SALL1 regulates commitment of odontoblast lineages by interacting with RUNX2 to remodel open chromatin regions.

Mouse dental papilla cells (mDPCs) derive from cranial neural crest cells and maintain mesenchymal stem cell characteristics. The differentiation of neural crest cells into odontoblasts is orchestrated by transcription factors regulating the expression of genes whose enhancers are initially inaccessible. However, the identity of the transcription factors driving the emergence of odontoblast lineages remains elusive. In this study, we identified SALL1, a transcription factor that was particularly expressed in preodontoblasts, polarizing odontoblasts, and secretory odontoblasts in vivo. Knockdown of Sall1 in mDPCs inhibited their odontoblastic differentiation. In order to identify the regulatory network of Sall1, RNA sequencing and an assay for transposase-accessible chromatin with high-throughput sequencing were performed to analyze the genome-wide direct regulatory targets of SALL1. We found that inhibition of Sall1 expression could decrease the accessibility of some chromatin regions associated with odontoblast lineages at embryonic day 16.5, whereas these regions remained unaffected at postnatal day 0.5, suggesting that SALL1 regulates the fate of mDPCs by remodeling open chromatin regions at the early bell stage. Specifically, we found that SALL1 could directly increase the accessibility of cis-regulatory elements near Tgf-ß2 and within the Runx2 locus. Moreover, coimmunoprecipitation and proximal ligation assays showed that SALL1 could establish functional interactions with RUNX2. Taken together, our results demonstrated that SALL1 positively regulates the commitment of odontoblast lineages by interacting with RUNX2 and directly activating Tgf-ß2 at an early stage.

Intra-Individual Variability of Human Dental Pulp Stem Cell Features Isolated from the Same Donor.

It is primarily important to define the standard features and factors that affect dental pulp stem cells (DPSCs) for their broader use in tissue engineering. This study aimed to verify whether DPSCs isolated from various teeth extracted from the same donor exhibit intra-individual variability and what the consequences are for their differentiation potential. The heterogeneity determination was based on studying the proliferative capacity, viability, expression of phenotypic markers, and relative length of telomere chromosomes. The study included 14 teeth (6 molars and 8 premolars) from six different individuals ages 12 to 16. We did not observe any significant intra-individual variability in DPSC size, proliferation rate, viability, or relative telomere length change within lineages isolated from different teeth but the same donor. The minor non-significant variances in phenotype were probably mainly because DPSC cell lines comprised heterogeneous groups of undifferentiated cells independent of the donor. The other variances were seen in DPSC lineages isolated from the same donor, but the teeth were in different stages of root development. We also did not observe any changes in the ability of cells to differentiate into mature cell lines-chondrocytes, osteocytes, and adipocytes. This study is the first to analyze the heterogeneity of DPSC dependent on a donor.

BMP signaling orchestrates a transcriptional network to control the fate of mesenchymal stem cells in mice.

Signaling pathways are used reiteratively in different developmental processes yet produce distinct cell fates through specific downstream transcription factors. In this study, we used tooth root development as a model with which to investigate how the BMP signaling pathway regulates transcriptional complexes to direct the fate determination of multipotent mesenchymal stem cells (MSCs). We first identified the MSC population supporting mouse molar root growth as Gli1+ cells. Using a Gli1-driven Cre-mediated recombination system, our results provide the first in vivo evidence that BMP signaling activity is required for the odontogenic differentiation of MSCs. Specifically, we identified the transcription factors Pax9, Klf4, Satb2 and Lhx8 as being downstream of BMP signaling and expressed in a spatially restricted pattern that is potentially involved in determining distinct cellular identities within the dental mesenchyme. Finally, we found that overactivation of one key transcription factor, Klf4, which is associated with the odontogenic region, promotes odontogenic differentiation of MSCs. Collectively, our results demonstrate the functional significance of BMP signaling in regulating MSC fate during root development and shed light on how BMP signaling can achieve functional specificity in regulating diverse organ development.

Pentimento: Neural Crest and the origin of mesectoderm.

