Etiological Mechanisms and Genetic/Biological Modulation Related to PTH1R in Primary Failure of Tooth Eruption.

Primary failure of eruption (PFE) is a rare disorder that is characterized by the inability of a molar tooth/teeth to erupt to the occlusal plane or to normally react to orthodontic force. This condition is related to hereditary factors and has been extensively researched over many years. However, the etiological mechanisms of pathogenesis are still not fully understood. Evidence from studies on PFE cases has shown that PFE patients may carry parathyroid hormone 1 receptor (PTH1R) gene mutations, and genetic detection can be used to diagnose PFE at an early stage. PTH1R variants can lead to altered protein structure, impaired protein function, and abnormal biological activities of the cells, which may ultimately impact the behavior of teeth, as observed in PFE. Dental follicle cells play a critical role in tooth eruption and root development and are regulated by parathyroid hormone-related peptide (PTHrP)-PTH1R signaling in their differentiation and other activities. PTHrP-PTH1R signaling also regulates the activity of osteoblasts, osteoclasts and odontoclasts during tooth development and eruption. When interference occurs in the PTHrP-PTH1R signaling pathway, the normal function of dental follicles and bone remodeling are impaired. This review provides an overview of PTH1R variants and their correlation with PFE, and highlights that a disruption of PTHrP-PTH1R signaling impairs the normal process of tooth development and eruption, thus providing insight into the underlying mechanisms related to PTH1R and its role in driving PFE.

Charting the cellular biogeography in colitis reveals fibroblast trajectories and coordinated spatial remodeling.

Gut inflammation involves contributions from immune and non-immune cells, whose interactions are shaped by the spatial organization of the healthy gut and its remodeling during inflammation. The crosstalk between fibroblasts and immune cells is an important axis in this process, but our understanding has been challenged by incomplete cell-type definition and biogeography. To address this challenge, we used multiplexed error-robust fluorescence in situ hybridization (MERFISH) to profile the expression of 940 genes in 1.35 million cells imaged across the onset and recovery from a mouse colitis model. We identified diverse cell populations, charted their spatial organization, and revealed their polarization or recruitment in inflammation. We found a staged progression of inflammation-associated tissue neighborhoods defined, in part, by multiple inflammation-associated fibroblasts, with unique expression profiles, spatial localization, cell-cell interactions, and healthy fibroblast origins. Similar signatures in ulcerative colitis suggest conserved human processes. Broadly, we provide a framework for understanding inflammation-induced remodeling in the gut and other tissues.

Hedgehog activation promotes osteogenic fates of growth plate resting zone chondrocytes through transient clonal competency.

The resting zone of the postnatal growth plate is organized by slow-cycling chondrocytes expressing parathyroid hormone-related protein (PTHrP), which include a subgroup of skeletal stem cells that contribute to the formation of columnar chondrocytes. The PTHrP-Indian hedgehog feedback regulation is essential for sustaining growth plate activities; however, molecular mechanisms regulating cell fates of PTHrP+ resting chondrocytes and their eventual transformation into osteoblasts remain largely undefined. Here, in a mouse model, we specifically activated Hedgehog signaling in PTHrP+ resting chondrocytes and traced the fate of their descendants using a tamoxifen-inducible Pthrp-creER line with patched-1-floxed and tdTomato reporter alleles. Hedgehog-activated PTHrP+ chondrocytes formed large, concentric, clonally expanded cell populations within the resting zone ("patched roses") and generated significantly wider columns of chondrocytes, resulting in hyperplasia of the growth plate. Interestingly, Hedgehog-activated PTHrP+ cell descendants migrated away from the growth plate and transformed into trabecular osteoblasts in the diaphyseal marrow space in the long term. Therefore, Hedgehog activation drives resting zone chondrocytes into transit-amplifying states as proliferating chondrocytes and eventually converts these cells into osteoblasts, unraveling a potentially novel Hedgehog-mediated mechanism that facilitates osteogenic cell fates of PTHrP+ skeletal stem cells.

