Impaired Tertiary Dentin Secretion after Shallow Injury in Tgfbr2-Deficient Dental Pulp Cells Is Rescued by Extended CGRP Signaling.

The transforming growth factor ß (TGFß) superfamily is a master regulator of development, adult homeostasis, and wound repair. Dysregulated TGFß signaling can lead to cancer, fibrosis, and musculoskeletal malformations. We previously demonstrated that TGFß receptor 2 (Tgfbr2) signaling regulates odontoblast differentiation, dentin mineralization, root elongation, and sensory innervation during tooth development. Sensory innervation also modulates the homeostasis and repair response in adult teeth. We hypothesized that Tgfbr2 regulates the neuro-pulpal responses to dentin injury. To test this, we performed a shallow dentin injury with a timed deletion of Tgfbr2 in the dental pulp mesenchyme of mice and analyzed the levels of tertiary dentin and calcitonin gene-related peptide (CGRP) axon sprouting. Microcomputed tomography imaging and histology indicated lower dentin volume in Tgfbr2cko M1s compared to WT M1s 21 days post-injury, but the volume was comparable by day 56. Immunofluorescent imaging of peptidergic afferents demonstrated that the duration of axon sprouting was longer in injured Tgfbr2cko compared to WT M1s. Thus, CGRP+ sensory afferents may provide Tgfbr2-deficient odontoblasts with compensatory signals for healing. Harnessing these neuro-pulpal signals has the potential to guide the development of treatments for enhanced dental healing and to help patients with TGFß-related diseases.

Supporting post-pandemic recovery: a qualitative study of the capabilities, opportunities and motivations to deliver oral health behaviour change messages to parents of young children in community settings.

BACKGROUND: The COVID-19 pandemic exacerbated vulnerabilities and inequalities in children's oral health, and treatment activity virtually ceased during periods of lockdown. Primary care dentistry is still in the post-pandemic recovery phase, and it may be some years before normal service is resumed in NHS dentistry. However, opportunities to support the dental workforce through offering some preventative care in outreach settings may exist. This has the additional benefit of potentially reaching children who do not routinely see a dentist. The aim of this research was therefore to explore views around upskilling practitioners working in early years educational and care settings to support families of pre-school aged children to adopt and maintain preventative oral health behaviours. METHODS: Using the Capability, Opportunity and Motivation model of behaviour (COM-B) to structure our data collection and analysis, we conducted semi-structured interviews with 16 practitioners (dental and non-dental) and analysed the data using deductive framework analysis. RESULTS: The data were a good fit with the COM-B model, and further themes were developed within each construct, representing insights from the data. CONCLUSION: Early years practitioners can reach vulnerable children who are not usually brought to see a dentist, and have the capability, opportunity and motivation to support the oral health behaviours of families of children in their care. Further research is needed to identify training needs (oral health and behaviour change knowledge and skills), acceptability to parents, and supporting dental practice teams to work in partnership with early years settings.

Unusual Presentation of Pediatric Scurvy: A Necrotic Gastrostomy Tube Site in a 14-Year-Old Boy.

BACKGROUND Despite being considered a disease of the past, pediatric scurvy is increasingly reported in developed countries, especially among children with autism spectrum disorder, developmental delays, or a restrictive diet. Pediatric patients typically present with lower extremity pain or refusal to walk. This case study features an atypical presentation of scurvy in a non-ambulatory patient. CASE REPORT A 14-year-old boy with arthrogryposis multiplex congenita displayed a late-stage scurvy symptom: a necrotic gastrostomy tube site, indicative of poor wound healing due to vitamin C deficiency. The usual telltale symptoms of scurvy were camouflaged due to his non-ambulatory status, which may have contributed to a delayed presentation. Nevertheless, a comprehensive clinical evaluation, incorporating diet history, gingival symptoms, petechiae, and characteristic radiological signs, eventually led to the correct diagnosis. Although acute osteomyelitis was initially suspected, it was subsequently ruled out. Upon initiation of vitamin C therapy, the patient's symptoms subsided within a few days, and the necrotic tissue surrounding the gastrostomy tube healed completely within two weeks. CONCLUSIONS The highlighted case underscores the importance of including scurvy in the differential diagnosis for pediatric patients with lower extremity pain without fever. A detailed dietary history focusing on vitamin C intake is crucial during clinical evaluation. Early initiation of vitamin C therapy, when scurvy is suspected, may prevent unnecessary and extensive diagnostic workup for other potential causes, offering timely relief to the patient.

Tgfbr2 in Dental Pulp Cells Guides Neurite Outgrowth in Developing Teeth.