The Neural Crest, a transient epithelium in vertebrate embryos, is the source of putative stem cells known to give rise to neuronal, glial and endocrine components of the peripheral (sensory, autonomic and enteric) nervous system (PNS) and pigment cells in the skin. The Neural Crest is also widely believed to be the source of mesectodermal derivatives (skeletogenic, odontogenic, connective tissue and smooth muscle mesenchyme) in the vertebrate head [see (Bronner and LeDouarin, 2012; Le Douarin, 2012; Le Douarin and Kalcheim, 1999); see also (Hörstadius, 1950; Weston, 1970)]. This conventional understanding of the broad developmental potential of the Neural Crest has been challenged over the past few years (Breau et al., 2008; Lee et al., 2013a, 2013b; Weston et al., 2004), based on recognition that the definition of the embryonic epithelia that comprise the Neural Crest may be imprecise. Indeed, the definition of the embryonic tissues understood to constitute the Neural Crest has changed considerably since it was first described by Wilhelm His 150 years ago (His, 1868). Today, the operational definition of the Neural Crest is inconsistent and functionally ambiguous. We believe that more precise definitions of the embryonic tissues involved in Neural Crest development would be useful to understand (1) the range of cellular phenotypes that actually segregate from it, (2) when this lineage diversification occurs, and (3) how diversification is regulated. In this idiosyncratic review, we aim to explain our concerns with the current definitions in this field, and in the chiastic words of Samuel Johnson (1781), "… make new things familiar and familiar things new".(1) Then, we will try to distinguish the developmental events crucial to the regulation of Neural Crest development at both cranial and trunk axial levels of vertebrate embryos, and address some of the implicit assumptions that underlie the conventional interpretation of experimental results on the origin and fates of Neural Crest-derived cells. We hope our discussion will resolve some ambiguities regarding both the range of derivatives in the Neural Crest lineage and the conventional understanding that cranial mesectodermal derivatives share a common Neural Crest-derived lineage precursor with components of the PNS.

Differential Neuronal Plasticity of Dental Pulp Stem Cells From Exfoliated Deciduous and Permanent Teeth Towards Dopaminergic Neurons.

Based on early occurrence in chronological age, stem-cells from human exfoliated deciduous teeth (SHED) has been reported to possess better differentiation-potential toward certain cell-lineage in comparison to stem-cells from adult teeth (DPSCs). Whether this same property between them extends for the yield of functional central nervous system neurons is still not evaluated. Hence, we aim to assess the neuronal plasticity of SHED in comparison to DPSCs toward dopaminergic-neurons and further, if the difference is reflected in a differential expression of sonic-hedgehog (SHH)-receptors and basal-expressions of tyrosine-hydroxylase [TH; through cAMP levels]. Human SHED and DPSCs were exposed to midbrain-cues [SHH, fibroblast growth-factor8, and basic fibroblast growth-factor], and their molecular, immunophenotypical, and functional characterization was performed at different time-points of induction. Though SHED and DPSCs spontaneously expressed early-neuronal and neural-crest marker in their naïve state, only SHED expressed a high basal-expression of TH. The upregulation of dopaminergic transcription-factors Nurr1, Engrailed1, and Pitx3 was more pronounced in DPSCs. The yield of TH-expressing cells decreased from 49.8% to 32.16% in SHED while it increased from 8.09% to 77.47% in DPSCs. Dopamine release and intracellular-Ca(2+) influx upon stimulation (KCl and ATP) was higher in induced DPSCs. Significantly lower-expression of SHH-receptors was noted in naïve SHED than DPSCs, which may explain the differential neuronal plasticity. In addition, unlike DPSCs, SHED showed a down-regulation of cyclic adenosine-monophosphate (cAMP) upon exposure to SHH; possibly another contributor to the lesser differentiation-potential. Our data clearly demonstrates for the first time that DPSCs possess superior neuronal plasticity toward dopaminergic-neurons than SHED; influenced by higher SHH-receptor and lower basal TH expression. J. Cell. Physiol. 231: 2048-2063, 2016. © 2016 Wiley Periodicals, Inc.

