HSDM Theses and Dissertations

Background and Objective: Periodontitis is an inflammatory disease caused by bacterial dysbiosis that leads to tooth support loss and eventually, loss of tooth. There have been studies using micro-CT, histology, and electron microscopy to elucidate the pathological effect of the pathogens in periodontal disease; however, the exact trace of these bacteria in inflamed periodontal tissue remains unclear. The purpose of this study was to develop a 3D imaging method that elucidates the distribution of P. gingivalis within periodontal ligament (PDL) upon infection, combining an optical clearing method, fluorescent-labeling via click chemistry, and multi-photon microscopy. Materials and methods: Periodontal disease was induced by using 5-0 silk ligature in C57BL/6 mice. P.g was labeled with fluorophore via click chemistry, then introduced to the sulcus by tying 7-0 silk ligature. Optical clearing of dissected hemi-mandibles was achieved by ECi-based clearing method. Multi-photon microscopy was utilized to obtain 3D images of 2-photon signals from the fluorophore and SHG signals from type I collagen in PDL. Fluorescent bacteria in 2-photon images were detected by trained Zen Intellesis module. Results: ECi-based clearing of hemi-mandible resulted in optimal transparency for microscope imaging, and preserved fluorophore signals labeled with click chemistry method. Multi-photon microscopy imaging allowed 3D visualization of fluorescent-labeled P.g and collagen fibers from PDL. Significantly more bacteria were detected by Zen Intellesis module in the infection group compared to the non-infection group. Conclusion: This study presents a 3D imaging method that allows tracing of bacteria in murine model without sectioning. This method has the potential to be effectively utilized in different research fields of dentistry, such as in endodontics, oral pathology, or implantology.

Background: Platelet rich plasma (PRP) has been utilized as an adjunct to various periodontal procedures with the goal of potentializing the regenerative outcome of surgical therapy. The biopolymer gelatin hydroxylphenylpropionic acid (Gtn-HPA) has recently been developed and proposed to be used as an injectable scaffold for tissue engineering. The aim of the present study was to characterize the influence of the Gtn-HPA hydrogel incorporating PRP on osteoprogenitor cells behavior and growth factor release in vitro. Methods: Platelet rich plasma was obtained from 6 human donors followed by platelet counting. The mRNA levels of osteoblast differentiation markers including OCN and COL1A1 plus the growth factors receptors FGFr, EGFr and PDGFr were analyzed in 2D cultures, hydrogel only, 10% PRP, 20% PRP, 10% FBS and 20% FBS in a 2% Gtn-HPA hydrogel (n=3) and seeded with osteoprogenitor cells. Furthermore, protein quantification for PDGF, TGF-B, EGF, FGF and IL-1B (n=5) were analyzed in the PRP lysate samples. Results: Platelet concentration increased between 5 to 9-fold in PRP compared to whole blood counts. Protein quantification was consistently greater for TGFB, PDGF and EGF. Gene expression analysis showed a tendency of upregulation of the osteoblast differentiation markers and growth factor receptor genes in the PRP groups, however a great variability between the donors was noticed. Conclusion: Within the limitations of this study, the use of human PRP in a Gtn-HPA hydrogel reveals this biopolymer as a promising and suitable carrier/scaffold with optimal growth factor release and enhanced gene expression favoring tissue regeneration.

Increased inflammation can disturb bone homeostasis leading to increased osteoclastogenesis and bone resorption. Studies have demonstrated that specialized pro-resolving lipid mediators such as lipoxins, can counteract inflammation and reduce bone loss. Despite the promising results, lipoxins may not be very stable and will require a more efficient delivery method that allows them to remain at the target site for a sufficient amount of time without disintegration. This study aims to develop a gelatin-based injectable hydrogel that will contain lipoxins for the treatment of inflammation-mediated bone resorption. Gelatin-hydroxyphenylpropionic acid (Gtn-HPA) cross-linked using horseradish peroxidase (HRP) and hydrogen peroxide (H2O2) was used as a hydrogel delivery vehicle. Aspirin- triggered Lipoxin A4 (ATL A4) was incorporated in the hydrogel. The impact of Lipoxin incorporation on the cross-linking of the hydrogel was investigated using a tube tilting method. The release profile of Lipoxins from the hydrogel was studied by incubating the hydrogel in PBS and measuring lipoxin release at predetermined timepoints. Finally, a neonatal rat calvarial organ culture model was used to test the impact of Lipoxins released from hydrogel on inflammation- mediated bone resorption. Our results indicate that Lipoxins can be successfully incorporated in the Gtn-HPA hydrogel. However, at higher doses this may result in increased gelation time. The incorporated Lipoxins can be released from the gelatin matrix and the release rate can be controlled by varying the gel weight percentage and degree of cross-linking. Softer gels release more of the original lipoxin content compared to stiffer gels. Lastly, the reduction of inflammation-mediated bone resorption from a single dose of Lipoxins released from the hydrogel was comparable to multiple doses of Lipoxins added directly to the medium in a neonatal rat calvarial bone organ culture model. The promising data from this study provides the foundation for the development of a new therapeutic modality for the treatment of conditions characterized by inflammation-mediated bone loss.