Endosteal stem cells at the bone-blood interface: A double-edged sword for rapid bone formation: Bone marrow endosteal stem cells provide a robust source of bone-making osteoblasts both in normal and abnormal bone formation.

Endosteal stem cells are a subclass of bone marrow skeletal stem cell populations that are particularly important for rapid bone formation occurring in growth and regeneration. These stem cells are strategically located near the bone surface in a specialized microenvironment of the endosteal niche. These stem cells are abundant in young stages but eventually depleted and replaced by other stem cell types residing in a non-endosteal perisinusoidal niche. Single-cell molecular profiling and in vivo cell lineage analyses play key roles in discovering endosteal stem cells. Importantly, endosteal stem cells can transform into bone tumor-making cells when deleterious mutations occur in tumor suppressor genes. The emerging hypothesis is that osteoblast-chondrocyte transitional identities confer a special subset of endosteal stromal cells with stem cell-like properties, which may make them susceptible for tumorigenic transformation. Endosteal stem cells are likely to represent an important therapeutic target of bone diseases caused by aberrant bone formation.

Mechanisms of Osteoclastogenesis in Orthodontic Tooth Movement and Orthodontically Induced Tooth Root Resorption.

Orthodontic tooth movement (OTM) is achieved by the simultaneous activation of bone resorption by osteoclasts and bone formation by osteoblasts. When orthodontic forces are applied, osteoclast-mediated bone resorption occurs in the alveolar bone on the compression side, creating space for tooth movement. Therefore, controlling osteoclastogenesis is the fundamental tenet of orthodontic treatment. Orthodontic forces are sensed by osteoblast lineage cells such as periodontal ligament (PDL) cells and osteocytes. Of several cytokines produced by these cells, the most important cytokine promoting osteoclastogenesis is the receptor activator of nuclear factor-κB ligand (RANKL), which is mainly supplied by osteoblasts. Additionally, osteocytes embedded within the bone matrix, T lymphocytes in inflammatory conditions, and PDL cells produce RANKL. Besides RANKL, inflammatory cytokines, such as interleukin-1, tumor necrosis factor-α, and prostaglandin E2 promote osteoclastogenesis under OTM. On the downside, excessive osteoclastogenesis activation triggers orthodontically-induced external root resorption (ERR) through pro-osteoclastic inflammatory cytokines. Therefore, understanding the mechanisms of osteoclastogenesis during OTM is essential in reducing the adverse effects of orthodontic treatment. Here, we review the current concepts of the mechanisms underlying osteoclastogenesis in OTM and orthodontically induced ERR.

Multi-omics analysis in developmental bone biology.

Single-cell omics and multi-omics have revolutionized our understanding of molecular and cellular biological processes at a single-cell level. In bone biology, the combination of single-cell RNA-sequencing analyses and in vivo lineage-tracing approaches has successfully identified multi-cellular diversity and dynamics of skeletal cells. This established a new concept that bone growth and regeneration are regulated by concerted actions of multiple types of skeletal stem cells, which reside in spatiotemporally distinct niches. One important subtype is endosteal stem cells that are particularly abundant in young bone marrow. The discovery of this new skeletal stem cell type has been facilitated by single-cell multi-omics, which simultaneously measures gene expression and chromatin accessibility. Using single-cell omics, it is now possible to computationally predict the immediate future state of individual cells and their differentiation potential. In vivo validation using histological approaches is the key to interpret the computational prediction. The emerging spatial omics, such as spatial transcriptomics and epigenomics, have major advantage in retaining the location of individual cells within highly complex tissue architecture. Spatial omics can be integrated with other omics to further obtain in-depth insights. Single-cell multi-omics are now becoming an essential tool to unravel intricate multicellular dynamics and intercellular interactions of skeletal cells.

Growth plate resting zone chondrocytes acquire transient clonal competency upon Hedgehog activation and efficiently transform into trabecular bone osteoblasts.

