The aim of the research presented in this thesis is to test the applicability of some of the techniques of dental anthropology to begin to provide answers to certain questions facing British archaeology. The question directly confronted in this thesis is how the change in fifth century Britain, manifested by a change in cultural material from archaeological sites, came to pass. The transition from the Romano-British period to the Anglo-Saxon period in the country now known as England is often assumed to have occurred as a result of invasions from people known as Angles, Saxons and Jutes. A common belief is that these Continental invaders wiped out the local population. The resultant replacement of the earlier culture with a 'Germanic' culture is due to these invasions. The competing hypothesis is that of biological continuity with cultural replacement. Either of these hypotheses can be supported when one examines only cultural aspects of the populations. Pottery, clothing, building and burial styles, as well as the history of the English language, have all been used to support versions of both hypotheses. It is at least theoretically possible for all of these cultural trappings to change without any biological contribution from an outside source. To ascertain which hypothesis more accurately describes the events of the fifth century in Britain, one must first know how the populations from the later period are related to those from the earlier period. To do that, one must assess the biological profiles of each population and compare them. The remains of a total of 799 people from seven sites dated to the Romano-British and Anglo-Saxon periods are evaluated using the Arizona State University Dental Anthropology Scoring method. Six of the sites were chosen in pairs, one from the Romano-British period and one from the Anglo-Saxon period in each pair, in order to test for continuity or discontinuity across time. The site pairs were spread across Southern England to test for changes across geography. Several statistical methods are used to explore the data. The results of two different distance measures show that people buried in Anglo-Saxon sites are closely related to people buried in nearby Romano-British sites. These results clearly support the hypothesis of biological continuity in the face of cultural change.

Dental pulp progenitor cells (DPPCs) are among many stem cell sources potentially beneficial for tissue engineering. DPPCs offer advantages over other mesenchymal stem cell sources, due to their accessibility and multi-lineage differentiation. However, distinct DPPC clones exist within dental pulp, with contrasting proliferative/regenerative capabilities. This is a key consideration for the exploitation of DPPCs, in terms of the abilities of isolated clones to undergo sufficient in vitro proliferative expansion, while maintaining their regenerative potential. This Thesis supports the heterogeneous nature of DPPCs, demonstrating significant variations in population doublings (PDs) and senescence, with highly proliferative clones exhibiting greater proliferation (>80PDs) under normal and oxidative stress (H2O2 treatment) conditions, compared to low proliferative clones (<40PDs) demonstrating altered morphology, increased SA-β-galactosidase staining and senescence marker (p53/p16) expression. Although negative for human telomerase expression, highly proliferative clones possessed longer telomeres (>18kb); maintained stem cell marker expression (CD73, CD90, CD105) and osteogenic/chondrogenic differentiation in culture. In contrast, low proliferative clones exhibited shorter telomeres (<6kb), reduced marker expression and increased adipogenesis. DPPC heterogeneity was further evident upon Raman Spectroscopy analysis, which distinguished between undifferentiated high and low proliferative clones and undifferentiated DPPCs and clones following chondrogenic differentiation. High and low proliferative clonal behaviour was also assessed in type I collagen gels, demonstrating increased contraction and reduced cell proliferation in detached versus attached gels, irrespective of clonal type. Increased contraction was due to increased MMP-2 expression by both clonal types, while highly proliferative clones also expressed MMP-9, especially at late PDs. These findings indicate significant variations in DPPC clonal proliferative/differentiation capabilities, partly explained by telomere length/cellular ageing differences. Identification of clonal populations with contrasting tissue regeneration capacities supports the development of screening strategies (e.g. Raman Spectroscopy) for the selective isolation of highly proliferative clones from dental pulp for therapeutic use, or assessing DPPC differentiation in 3D culture.

