an odontoblast-like differentiation of DPSCs. Moreover, FGF2 can synergistically upregulate the effects of TGFbeta(1) on the odontoblastic differentiation of DPSCs, as indicated by the increased alkaline phosphatase activity, the polarized cell appearance and secretary ultrastructural features, the formation of mineralized nodules and the gene/protein expression of dentin sialoprotein and dentin matrix protein-1. Together, FGF2 acted primarily on the cell proliferation, while TGFbeta(1) and FGF2+TGFbeta(1) mainly stimulated the odontoblastic differentiation of DPSCs. This study provides interesting progress in the odontoblastic differentiation of DPSCs induced by FGF2 and TGFbeta(1).

Departamento di Scienze Odontostomatologiche, Universitá La Sapienze, Romae, Italy.

OBJECTIVE: Bone tissue engineering is a promising approach for bone reconstruction in oral-maxillofacial surgery. This study investigates the suitability of oral skeletal tissues as convenient and accessible sources of osteogenic progenitors as an alternative to the iliac crest bone marrow. STUDY DESIGN: Samples of maxilla tuberosity (MT) and maxillary and mandibular periosteum (MP) were obtained during routine oral surgery, and donor site morbidity was assessed using a "split-mouth" approach. Cells isolated from MT (bone marrow stromal cells; MT-BMSCs) and from MP (periosteal cells; M-PCs), were analyzed for clonogenicity, phenotype, expression of osteogenic markers, and ability to form bone in vivo. RESULTS: Both MT-BMSCs and M-PCs included clonogenic cells, showed comparable phenotypic profiles, and expressed early osteogenic markers. Most importantly, both cell populations formed bone upon ectopic in vivo transplantation. CONCLUSION: MT-BMSCs and M-PCs behaved as osteoprogenitor cells in vitro and in vivo. MT and MP may be considered as suitable sources of cells for bone tissue engineering in humans.

Department of Oral and Maxillofacial Science and.

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.

Department of Pediatric Dentistry, Peking University School & Hospital of Stomatology, Beijing 100081, China.

OBJECTIVE: To isolate and culture the pulp cells from human young permanent teeth (pDPC), and to observe their biological characteristics and the expression of some specific markers, and to induce these pulp cells to differentiate into osteoblast, adipocyte, neuron and chondrocyte lineages. METHODS: Pulp cells were isolated and cultured from orthodontic extracted premolars of children. The attached cells after at least 3 passages were used for the following experiments: 1. Morphology and ultrastructure analysis; 2. Cell cycle and phenotype were analyzed by flowcytometry; 3. Growth curve were recorded; 4. pDPC were induced to differentiate into osteoblast, adipocyte, neuron in vitro, and were identified by histochemical methods and RT-PCR. RESULTS: 1. Attached pDPCs were fibroblast-like cells, which were distinguished from BMSC. 2. The cell organs in dDPCs were well developed. 3. pDPCs were highly positive for CD90, CD44, CD147, which are mesenchymal stem-cell markers, but were negative for other markers including CD34, CD38, CD45, HLA-DR. 4. pDPCs showed high growth rate. 5. pDPCs could be induced to differentiate into osteoblast, adipocyte, and neuron lineages, but not chondrocyte lineages. CONCLUSION: pDPCs were characterized by their ability to proliferate with high growth rate in vitro. The expression of some BMSC markers in these cells were observed. They showed the potential to differentiate into multiple mesenchymal lineages such as osteoblast, adipocyte, neuron lineages under specific conditions in vitro.

Principal Investigator: Jeffrey D. Hartgerink, Ph.D. (Rice), Co-Investigator: Rena D'Souza, D.D.S., Ph.D. (UTHSC-H)

General BioTechnology, LLC, Indianapolis, Indiana.

Recent studies have shown that mesenchymal stem cells (MSC) with the potential for cell-mediated therapies and tissue engineering applications can be isolated from extracted dental tissues. Here, we investigated the collection, processing, and cryobiological characteristics of MSC from human teeth processed under current good tissue practices (cGTP). Viable dental pulp-derived MSC (DPSC) cultures were isolated from 31 of 40 teeth examined. Of eight DPSC cultures examined more thoroughly, all expressed appropriate cell surface markers and underwent osteogenic, adipogenic, and chondrogenic differentiation in appropriate differentiation medium, thus meeting criteria to be called MSC. Viable DPSC were obtained up to 120 h postextraction. Efficient recovery of DPSC from cryopreserved intact teeth and second-passage DPSC cultures was achieved. These studies indicate that DPSC isolation is feasible for at least 5 days after tooth extraction, and imply that processing immediately after extraction may not be required for successful banking of DPSC. Further, the recovery of viable DPSC after cryopreservation of intact teeth suggests that minimal processing may be needed for the banking of samples with no immediate plans for expansion and use. These initial studies will facilitate the development of future cGTP protocols for the clinical banking of MSC.

