Saudi Cultural Missions Theses & Dissertations
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Item Restricted EDTA Irrigation Protocols Influence the Liberation of Dentine Matrix Components: a Proteomics Insight(Saudi Digital Library, 2026) AlOthman, Shihanah Ali; Nicholas, Longridge; Kazuhiro, Yamamoto; Kevin, Hamill; Emad, Moawad; Fadi, JaradBackground: Vital pulp treatment and regenerative procedures aim to preserve dental pulp vitality and support tissue healing using biomaterials and endogenous stem/progenitor cells. However, clinical success is limited by an incomplete understanding of pulp-dentine biology, particularly the effects of irrigation solutions on dentine matrix proteins and dental pulp stem cells. Aim: This thesis aimed to determine optimal ethylenediamine tetra acetic acid (EDTA) irrigation conditions for releasing bioactive dentine proteins, using proteomic and protein detection techniques to assess their potential effects on dental pulp healing and regeneration. Methods: Fresh bovine incisors were sectioned at the cemento-enamel junction, the pulp tissue was removed, and the inverted crowns were mounted in tissue culture plates. The pulp chambers served as reservoirs for a series of irrigation-based experiments designed to investigate the extraction and identification of dentine extracellular matrix (ECM) components under varying conditions. Different irrigation protocols were applied using EDTA. The first experiment assessed the effect of irrigation duration, with EDTA applied for 20 minutes, 2 hours, and 24 hours to compare the quantity and profile of released dentine matrix proteins. The second experiment compared protein extraction using EDTA and sodium dodecyl sulphate gel (SDS), a laboratory detergent known for its strong protein-solubilising properties. The third experiment evaluated the release of water-soluble proteins through sequential water washing of dentine surfaces followed by EDTA irrigation on the same samples, allowing assessment of cumulative protein extraction efficiency. The extraction lysates from these three experiments were analysed using sodium dodecyl-sulphate polyacrylamide gel electrophoresis (SDS-PAGE), and in the first experiment, semi- quantification was performed using ImageJ. The final experiments aimed to identify and characterise the extracted proteins. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was employed to analyse proteins. Samples were treated with EDTA and water under different temperature conditions (4 °C, 37 °C, and 60 °C). Each condition was prepared in triplicate (n = 18), and the total irrigation time was 30 minutes for all samples. Analysis using MaxQuant identified 2,557 proteins, which were refined using Perseus filters to exclude entries identified only by site, reverse hits, or contaminants. Only proteins detected in at least two replicates were retained, yielding a total of 1,878 proteins for inclusion in this study. Western blot analysis was subsequently conducted to validate the LC–MS/MS findings and to confirm the presence of transforming growth factor beta 1 (TGF-β1) in water- and EDTA-treated dentine samples at 4 °C. Results: Increasing the duration of EDTA irrigation resulted in greater protein extraction from the dentine matrix. An irrigation period of 20 minutes was sufficient to extract detectable dentine matrix proteins; however, a 2 hours application released more proteins, and the 24 hours irrigation period produced the highest yield among all time intervals assessed. A potent agent such as SDS extracted higher amounts of protein within a shorter application time compared with EDTA irrigation. Water-soluble proteins were readily released when dentine tissue was exposed to a liquid, likely comprising a mixture of dentine matrix and pulp proteins, as the dentine-pulp complex functions as a unified biological system. Temperature also influenced protein release, with EDTA at 4 °C yielding the greatest quantity and diversity of proteins. Forty-one proteins were commonly upregulated in EDTA-treated samples compared with water at all temperatures, including TGF-β1, IGFBP5, DSPP, BGN, COL11A2, and CTHRC1. Seventeen proteins were consistently downregulated in EDTA-treated samples relative to water across all temperature conditions. Notably, at least thirteen growth factors were exclusively identified in EDTA extracts but were absent in water-treated groups. Finally, western blot analysis confirmed the presence of TGF-β1 at multiple molecular weights across all EDTA conditions and in water extracts at 4 °C. Conclusion of the thesis: EDTA irrigation effectively releases bioactive dentine matrix components, with protein yield increasing proportionally to exposure duration. While a 20 minute application extracted detectable protein levels, prolonged irrigation (up to 24 hours) notably enhanced protein recovery. Temperature also influenced extraction efficiency, with EDTA at 4 °C producing the greatest protein quantity and diversity. Proteomic analysis identified forty-one proteins consistently upregulated in EDTA-treated samples compared with water, including several growth factors such as TGF-β1, IGFBP5, DSPP, and BGN. Western blot validation confirmed the presence of TGF-β1 across all EDTA conditions. Another notable finding was that EDTA released at least thirteen growth factors that were not detected in the