Genetic lineage labeling in zebrafish uncovers novel neural crest contributions to the head, including gill pillar cells.

At the protochordate-vertebrate transition, a new predatory lifestyle and increased body size coincided with the appearance of a true head. Characteristic innovations of this head are a skull protecting and accommodating a centralized nervous system, a jaw for prey capture and gills as respiratory organs. The neural crest (NC) is a major ontogenetic source for the 'new head' of vertebrates and its contribution to the cranial skeleton has been intensively studied in different model organisms. However, the role of NC in the expansion of the respiratory surface of the gills has been neglected. Here, we use genetic lineage labeling to address the contribution of NC to specific head structures, in particular to the gills of adult zebrafish. We generated a sox10:ER(T2)-Cre line and labeled NC cells by inducing Cre/loxP recombination with tamoxifen at embryonic stages. In juvenile and adult fish, we identified numerous established NC derivatives and, in the cranium, we precisely defined the crest/mesoderm interface of the skull roof. We show the NC origin of the opercular bones and of multiple cell types contributing to the barbels, chemosensory organs located in the mouth region. In the gills, we observed labeled primary and secondary lamellae. Clonal analysis reveals that pillar cells, a craniate innovation that mechanically supports the filaments and forms gill-specific capillaries, have a NC origin. Our data point to a crucial role for the NC in enabling more efficient gas exchange, thus uncovering a novel, direct involvement of this embryonic tissue in the evolution of respiratory systems at the protochordate-vertebrate transition.

Designing materials to direct stem-cell fate.

Proper tissue function and regeneration rely on robust spatial and temporal control of biophysical and biochemical microenvironmental cues through mechanisms that remain poorly understood. Biomaterials are rapidly being developed to display and deliver stem-cell-regulatory signals in a precise and near-physiological fashion, and serve as powerful artificial microenvironments in which to study and instruct stem-cell fate both in culture and in vivo. Further synergism of cell biological and biomaterials technologies promises to have a profound impact on stem-cell biology and provide insights that will advance stem-cell-based clinical approaches to tissue regeneration.

SMAD4-mediated WNT signaling controls the fate of cranial neural crest cells during tooth morphogenesis.

TGFß/BMP signaling regulates the fate of multipotential cranial neural crest (CNC) cells during tooth and jawbone formation as these cells differentiate into odontoblasts and osteoblasts, respectively. The functional significance of SMAD4, the common mediator of TGFß/BMP signaling, in regulating the fate of CNC cells remains unclear. In this study, we investigated the mechanism of SMAD4 in regulating the fate of CNC-derived dental mesenchymal cells through tissue-specific inactivation of Smad4. Ablation of Smad4 results in defects in odontoblast differentiation and dentin formation. Moreover, ectopic bone-like structures replaced normal dentin in the teeth of Osr2-IresCre;Smad4(fl/fl) mice. Despite the lack of dentin, enamel formation appeared unaffected in Osr2-IresCre;Smad4(fl/fl) mice, challenging the paradigm that the initiation of enamel development depends on normal dentin formation. At the molecular level, loss of Smad4 results in downregulation of the WNT pathway inhibitors Dkk1 and Sfrp1 and in the upregulation of canonical WNT signaling, including increased ß-catenin activity. More importantly, inhibition of the upregulated canonical WNT pathway in Osr2-IresCre;Smad4(fl/fl) dental mesenchyme in vitro partially rescued the CNC cell fate change. Taken together, our study demonstrates that SMAD4 plays a crucial role in regulating the interplay between TGFß/BMP and WNT signaling to ensure the proper CNC cell fate decision during organogenesis.

Matrix mechanics and regulation of the fibroblast phenotype.