Introduction: Having an accurate diagnosis following a traumatic dental injury (TDI) is fundamental for appropriate, conservative treatment that will allow minimal destruction and maximum regain of function. However, following a TDI, the inflammation from the injury makes diagnosis unpredictable. The current recommendation is to delay treatment up to 8 weeks until a diagnosis can be determined, however a delay in treatment can lead to complications, such as the development of infection or the progression of external inflammatory root resorption (EIR). The gingival crevicular fluid (GCF) is readily accessible and has been shown contain biomarkers that can be used for diagnosis of the pulp status and of resorption. The aim of the study is to determine if biomarker levels measured in GCF are associated with infection or EIR of immature, permanent teeth after a TDI. Methods: Patients presenting to the Department of Dentistry at Boston Children’s Hospital with a TDI to an immature, permanent tooth will be followed over the course of 1 year to determine if the level of biomarkers present can predict the development of infection or EIR. The GCF will be collected using paper points at different intervals. Samples will be analyzed for 10 proinflammatory cytokines (IFN-γ, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, TNF-α) using a highly sensitive multiplex antibody-based array, Meso Scale Discovery (MSD). One biomarker of bone resorption, dentin sialoprotein (DSP), will be screened by using an ELISA panel. Results: IRB approval was obtained at HSDM and BCH, utilizing Smart IRB with HSDM as the primary site. The study was launched in March of 2024 and multiple patients have been recruited. Patient recruitment will continue over the next one year. After completion, we will compare biomarker expression in teeth of participants that developed disease versus matched controls that did not. The data will be statistically analyzed by assessing for normality, then a t-test or Wilcoxin-signed rank test will be used to test statistical differences in specific biomarkers that either predict the development of pathology or were present at a higher level at the time disease was identified. This method will be used to narrow the group of biomarkers that were consistently associated with disease. Conclusions: There will be a difference in the biomarkers of the GCF in a healthy tooth versus an immature, permanent tooth that has undergone a TDI and develops infection or EIR within the first year after the TDI.

Systemic administration of ABL significantly reduces the risk of fragility fractures in the femur, hip, and vertebrae, but no data exists on the likely positive effects that could be exerted on alveolar bone of the jaws. Considering the mechanism of action in which ABL directly influences osteoblastic cell lines and the previous clinical data reporting a substantial treatment effect of TPD in the regeneration of periodontal defects, the osteoanabolic effect of ABL could be leveraged to enhance procedures that aim to regenerate alveolar bone.

The preclinical investigations in the present report evaluate the potential of systemic ABL therapy to increase alveolar bone formation and bone mineral density (BMD), in applications of tooth extraction socket healing, implant osseointegration, and peri-implant regeneration. A series of two experiments were performed in female, Sprague-Dawley rats to assess the effects of ABL under conditions simulating: 1) osteoporosis, in animals that received ovariectomy surgery to induce bone loss via estrogen-deficiency and; 2) in healthy, young animals with normal bone metabolism. These experiments assess the biologic possibility that ABL may increase alveolar bone quality prior to implant placement, enhance primary dental implant fixation, and accelerate healing in dental implant therapy.