water-treated samples, highlighting its superior ability to extract matrix-bound bioactive molecules. These findings indicate that EDTA efficiently liberates bioactive dentine components associated with tissue repair and regeneration. Maintaining EDTA at 4 °C and applying it as a final rinse in vital pulp and regenerative endodontic procedures may provide a biologically favourable environment that supports healing and enhances treatment outcomes.9 0Item Embargo Comparative Study of the Performance of an Artificial Intelligence Platform in Detecting Periapical Radiolucencies Across Different Imaging Modalities(Saudi Digital Library, 2025) Allihaibi, Marwa; Koller, Garrit; Mannocci, FrancescoAim: This thesis aimed to evaluate the diagnostic accuracy of a commercial artificial intelligence (AI) platform in detecting periapical radiolucencies (PARLs) across different imaging modalities. The evaluation included preoperative assessment of teeth requiring primary endodontic treatment with comparison against dental professionals, radiographic healing assessment at follow-up, and assessment of teeth referred for apical microsurgery. Methods: Five retrospective diagnostic accuracy studies were conducted to evaluate the commercial AI platform Diagnocat (versions 1.0 and 2.0) for PARL detection across multiple imaging modalities. The studies utilised radiographic data from patients treated at Guy's and St Thomas' NHS Foundation Trust between 2012-2023. The study sample included: (1) 339 teeth indicated for primary root canal treatment, assessed on periapical radiographs (PARs) and compared with two experienced endodontists; (2) 376 teeth assessed at minimum one-year follow-up on PARs for radiographic healing outcomes, compared with two endodontists; (3) 134 molars evaluated on cone-beam computed tomography (CBCT) for preoperative and postoperative assessment; (4) 177 posterior teeth requiring primary endodontic treatment, assessed on PARs and compared with eleven general dental practitioners (GDPs); and (5) 116 anterior teeth referred for apical microsurgery, evaluated on both PARs and CBCT. Reference standards varied by study design: CBCT for PAR validation, expert consensus for CBCT assessment, and histopathology for cases referred for apical microsurgery. Statistical analyses included calculation of sensitivity, specificity and accuracy with 95% confidence intervals. McNemar's test assessed diagnostic performance differences. Subgroup analyses examined performance across anatomical variables. Results: Across five retrospective studies, Diagnocat demonstrated significant performance variability dependent on imaging modality, anatomical location, and treatment status. On PARs, for non-root-filled teeth requiring primary root canal treatment, sensitivity was 47.9% and specificity 95.4%, indicating reliable exclusion of disease but missing over half of actual lesions. In root-filled teeth assessed at one-year follow-up, sensitivity increased to 67.3% while specificity decreased to 82.3%, suggesting altered diagnostic thresholds based on treatment status. Performance on CBCT scans of molars showed marked improvement, achieving 93.9% sensitivity and 65.2% specificity in preoperative cases, and 88.6% sensitivity and 63.3% specificity in follow-up cases. While three-dimensional (3D) imaging substantially enhanced sensitivity for posterior teeth, it was accompanied by reduced specificity, indicating potential for overdiagnosis. Anatomical analysis revealed consistent underperformance in maxillary teeth and specific roots on PARs, limitations that were largely resolved on CBCT for posterior teeth. In contrast, anterior teeth demonstrated persistently poor performance regardless of imaging modality, achieving only 63.8% sensitivity on PARs and 57.5% on CBCT despite histopathological confirmation of periapical pathology. Cross-modality consistency was poor, with only 43.8% of lesions detected on both imaging modalities. Compared to clinicians, Diagnocat showed lower sensitivity (47.9% vs 65.3%) but comparable specificity (95.4% vs 97.7%) when assessed against endodontists in non-root-filled teeth. In root-filled teeth, this pattern reversed, with the AI achieving higher sensitivity (67.3% vs 49.3%) but lower specificity (82.3% vs 92.5%). When compared with GDPs, Diagnocat demonstrated lower sensitivity (44.9% vs 80.8%) but markedly superior specificity (94.3% vs 47.5%). Re-evaluation with version 2.0 showed no improvement in PARL detection across 1,308 PARs and 268 CBCT scans. Conclusion: This thesis demonstrated that multiple factors critically determine AI diagnostic accuracy for PARL detection, including imaging modality, anatomical location, and treatment status, thus highlighting fundamental limitations in training data representation and model development. AI platforms require comprehensive training on datasets with balanced anatomical representation and the incorporation of three-dimensional imaging before being considered for reliable implementation in endodontic diagnosis.17 0Item Open Access In vitro Antibiofilm Activity of Cold Atmospheric Plasma on Enterococcus Faecalis(Univeristy of Liverpool, 2024) ALqahtani, Omar; Jarad, Fadi; Walsh, James; Longridge, Nicholas; Horsburgh, MalApical periodontitis is an infection primarily caused by bacteria that invade the dental pulp and periapical tissues, necessitating