The oral cavity hosts a variety of different fibroblast populations that are generally responsible for maintaining homeostasis of the soft connective tissue. In addition to regulating the turnover and structural arrangement of collagen and other proteins of the extracellular matrix, fibroblasts perform a number of specialized functions. Certain fibroblast subpopulations in the gingiva, oral mucosa and periodontal ligament serve as progenitor cells with multilineage differentiation and self-renewal characteristics. In the periodontal ligament, fibroblasts further appear to function as mechanosensing entities that regulate collagen-secretory and collagen-remodeling activities according to the level of strain in the ligament. Mechanical challenge also plays an important role during the activation of periodontal fibroblasts in response to injury. Dysregulation of this activation process can lead either to poor healing and chronic wounds or to overly healed wounds with fibrosis. This review will elaborate on the roles of mechanical factors and mechanoperception in fibroblast activation, the molecular features of activated fibroblasts and the regulation mechanisms of fibroblast contraction. Pharmacological interference at each level is currently being pursued to improve the outcome of healing of injured periodontal tissue.

The effect of melatonin on Hippo signaling pathway in dental pulp stem cells.

Dental pulp stem cells (DPSCs) have a high capacity to differentiate into the neuronal cell lineage. Meanwhile, both Hippo signaling and melatonin are key regulators in neuronal differentiation of neuronal progenitor cells. Recently emerging evidences suggest the possible interaction between melatonin and Hippo signaling in different cell lines. But underlying mechanisms involved in the initiation or progression of neurogenic differentiation in DPSCs through this connection need to be explored. Therefore, the scope of this study is to investigate the effect of melatonin on Hippo signaling pathway through the expression of its downstream effector (YAP/p-YAPY357) after the neuronal differentiation of DPSCs. In regard with this, DPSCs were incubated with growth and dopaminergic neuronal differentiation medium with or without melatonin (10 µM) for 21 days. The morphological changes were followed by phase contrast microscopy and differentiation of DPSCs was evaluated by immunofluorescence labelling with NeuN, GFAP, and tyrosine hydroxylase. Furthermore, we evaluated the presence of neural progenitor cells by nestin immunoreactivity. Hippo signaling pathway was investigated by evaluating the immunoreactivity of YAP and p-YAPY357. Our results were also supported by western-blot analysis and SOX2, PCNA and caspase-3 were also evaluated. The positive immunoreactivity for NeuN, tyrosine hydroxylase and negative immunoreactivity for GFAP showed the successful differentiation of DPSCs to neurons, not glial cells. Melatonin addition to dopaminergic media induced tyrosine hydroxylase and decreased significantly nestin expression. The expressions of PCNA and caspase-3 were also decreased significantly with melatonin addition into growth media. Melatonin treatment induced phosphorylation of YAPY357 and reduced YAP expression. In conclusion, melatonin has potential to induce neuronal differentiation and reduce the proliferation of DPSCs by increasing phosphorylation of YAPY357 and eliminating the activity of YAP, which indicates the active state of Hippo signaling pathway.

Effect of cryopreservation on biological and immunological properties of stem cells from apical papilla.

Stem cells from apical papilla (SCAP) are a novel population of multipotent stem cells that, although similar to dental pulp stem cells, are a discrete source of dental stem cells. SCAP have potential roles in root development, apexogenesis, pulp/dentin regeneration, and bioroot engineering. However, procedures to store and preserve SCAP for future clinical applications have not been explored. In this study, we compared human freshly isolated SCAP (fSCAP) with cryopreserved SCAP (cSCAP) in terms of cell viability, colony-forming efficiency, cell proliferation rate, multilineage differentiation potential, profiles of mesenchymal stem cell (MSC) markers, karyotype analysis, and immunological assays. cSCAP showed a similar viable cell ratio, colony-forming efficiency, cell proliferation rate, multilineage differentiation potential, MSC surface markers, apoptotic rate, and G-banded karyotype when compared to fSCAP. There was no significant difference between fSCAP and cSCAP with regard to immune properties. In addition, cSCAP of miniature pig possessed the similar proliferation rate, differentiation potential, and immunomodulatory function as seen in fSCAP. This study demonstrates that cryopreservation does not affect the biological and immunological properties of SCAP, supporting the feasibility of SCAP cryopreservation in nitrogen.

In vitro osteoclastogenesis on textile chitosan scaffold.