Contemporary orthodontic practices slowly adopt modern technologies, new treatment tools, and soft-tissue-based diagnostic approaches. Classic diagnostic tools, especially those concerning the cephalometric positioning of the incisors, are gradually being discarded from routine use. However, it is important to understand whether any of the classic treatment planning tools have a direct relation to perceived success in facial harmony. In this study, we aimed to evaluate the perceived success in posttreatment facial outcomes in a randomized orthodontic population using several components of the Steiner analysis. Inclusion criteria included an ANB angle ranging from -1 to 6 degrees and an SN-GoGn angle ranging between 26 and 39 degrees at the beginning of the treatment. In addition, to be enrolled in the study, the selected patients were required to have a Class I canine relationship, normal overjet and overbite, standard curves for Wilson and Spee, and absence of visible spacing at the end of the treatment. The records of 119 qualifying orthodontic patients were included in the study. Ten orthodontists were presented with pre-and posttreatment cephalometric radiographs of the sample and asked to review the facial outcomes of the treatment using a ten-point visualized analog scale (VAS). Based on the evaluations of the panel, three groups were formed: ideal (n=20), average (n=82), and below average (n=17). Cephalometric variables from the Steiner analysis were compared between the three groups using a one-way analysis of variance (ANOVA). In addition, Pearson Correlation coefficients were calculated between the evaluations of the judges and the cephalometric variables. The level of significance was set at p.01. Of the cephalometric variables used in this study, the measurements related to the position and inclination of the mandibular incisors and the position of the lower lip were significantly different between the three groups (p.01). Bonferroni post hoc comparisons showed that when the below average group was compared to the other two groups (p.01), significant differences occurred for L1-NB (mm and degrees) and the lower Lip to E line measurements. In addition, there was a significant difference in IMPA measurement between the below average and average groups (p.01). Significant negative correlations occurred between the judges' esthetic scores and the cephalometric variables, including all the mandibular incisor measurements and the posttreatment lower-lip position. Increased lower incisor proclination and protrusion both before and after treatment as well as posttreatment lower lip protrusion had strong affiliations with poor facial outcomes.

The objective of this study was to evaluate preoperative, intraoperative, and postoperative factors influencing outcomes of primary root canal treatment (RCT) performed by postgraduate residents at Harvard School of Dental Medicine (HSDM) Advanced Graduate Education (AGE) Endodontics between 2018 and 2024.

Patient charts were reviewed from the axiUm electronic dental record system. 367 cases with both treatment and recall codes were identified, and 250 had confirmed recall visits with appropriate periapical radiographs. After excluding cases with less than six months of follow-up, 122 cases were included, comprising 89 retrospective and 33 prospective recalls. Periapical status was assessed using the Periapical Index by two blinded examiners, and cases were classified as success (healed/healing) or failure (non-healed) based on radiographic and clinical criteria. Univariable associations were evaluated using chi-square or Fisher’s exact tests and t-tests, and selected variables were analyzed in a Firth penalized logistic regression model. Time-to-failure was analyzed using Kaplan–Meier and Cox proportional hazards models.

The overall success rate was 80.3% (98/122), with a tooth survival rate of 99.2% (121/122). Inter-examiner agreement was high (κ = 0.85). In univariable analyses, satisfactory coronal restoration, apical extent of gutta-percha filling, absence of symptoms at first recall, and higher provider year were associated with success. In the multivariable Firth model, absence of symptoms at recall (OR 0.01; 95% CI 0.001–0.06; p 0.001) and satisfactory coronal restoration (OR 5.11; 95% CI 1.17–30.48; p = 0.029) remained significant predictors, whereas provider year was not independently associated with outcome. The model demonstrated excellent discrimination (AUC = 0.936). In the Cox model, satisfactory coronal restoration was associated with a 75% reduction in hazard of failure (HR 0.25; 95% CI 0.09–0.70; p = 0.01).

Within this postgraduate training setting, primary RCT demonstrated favorable outcomes. Coronal restoration quality and symptoms at first recall visit were the strongest predictors of success, highlighting the importance of achieving an adequate coronal seal and closely monitoring symptomatic patients. Longer-term, standardized follow-up is needed to better assess treatment durability and prognostic factors.

This study aimed to compare the accuracy of a partially digital cross-mounting workflow utilizing direct scans of interocclusal records to a conventional workflow through the analysis of deviations in sequentially cross-mounted casts. Reference casts, including maxillary and mandibular full-arch prepared casts and interim prostheses, were utilized to create four reference casts for cross-mounting. In the conventional approach, 15 sets of these casts were printed, and polyvinyl siloxane (PVS) interocclusal records were employed for sequential cross-mounting. The partially digital group used the same PVS interocclusal records, which were scanned and digitally cross-mounted via design software. Mean deviations in both groups from the reference cast were analyzed using 3D inspection software. Statistical tests, including paired t-tests and analysis of variance (ANOVA), were conducted to compare discrepancies and evaluate differences in anterior and posterior regions (α = 0.05). Results demonstrated similar ranges of discrepancies in both groups, with mean deviations of 201.58 ± 136.98 mm in the conventional workflow and 248.69 ± 164.71 mm in the partially digital workflow. No statistically significant difference was observed between the two groups (P = 0.091). Error propagation analysis revealed no significant difference within the conventional group (P = 0.148), while a significant difference was found at each step of sequential mounting in the partially digital group (P 0.001). Additionally, a significant difference between anterior and posterior deviations was observed in the partially digital group (P 0.001), but not in the conventional group (P = 0.143).In conclusion, this study found no statistically significant difference between conventional and partially digital cross-mounting workflows. However, within the partially digital group, a notable difference in deviation emerged across cross-mounting steps, particularly with increased deviation in the anterior region. These findings emphasize the importance of considering specific aspects of the digital workflow to optimize accuracy in dental applications.

