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


BACKGROUND
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.


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.


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.
Laino
Gregorio
G
Dipartimento di Medicina Sperimentale, Sezione Istologia ed Embriologia, Secondo Ateneo di Napoli, Napoli, Italy.
d'Aquino
Riccardo
R
Graziano
Antonio
A
Lanza
Vladimiro
V
Carinci
Francesco
F
Naro
Fabio
F
Pirozzi
Giuseppe
G
Papaccio
Gianpaolo
G
D000328:Adult; D000469:Alkaline Phosphatase / Q000032:analysis; D000818:Animals; D018952:Antigens, CD34 / Q000032:analysis; D001861:Bone Regeneration; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D024061:Collagen Type III / Q000032:analysis; D003782:Dental Pulp / Q000166:cytology*; D005353:Fibronectins / Q000032:analysis; D006801:Humans; D018960:Hyaluronan Receptors / Q000032:analysis; D058574:Integrin-Binding Sialoprotein; D017493:Leukocyte Common Antigens / Q000032:analysis; D010006:Osteoblasts / Q000502:physiology; D015675:Osteocalcin / Q000032:analysis; D010011:Osteocytes / Q000166:cytology* / Q000502:physiology; D015676:Osteonectin / Q000032:analysis; D019009:Proto-Oncogene Proteins c-kit / Q000032:analysis; D051381:Rats; D012795:Sialoglycoproteins / Q000032:analysis; D033581:Stem Cell Transplantation; D013234:Stem Cells / Q000276:immunology / Q000502:physiology*
D018952:Antigens, CD34; D024061:Collagen Type III; D005353:Fibronectins; D018960:Hyaluronan Receptors; C546871:IBSP protein, human; D058574:Integrin-Binding Sialoprotein; D015676:Osteonectin; D012795:Sialoglycoproteins; D015675:Osteocalcin; D019009:Proto-Oncogene Proteins c-kit; D000469:Alkaline Phosphatase; D017493:Leukocyte Common Antigens