Textile chitosan fibre scaffolds were evaluated in terms of interaction with osteoclast-like cells, derived from human primary monocytes. Part of the scaffolds was further modified by coating with fibrillar collagen type I in order to make the surface biocompatible. Monocytes were cultured directly on the scaffolds in the presence of macrophage colony stimulating factor (M-CSF) and receptor activator of nuclear factor kappaB ligand (RANKL) for up to 18 days. Confocal laser scanning microscopy (CLSM) as well as scanning electron microscopy (SEM) revealed the formation of multinuclear osteoclast-like cells on both the raw chitosan fibres and the collagen-coated scaffolds. The modified surface supported the osteoclastogenesis. Differentiation towards the osteoclastic lineage was confirmed by the microscopic detection of cathepsin K, tartrate resistant acid phosphatase (TRAP), acidic compartments using 3-(2,4-dinitroanillino)-3'-amino-N-methyldipropylamine (DAMP), immunological detection of TRAP isoform 5b, and analysis of gene expression of the osteoclastic markers TRAP, cathepsin K, vitronectin receptor, and calcitonin receptor using reverse transcription-polymerase chain reaction (RT-PCR). The feature of the collagen-coated but also of the raw chitosan fibre scaffolds to support attachment and differentiation of human monocytes facilitates cell-induced material resorption--one main requirement for successful bone tissue engineering.

TGF-beta mediated Dlx5 signaling plays a crucial role in osteo-chondroprogenitor cell lineage determination during mandible development.

Transforming growth factor-beta (TGF-beta) signaling is crucial for mandible development. During its development, the majority of the mandible is formed through intramembranous ossification whereas the proximal region of the mandible undergoes endochondral ossification. Our previous work has shown that TGF-beta signaling is required for the proliferation of cranial neural crest (CNC)-derived ectomesenchyme in the mandibular primordium where intramembranous ossification takes place. Here we show that conditional inactivation of Tgfbr2 in CNC cells results in accelerated osteoprogenitor differentiation and perturbed chondrogenesis in the proximal region of the mandible. Specifically, the appearance of chondrocytes in Tgfbr2(fl/fl);Wnt1-Cre mice is delayed and they are smaller in size in the condylar process and completely missing in the angular process. TGF-beta signaling controls Sox9 expression in the proximal region, because Sox9 expression is delayed in condylar processes and missing in angular process in Tgfbr2(fl/fl);Wnt1-Cre mice. Moreover, exogenous TGF-beta can induce Sox9 expression in the mandibular arch. In the angular processes of Tgfbr2(fl/fl);Wnt1-Cre mice, osteoblast differentiation is accelerated and Dlx5 expression is elevated. Significantly, deletion of Dlx5 in Tgfbr2(fl/fl);Wnt1-Cre mice results in the rescue of cartilage formation in the angular processes. Finally, TGF-beta signaling-mediated Scleraxis expression is required for tendonogenesis in the developing skeletal muscle. Thus, CNC-derived cells in the proximal region of mandible have a cell intrinsic requirement for TGF-beta signaling.

Onset of keratin 17 expression coincides with the definition of major epithelial lineages during skin development.

The type I keratin 17 (K17) shows a peculiar localization in human epithelial appendages including hair follicles, which undergo a growth cycle throughout adult life. Additionally K17 is induced, along with K6 and K16, early after acute injury to human skin. To gain further insights into its potential function(s), we cloned the mouse K17 gene and investigated its expression during skin development. Synthesis of K17 protein first occurs in a subset of epithelial cells within the single-layered, undifferentiated ectoderm of embryonic day 10.5 mouse fetuses. In the ensuing 48 h, K17-expressing cells give rise to placodes, the precursors of ectoderm-derived appendages (hair, glands, and tooth), and to periderm. During early development, there is a spatial correspondence in the distribution of K17 and that of lymphoid-enhancer factor (lef-1), a DNA-bending protein involved in inductive epithelial-mesenchymal interactions. We demonstrate that ectopic lef-1 expression induces K17 protein in the skin of adult transgenic mice. The pattern of K17 gene expression during development has direct implications for the morphogenesis of skin epithelia, and points to the existence of a molecular relationship between development and wound repair.