Many times the relationship between dental materials and clinical outcomes is poorly understood because so little is known about the biocompatibility of materials and the biological pathways they may affect. In endodontics, there is no doubt a relationship between materials like canal sealers and the tissues they come in contact with, such as dental pulp, nerve, bone, and periodontal ligament—but how this relationship manifests in clinical outcomes such as post-operative pain is unknown. This study seeks to begin to uncover these pathways by examining the toxic effects of bioceramic, resin, zinc-oxide eugenol, and silicone-based endodontic sealers on cells and tissues. An in vitro model testing three cell types, human dental pulp stem cells (hDPSCs), neuron-like cells (NBs), and murine peripheral neurons (MPNs), exposed to sealers in a dose-response fashion was used to determine the relative toxicity of sealers. An in vitro model testing inflammatory and neurotoxicity biomarker gene expression after murine mental nerve exposure to sealer provided further evidence to the toxicities of these compounds and elucidated some of the biologic pathways they may trigger from the mental nerve to the trigeminal ganglia. Results from both models generally agree that AH Plus is the most toxic and that GuttaFlow 2 exhibits nearly no toxicity, but more importantly the results of this study show how in vitro modeling can complement more complex in vivo modeling in biomaterials science.

Background: Through the progression of the COVID-19 global health crisis, dental education was under transformation. The work environment for dental faculty was characterized by new educational and administrative challenges with ever-changing social distancing protocols. Objectives: The purpose of the study was to assess the prevalence of burnout among full-time dental faculty in the US during the COVID-19 pandemic. Method: A cross-sectional study was conducted using the adapted Copenhagen Burnout Inventory. An electronic Qualtrics survey (Qualtrics®, Provo, UT) of 27 questions was administered to the deans of all US dental schools to collect data from their full-time faculty members. Descriptive and multivariable logistic analysis was conducted. Results: A total of 124 participants responded to the survey. The mean personal burnout score measured for this sample was 47.04, and the work-related score was 43.53. The junior faculty scored significantly higher on burnout than senior faculty members. During the pandemic, most participants rated “lack of social interaction and “increased household responsibilities” as disruptive to personal life, while “administrative activities” and “pursuing grants” affected their work life during the pandemic. Conclusion: Dental faculty experienced high levels of burnout during the pandemic. Therefore, dental institutions must establish supportive measures for their faculty and promote targeted faculty wellness programs.

Evolving advances in the field of implant dentistry aim to continue improving the predictability of implant therapy in replacing missing teeth. The aim of this research project is to investigate the influence of implant body thread design on the accuracy of implant placement using computer-aided implant surgery, specifically comparing the impact of two implant designs. It was hypothesized that the more aggressive thread design of the BLX implant will lead to greater 3D deviation in both healed (type 4) and extraction socket (type 1) sites, compared to the less aggressive BLC design. The study used acrylic bone models simulating a human maxillary arch with missing teeth, and implants were placed using a complete digital workflow and the deviation between planned and actual implant positions were measured using a digital software. The results revealed BLX implants were more accurate in both type 1 and type 4 sites, and also showed that type 4 had less deviation than type 1 scenarios. Limitations of the study include the use of bone models and alloy implant replicas, which do not fully replicate human bone or titanium implant surfaces, though these limitations were standardized to isolate the variables of interest, such as thread design. Although the results showing statistical significance, more clinical research is needed to corelate clinical relevance and formulate clinical recommendations.