Dept. of Stomatology and Oral Science, Division of Oral Surgery, University G. d'Annunzio, Chieti, Ialy. [email protected]

In this study we investigated the in vitro behaviour, morphostructure and extracellular matrix synthesis of human dental follicular stem cells (hDFSCs) isolated from human dental bud, which resulted to be positive for mesenchymal markers (CD29, CD90, CD146 and CD166) by FACS analysis. Cells were analysed by light and electronic microscopy to evaluate their biological response either at week 1, that is before differentiation, or at weeks 3-6, when they had been cultured in osteogenic medium onto a highly porous natural scaffold material (Bio-Oss). Microscopy analysis of primary culture cells showed they had a mesenchymal stem cell-like morphostructure, spindle shaped, similar to the culture of mesenchymal stem cells derived from adult bone marrow. Also, after osteogenic differentiation, these analyses indicate typical osteoblast morphostructure and reveale a tri-dimensional organization of the cells and deposition of extracellular matrix (ECM) in close contact with biomaterial. This approach would allow to personalize the scaffold for bone tissue engineering in order to accelerate the process of osteogenesis.

J Dent Res. 2005 Oct;84(10):907-12.

Seo BM, Miura M, Sonoyama W, Coppe C, Stanyon R, Shi S.

Craniofacial and Skeletal Diseases Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD 20892, USA.

Inserm, JPA Center, Inserm U837, Lille, cedex, France; [email protected].

Adult tissues contain highly proliferative, clonogenic cells that meet criteria of multipotent stem cells and are potential sources for autologous reparative and reconstructive medicine. We demonstrated that human dental pulp contains self renewing human dental pulp stem cells (hDPSCs) capable of differentiating into mesenchymal-derived odontoblasts, osteoblasts, adipocytes, chondrocytes and striated muscle, and interestingly, also into non-mesenchymal melanocytes. Furthermore, we showed that hDPSC cultures include cells with the label-retaining and sphere-forming abilities, traits attributed to multipotent stem cells, and provide evidence that these might be multipotent neural crest stem cells.

Faculty of Industrial Science and Technology Tokyo University of Science, Department of Biological Science and Technology, Noda, Chiba 278-8510, Japan.

BACKGROUND: The ultimate goal of regenerative therapy is to develop fully functioning bioengineered organs that can replace organs lost or damaged due to disease, injury or aging. Dental regenerative medicine has made the most progress and is the most useful model for the consideration of strategies in future organ replacement therapies.

OBJECTIVE: This review describes strategies that have been pursued to date and experiments currently being conducted to bioengineer teeth in anticipation of the production of fully functional organs.

METHODS: To realize the practical application of 'bioengineered tooth' transplantation therapy, four major hurdles must be overcome. The present status of the hurdles to this therapy are described and discussed in this review.

RESULTS/CONCLUSION: The bioengineering techniques developed for tooth regeneration will in the future make substantial contributions to the ability to grow primordial organs in vitro and also to grow fully functioning organs, such as the liver, kidney and heart.

Department of Stomatology, Jinzhou Central Hospital, Jinzhou 121000, People's Republic of China. [email protected]

This study examined the biological characteristics of mesenchymal stem cells (MSCs) grown on sand-blasted, large-grit, acid-etched (SLA) surface and hydroxyapatite (HA) coating on the SLA (HA/SLA) surface of titanium dental implants. The HA/SLA surfaces of titanium dental implants were formed by the ion beam assisted deposition (IBAD) method. Rabbit bone marrow derived mesenchymal stem cells cultured in vitro were seeded onto the surface of SLA and HA/SLA; the growth states of MSCs on the two samples were observed by a scanning electron microscope; the proliferation index, alkaline phosphatase (ALP) activity, osteocalcin (OCN) content of MSCs and mRNA relative expression level of osteopontin (opn) were compared between two groups. MSCs were found to be easier to adhere to the HA/SLA surface compared to the SLA surface. At the same time, the ALP activity and the OCN content of MSCs grown on the HA/SLA surface were obviously higher, and the relative expression level of opn mRNA was 4.78 times higher than that on the SLA surface. The HA coating formed by the IBAD method on the SLA surface of titanium dental implants significantly improves proliferation and well-differentiated osteoblastic phenotype of MSCs, which indicates a promising method for the surface modification of titanium dental implants.