The resting zone of the postnatal growth plate is organized by slow-cycling chondrocytes expressing parathyroid hormone-related protein (PTHrP), which include a subgroup of skeletal stem cells that contribute to the formation of columnar chondrocytes. The PTHrP-indian hedgehog (Ihh) feedback regulation is essential for sustaining growth plate activities; however, molecular mechanisms regulating cell fates of PTHrP + resting chondrocytes and their eventual transformation into osteoblasts remain largely undefined. Here, in a mouse model, we utilized a tamoxifen-inducible PTHrP-creER line with Patched-1 ( Ptch1 ) floxed and tdTomato reporter alleles to specifically activate Hedgehog signaling in PTHrP + resting chondrocytes and trace the fate of their descendants. Hedgehog-activated PTHrP + chondrocytes formed large concentric clonally expanded cell populations within the resting zone (' patched roses ') and generated significantly wider columns of chondrocytes, resulting in hyperplasia of the growth plate. Interestingly, Hedgehog-activated PTHrP + cell-descendants migrated away from the growth plate and eventually transformed into trabecular osteoblasts in the diaphyseal marrow space in the long term. Therefore, Hedgehog activation drives resting zone chondrocytes into transit-amplifying states as proliferating chondrocytes and eventually converts these cells into osteoblasts, unraveling a novel Hedgehog-mediated mechanism that facilitates osteogenic cell fates of PTHrP + skeletal stem cells.

Hes1 marks peri-condensation mesenchymal cells that generate both chondrocytes and perichondrial cells in early bone development.

Bone development starts with condensations of undifferentiated mesenchymal cells that set a framework for future bones within the primordium. In the endochondral pathway, mesenchymal cells inside the condensation differentiate into chondrocytes and perichondrial cells in a SOX9-dependent mechanism. However, the identity of mesenchymal cells outside the condensation and how they participate in developing bones remain undefined. Here we show that mesenchymal cells surrounding the condensation contribute to both cartilage and perichondrium, robustly generating chondrocytes, osteoblasts, and marrow stromal cells in developing bones. Single-cell RNA-seq analysis of Prrx1-cre-marked limb bud mesenchymal cells at E11.5 reveals that Notch effector Hes1 is expressed in a mutually exclusive manner with Sox9 that is expressed in pre-cartilaginous condensations. Analysis of a Notch signaling reporter CBF1:H2B-Venus reveals that peri-condensation mesenchymal cells are active for Notch signaling. In vivo lineage-tracing analysis using Hes1-creER identifies that Hes1+ early mesenchymal cells surrounding the SOX9+ condensation at E10.5 contribute to both cartilage and perichondrium at E13.5, subsequently becoming growth plate chondrocytes, osteoblasts of trabecular and cortical bones, and marrow stromal cells in postnatal bones. In contrast, Hes1+ cells in the perichondrium at E12.5 or E14.5 do not generate chondrocytes within cartilage, contributing to osteoblasts and marrow stromal cells only through the perichondrial route. Therefore, Hes1+ peri-condensation mesenchymal cells give rise to cells of the skeletal lineage through cartilage-dependent and independent pathways, supporting the theory that early mesenchymal cells outside the condensation also play important roles in early bone development.

Bone marrow endosteal stem cells dictate active osteogenesis and aggressive tumorigenesis.

The bone marrow contains various populations of skeletal stem cells (SSCs) in the stromal compartment, which are important regulators of bone formation. It is well-described that leptin receptor (LepR)+ perivascular stromal cells provide a major source of bone-forming osteoblasts in adult and aged bone marrow. However, the identity of SSCs in young bone marrow and how they coordinate active bone formation remains unclear. Here we show that bone marrow endosteal SSCs are defined by fibroblast growth factor receptor 3 (Fgfr3) and osteoblast-chondrocyte transitional (OCT) identities with some characteristics of bone osteoblasts and chondrocytes. These Fgfr3-creER-marked endosteal stromal cells contribute to a stem cell fraction in young stages, which is later replaced by Lepr-cre-marked stromal cells in adult stages. Further, Fgfr3+ endosteal stromal cells give rise to aggressive osteosarcoma-like lesions upon loss of p53 tumor suppressor through unregulated self-renewal and aberrant osteogenic fates. Therefore, Fgfr3+ endosteal SSCs are abundant in young bone marrow and provide a robust source of osteoblasts, contributing to both normal and aberrant osteogenesis.

