The potential of enriched mesenchymal stem cells with neural crest cell phenotypes as a cell source for regenerative dentistry

Abstract

Effective regenerative treatments for periodontal tissue defects have recently been demonstrated using mesenchymal stromal/stem cells (MSCs). Furthermore, current bioengineering techniques have enabled de novo fabrication of tooth-perio dental units in mice. These cutting-edge technologies are expected to address unmet needs within regenerative dentistry. However, to achieve efficient and stable treatment outcomes, preparation of an appropriate stem cell source is essential. Many researchers are investigating the use of adult stem cells for regenerative dentistry; bone marrow-derived MSCs (BM-MSCs) are particularly promising and presently used clinically. However, current BM-MSC isolation techniques result in a heterogeneous, non-reproducible cell population because of a lack of identified distinct BM-MSC surface markers. Recently, specific subsets of cell surface markers for BM-MSCs have been reported in mice (PDGFRα⁺ and Sca-1⁺) and humans (LNGFR⁺, THY-1⁺ and VCAM-1⁺), facilitating the isolation of unique enriched BM-MSCs (so-called "purified MSCs"). Notably, the enriched BM-MSC population contains neural crest-derived cells, which can differentiate into cells of neural crest- and mesenchymal lineages. In this review, characteristics of the enriched BM-MSCs are outlined with a focus on their potential application within future regenerative dentistry.

Keywords: Bone marrow-derived mesenchymal stem cell; Enriched/purified mesenchymal stem cell; Flow cytometric isolation; Neural crest cell; Regenerative dentistry.

Figure 1

Schematic diagram illustrating neural crest cell (NCC) location and progression. In the early embryo, the neural tube is formed and epithelial cells at the dorsal region of the neural tube undergo an epithelial-to-mesenchymal transition to become migratory NCCs. NCCs migrating from cranial and trunk regions of the neural tube differentiate into many diverse derivatives of mesenchymal- and ectodermal-lineage cells and tissues. In craniofacial development, cranial NCCs contribute to ectodermal tissues and mesenchymal tissues, including dental mesenchyme components, such as the dentin, dental pulp, periodontal ligament, gingival connective tissue, cementum, and alveolar bone. Trunk NCCs differentiate mainly into neurons and glial cells of the peripheral nervous system. A small mesenchymal contribution by trunk NCCs has also been reported.

Figure 2

Schematic diagram illustrating mBM-MSC isolation by flow cytometry , . Crushed bone fragments from adult mouse tibia are incubated in cell culture medium with collagenase to obtain a cell suspension. Cells are stained with monoclonal antibodies against CD45, Ter119, PDGFRα, and Sca-1. After staining, cells are sorted using a flow cytometer (see details in the protocol article [72]). Sorted Sca-1⁻/Ter119⁻/PDGFRα⁺/Sca-1⁺ cells (PαS mBM-MSCs: enriched mBM-MSCs) differentiate into cells of mesenchymal lineage (osteoblasts, adipocytes, and chondrocytes) and neural crest lineage (neurons, glia, and smooth muscle cells). Phase: phase contrast microscope image; Os: osteoblasts (alkaline phosphatase); Ad: adipocytes (Oil Red O); Ch: chondrocytes (toluidine blue); Neu: neurons (βIII tubulin); Gl: glia (glial fibrillary acidic protein: GFAP); SM: smooth muscle cells (α-smooth muscle actin); ND: not detected.

Figure 3

Schematic diagram illustrating the prospects of using enriched hBM-MSCs from bone marrow for craniofacial tissue/organ regeneration. Craniofacial mesenchyme develops from migrating cranial NCCs. A certain portion of trunk NCCs migrates to limb bone marrow, where these cells reside as a part of adult BM-MSCs. Therefore, enriched hBM-MSCs, which are isolated from the BM-MSC population using a flow cytometer (LNGFR⁺, THY1⁺, and VCAM-1⁺ cells: LTV hBM-MSCs), show an NCC phenotype, and they are expected to be a useful cell source for craniofacial regenerative medicine.