Associate Professor of Periodontology, Dental Faculty, Tehran University of Medical Sciences, Tehran, Iran.

OBJECTIVE: Implant placement in the posterior maxilla may often be contraindicated because of insufficient bone volume and presence of the maxillary sinus. In these situations, sinus floor augmentation frequently has been proposed as the best treatment. This clinical study was based on the hypothesis that the clinical effectiveness of adult mesenchymal stem cells (MSCs) loaded to the biphasic scaffold. METHODS: In this report, the clinical and radiographic results are presented on 6 consecutively treated patients using MSCs in combination with biphasic hydroxyl apatite/ beta-tricalcium phosphate (HA/TCP) for sinus elevation. All the patients in the study had less than 3 mm initial bone height in the posterior maxillary area (IBH). MSCs were cultured and expanded from bone marrow aspirate for each patient. Three months after sinus elevation, radiographic evaluation was performed for the patients and the secondary bone height was measured (SBH(1)). In the second stage surgery, 30 implants were placed. Trephine bur was used as a pilot drill and a core biopsy was obtained from each implant site. Prosthetic rehabilitation of the patients was performed after 4 months. Secondary bone height was measured 9 months after implant placement (SBH(2)). RESULTS: Of 30 implants, 28 (93%) were considered clinically successful. Two implants were removed due to mobility at the time of surgical exposure. Histologic evaluation of the biopsy specimens revealed numerous areas of osteoid and bone formation HA/TCP, with no evidence of inflammatory cell infiltrate. Mean bone regenerate was 41.34%. Clinically, no complications were observed, and all implants were considered clinically osseointegrated after 4 months. Mean bone height was measured 3 and 12 months after sinus grafting (mean of SBH(1)= 12.08 mm and mean of SBH(2)= 10.08 mm). CONCLUSIONS: These clinical and histological findings suggest that sinus grafting with HA/TCP in combination with MSCs provide a viable therapeutic alternative for implant placement. The findings suggest that the addition of MSCs to bone derivative/substitute materials may enhance bone formation in the maxillary sinus area. Of course more studies with the control groups are needed for the evaluation of this method as a clinical solution for the patients.

Laboratory of Molecular Signaling, Department of Biologic and Materials Sciences, School of Dentistry, The University of Michigan, Ann Arbor, MI 48109, USA.

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

Department of Comparative Biomedical Sciences, Louisiana State University, School of Veterinary Medicine, Skip Bertman Drive, Baton Rouge, LA 70803, USA

^* corresponding author, [email protected]

Craniofacial and Skeletal Diseases Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD 20892, USA.

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.

Department of Dental Pathology, Operative Dentistry and Endodontics, Royal Dental College, Faculty of Health Sciences, University of Aarhus, DK-8000 Aarhus C, Denmark. [email protected]

Recent studies have shown that the pulp of human teeth contains a population of cells with stem cell properties and it has been suggested that these cells originate from pericytes. Molecules of the Notch signaling pathway regulate stem cell fate specification, while Rgs5 represents an excellent marker for pericytes. Pathological conditions such as dental trauma and carious lesion stimulate pulp stem cells to elaborate reparative dentin. Previous studies have shown that genes involved in the Notch pathway are activated in response to pulp injury in rodent and humans. To demonstrate the importance of pericytes as a source of stem cells during dental repair, we have studied Rgs5 and Notch3 mRNA expression by in situ hybridization in developing, adult intact and injured rodent teeth. Furthermore, we have examined the distribution of Notch3 protein in carious and injured human teeth using immunohistochemistry. Overlapping expression patterns of Rgs5 and Notch3 were observed during rodent tooth development as well as immediately after injury. Both genes were expressed in vascular structures during development and in perivascular and single capillary cells of injured teeth. However, the expression patterns of Rgs5 and Notch3 were different during tooth repair, with relatively extensive Rgs5 expression along the pericyte-vascular smooth muscle cell axis in central pulp arterioles. These results show co-expression of Rgs5 and Notch3 in pericytes of developing and injured teeth and furthermore indicate the importance of vascular-derived stem cells during pulp healing.