Objective The link between diet and oral health is well-established, yet the role of inflammation in this relationship has been underexplored. This study investigates the association between the energy-adjusted Dietary Inflammatory Index (E-DII) and various oral health outcomes including caries prevalence, missing teeth, and overall caries experience, while accounting for demographic and behavioral factors using data from the 2015–2018 National Health and Nutrition Examination Survey (NHANES). Materials and Methods Dental outcomes assessed included active untreated caries, measured as the number and prevalence of: coronal and root caries; and number of missing teeth due to caries or periodontal disease complete edentulism; and overall caries experience measured by the DMFT index, which quantifies decayed, missing, and filled teeth. Logistic regression and Poisson regression models were used to evaluate associations between E-DII and oral health outcomes, adjusting for age, race, gender, education, income, smoking status, and time since last dental visit. Sensitivity analyses were conducted to assess the robustness of the findings by modifying the E-DII and applying different analytical approaches. Results The analysis included 8,131 U.S. adults aged 20 years or older who had completed dental examinations and two-day dietary recalls, representing a weighted population of approximately 231 million adults. Participants were categorized into quartiles based on their E-DII score, with higher scores indicating a more pro-inflammatory diet. The distribution of E-DII quartiles varied by demographic factors. Higher E-DII scores were associated with lower education and income levels. More than one-third of non-Hispanic Black adults (34.4%) were in the highest E-DII quartile, whereas 43.2% of non-Hispanic Asians were in the lowest quartile. A higher proportion of current smokers and individuals without recent dental visits were also in the highest E-DII quartile. Higher E-DII quartiles were significantly associated with poorer oral health outcomes. Individuals in the highest E-DII quartile had greater odds of experiencing coronal caries (adjusted OR: 2.17, 95% CI: 1.62–2.91), root caries (adjusted OR: 2.10, 95% CI: 1.37–3.22), and missing teeth (adjusted OR: 1.53, 95% CI: 1.13–2.06) compared to those in the lowest quartile. The mean number of decayed teeth increased progressively with higher E-DII quartiles, with an adjusted mean ratio of 1.79 (95% CI: 1.29–2.47) for the highest quartile. Sensitivity analyses, including a modified E-DII that excluded carbohydrates and alcohol, confirmed these associations. Further analysis using a binary classification of E-DII and continuous modeling supported the relationship between pro-inflammatory diets and poor oral health. Each unit increase in continuous E-DII was associated with increased odds of coronal caries (adjusted OR: 1.30, 95% CI: 1.16–1.44) and missing teeth (adjusted OR: 1.16, 95% CI: 1.05–1.27). Conclusion This study highlights the significant impact of pro-inflammatory diets on oral health. A higher E-DII, indicating a more inflammatory diet, was consistently associated with worse oral health outcomes, including greater odds of coronal and root caries, missing teeth, and a higher number of decayed teeth. These findings suggest that dietary modifications could serve as a potential strategy for preventing oral diseases and improving overall oral health.

Objective To investigate whether the genetic deletion of the serine protease, high temperature requirement protein A1 (HtrA1), will attenuate the progression of articular cartilage degeneration in mouse osteoarthritis (OA) models.
Methods We used two mouse models for our study. The first is a genetic model of OA and includes two strains of mice. The first strain is a heterozygous genetic deletion of type-XI collagen that results in OA in mice (Col11a1+/-). The second strain is a homozygous HtrA1 knockout (HtrA1-/-). By crossing the two strains, we can analyze a possible chondral-protective effect of HtrA1-deficiency on the progression of OA. The second model is a surgical/injury model of OA, in which we performed microsurgery on the knee joints of HtrA1-/- mice to destabilize the medial meniscus (DMM). This technique undermines the joint, and rapidly induces OA in the mice. By surgically inducing osteoarthritis in HtrA1-/- knockout mice, we can simulate the effect of HtrA1-deficiency on the progression of OA due to surgery or injury. Conditions of the articular cartilage from the knee joints of the Col11a1+/-;HtrA1-/- mice, Col11a1+/- mice, HtrA1-/- mice, and the surgical mice were then examined by histology, graded on a standard scoring system, and characterized by immunohistochemistry.
Results We examined the HtrA1-/- knockout mice and found no overt phenotype abnormalities. We also found that the deletion of HtrA1 delays the progressive process of articular cartilage degeneration in Col11a1+/- mice. In our surgical model, the degenerative progression towards OA was dramatically delayed in HtrA1-/- mice when compared to the controls. Conclusion Deletion of HtrA1 delays the progression of articular cartilage degeneration in OA models induced either by collagen type-XI haploinsufficiency or by DMM surgery. Therefore, the development of antagonistic drugs that specifically target HTRA1 may be an effective method to treat OA in the future.