Objectives: The primary objective was to assess if a correlation exists between CVM and statural height growth velocity. The secondary objective was to assess if a correlation exists between CVM and mandibular growth velocity. Design/Setting: A prospective longitudinal study undertaken at Liverpool University. Subjects: Participants were aged between 8-18 years, of either gender and enrolled from the orthodontic waiting list at Liverpool University Dental Hospital. Methods: Standing height was measured every 6 weeks with subjects barefoot and in natural head position. Lateral cephalograms were taken at the start of treatment, on completing functional appliance therapy and prior to debond. Mandibular growth was assessed using the area of the triangle condylion-gnathion-gonion. Intra- and inter-observer reliability of CVM staging, cephalometric and statural height measurements were assessed using Cohen’s weighted kappa, intra-class correlation coefficient, and Bland and Altman plots respectively. ANOVA was used to test for statistically significant differences between the CVM stages. Results: 108 participants were included for analysis. The peak in statural height growth velocity occurred at CVM stage 3 (P=0.001). The peak in mandibular growth occurred at CVM stage 3, although this was not statistically significant. Conclusions: The findings of this study demonstrate that CVM staging is valid for identifying the pubertal peak in statural height. The peak in mandibular growth as assessed by the triangle Co-Go-Gn occurred at CVM Stage 3, but this was not statistically significant. Ethical approval was granted from Liverpool East Research Ethics Committee on 30th October 2013 with reference number 13/NW/0408 and protocol number UoL000751.

Titanium (Ti) dental implants are a successful treatment modality to replace missing teeth. Success is traditionally defined as the retention of the Ti dental implant but fails to account for peri-implant inflammatory diseases such as peri-implant mucositis and peri-implantitis. Peri-implant diseases are caused by the formation of pathogenic bacteria biofilms on the implant surface and disease progression can lead to dysfunctional and unaesthetic outcomes. There is no universally accepted treatment or management protocol for peri-implant disease. The objectives were to develop methods to prevent bacterial adhesion to Ti implant surfaces or treat existing biofilms. The relationship between bacterial adhesion of common early coloniser bacteria and topological features on dental implant surfaces was studied. Reproducible model systems were identified to be used in studies of biofilm formation and disruption. Early bacterial adhesion was investigated on engineered Ti surfaces created using Scanning-Laser-Melting or on Ti nanotubule surfaces. Photoactivation of Ti oxide films was investigated on thermally or anodically oxidised Ti and demonstrated the potential to pre-treat implant surfaces to reduce bacterial attachment. Finally chemical disinfection of Ti surfaces with a novel Eucalyptus Oil (EO) based formulation was demonstrated to increase the permeation of bactericidal agents into immature biofilms formed on Ti surfaces.

Tooth development is complex and dependent on epithelial-mesenchymal interactions involving key molecular signalling pathways. Preliminary data indicate that the pleiotropic growth factor adrenomedullin (ADM) is expressed during tooth development. Furthermore, in osteoblasts, cells which share structural and functional similarities to odontoblasts, ADM increases proliferation in vitro and can promote mineralised bone volume and strength in vivo. Immunohistochemical analysis of ADM demonstrated expression during key stages in tooth development in particular in cells responsible for signalling odontoblast differentiation and subsequently in secretory odontoblasts. Similarities with the temporo-spatial expression profile of TGF-β1 were also observed. In vitro analysis using the developmentally derived dental cell lines, MDPC-23 and OD-21, demonstrated ADM stimulated a biphasic response in dental cell numbers with peak stimulation at 10-11M and that it stimulated mineral deposition at levels comparable to that of the known mineralising agent dexamethasone. Analysis of tooth tissue volume and key mandibular measurements in Swiss mice systemically treated with ADM using techniques including micro-Computer Tomography did not identify significant differences in craniofacial mineralised tissue structures compared to sham treated controls. The data presented here along with the known pleiotropic properties of ADM indicate it may be an important regulator of tooth development particularly in the processes of cell proliferation, differentiation and mineralisation. However, in adult animals systemic ADM supplementation appears to have limited affect on mandibular bone and dentine synthesis.

