Masako Miura

Investigation of multipotent postnatal stem cells from human periodontal ligament

BACKGROUNDnPeriodontal diseases that lead to the destruction of periodontal tissues--including periodontal ligament (PDL), cementum, and bone--are a major cause of tooth loss in adults and are a substantial public-health burden worldwide. PDL is a specialised connective tissue that connects cementum and alveolar bone to maintain and support teeth in situ and preserve tissue homoeostasis. We investigated the notion that human PDL contains stem cells that could be used to regenerate periodontal tissue.nnnMETHODSnPDL tissue was obtained from 25 surgically extracted human third molars and used to isolate PDL stem cells (PDLSCs) by single-colony selection and magnetic activated cell sorting. Immunohistochemical staining, RT-PCR, and northern and western blot analyses were used to identify putative stem-cell markers. Human PDLSCs were transplanted into immunocompromised mice (n=12) and rats (n=6) to assess capacity for tissue regeneration and periodontal repair. Findings PDLSCs expressed the mesenchymal stem-cell markers STRO-1 and CD146/MUC18. Under defined culture conditions, PDLSCs differentiated into cementoblast-like cells, adipocytes, and collagen-forming cells. When transplanted into immunocompromised rodents, PDLSCs showed the capacity to generate a cementum/PDL-like structure and contribute to periodontal tissue repair.nnnINTERPRETATIONnOur findings suggest that PDL contains stem cells that have the potential to generate cementum/PDL-like tissue in vivo. Transplantation of these cells, which can be obtained from an easily accessible tissue resource and expanded ex vivo, might hold promise as a therapeutic approach for reconstruction of tissues destroyed by periodontal diseases.

SHED: Stem cells from human exfoliated deciduous teeth

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.

Accumulated chromosomal instability in murine bone marrow mesenchymal stem cells leads to malignant transformation

Despite recent emerging evidence suggesting that cancer stem cells subsist in a variety of tumors, it is not yet fully elucidated whether postnatal stem cells are directly involved in tumorigenesis. We used murine bone marrow–derived mesenchymal stem cells (BMMSCs) as a model to test a hypothesis that tumorigenesis may originate from spontaneous mutation of stem cells. In this study, we demonstrated that murine BMMSCs, after numerous passages, obtained unlimited population doublings and proceeded to a malignant transformation state, resulting in fibrosarcoma formation in vivo. Transformed BMMSCs colonized to multiple organs when delivered systemically through the tail vein. Fibrosarcoma cells formed by transformed BMMSCs contained cancer progenitors, which were capable of generating colony clusters in vitro and fibrosarcoma in vivo by the second administration. The mechanism by which BMMSCs transformed to malignant cells was associated with accumulated chromosomal abnormalities, gradual elevation in telomerase activity, and increased c‐myc expression. Moreover, BMMSCs and their transformed counterpart, fibrosarcoma‐forming cells, demonstrated different sensitivity to anti‐cancer drugs. BMMSCs/fibrosarcoma transformation system may provide an ideal system to elucidate the mechanism of how stem cells become cancer cells and to screen anti‐sarcoma drugs.

Recovery of Stem Cells from Cryopreserved Periodontal Ligament

Human post-natal stem cells possess a great potential to be utilized in stem-cell-mediated clinical therapies and tissue engineering. It is not known whether cryopreserved human tissues contain functional post-natal stem cells. In this study, we utilized human periodontal ligament to test the hypothesis that cryopreserved human periodontal ligament contains retrievable post-natal stem cells. These cryopreserved periodontal ligament stem cells maintained normal periodontal ligament stem cell characteristics, including expression of the mesenchymal stem cell surface molecule STRO-1, single-colony-strain generation, multipotential differentiation, cementum/periodontal-ligament-like tissue regeneration, and a normal diploid karyotype. Collectively, this study provides valuable evidence demonstrating a practical approach to the preservation of solid-frozen human tissues for subsequent post-natal stem cell isolation and tissue regeneration. The present study demonstrates that human post-natal stem cells can be recovered from cryopreserved human periodontal ligament, thereby providing a practical clinical approach for the utilization of frozen tissues for stem cell isolation.

Mesenchymal Stem Cell‐Organized Bone Marrow Elements: An Alternative Hematopoietic Progenitor Resource

Bone marrow‐derived mesenchymal stem cells (BMMSCs) are multipotent postnatal stem cells that have been used for the treatment of bone defects and graft‐versus‐host diseases in clinics. In this study, we found that subcutaneously transplanted human BMMSCs are capable of organizing hematopoietic progenitors of recipient origin. These hematopoietic cells expressed multiple lineages of hematopoietic cell associated markers and were able to rescue lethally irradiated mice, with successful engraftment in the recipient, suggesting a potential bone marrow (BM) resource for stem cell therapies. Furthermore, we found that platelet‐derived growth factor (PDGF) promotes the formation of BMMSC‐generated BM niches through upregulation of β‐catenin, implying that the PDGF pathway contributes to the formation of ectopic BM. These results indicate that the BMMSC‐organized BM niche system represents a unique hematopoietic progenitor resource possessing potential clinical value.