Charting the cellular biogeography in colitis reveals fibroblast trajectories and coordinated spatial remodeling.

Gut inflammation involves contributions from immune and non-immune cells, whose interactions are shaped by the spatial organization of the healthy gut and its remodeling during inflammation. The crosstalk between fibroblasts and immune cells is an important axis in this process, but our understanding has been challenged by incomplete cell-type definition and biogeography. To address this challenge, we used MERFISH to profile the expression of 940 genes in 1.35 million cells imaged across the onset and recovery from a mouse colitis model. We identified diverse cell populations; charted their spatial organization; and revealed their polarization or recruitment in inflammation. We found a staged progression of inflammation-associated tissue neighborhoods defined, in part, by multiple inflammation-associated fibroblasts, with unique expression profiles, spatial localization, cell-cell interactions, and healthy fibroblast origins. Similar signatures in ulcerative colitis suggest conserved human processes. Broadly, we provide a framework for understanding inflammation-induced remodeling in the gut and other tissues.

Effects of antiresorptive medications on tooth root formation and tooth eruption in paediatric patients.

Tooth eruption is a pivotal milestone for children's growth and development. This process involves with the formation of the tooth root, the periodontal ligament (PDL) and the alveolar bone, as the tooth crown penetrates the bone and gingiva to enter the oral cavity. This review aims to outline current knowledge of the adverse dental effects of antiresorptive medications. Recently, paediatric indications for antiresorptive medications, such as bisphosphonates (BPs), have emerged, and these agents are increasingly used in children and adolescents to cure pathological bone resorption associated with bone diseases and cancers. Since tooth eruption is accompanied by osteoclastic bone resorption, it is expected that the administration of antiresorptive medications during this period affects tooth development. Indeed, several articles studying human patient cohorts and animal models report the dental defects associated with the use of these antiresorptive medications. This review shows the summary of the possible factors related to tooth eruption and introduces the future research direction to understand the mechanisms underlying the dental defects caused by antiresorptive medications.

Control of craniofacial development by the collagen receptor, discoidin domain receptor 2.

Development of the craniofacial skeleton requires interactions between progenitor cells and the collagen-rich extracellular matrix (ECM). The mediators of these interactions are not well-defined. Mutations in the discoidin domain receptor 2 gene (DDR2), which encodes a non-integrin collagen receptor, are associated with human craniofacial abnormalities, such as midface hypoplasia and open fontanels. However, the exact role of this gene in craniofacial morphogenesis is not known. As will be shown, Ddr2-deficient mice exhibit defects in craniofacial bones including impaired calvarial growth and frontal suture formation, cranial base hypoplasia due to aberrant chondrogenesis and delayed ossification at growth plate synchondroses. These defects were associated with abnormal collagen fibril organization, chondrocyte proliferation and polarization. As established by localization and lineage-tracing studies, Ddr2 is expressed in progenitor cell-enriched craniofacial regions including sutures and synchondrosis resting zone cartilage, overlapping with GLI1 + cells, and contributing to chondrogenic and osteogenic lineages during skull growth. Tissue-specific knockouts further established the requirement for Ddr2 in GLI +skeletal progenitors and chondrocytes. These studies establish a cellular basis for regulation of craniofacial morphogenesis by this understudied collagen receptor and suggest that DDR2 is necessary for proper collagen organization, chondrocyte proliferation, and orientation.