This research aimed to investigate the molecular regulation of odontoblast behaviour during primary, secondary and tertiary (reactionary and reparative) dentinogenesis. Using gene microarray analysis and sq-RT-PCR, evidence is presented that the changes in secretory activity of odontoblasts reflect differential transcriptional control and that common regulatory processes may exist between dentine and bone. The p38 gene was shown to be highly expressed in odontoblasts during active primary dentinogenesis, but was drastically down-regulated as cells become quiescent in secondary dentinogenesis. Based on the hypothesis that parallels between development and healing processes in teeth exist, the results suggested that the p38- MAPKinase pathway may be activated during odontoblast stimulation in tertiary dentinogenesis by both p38 phosphorylation and enhanced nuclear translocation, supporting a recapitulation of events from primary dentinogenesis during tertiary dentinogenesis. The feasibility of use of the mouse as an in vivo model for studying pulpal healing in response to restorative procedures was also assessed. This approach provides a novel opportunity to exploit use of genetically modified animals to explore cellular and molecular processes during reparative events. Lastly, a transgenic mouse model was used to analyse the possible role of Msx2 transcription factor in odontoblast differentiation. The nature of the tooth phenotype in Msx2 null mutant animals was subsequently analysed. This study has increased our understanding of the regulation of dentinogenic events, which may allow translation into new therapies aimed at maintenance of the vitality of the pulp. L’objectif de ce travail a été d’explorer les régulations moléculaires au sein de l’odontoblaste au cours des dentinogénèses primaire, secondaire et tertiaire. Les résultats obtenus à partir de micro-puces et de PCR semi quantitative ont permis de mettre en évidence un contrôle différentiel transcriptionnel de l’activité sécrétrice de l’odontoblaste. Des processus de régulation commun entre la dentine et l’os sont également discutés. La forte expression du gène p38 au cours de la dentinogénèse primaire, est drastiquement diminuée dans l’odontoblaste mature. Les résultats suggèrent également que la voie de signalisation p38-MAPKinase pourrait être activée pendant la dentinogénèse tertiaire, par la phosphorylation de la protéine p38 et sa translocation nucléaire, confirmant ainsi la récapitulation d’un processus du développement initiale de la dent dans celui de la cicatrisation pulpaire. L’utilisation de la souris comme nouveau modèle de laboratoire pour étudier la cicatrisation pulpaire est également décrite. Ce nouveau modèle constitue une opportunité réelle car il permet d’utiliser les animaux génétiquement modifiés pour explorer les processus cellulaires et moléculaires impliqués dans la réparation pulpaire. Enfin, un modèle de souris transgénique a été utilisé afin d’analyser l’éventuel rôle du facteur de transcription Msx2 dans le processus de différentiation odontoblastique. Le phénotype dentaire du mutant nul Msx2 -/- est analysé en détail. Ce travail permet de compléter les connaissances sur la régulation moléculaire de la dentinogénèse, étape importante pour le développement de nouvelles thérapeutiques en terme de conservation de la vitalité pulpaire.

Glial cell line-derived neurotrophic factor (GDNF) is a growth factor promoting survival, proliferation and differentiation of neural crest cells. Neural crest cells play an important role within mesenchymal tissues during dental pulp and calvarial bone development. GDNF also has a role within non-neuronal tissues and is expressed during dental development. GDNF null mutations prevent the formation of the mineralised hard tissues of the tooth. The hypothesis for this study was that GDNF affects mesenchymal dental pulp cells (DPC), promoting the regenerative responses of mineralised tissues. This study utilised cell culture models to investigate the direct effects of GDNF on the proliferation and differentiation of dental pulp cells, bone marrow mesenchymal stromal/stem cells (BMSC) and calvarial osteoblasts. This research demonstrated that these culture models expressed GDNF and its receptors GFR\(\alpha\)1 and RET. GDNF was shown to directly stimulate DPC and osteoblast-like cell proliferation and differentiation. Moreover, GDNF was cytoprotective when DPCs were cultured under conditions reflecting aspects of inflammation, which may occur during repair. These conditions included supplementation with the pro-inflammatory cytokine TNF\(\alpha\) and culture under serum-starved conditions. It is proposed that GDNF may play an important regulatory role in dental pulp homeostasis and bone metabolism.

This investigation was aimed at measuring the changes in dielectric properties of glass ionomer cements during their setting reaction in order to observe if there is a correlation between these properties and the cement curing. Commercial glass ionomer cements were prepared and their setting process was monitored over a 24 hour period using FT-IR and direct contact impedance measurement. An impedance bridge with a dielectric test assembly, based on previous work by Braden et al, was used to measure the dielectric properties of a number of different glass ionomer cements using a simple design. Using the dielectric properties of the glass ionomer cements, it could be possible to develop a micro-electro-mechanical sensor (MEMS) based on this design, which could be implanted into a dental restoration and interrogated remotely. During the curing of the cements examined, the dielectric data collected from the co-planar assembly showed a change in impedance over the course of the setting of the cement, which when compared to FT-IR spectra over the same period, showed a correlation between the dielectric properties and the chemical changes within the cement.