University of Maryland, College of Dental Surgery, Dental School, Department of Endodontics, Prosthodontics and Operative Dentistry, Baltimore, Maryland 21201, USA. [email protected]

Some clinical case reports have shown that immature permanent teeth with periradicular periodontitis or abscess can undergo apexogenesis after conservative endodontic treatment. A call for a paradigm shift and new protocol for the clinical management of these cases has been brought to attention. Concomitantly, a new population of mesenchymal stem cells residing in the apical papilla of permanent immature teeth recently has been discovered and was termed stem cells from the apical papilla (SCAP). These stem cells appear to be the source of odontoblasts that are responsible for the formation of root dentin. Conservation of these stem cells when treating immature teeth may allow continuous formation of the root to completion. This article reviews current findings on the isolation and characterization of these stem cells. The potential role of these stem cells in the following respects will be discussed: (1) their contribution in continued root maturation in endodontically treated immature teeth with periradicular periodontitis or abscess and (2) their potential utilization for pulp/dentin regeneration and bioroot engineering.

University of Maryland, College of Dental Surgery, Dental School, Department of Endodontics, Prosthodontics and Operative Dentistry, 650 West Baltimore Street, Baltimore, MD 21201, United States.

OBJECTIVE: This article will review the new concept of regenerative endodontics in the management of immature permanent teeth. The potential role of stem cells to regenerate immature permanent teeth after conservative treatment will be discussed. DATA AND SOURCES: Two sets of data source are focused in this review: (i) the characterization of various dental stem cells discovered since 2000 and (ii) recent clinical case reports showing that after conservative treatment, severely infected immature teeth with periradicular periodontitis and abscess can undergo healing and apexogenesis or maturogenesis. RESULTS: A new protocol of treating endodontically involved immature permanent teeth based on published articles to date is summarized in the review. The key procedures of the protocol are (1) minimal or no instrumentation of the canal while relying on a gentle but thorough irrigation of the canal system, (2) the disinfection is augmented with intra-canal medication of a triple-antibiotic paste between appointments, and (3) the treated tooth is sealed with mineral trioxide aggregate (MTA) and glass ionomer/resin cement at the completion of the treatment. Periodical follow-ups will take place to observe any continued maturation of the root. CONCLUSION: While more clinical research is needed, regenerative endodontics promotes a paradigm shift in treating endodontically involved immature permanent teeth from performing apexification procedures to conserving any dental stem cells that might remain in the disinfected viable tissues to allow tissue regeneration and repair to achieve apexogenesis/maturogenesis.

Abstract:
Harvesting bone for autologous grafting is a daily problem encountered by craniofacial and oral surgeons. Stem cells derived from human dental pulp are able to differentiate in osteoblasts and are a potential source of autologous bone produced in vitro. However, as stem cells are characterized by self-renewing and commitment in several cellular subtypes (ie, pluripotential differentiation), some concerns may arise as regards their potential uncontrolled proliferation.

To screen the behavior of osteoblasts derived from human pulpar stem cells (ODHPSCs), we used microarray techniques to identify genes that are differently regulated in ODHPSC in comparison to normal osteoblasts (NOs). Osteoblasts derived from human pulpar stem cells were obtained from human dental pulp, and cells were selected using a cytometer. The cell profile was c-kit+/CD34+/STRO-1+/CD45-. These cells were capable of differentiation of osteoblasts in vitro.

By using DNA microarrays containing 19,200 genes, we identified in ODHPSC some genes whose expression was significantly up- and downregulated compared to NO. The differentially expressed genes have different functional activities: (a) cell differentiation, (b) developmental maturation, (c) cell adhesion, and (d) production of cytoskeleton elements.

Laboratório de Genética, Instituto Butantan, São Paulo, Brasil. [email protected]

We report the isolation of a population of immature dental pulp stem cells (IDPSC), which express embryonic stem cell markers Oct-4, Nanog, SSEA-3, SSEA-4, TRA-1-60 and TRA-1-81 as well as several other mesenchymal stem cell markers during at least 25 passages while maintaining the normal karyotype and the rate of expansion characteristic of stem cells. The expression of these markers was maintained in subclones obtained from these cells. Moreover, in vitrothese cells can be induced to undergo uniform differentiation into smooth and skeletal muscles, neurons, cartilage, and bone under chemically defined culture conditions. After in vivo transplantation of these cells into immunocompromised mice, they showed dense engraftment in various tissues. The relative ease of recovery and the expression profiles of various markers justify further exploration of IDPSC for clinical therapy. Copyright 2007 S. Karger AG, Basel.

Suchánek J, Soukup T, Ivancaková R, Karbanová J, Hubková V, Pytlík R, Kucerová L.

Charles University in Prague, Faculty of Medicine in Hradec Králové and University Hospital Hradec Králové, Department of Dentistry, Czech Republic. [email protected]  Acta Medica (Hradec Kralove). 2007;50(3):195-201.