Background The goal of this study is to understand role of transforming growth factor beta 1 (TGF-β1) in development of osteoarthritis (OA). Results from studies indicate that the genetic inactivation of Smad-3, or the disruption of the interaction of Tgf-β1 with its receptor Tgf-β type II receptor (Tgfbr2), in germline cells results in OA-like knee joints in mice at one month of age. However, other studies suggest that the increased expression of Tgf-β1 in mature knee joints causes OA in animal models. A human genetic study reports that a two-nucleotide deletion, 741-742del AT, and/or a nucleotide change, 859C>T or 782C>T in SMAD-3 are associated with early-onset OA. This observation is consistent with the finding that the lack of Tgf-β1 signaling in the germline cell results in OA in developing joints and that increased Tgf-β1 signaling causes OA in mature joints. The plausible explanation for this “conflicting” role of TGF-β1 in the pathogenesis of OA is that the effective TGF-β1 signaling acts in either a dose-dependent or a developmental stage-dependent manner. The present study addresses the question as to whether inhibition of Tgf-β1 signaling prevents mature knee joints from being degenerated in mouse models of OA.

Methods

1. Using conditional knock out techniques with aggrecan-CreERt2 mice and floxed Tgfbr2 mice, Tgfbr2 was removed from articular cartilage of knee joints in 2-month-old mice. Mice without Tgfbr2 were kept for another 6 months or longer. Knee joints from the mice (n=8) and their corresponding control (n=4) were collected for morphological analysis.
2. Mice without Tgfbr2 at two months old were subjected to DMM to induce articular cartilage degeneration. Knee joints from the mice at 4 and 8 weeks post surgery (n=8 in each group) were collected for morphological analysis.

Results

1. We did not find the initiation and acceleration of articular cartilage degeneration by the genetic inactivation of Tgfbr2 in knee joints of mice at the age of 9 months or older. We also did not find hypertrophic chondrocytes in the articular cartilage of the mice.
2. We found that removal of Tgfbr2 in articular cartilage of knee joints delayed articular cartilage degeneration, at least 6 weeks, compared to that in wild-type littermates.

Conclusion Inhibition of Tgf-β1 signaling attenuated articular cartilage degeneration in mature knee joints of mouse models of OA. Therefore, inhibition activity of TGF-β1, not application of TGF-β1, may be considered in treatment of OA in mature joints.

Current modeling postulates that the development of cancer is driven by the accumulation of genetic mutations and epigenetic modifications, resulting in a clonal population of cells with deregulated growth characteristics. Three epigenetic mechanisms have been well documented, including DNA methylation, histone modifications and non-coding RNAs. Of these, DNA methylation is the best studied epigenetic alteration in cancer. Characterization of the expression of the epigenetic marker 5-hydroxymethylcytosine (5-hmC) in human and experimental melanoma and its precursors has established it as an important functionally relevant epigenetic biomarker in cancer. We therefore posited that 5-hmC in oral premalignant lesions and oral squamous cell carcinomas represents an opportunity to identify a novel epigenetic molecular pathway that may serve as a biomarker for early diagnosis of oral neoplasms and subsequent improvements in the prognosis of oral cancer. A total of sixty-six histologic samples (N=66) were obtained from the Harvard School of Dental Medicine affiliated bio-archives. Immunohistochemistry for 5-hmC was performed on nine cases diagnosed as fibromas (F, nF=9) with uninvolved margins used to evaluate non-inflamed/non-cancerous mucosa; nine cases diagnosed as frictional keratosis (FK, nFK=9); ten cases diagnosed as lichen planus (LP, nLP=10); fifteen cases diagnosed as moderate-to-severe oral epithelial dysplasia (OED, nOED=15) and twenty-three cases diagnosed as oral squamous cell carcinoma (oSCC, noSCC=23). Human cell lines of oral keratinocytes from benign oral mucosa, oral epithelial dysplasia and four types of oral squamous cell carcinoma were also evaluated to detect the expression of 5-hmC genomic DNA. Finally, immunohistochemistry for 5-hmC was also performed in histologic samples of a murine model where oral squamous cell carcinoma was induced with the carcinogen 4-nitroquinoline-1-oxide (N=10). Five samples consisted of non-inflamed/non-neoplastic normal dorsal tongue mucosa (T, nT=5) and five consisted of oral squamous cell carcinoma (mSCC, nmSCC=5). Progressive loss from benign and reactive/inflammatory oral mucosal lesions to oral epithelial dysplasia and oral squamous cell carcinoma was observed in patient samples. Likewise, distinct patterns with regard to the distribution of staining was noted in benign non-inflamed lesions versus reactive inflammatory lesions such as frictional keratosis and lichen planus, providing further explanation germane for the understanding of the responses of the oral mucosa to tissue injury. The finding in human tissue of loss of 5-hmC immunoreactivity within dysplasia and carcinoma were substantiated in human cell lines at a DNA level, and further validated in an in vivo experimental model of murine oral SCC. In aggregate, our results confirm the potential importance of loss of 5-hmC as a novel epigenetic mark in oral SCC at both diagnostic and biological levels, and establish a murine model of oral SCC induction as a potentially informative approach to understanding the role of epigenetic regulation in oral carcinogenesis.