Transforming growth factor ß (TGFß) plays an important role in tooth morphogenesis and mineralization. During postnatal development, the dental pulp (DP) mesenchyme secretes neurotrophic factors that guide trigeminal nerve fibers into and throughout the DP. This process is tightly linked with dentin formation and mineralization. Our laboratory established a mouse model in which Tgfbr2 was conditionally deleted in DP mesenchyme using an Osterix promoter-driven Cre recombinase (Tgfbr2 cko ). These mice survived postnatally with significant defects in bones and teeth, including reduced mineralization and short roots. Hematoxylin and eosin staining revealed reduced axon-like structures in the mutant mice. Reporter imaging demonstrated that Osterix-Cre activity within the tooth was active in the DP and derivatives, but not in neuronal afferents. Immunofluorescence staining for ß3 tubulin (neuronal marker) was performed on serial cryosections from control and mutant molars on postnatal days 7 and 24 (P7, P24). Confocal imaging and pixel quantification demonstrated reduced innervation in Tgfbr2 cko first molars at both stages compared to controls, indicating that signals necessary to promote neurite outgrowth were disrupted by Tgfbr2 deletion. We performed mRNA-Sequence (RNA-Seq) and gene onotology analyses using RNA from the DP of P7 control and mutant mice to investigate the pathways involved in Tgfbr2-mediated tooth development. These analyses identified downregulation of several mineralization-related and neuronal genes in the Tgfbr2 cko DP compared to controls. Select gene expression patterns were confirmed by quantitative real-time PCR and immunofluorescence imaging. Lastly, trigeminal neurons were co-cultured atop Transwell filters overlying primary Tgfbr2 f/f DP cells. Tgfbr2 in the DP was deleted via Adenovirus-expressed Cre recombinase. Confocal imaging of axons through the filter pores showed increased axonal sprouting from neurons cultured with Tgfbr2-positive DP cells compared to neurons cultured alone. Axon sprouting was reduced when Tgfbr2 was knocked down in the DP cells. Immunofluorescence of dentin sialophosphoprotein in co-cultured DP cells confirmed reduced mineralization potential in cells with Tgfbr2 deletion. Both our proteomics and RNA-Seq analyses indicate that axonal guidance cues, particularly semaphorin signaling, were disrupted by Tgfbr2 deletion. Thus, Tgfbr2 in the DP mesenchyme appears to regulate differentiation and the cells' ability to guide neurite outgrowth during tooth mineralization and innervation.

Skeletal Deformities in Osterix-Cre;Tgfbr2f/f Mice May Cause Postnatal Death.

Transforming growth factor ß (TGFß) signaling plays an important role in skeletal development. We previously demonstrated that the loss of TGFß receptor II (Tgfbr2) in Osterix-Cre-expressing mesenchyme results in defects in bones and teeth due to reduced proliferation and differentiation in pre-osteoblasts and pre-odontoblasts. These Osterix-Cre;Tgfbr2f/f mice typically die within approximately four weeks for unknown reasons. To investigate the cause of death, we performed extensive pathological analysis on Osterix-Cre- (Cre-), Osterix-Cre+;Tgfbr2f/wt (HET), and Osterix-Cre+;Tgfbr2f/f (CKO) mice. We also crossed Osterix-Cre mice with the ROSA26mTmG reporter line to identify potential off-target Cre expression. The findings recapitulated published skeletal and tooth abnormalities and revealed previously unreported osteochondral dysplasia throughout both the appendicular and axial skeletons in the CKO mice, including the calvaria. Alterations to the nasal area and teeth suggest a potentially reduced capacity to sense and process food, while off-target Cre expression in the gastrointestinal tract may indicate an inability to absorb nutrients. Additionally, altered nasal passages and unexplained changes in diaphragmatic muscle support the possibility of hypoxia. We conclude that these mice likely died due to a combination of breathing difficulties, malnutrition, and starvation resulting primarily from skeletal deformities that decreased their ability to sense, gather, and process food.

A feasibility and acceptability study of an e-training intervention to facilitate health behaviour change conversations in dental care settings.

Introduction Health behaviours result in oral health problems. Behaviour change techniques, informed by behaviour science, are rarely utilised by dental care professionals (DCPs) within routine care.Aim To develop a theory-informed intervention to support DCPs' behaviour change conversations and evaluate its feasibility and acceptability.Intervention A behaviour change toolkit (Toothpicks) comprising 33 behaviour change techniques relevant to dentistry, delivered within an interactive online course.Design Development and mixed-methods evaluation of the intervention.Materials and methods Participants' motivation to discuss behaviour change with patients was measured before and after training using a validated questionnaire. Acceptability was assessed through semi-structured interviews.Results DCPs' (N = 32) motivation increased significantly post-training. Participants found the training acceptable and reported subsequently implementing techniques into their practice. Potential barriers preventing implementation to clinical practice include perceived lack of opportunities to effect change within the constraints of the clinical context.Conclusions Acceptable theory-informed training that is acceptable and accessible can be developed that increases DCPs' motivation to discuss behaviour change with patients. Further research is necessary to establish the longer-term impact of brief behaviour change training on DCPs' clinical practice and patient health behaviours.

A Co-Culture Method to Study Neurite Outgrowth in Response to Dental Pulp Paracrine Signals.