The fate of cranial neural crest cells in the Australian lungfish, Neoceratodus forsteri.

The cranial neural crest has been shown to give rise to a diversity of cells and tissues, including cartilage, bone and connective tissue, in a variety of tetrapods and in the zebrafish. It has been claimed, however, that in the Australian lungfish these tissues are not derived from the cranial neural crest, and even that no migrating cranial neural crest cells exist in this species. We have earlier documented that cranial neural crest cells do migrate, although they emerge late, in the Australian lungfish. Here, we have used the lipophilic fluorescent dye, DiI, to label premigratory cranial neural crest cells and follow their fate until stage 43, when several cranial skeletal elements have started to differentiate. The timing and extent of their migration was investigated, and formation of mandibular, hyoid and branchial streams documented. Cranial neural crest was shown to contribute cells to several parts of the head skeleton, including the trabecula cranii and derivatives of the mandibular arch (e.g., Meckel's cartilage, quadrate), the hyoid arch (e.g., the ceratohyal) and the branchial arches (ceratobranchials I-IV), as well as to the connective tissue surrounding the myofibers in cranial muscles. We conclude that cranial neural crest migration and fate in the Australian lungfish follow the stereotyped pattern documented in other vertebrates.

Structure and development of the subesophageal zone of the Drosophila brain. I. Segmental architecture, compartmentalization, and lineage anatomy.

The subesophageal zone (SEZ) of the Drosophila brain houses the circuitry underlying feeding behavior and is involved in many other aspects of sensory processing and locomotor control. Formed by the merging of four neuromeres, the internal architecture of the SEZ can be best understood by identifying segmentally reiterated landmarks emerging in the embryo and larva, and following the gradual changes by which these landmarks become integrated into the mature SEZ during metamorphosis. In previous works, the system of longitudinal fibers (connectives) and transverse axons (commissures) has been used as a scaffold that provides internal landmarks for the neuromeres of the larval ventral nerve cord. We have extended the analysis of this scaffold to the SEZ and, in addition, reconstructed the tracts formed by lineages and nerves in relationship to the connectives and commissures. As a result, we establish reliable criteria that define boundaries between the four neuromeres (tritocerebrum, mandibular neuromere, maxillary neuromere, labial neuromere) of the SEZ at all stages of development. Fascicles and lineage tracts also demarcate seven columnar neuropil domains (ventromedial, ventro-lateral, centromedial, central, centrolateral, dorsomedial, dorsolateral) identifiable throughout development. These anatomical subdivisions, presented in the form of an atlas including confocal sections and 3D digital models for the larval, pupal and adult stage, allowed us to describe the morphogenetic changes shaping the adult SEZ. Finally, we mapped MARCM-labeled clones of all secondary lineages of the SEZ to the newly established neuropil subdivisions. Our work will facilitate future studies of function and comparative anatomy of the SEZ.

Adenylate Cyclase Type III Is Not a Ubiquitous Marker for All Primary Cilia during Development.

Adenylate cyclase type III (AC3) is localized in plasma membrane of neuronal primary cilium and can be used as a marker of this cilium. AC3 has also been detected in some other primary cilia such as those of fibroblasts, synoviocytes or astrocytes. Despite the presence of a cilium in almost all cell types, we show that AC3 is not a common marker of all primary cilia of different human and mouse tissues during development. In peripheral organs, AC3 is present mainly in primary cilia in cells of the mesenchymal lineage (fibroblasts, chondroblasts, osteoblasts-osteocytes, odontoblasts, muscle cells and endothelial cells). In epithelia, the apical cilium of renal and pancreatic tubules and of ductal plate in liver is AC3-negative whereas the cilium of basal cells of stratified epithelia is AC3-positive. Using fibroblasts cell culture, we show that AC3 appears at the plasma membrane of the primary cilium as soon as this organelle develops. The functional significance of AC3 localization at the cilium membrane in some cells but not others has to be investigated in relationship with cell physiology and expression at the cilium plasma membrane of specific upstream receptors.