Background: Traumatic dental injuries (TDIs) present a significant public health challenge in pediatric populations, with prevalence ranging from 20% to 30%. Diagnostic uncertainty complicates the management of permanent teeth following trauma. Transient denervation frequently renders traditional thermal sensibility testing unreliable during the initial months post-injury, while external inflammatory root resorption can progress rapidly before radiographic detection. There is a critical need for non-invasive predictive tools to identify teeth at high risk for delayed complications.

Objective: This prospective cohort pilot study aimed to determine if inflammatory cytokine and chemokine levels in the gingival crevicular fluid (GCF) of traumatized permanent teeth can serve as early biomarkers for adverse events, specifically pulp necrosis and external inflammatory root resorption.

Methods: This study recruited a cohort of 27 pediatric patients presenting to Boston Children’s Hospital with TDIs in their permanent dentition, resulting in a total of 64 traumatized teeth for analysis. GCF was collected non-invasively for proteomic analysis using the Olink Target 48 cytokine assay to quantify key inflammatory mediators. Clinical variables and trauma types were recorded to assess correlations with adverse endodontic outcomes. Statistical analysis in R used Fisher’s Exact Tests and Firth’s Logistic Regression to assess associations between clinical parameters and adverse outcomes, supplemented by Kaplan-Meier survival curves for event-free probability. GCF proteomic data were analyzed via Benjamini-Hochberg adjusted t-tests to compare protein levels across injury and outcome categories, with longitudinal trends plotted for a subset of cases.

Results: In this pediatric cohort (N=27), adverse events occurred in 17.2% of traumatized teeth (n=27). There was an average of 2.3 follow-ups per patient. While the majority of cases involved single trauma types, sustaining multiple simultaneous injuries significantly increased the risk of requiring treatment (OR = 4.65). GCF analysis revealed significantly elevated pro-inflammatory and resorptive biomarkers (adjusted p 0.05), including colony-stimulating factors, chemokines, and tissue degradation mediators. These molecular shifts correlated most strongly with crown fractures and minor displacements (). Exploratory analysis further identified a potential immunological transition based on complication type: isolated necrosis showed a trend toward acute pro-inflammatory signaling (IL-17C, IL-18, IL-15), whereas concurrent resorption was associated with elevated levels of the anti-inflammatory cytokine IL-27, suggesting a shift from an acute inflammation phase to a chronic phase.

Conclusion: Gingival crevicular fluid analysis identified a distinct profile of changes in biomarkers associated with differences in trauma type and the development of adverse outcomes in traumatized teeth. While the restricted sample size and overlapping injury profiles limited our ability to isolate specific clinical triggers or a definitive timeline for these shifts, these findings underscore the potential for non-invasive GCF profiling to predict delayed endodontic complications and inform future diagnostic thresholds.

Objectives – To investigate the anatomy of the mouse palate, the midpalatal suture, and the cellular characteristics in the sutures before and immediately after midpalatal suture expansion, and as well as, to study the effects of compression force on the midpalatal suture.

Materials and Methods – Wild-type C57BL ⁄ 6 male mice, aged between 6 weeks and 12 months, were chosen for all the experiments. The complete palate of the non-operated group and the midpalatal suture-expanded or -compressed group at different ages was used for histological, micro-CT, immunohistochemistry, and sutural cell analyses. Animals in the experimental group for compression were subjected to palatal suture compression force by closing loops for the periods of 1, 3, 5, 7, 14, and 28 days.