effective management to preserve tooth function and aesthetics through endodontic procedures. E. faecalis is one of the most significant bacteria implicated in endodontic treatment failure. Effective disinfection of the root canal system is crucial for reducing the bacterial load and ensuring long-term treatment success. Traditionally, NaOCl has been the gold standard for root canal disinfection due to its broad-spectrum antimicrobial properties. However, NaOCl presents several significant drawbacks, including potential safety concerns, adverse effects on dentine mechanical properties, and limited penetration into the dentinal tubules. These limitations necessitate the exploration of alternative or adjunctive disinfection methods that can offer enhanced efficacy and safety. Atmospheric plasma jet (APJ) technology has emerged as a promising alternative in various fields due to its potent antimicrobial properties. APJ has been investigated in areas such as food safety, industrial sanitation, medical sterilisation, and more recently, dentistry. The APJ generates reactive oxygen species (ROS), which are believed to play a critical role in its antimicrobial activity. Although the precise mode of action of APJ is not yet fully understood, its broad-spectrum antimicrobial effectiveness highlights its potential as a significant area of interest for researchers aiming to enhance endodontic treatment disinfection. The primary aim of this research was to investigate the antimicrobial efficacy of APJ against E. faecalis biofilms on hydroxyapatite (HA) discs. A secondary aim was to determine whether adding NaOCl 0.5% to APJ treatment would enhance its antimicrobial efficacy, thereby testing the synergistic potential of the combined treatment. In addition, phosphate-buffered saline (PBS) was used as the liquid medium and tested in two ways to evaluate its antimicrobial efficacy. The first method involved applying APJ to PBS without the HA disc sample for a determined period. The sample was then inserted into the activated PBS without further application of APJ to observe any antimicrobial activities generated by the APJ. This group was referred to as Plasma-Activated PBS (PAPBS). The second method involved exposing the sample to continuous APJ application while it was in the PBS medium. This approach aimed to enhance the disinfection process by utilising both APJ and the ROS generated in the PBS. The PBS media treated the sample with APJ during its time in PBS, resulting in a group termed Plasma-Enhanced PBS (PEPBS). The objective of comparing these two methods was to determine which technique would result in more significant bacterial reduction, thus guiding future clinical applications. The methodology involved culturing E. faecalis biofilms on HA discs in10 BHI broth under anaerobic conditions at 37°C. Biofilms were grown for different periods: 24 hours, 1 week, and 3 weeks, to represent varying stages of biofilm maturity. Treatment groups comprised a negative control PBS, a positive control (1% NaOCl), and various combinations involving APJ alone, PEPBS, PAPBS and APJ with 0.5% NaOCl. Biofilms were exposed to the antimicrobial agents for 1, 5, and 10 min. Bacterial reduction was assessed using CFU counting. For statistical analysis, a three-way ANOVA was employed to evaluate the significance of the results. The results demonstrated that while 1% NaOCl exhibited the highest reduction rate for immature biofilms, the mature biofilm showed higher resistance. In contrast, APJ showed superior performance in killing mature biofilms. APJ alone, PEPBS, and PAPBS provided significant differences compared to the control group. Notably, the combination of APJ and 0.5% NaOCl achieved 100% eradication of E. faecalis biofilms regardless of their age, highlighting a synergistic effect that significantly enhances antimicrobial efficacy, indicating the potential of APJ to improve the performance of various irrigants.21 0Item Restricted Clinical Investigation of the Impact of Endodontic Disinfection on the Bacteriome of Root Canal Infection Using Next-Generation Sequencing on the Illumina MiSeq Platform(University of Maryland, Baltimore, 2024-07-01) Alquria, Theeb; Martinho, FredericoThe primary cause of root canal infection is bacteria and their by-products, making disinfection of the root canal system a key goal in endodontic therapy. However, the complex anatomy of root canal systems, particularly the isthmus and its ramifications, poses challenges for effective disinfection. Currently, no disinfection protocol can eliminate all bacterial contents from root canal infections, driving the ongoing search for an optimal disinfection approach. Recently, next-generation sequencing (NGS), particularly the Illumina MiSeq platform, has been widely explored in endodontic infections due to its low sequencing error rates, cost-effectiveness, and high-quality reads. Leveraging advanced sequencing techniques to reveal the bacteriome of root canal infections and assess the impact of current disinfection methods could enable the development of more targeted and effective disinfection protocols. This dissertation presents an interventional clinical study aiming to investigate the diversity and composition of the bacteriome in primary endodontic infection (PEI) with apical periodontitis (AP) and evaluate the impact of root canal disinfection