Recent progress in stem cell differentiation directed by material and mechanical cues.

Stem cells play essential roles in tissue regeneration in vivo via specific lineage differentiation induced by environmental factors. In the past, biochemical signals were the focus of induced stem cell differentiation. As reported by Engler et al (2006 Cell 126 677-89), biophysical signal mediated stem cell differentiation could also serve as an important inducer. With the advancement of material science, it becomes a possible strategy to generate active biophysical signals for directing stem cell fate through specially designed material microstructures. In the past five years, significant progress has been made in this field, and these designed biophysical signals include material elasticity/rigidity, micropatterned structure, extracellular matrix (ECM) coated materials, material transmitted extracellular mechanical force etc. A large number of investigations involved material directed differentiation of mesenchymal stem cells, neural stem/progenitor cells, adipose derived stem cells, hematopoietic stem/progenitor cells, embryonic stem cells and other cells. Hydrogel based materials were commonly used to create varied mechanical properties via modifying the ratio of different components, crosslinking levels, matrix concentration and conjugation with other components. Among them, polyacrylamide (PAM) and polydimethylsiloxane (PDMS) hydrogels remained the major types of material. Specially designed micropatterning was not only able to create a unique topographical surface to control cell shape, alignment, cell-cell and cell-matrix contact for basic stem cell biology study, but also could be integrated with 3D bioprinting to generate micropattered 3D structure and thus to induce stem cell based tissue regeneration. ECM coating on a specific topographical structure was capable of inducing even more specific and potent stem cell differentiation along with soluble factors and mechanical force. The article overviews the progress of the past five years in this particular field.

Periodontal Ligament and Alveolar Bone in Health and Adaptation: Tooth Movement.

The periodontal ligament (PDL) and alveolar bone are two critical tissues for understanding orthodontic tooth movement. The current literature is replete with descriptive studies of multiple cell types and their matrices in the PDL and alveolar bone, but is deficient with how stem/progenitor cells differentiate into PDL and alveolar bone cells. Can one type of orthodontic force with a specific magnitude and frequency activate osteoblasts, whereas another force type activates osteoclasts? This chapter will discuss the biology of not only mature cells and their matrices in the periodontal ligament and alveolar bone, but also stem/progenitor cells that differentiate into fibroblasts, osteoblasts and osteoclasts. Key advances in tooth movement rely on further understanding of osteoblast and fibroblast differentiation from mesenchymal stem/progenitor cells, and osteoclastogenesis from the hematopoietic/monocyte lineage.

CD146 Expression Influences Periapical Cyst Mesenchymal Stem Cell Properties.

Recent studies have identified a new human dental derived progenitor cell population with multi-lineage differentiation potential referred to as human periapical cyst mesenchymal stem cells (hPCy-MSCs). In the present study, we compared two subpopulations of hPCy-MSCs characterised by the low or high expression of CD146 to establish whether this expression can regulate their stem cell properties. Using flow cytometry, we evaluated the stem cell marker profile of hPCy-MSCs during passaging. Furthermore, CD146Low and CD146High cells were sorted by magnetic beads and subsequently both cell populations were evaluated for differences in their proliferation, self-renewal, stem cell surface markers, stemness genes expression and osteogenic differentiation potential.We found that hPCy-MSCs possessed a stable expression of several mesenchymal stem cell surface markers, whereas CD146 expression declined during passaging.In addition, sorted CD146Low cells proliferated significantly faster, displayed higher colony-forming unit-fibroblast capacity and showed higher expression of Klf4 when compared to the CD146High subset. Significantly, the osteogenic potential of hPCy-MSCs was greater in the CD146Low than in CD146High population. These results demonstrate that CD146 is spontaneously downregulated with passaging at both mRNA and protein levels and that the high expression of CD146 reduces the proliferative, self-renewal and osteogenic differentiation potential of hPCy-MSCs. In conclusion, our study demonstrates that changes in the expression of CD146 can influence the stem cell properties of hPCy-MSCs.