We each have unique facial features that are key to our identities. These features are inherited, but the mechanisms are poorly understood. People with the genetic disease spondylo-meta-epiphyseal dysplasia, or SMED, have characteristic facial and skull abnormalities including a flattened face and shortened skull. SMED is associated with mutations that inactivate the gene encoding a protein called discoidin domain receptor 2 (DDR2), which is a receptor for collagen. Collagen is the major structural protein in the human body, supporting the structure of cells and tissues. It also controls cell behaviors including growth, migration and differentiation, and it helps form tissues such as cartilage or bone. At least some of the effects of collagen on cells depend on its interaction with DDR2. Since the facial and skull abnormalities in mice with mutations that stop DDR2 from working correctly resemble those of SMED patients, these mice can be used to understand the cellular basis for this disease, as well as the role of DDR2 in the embryonic development of the face and skull. Therefore, Mohamed et al. set out to understand how loss of DDR2 causes the characteristic facial and skull defects associated with SMED. Mohamed et al. used mice that had been genetically modified so that DDR2 could be inactivated in skeletal progenitor cells, cartilage cells and bone cells (osteoblasts). Examining these mice, they found that the shortened skulls and flat face characteristic of mice lacking DDR2 are due to bones at the skull base failing to elongate correctly due to defects in the growth centers that depend on cartilage. Mohamed et al. also discovered that the cells that normally produce DDR2 are the progenitors of cartilage and bone-forming cells, which partly explains why lacking this protein leads to issues in growth of these tissues. In addition to shedding light on the causes of SMED, Mohamed et al.'s results also provide general insights into the mechanisms controlling the formation of facial and skull bones that depend on interactions between cells and collagen. This information may help explain how other abnormalities in the face and skull emerge, and provide a basis for how the shape of the skull has changed during human evolution. In the future, it may be possible to manipulate the activity of DDR2 to correct skull defects.

The mechanism of bone repair: Stem cells in the periosteum dedicated to bridging a large gap.

Bone repair requires the mobilization of stem cells from outer periosteum and inner bone marrow. A study by Jeffery et al.1 shows that periosteal stem cells are dedicated to repairing a large defect and regenerating both bone and marrow stroma.

The fate of early perichondrial cells in developing bones.

In endochondral bone development, bone-forming osteoblasts and bone marrow stromal cells have dual origins in the fetal cartilage and its surrounding perichondrium. However, how early perichondrial cells distinctively contribute to developing bones remain unidentified. Here we show using in vivo cell-lineage analyses that Dlx5+ fetal perichondrial cells marked by Dlx5-creER do not generate cartilage but sustainably contribute to cortical bone and marrow stromal compartments in a manner complementary to fetal chondrocyte derivatives under the regulation of Hedgehog signaling. Postnatally, Dlx5+ fetal perichondrial cell derivatives preferentially populate the diaphyseal marrow stroma with a dormant adipocyte-biased state and are refractory to parathyroid hormone-induced bone anabolism. Therefore, early perichondrial cells of the fetal cartilage are destined to become an adipogenic subset of stromal cells in postnatal diaphyseal bone marrow, supporting the theory that the adult bone marrow stromal compartments are developmentally prescribed within the two distinct cells-of-origins of the fetal bone anlage.

Primary failure of tooth eruption: Etiology and management.

Primary failure of eruption (PFE) is a rare disorder defined as incomplete tooth eruption despite the presence of a clear eruption pathway. PFE is known to be caused by rare variants in the parathyroid hormone 1 receptor gene (PTH1R). Although several PTH1R variants have been reported, the etiology of PFE remains unclear. However, important studies that help elucidate the pathology of PFE have recently been published. The purpose of this review is to summarize current treatment options, clinical symptoms or phenotypes for diagnosis, genetic information including solid evidence in mouse disease models and disease-specific induced pluripotent stem cells, thus approaching the etiology of PFE from the perspective of the latest research.

Diverse stem cells for periodontal tissue formation and regeneration.

The periodontium is comprised of multiple units of mineralized and nonmineralized tissues including the cementum on the root surface, the alveolar bone, periodontal ligament (PDL), and the gingiva. PDL contains a variety of cell populations including mesenchymal stem/progenitor cells (MSCs) termed PDLSCs, which contribute to periodontal regeneration. Recent studies utilizing mouse genetic models shed light on the identities of these mesenchymal progenitors in their native environment, particularly regarding how they contribute to homeostasis and repair of the periodontium. The current concept is that mesenchymal progenitors in the PDL are localized to the perivascular niche. Single-cell RNA sequencing (scRNA-seq) analyses reveal heterogeneity and cell-type specific markers of cells in the periodontium, as well as their developmental relationship with precursor cells in the dental follicle. The characteristics of PDLSCs and their diversity in vivo are now beginning to be unraveled thanks to insights from mouse genetic models and scRNA-seq analyses, which aid to uncover the fundamental properties of stem cells in the human PDL. The new knowledge will be highly important for developing more effective stem cell-based regenerative therapies to repair periodontal tissues in the future.