Radboud University Nijmegen Medical Centre, Periodontology & Biomaterials, Nijmegen, The Netherlands.

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.

Stiftung Caesar, Center of Advanced European Studies and Research, Ludwig Erhard Allee 2, 53175 Bonn, Germany. [email protected]

The dental follicle is an ectomesenchymal tissue surrounding the developing tooth germ. It is believed that this tissue contains stem cells and lineage committed progenitor cells or precursor cells (PCs) for cementoblasts, periodontal ligament cells, and osteoblasts. In this study, we report the isolation of PCs derived from dental follicle of human third molar teeth. These fibroblast-like, colony forming and plastic adherent cells expressed putative stem cell markers Notch-1 and Nestin. We compared gene expressions of PCs, human mesenchymal stem cells (hMSCs), periodontal ligament cells (PDL-cells) and osteoblasts (MG63) for delimitation of PCs. Interestingly, PCs expressed higher amounts of insulin-like growth factor-2 (IGF-2) transcripts than hMSCs. Differentiation capacity was demonstrated under in vitro conditions for PCs. Long-term cultures with dexamethasone produced compact calcified nodules or appeared as plain membrane structures of different dimensions consisting of a connective tissue like matrix encapsulated by a mesothelium-like cellular structure. PCs differentially express osteocalcin (OCN) and bone sialoprotein (BS) after transplantation in immunocompromised mice but without any sign of cementum or bone formation. Therefore, our results demonstrate that cultured PCs are unique undifferentiated lineage committed cells residing in the periodontium prior or during tooth eruption.

Dipartimento di Medicina Sperimentale, Sezione Istologia ed Embriologia, Secondo Ateneo di Napoli, Napoli, Italy.

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

Transplantation. 2005 Sep 27; 80(6):836-42.

Pierdomenico L, Bonsi L, Calvitti M, Rondelli D, Arpinati M, Chirumbolo G, Becchetti E, Marchionni C, Alviano F, Fossati V, Staffolani N, Franchina M, Grossi A, Bagnara GP.

Department of Histology, Embryology, and Applied Biology, University of Bologna, Bologna, Italy.

Department of Periodontology and Biomaterials, Radboud University Nijmegen Medical Centre, PO Box 9101, 6500 HB Nijmegen, The Netherlands. J Tissue Eng Regen Med. 2008 Mar 14

Journal of Cellular Physiology. 2006 Aug; 208(2):319-25.

Papaccio G, Graziano A, d'Aquino R, Graziano MF, Pirozzi G, Menditti D, De Rosa A, Carinci F, Laino G.

Ballini A, De Frenza G, Cantore S, Papa F, Grano M, Mastrangelo F, Tetè S, Grassi FR.

Department of Internal Medicine and Public Health, Section of Medical Genetics, University of Bari, Italy.

Grace Dental Clinic, Kaohsiung, Taiwan.

Background: Dental pulp stem cells (DPSCs) were primarily derived from the pulp tissues of primary incisors and permanent third molar teeth, whereas no report to our knowledge has yet been documented on deriving DPSCs from the other tooth types. The aim of this study is to present a novel approach of harvesting stem cells from a supernumerary tooth (a mesiodens). Materials and methods: The pulp tissues from a mesiodens of a 20-year-old healthy male patient and the left lower deciduous canine of a healthy 10-year-old boy (the positive control) were extracted and cultured for DPSCs, which were examined with stem cells (Oct-4, Nanog and Rex-1) and differentiation (Osteonectin and Nestin) markers. Furthermore, DPSCs were directionally differentiated to osteogenic and adipogenic cell lineages. Results: Dental pulp stem cells derived from the mesiodens were capable of differentiating into adipogenic and osteogenic lineages. The mesioden's DPSCs also expressed stem cell and differentiation markers, which suggested their stem cell origin and differentiation capability. All the aforementioned results for the mesiodens were consistent with those of the DPSCs derived from the positive control. Conclusion: We have demonstrated the feasibility of deriving DPSCs from a usually discarded tissue such as a supernumerary tooth.

Dipartimento di Discipline Odontostomatologiche, Ortodontiche e Chirurgiche, Università Secondo Ateneo di Napoli, Napoli, Italy.