Tooth innervation allows teeth to sense pressure, temperature and inflammation, all of which are crucial to the use and maintenance of the tooth organ. Without sensory innervation, daily oral activities would cause irreparable damage. Despite its importance, the roles of innervation in tooth development and maintenance have been largely overlooked. Several studies have demonstrated that DP cells secrete extracellular matrix proteins and paracrine signals to attract and guide TG axons into and throughout the tooth. However, few studies have provided detailed insight into the crosstalk between the DP mesenchyme and neuronal afferents. To address this gap in knowledge, researchers have begun to utilize co-cultures and a variety of techniques to investigate these interactions. Here, we demonstrate the multiple steps involved in co-culturing primary DP cells with TG neurons dispersed on an overlying transwell filter with large diameter pores to allow axonal growth through the pores. Primary DP cells with the gene of interest flanked by loxP sites were utilized to facilitate gene deletion using an Adenovirus-Cre-GFP recombinase system. Using TG neurons from the Thy1-YFP mouse allowed for precise afferent imaging, with expression well above background levels by confocal microscopy. The DP responses can be investigated via protein or RNA collection and analysis, or alternatively, through immunofluorescent staining of DP cells plated on removable glass coverslips. Media can be analyzed using techniques such as proteomic analyses, although this will require albumin depletion due to the presence of fetal bovine serum in the media. This protocol provides a simple method that can be manipulated to study the morphological, genetic, and cytoskeletal responses of TG neurons and DP cells in response to the controlled environment of a co-culture assay.

Managing chronic orofacial pain: A qualitative study of patients', doctors', and dentists' experiences.

OBJECTIVE: Persistent pain in the face, mouth, and jaws is a common presentation to dental and medical services. The aetiology remains unclear, but a growing evidence base recognizes the effectiveness of psychological rather than biomedical interventions. To understand how this approach might be implemented into clinical practice, knowledge is needed of patients' and clinicians' experience of chronic orofacial pain (COFP). The aim of this study was to explore the experience and understanding of COFP by patients and primary and secondary care medical and dental practitioners. DESIGN: Qualitative interview study. METHODS: Audio-recorded semi-structured interviews with a purposive sample of 12 dentists, 11 general practitioners, and seven patients were thematically analysed. RESULTS: Clinicians and patients recognized the role that psychological factors could play in the development and maintenance of COFP, yet management and self-management strategies were largely limited to biomedical interventions. Achieving a diagnosis proved problematic but largely functional for both parties. GPs and dentists viewed COFP as a non-dental problem and felt inadequately equipped to manage the condition. GPs, unlike dentists, felt obligated to support patients using techniques for managing patients with other long-term conditions. Frustration at the current inadequacy of COFP management often led to conflict with (or disengagement from) the clinician-patient relationship. CONCLUSIONS: Current management of COFP is ineffective and unsatisfactory for patients and practitioners, which impacts on their relationship. Fundamental barriers to accessing and implementing psychological interventions for COFP arise from ineffective communication between physicians and patients, and between medical and dental practitioners. Statement of contribution What is already known on this subject? COFP is characterized by persistent pain in the face, mouth, or jaws that are not the result of organic disease or physical trauma. Patients with COFP present to both medical and dental services and receive sub-optimal care. No studies have examined the experiences of managing this problem from the perspectives of dentists, general practitioners and patients. What does this study add? Patients, dentists, and GPs recognize the role that psychological factors have in maintaining and addressing facial pain symptoms, yet principally manage it through biomedical interventions. Challenges exist over arriving at a diagnosis and managing the problem, and challenges are exacerbated by poor communication between doctors and medical services. Improvements are needed in liaison between medical and dental services and further training to support primary care clinicians to facilitate a stepped care approach to managing COFP.

Biocompatible tissue scaffold compliance promotes salivary gland morphogenesis and differentiation.

Substrate compliance is reported to alter cell phenotype, but little is known about the effects of compliance on cell development within the context of a complex tissue. In this study, we used 0.48 and 19.66 kPa polyacrylamide gels to test the effects of the substrate modulus on submandibular salivary gland development in culture and found a significant decrease in branching morphogenesis in explants grown on the stiff 19.66 kPa gels relative to those grown on the more physiologically compliant 0.48 kPa gels. While proliferation and apoptosis were not affected by the substrate modulus, tissue architecture and epithelial acinar cell differentiation were profoundly perturbed by aberrant, high stiffness. The glands cultured on 0.48 kPa gels were similar to developing glands in morphology and expression of the differentiation markers smooth muscle alpha-actin (SM α-actin) in developing myoepithelial cells and aquaporin 5 (AQP5) in proacinar cells. At 19.66 kPa, however, tissue morphology and the expression and distribution of SM α-actin and AQP5 were disrupted. Significantly, aberrant gland development at 19.66 kPa could be rescued by both mechanical and chemical stimuli. Transfer of glands from 19.66 to 0.48 kPa gels resulted in substantial recovery of acinar structure and differentiation, and addition of exogenous transforming growth factor beta 1 at 19.66 kPa resulted in a partial rescue of morphology and differentiation within the proacinar buds. These results indicate that environmental compliance is critical for organogenesis, and suggest that both mechanical and chemical stimuli can be exploited to promote organ development in the contexts of tissue engineering and organ regeneration.