on the endodontic bacteriome using NGS on the Illumina MiSeq Platform. First, we characterized the bacteriome in PEI with AP, identified core and rare bacteriome species, and analyzed community diversity metrics using the Illumina MiSeq platform. Our results showed that Bacteroidetes, Firmicutes, Synergistetes, Fusobacteria, and Actinobacteria were the most abundant bacterial phyla. We identified 113 genera and 215 species. Analysis revealed differences in abundant taxa among distinct age, gender, symptomatology, and lesion size groups. These findings suggest that the bacteriome in PEI with AP is complex and has high microbial heterogeneity among patients. Moreover, age, gender, symptomatology, and lesion size might play a role in the abundant taxa present in PEI with AP. Second, we determined quantitatively and qualitatively the impact of chemomechanical preparation (CMP) using 2.5% sodium hypochlorite (NaOCl) on the bacteriome found in PEI with AP using the Illumina MiSeq platform. Despite a significant decrease in bacterial abundance, our findings demonstrated a distinct community composition and increased alpha diversity after CMP. We observed differential enrichment of specific taxa, including Stenotrophomonas_unclassified, Enterococcus_unclassified, and Actinomyces_unclassified, suggesting lower effectiveness of CMP using 2.5% NaOCl against these taxa. Findings from this dissertation highlight the complexity and heterogeneity of the bacteriome in PEI with AP, emphasizing the influence of patient-related factors on microbial diversity. The research highlighted the limited effectiveness of current endodontic disinfection protocols, specifically the use of 2.5% NaOCl, in reducing bacterial abundance while revealing limitations against certain taxa. These insights provide a foundation for developing more targeted and effective disinfection strategies, potentially leading to improved outcomes in endodontic therapy.16 0Item Restricted Tooth Loss in Root Canal Treated Teeth: A Six-year Cross-sectional study(Saudi Digital Library, 2023) Alrumaih, Abdulaziz; Mannocci, FrancescoAim: The aim of this cross-sectional study was to assess and compare tooth loss rates in root canal treated and non-root canal treated teeth with different restorations, and to determine the influence of restoration type on tooth survival rates in a group of patients attending a large NHS hospital. Methodology: This study analysed 1030 orthopantomograms (OPGs) from 515 patients, including baseline (T0) and follow-up (T1) OPGs. Teeth were categorised as root canal treated (RCT) or non-root canal treated (non-RCT) and further subcategorised based on the radiographic restoration type. Comparisons were made between T1 and T0 to assess tooth loss rates. Variables recorded included tooth type, the number of teeth restored at T0, and the number of teeth crowned at T1. Statistical analyses used Wilcoxon's test, Spearman's correlation coefficient, and multi-level logistic regression with generalised estimation equations (GEE) at α = 0.05 significance level. Results: The overall tooth survival rate was 75.5%, with non-RCT teeth showing a higher survival rate of 78.11% compared to RCT teeth, which had a slightly lower survival rate of 64.73%. There was higher tooth loss in 3-s restorations (RCT and non-RCT) compared to 1-s and 2-s (P<0.05). RCT 3-s restorations had an 84% increased failure risk compared with non-root canal treated teeth (OR=1.84, P=0.003). Crowning rate for RCT teeth in the time interval taken into consideration was 4.8%. Gender had no significant association, except in non-RCT 2-s restorations where females had lower failure rates than males (P=0.025). Tooth position (anterior vs. posterior teeth) did not affect survival rates (P>0.05). Conclusion: This study reveals higher tooth loss rates in root canal treated teeth, especially with three surfaces restorations. Age is identified as a risk factor for failure, particularly in root canal treated teeth with crowns.12 0Item Restricted The Effect of Simvastatin Scaffolds on human Periodontal Ligament Cells as a Potential Endodontic Regenerative Technique(2023) Almohimeed, Khawlah; Crawford, Aileen; Martin, NicolasRegenerative endodontic procedures have received considerable critical attention in the field of restorative and paediatric dentistry. Vital pulp is the best filling material a tooth can have, thus, maintaining pulp vitality is crucial in this respect. This is a preliminary study is to investigate the dose and timely release of 2% simvastatin from PLGA scaffolds as well as assess viability of human periodontal ligament cells (hPDL). The release of simvastatin in DMEM was much higher than PBS in the first 24 hours. Due to limitations in accessing the lab during Covid-19 Pandemic, spectrophotometer readings have only been obtained for a 96-hour incubation period on the Simvastatin PLGA scaffolds. HPDL showed viability after 96-hour incubation in 2% simvastatin PLGA scaffolds. the conclusion was that our newly fabricated 2% simvastatin PLGA scaffolds showed biocompatibility. Within the limitations of this study, we can conclude that using the newly fabricated 2% simvastatin PLGA scaffolds-maintained cell viability for 96 hours although it was lower than desired. Further research is needed in this respect especially that the present study contains preliminary data.31 0