Stromal stem cells from human dental pulp (SBP-DPSCs) were used to study osteogenic differentiation in vitro and in vivo. We previously reported that SBP-DPSCs are multipotent stem cells able to differentiate into osteoblasts, which synthesize three-dimensional woven bone tissue chips in vitro. In this study, we followed the temporal expression pattern of specific markers in SBP-DPSCs and found that, when differentiating into osteoblasts, they express, besides osteocalcin, also flk-1 (VEGF-R2). In addition, 30% of them expressed specific antigens for endothelial cells, including CD54, von-Willebrand (domain 1 and 2), CD31 (PECAM-1) and angiotensin-converting enzyme. Interestingly, we found endotheliocytes forming vessel walls, observing that stem cells synergically differentiate into osteoblasts and endotheliocytes, and that flk-1 exerts a pivotal role in coupling osteoblast and endotheliocyte differentiation. When either SBP-DPSCs or bone chips obtained in vitro were transplanted into immunocompromised rats, they generated a tissue structure with an integral blood supply similar to that of human adult bone; in fact, a large number of HLA-1+ vessels were observed either within the bone or surrounding it in a periosteal layer. This study provides direct evidence to suggest that osteogenesis and angiogenesis mediated by human SBP-DPSCs may be regulated by distinct mechanisms, leading to the organization of adult bone tissue after stem cell transplantation.

Centre of Molecular Genetics, CARISBO Foundation, Institute of Histology and General Embryology, School of Medicine, University of Bologna, Bologna, Italy.

Stem cells derived from human dental pulp are able to differentiate into osteoblasts and are a potential source of autologous bone. The aim of this study was to compare genes differentially expressed in osteoblastoids from human dental pulp (OHDP) to osteosarcoma cells (OCs). Human dental pulp was extracted and immersed in a digestive solution. Cells were cultured and selected using c-kit, CD34, CD45 and STRO-1 antibodies. In parallel, two OCs (i.e., SAOS2 and TE85) were cultured. RNA was extracted from different populations of cells and cDNA was used for the hybridisation of human 19.2K DNA microarrays. We identified several differences in gene expression between OHDP and OCs. Some down-regulated OHDP genes, such as RUNX1, MAP4K4 and PRDM2, are involved in bone development, cell motility and transcript regulation. Gene expression in OHDP is significantly different from that in OCs, suggesting differences in cell function and activity between these cells.

Institute of Human Genetics, University of Regensburg, Regensburg, Germany.

Complex human tissues harbour stem cells and/or precursor cells, which are responsible for tissue development or repair. Recently, dental tissues such as periodontal ligament (PDL), dental papilla or dental follicle have been identified as easily accessible sources of undifferentiated cells. The dental stem cell biology might provide meaningful insights into the development of dental tissues and cellular differentiation processes. Dental stem cells could also be feasible tools for dental tissue engineering. Constructing complex structures like a periodontium, which provides the functional connection between a tooth or an implant and the surrounding jaw, could effectively improve modern dentistry. Dental precursor cells are attractive for novel approaches to treat diseases like periodontitis, dental caries or to improve dental pulp healing and the regeneration of craniofacial bone and teeth. These cells are easily accessible and, in contrast to bone-marrow-derived mesenchymal stem cells, are more closely related to dental tissues. This review gives a short overview of stem cells of dental origin.

Department of Craniofacial Development, Dental Institute, Floor 27, Guy’s Hospital, Kings College London, London Bridge, London, SE1 9RT, UK, [email protected].

The notion that teeth contain stem cells is based on the well-known repairing ability of dentin after injury. Dental stem cells have been isolated according to their anatomical locations, colony-forming ability, expression of stem cell markers, and regeneration of pulp/dentin structures in vivo. These dental-derived stem cells are currently under increasing investigation as sources for tooth regeneration and repair. Further attempts with bone marrow mesenchymal stem cells and embryonic stem cells have demonstrated the possibility of creating teeth from non-dental stem cells by imitating embryonic development mechanisms. Although, as in tissue engineering of other organs, many challenges remain, stem-cell-based tissue engineering of teeth could be a choice for the replacement of missing teeth in the future.

Department of Oral and Maxillofacial Surgery, West China College of Stomatology, Sichuan University, No. 14, 3rd Sec., Ren Min Nan Road, Sichuan Province, Chengdu 610041, China.

Tooth regeneration using tissue engineering concepts is a promising biological approach to solving problems of tooth loss in elderly patients. The seeding cells, however, for tooth regeneration such as odontoblasts from dental germ, stem cells from dental pulp and deciduous teeth, and ectomesenchymal cells from the first branchial arch are difficult, even impossible to harvest in clinic. Bone marrow mesenchymal stem cells have odontogenic capacity, but their differentiation abilities significantly decrease with the increasing age of the donors. Therefore, the cells mentioned above are not practical in the clinical application of tooth regeneration in the old. Adipose derived stem cells have many clinical advantages over bone marrow mesenchymal stem cells, and their differentiation potential can be maintained with aging. Here we propose the hypothesis that adipose derived stem cells could be induced into odontogenic lineage and might be used as suitable seeding cells for tooth regeneration to replace the lost tooth of elderly patients.