Dental pulp contains mesenchymal stem cells (MSCs) similar to those present within bone marrow. Several factors are postulated to contribute to the signalling involved in regulating these cells. This project aimed to investigate the role of pulp and dentine extracellular matrix (pECM/dECM) in the regulation of pulp cell behaviour during health and disease. pECM/dECM molecules were extracted using 10% EDTA, pH7.2 followed by 0.5M-NaCl, pH11.7 and 0.1M tartaric acid, pH2.0, respectively containing protease inhibitors. Proteomic analysis demonstrated the complexity of the ECM extractions. pECM-coated cultureware reduced pulp cell proliferation rates and increased stem cell marker expression compared with controls. Pulp cells exhibited multipotential capacity, with pECM-coated culture surfaces enhancing differentiation activity. pECM and dECM promoted pulp cell migration through an active rho dependent pathway and the chemotactic effects of these ECM molecules were enhanced following acidic/proteolytic degradation. Recruited cells exhibited increased stem cell marker expression. dECM and pECM possessed demonstrable bacteriostatic activity against three anaerobic bacteria associated with dental disease. Dental pulp cells were shown to be viable and capable of secreting mineral when encapsulated within a pECM/alginate scaffold and exposed to dECM molecules. Dental ECMs play important roles in regulating cellular and tissue responses during health and disease.

Periodontitis is a widely spread disease, affecting about 80% of the worldwide population, resulting in teeth loss, a heavy impact on patients in terms of function and aesthetic. Type 2 Diabetes Mellitus (T2DM) is described to be linked to the exacerbation of periodontitis and delayed healing. This link between these two diseases, however, is not fully evaluated and the mechanisms are yet to be fully elucidated. Osteopontin (OPN) is described to inhibit mineral crystal formation. Herein, it has been hypothesized that increased OPN in diabetic healing bone may be the causative factor of delayed healing in periodontitis and subsequent deterioration, leading to teeth loss. This project aims to gain a greater understanding of the effect of high glucose (HG) levels on mesenchymal stem cells (MSCs) and macrophages, their ability to synthesise OPN and hence, its effects on MSCs to synthesise new bone tissue. Further, the influence of Porphyromonas gingivalis lipopolysaccharides (Pg-LPS) on these cells was analyzed, in an attempt to create a model to study the healing in the presence of periodontitis and T2DM. Investigating the MSCs isolated from the rat compact bone (CB-MSCs) during the growth in culture, revealed two main populations; a heterogenous population appeared with predominantly mature characteristics at PD15. This population then demonstrated a change in its heterogeneity and became more immature in nature at PD50. These two main populations differed in their growth rate and capability of osteodifferentiation. HG environments exerted significant decreases in osteogenic differentiation on PD15, but not PD50. Addition of pg-LPS showed inhibitory effects on osteodifferentiation on PD15 cells more than PD50. Conversely, in the combined presence of HG and pg-LPS, PD50 showed a significant decrease in osteodifferentiation. OPN levels demonstrated a gradual decrease in CB-MSCs in both normal and HG conditions. Investigating OPN levels secreted by macrophages, however, revealed interesting results. Synergistic effects of both HG and pg-LPS exhibited a significant increase in OPN levels in both pro-inflammatory M1 macrophages and in repair related M2 macrophages. In conclusion, HG was mainly reported to inhibit osteogenic differentiation of the mature cell population, whereas the immature population was found to be affected by combined pg-LPS and HG. OPN levels in HG conditions were shown to decrease along the osteodifferentiation period. However, macrophages showed increase secretion of OPN by the synergistic effects of both pg-LPS and HG in both M1 and M2 and by pg-LPS effects in M2 macrophages. These outcomes as far as we are aware, are novel and disclose a new mechanism of bone resorption in the case of T2DM patients concurrently with periodontal disease.