Cranial Base Synchondrosis Lacks PTHrP-Expressing Column-Forming Chondrocytes.

The cranial base contains a special type of growth plate termed the synchondrosis, which functions as the growth center of the skull. The synchondrosis is composed of bidirectional opposite-facing layers of resting, proliferating, and hypertrophic chondrocytes, and lacks the secondary ossification center. In long bones, the resting zone of the epiphyseal growth plate houses a population of parathyroid hormone-related protein (PTHrP)-expressing chondrocytes that contribute to the formation of columnar chondrocytes. Whether PTHrP+ chondrocytes in the synchondrosis possess similar functions remains undefined. Using Pthrp-mCherry knock-in mice, we found that PTHrP+ chondrocytes predominantly occupied the lateral wedge-shaped area of the synchondrosis, unlike those in the femoral growth plate that reside in the resting zone within the epiphysis. In vivo cell-lineage analyses using a tamoxifen-inducible Pthrp-creER line revealed that PTHrP+ chondrocytes failed to establish columnar chondrocytes in the synchondrosis. Therefore, PTHrP+ chondrocytes in the synchondrosis do not possess column-forming capabilities, unlike those in the resting zone of the long bone growth plate. These findings support the importance of the secondary ossification center within the long bone epiphysis in establishing the stem cell niche for PTHrP+ chondrocytes, the absence of which may explain the lack of column-forming capabilities of PTHrP+ chondrocytes in the cranial base synchondrosis.

Cranial Base Synchondrosis: Chondrocytes at the Hub.

The cranial base is formed by endochondral ossification and functions as a driver of anteroposterior cranial elongation and overall craniofacial growth. The cranial base contains the synchondroses that are composed of opposite-facing layers of resting, proliferating and hypertrophic chondrocytes with unique developmental origins, both in the neural crest and mesoderm. In humans, premature ossification of the synchondroses causes midfacial hypoplasia, which commonly presents in patients with syndromic craniosynostoses and skeletal Class III malocclusion. Major signaling pathways and transcription factors that regulate the long bone growth plate-PTHrP-Ihh, FGF, Wnt, BMP signaling and Runx2-are also involved in the cranial base synchondrosis. Here, we provide an updated overview of the cranial base synchondrosis and the cell population within, as well as its molecular regulation, and further discuss future research opportunities to understand the unique function of this craniofacial skeletal structure.

Toward Marrow Adipocytes: Adipogenic Trajectory of the Bone Marrow Stromal Cell Lineage.

Bone marrow contains precursor cells for osteoblasts and adipocytes in the stromal compartment. Bone marrow adipose tissue (BMAT) is an important constituent of the bone marrow that is particularly abundant in adults. BMAT is composed of the proximal "regulated" BMAT containing individual adipocytes interspersed within actively hematopoietic marrow, and the distal "constitutive" BMAT containing large adipocytes in the area of low hematopoiesis. Historically, bone marrow adipocytes were regarded as one of the terminal states of skeletal stem cells, which stand at the pinnacle of the lineage and possess trilineage differentiation potential into osteoblasts, chondrocytes and adipocytes. Recent single-cell RNA-sequencing studies uncover a discrete group of preadipocyte-like cells among bone marrow stromal cells (BMSCs), and recent mouse genetic lineage-tracing studies reveal that these adipocyte precursor cells possess diverse functions in homeostasis and regeneration. These adipogenic subsets of BMSCs are abundant in the central marrow space and can directly convert not only into lipid-laden adipocytes but also into skeletal stem cell-like cells and osteoblasts under regenerative conditions. It remains determined whether there are distinct adipocyte precursor cell types contributing to two types of BMATs. In this short review, we discuss the functions of the recently identified subsets of BMSCs and their trajectory toward marrow adipocytes, which is influenced by multiple modes of cell-autonomous and non-cell autonomous regulations.