University of Maryland, College of Dental Surgery, Dental School, Department of Endodontics, Prosthodontics and Operative Dentistry, 650 West Baltimore Street, Baltimore, MD 21201, United States.

OBJECTIVE: This article will review the new concept of regenerative endodontics in the management of immature permanent teeth. The potential role of stem cells to regenerate immature permanent teeth after conservative treatment will be discussed. DATA AND SOURCES: Two sets of data source are focused in this review: (i) the characterization of various dental stem cells discovered since 2000 and (ii) recent clinical case reports showing that after conservative treatment, severely infected immature teeth with periradicular periodontitis and abscess can undergo healing and apexogenesis or maturogenesis. RESULTS: A new protocol of treating endodontically involved immature permanent teeth based on published articles to date is summarized in the review. The key procedures of the protocol are (1) minimal or no instrumentation of the canal while relying on a gentle but thorough irrigation of the canal system, (2) the disinfection is augmented with intra-canal medication of a triple-antibiotic paste between appointments, and (3) the treated tooth is sealed with mineral trioxide aggregate (MTA) and glass ionomer/resin cement at the completion of the treatment. Periodical follow-ups will take place to observe any continued maturation of the root. CONCLUSION: While more clinical research is needed, regenerative endodontics promotes a paradigm shift in treating endodontically involved immature permanent teeth from performing apexification procedures to conserving any dental stem cells that might remain in the disinfected viable tissues to allow tissue regeneration and repair to achieve apexogenesis/maturogenesis.

Department of Stomatology and Oral Sciences and Ce.S.I, University of Chieti-Pescara, Italy.

Recent studies have shown that mesenchymal stem cells obtained from periodontal ligament (PDL-MSCs) are multipotent cells that have similar features of the bone marrow and dental pulp MSCs and are capable of proliferating and producing different types of tissue such as bone and tooth associated-tissues. Human PDL-MSCs expanded ex vivo were induced to osteogenesis, seeded in three-dimensional biocompatible scaffolds (fibrin sponge, bovine-derived substitutes) and examined using light, scanning and transmission electron microscopy. Morphological observations showed extensive growth of cellular biomass partially covering the scaffolds after 4 weeks of incubation in mineralization medium. These findings indicate that periodontal ligament can be an easily and efficient autologous source of stem cells with a high expansion capacity and ability to differentiate in osteogenic cells that can colonize and grow connected to bio-compatible scaffold. It can be suggested that the use of PDL-MSCs for generating graft biomaterials is advantageous for bone tissue engineering in regenerative dentistry. (c) 2008 Wiley Periodicals, Inc. J Biomed Mater Res 2008.

Dipartimento di Medicina Sperimentale, Sezione Istologia ed Embriologia, TESLab, Secondo Ateneo di Napoli, Napoli, Italy.

OBJECTIVES: The aim of this study was to select and provide enough stem cells for quick transplantation in bone engineering procedures, avoiding any in vitro expansion step. MATERIALS AND METHODS: Dental germ pulp, collected from 25 healthy subjects aged 13-20 years, were subjected to magnetic-activated cell sorting to select a CD34(+) stem cell population capable of differentiating into pre-osteoblasts. These cells were allowed to adhere to an absorbable polylactic-coglycolic acid scaffold for 30 min, without any prior expansion, and the CD34(+) cell-colonized scaffolds were then transplanted into immunocompromised rats, subcutaneously. RESULTS: After 60 days, analysis of recovered transplants revealed that they were formed of nodules of bone, of the same dimensions as the original scaffold. Bone-specific proteins were detected by immunofluorescence, within the nodules, and X-ray diffraction patterns revealed characteristic features of bone. In addition, presence of platelet endothelial cell adhesion molecule and von Willebrand factor immunoreactivity were suggestive of neo-angiogenesis and neovasculogenesis taking place within nodules. Importantly, these vessels were HLA-1(+) and, thus, clearly human in origin. CONCLUSIONS: This study indicates that CD34(+) cells obtained from dental pulp can be used for engineering bone, without the need for prior culture expanding procedures. Using autologous stem cells, this schedule could be used to provide the basis for bone regenerative surgery, with limited sacrifice of tissue, low morbidity at the collection site, and significant reduction in time needed for clinical recovery.

Dipartimento di Discipline Odontostomatologiche, Ortodontiche e Chirurgiche, Università Secondo Ateneo di Napoli, Napoli, Italy.

Stromal stem cells from human dental pulp (SBP-DPSCs) were used to study osteogenic differentiation in vitro and in vivo. We previously reported that SBP-DPSCs are multipotent stem cells able to differentiate into osteoblasts, which synthesize three-dimensional woven bone tissue chips in vitro. In this study, we followed the temporal expression pattern of specific markers in SBP-DPSCs and found that, when differentiating into osteoblasts, they express, besides osteocalcin, also flk-1 (VEGF-R2). In addition, 30% of them expressed specific antigens for endothelial cells, including CD54, von-Willebrand (domain 1 and 2), CD31 (PECAM-1) and angiotensin-converting enzyme. Interestingly, we found endotheliocytes forming vessel walls, observing that stem cells synergically differentiate into osteoblasts and endotheliocytes, and that flk-1 exerts a pivotal role in coupling osteoblast and endotheliocyte differentiation. When either SBP-DPSCs or bone chips obtained in vitro were transplanted into immunocompromised rats, they generated a tissue structure with an integral blood supply similar to that of human adult bone; in fact, a large number of HLA-1+ vessels were observed either within the bone or surrounding it in a periosteal layer. This study provides direct evidence to suggest that osteogenesis and angiogenesis mediated by human SBP-DPSCs may be regulated by distinct mechanisms, leading to the organization of adult bone tissue after stem cell transplantation.

Oral and Maxillofacial Department, Qilu Hospital of Shangdong University, Jinan 250012, Shandong province, China. [email protected]

PURPOSE: This study was aimed to investigate the localization of dental pulp stem cells (DPSCs) by comparing the characteristics of cultured dental pulp cells in coronal and root pulp. METHODS: Human dental coronal and root pulp cells were cultured in tissue-explant method, the cell culture successfulness, attachment efficiency, cell viality, morphology, proliferation pattern, and the mineralization ability were observed, the localization of DPSCs was investigated in the functional respect of DPSCs. RESULTS: The human dental root pulp cells have more culture successfulness,more attachment efficiency,more cell viality, more primary characteristics, and stronger induced mineralization ability than that of coronal pulp cells. Root and coronal pulp cells showed same proliferation patterns. CONCLUSIONS: DPSCs may exist in both dental root and coronal pulp, and the density of DPSCs in the root pulp may be higher than the coronal pulp.

Center for Craniofacial Molecular Biology, University of Southern California School of Dentistry, Los Angeles, California, USA.

Mesenchymal stem cells (MSCs) have been isolated from the pulp tissue of permanent teeth (dental pulp stem cells or DPSCs) and deciduous teeth (stem cells from human exfoliated deciduous teeth). We recently discovered another type of MSCs in the apical papilla of human immature permanent teeth termed stem cells from the apical papilla (SCAP). Here, we further characterized the apical papilla tissue and stem cell properties of SCAP using histologic, immunohistochemical, and immunocytofluorescent analyses. We found that the apical papilla is distinctive to the pulp in terms of containing less cellular and vascular components than those in the pulp. Cells in the apical papilla proliferated 2- to 3-fold greater than those in the pulp in organ cultures. Both SCAP and DPSCs were as potent in osteo/dentinogenic differentiation as MSCs from bone marrows, whereas they were weaker in adipogenic potential. The immunophenotype of SCAP is similar to that of DPSCs on the osteo/dentinogenic and growth factor receptor gene profiles. Double-staining experiments showed that STRO-1 coexpressed with dentinogenic markers such as bone sialophosphoprotein, osteocalcin, and growth factors FGFR1 and TGFbetaRI in cultured SCAP. Additionally, SCAP express a wide variety of neurogenic markers such as nestin and neurofilament M upon stimulation with a neurogenic medium. We conclude that SCAP are similar to DPSCs but a distinct source of potent dental stem/progenitor cells. Their implications in root development and apexogenesis are discussed.

Mund Kiefer Gesichtschir. 2007 Sep 6

Morsczeck C, Reichert TE, Völlner F, Gerlach T, Driemel O.

Institut für Humangenetik, Universität Regensburg, Franz-Josef-Strauß-Allee Regensburg, Germany

International Journal Immunopathology & Pharmacology. 2006 Jul-Sep; 19(3):451-60.

Trubiani O, Orsini G, Caputi S, Piatelli A.

Department of Stomatology and Oral Science, Ce.SI. Foundation G. d'Annunzio, Chieti, Italy.