Results – This study documents precise morphological and histological characteristics of the mouse palatal sutures. In addition to the opening of the midpalatal suture caused by expansion, both transverse and interpalatine sutures were also seen to be affected. Cellular density was decreased in different types of sutures following the application of expansion force. In respect to compression study, suture width, maxillary width, and bone volume to total volume were significantly decreased compared to control at 14 and 28 days following the application of compressive force. Compression force caused an increased cell apoptosis in midpalatal suture area. Osteoclast activity and expression of MMP-9 were increased, specifically on the nasal side.

Conclusions – The detailed morphology and histology of the mouse palate and the cellular changes that occur following midpalatal suture expansion or compression, as described here, will be helpful as a basis for further investigations of palatal suture tissue responses to mechanical force.

Introduction: Children of Kuwait have both a high obesity and dental caries prevalence, as well as excessive consumption of sugar-sweetened beverages (SSB). Obesity is a major risk factor for serious health conditions such as diabetes, cancer, and cardiovascular disease. Metabolic syndrome (SMets), the occurrence of 3 out 5 variables (obesity, hypertension, hyperglycemia, lowered high-density lipoprotein cholesterol (HDLC), and hypertriglyceridemia) is a risk factor that predicts future cardiovascular disease and type 2-diabetes. The aim of this analysis was to assess the impact of the consumption of three beverages (soda, milk, and juice) on the obesity and dental caries in Kuwaiti children. We also aim to investigate the changes overtime of the salivary biomarkers; insulin, CRP, phosphate, and uric acid. This study followed a group of children before having developed SMets and after its development, and compared them to children with no SMets characteristics. Methods and Materials: Data were obtained from the Kuwait Healthy Life Study. For the impact of beverage consumption on the severity of dental caries, 8,317 children were included from the baseline visit in 2012-13 (age =10). For the obesity 6,305 children were included, from both baseline and follow-up visit in 2014-15 (age =12). A subset of 94 children were included in the study of the salivary biomarker change over time, all the children had no SMets at baseline, 51 became SMets positive and 43 remain healthy at follow-up. A multivariate logistic regression analysis was conducted to examine the relation between developing obesity as a dependent binary variable, and beverages consumption amount as a categorical independent variable. For the severity of dental caries, a multivariate logistic regression was used to evaluate this association between the severity of dental caries as a binary dependent variable (low and high), and the categorical consumption (amount and pattern) of the three beverages as independent variables. For the salivary biomarkers, the changes in the salivary biomarker measurements from baseline to follow-up for each group were tested using Wilcoxon matched-pairs signed-ranks test. Results: A significant association between high soda and milk consumption with developing obesity (OR=1.68, P= 0.004, CI =1.19- 2.39) (OR=1.77, P=0.019, CI =1.10- 2.87), respectively. For the severity of dental caries, high soda consumption was significantly associated with severe dental caries (OR=1.20, P= 0.041, CI =1.01- 1.42). Moderate milk drinking showed a protective effect from having severe dental caries (OR=0.88, P= 0.007, CI =0.81- 0.97). The change in salivary biomarkers from baseline to follow-up, children in the healthy group had no significant change. Children who developed SMets had a significant increase in all salivary biomarkers; insulin became double the baseline levels (p =0.014), CRP were 120% higher (p =0.005), phosphate became 11% higher (p =0.030) and uric acid showed 17% elevation (p =0.009). Conclusion: High soda consumption was significantly associated with obesity and dental caries. High milk consumption was significantly associated with obesity but not with dental caries. Moderate milk consumption was protective from having severe dental caries, even if it was occasional use of flavored milk. Children who developed SMets showed a significant elevation in all the four salivary biomarkers while there was no significant change noted in the levels for children who did not develop SMets.

Background: Conventional bone augmentation for dental implants typically involves grafting materials, with or without barrier membranes, to preserve ridge dimensions. This study compares the changes in horizontal ridge width in patients undergoing alveolar ridge preservation (ARP) with a xenogenic (bovine) collagen plug, with and without a collagen membrane. Methods: A 6-month randomized clinical trial was conducted with 20 participants requiring ARP after tooth extraction. Participants were randomly assigned to two groups: Group I, which received a xenogenic plug alone, and Group II, which received a xenogeneic plug with a non-crosslinked collagen membrane. Both groups were allowed to heal by secondary intention. The outcomes of interest included clinical, radiographic, and ultrasonographic changes in hard and soft tissues, as well as histologic and histomorphometric evidence of new bone formation. Results: All enrolled participants completed the study, with a mean age of 54.25 ± 17.08 years, including 17 premolars and 3 . Baseline buccal bone thickness was similar between the groups (Group 1: 1.02 ± 0.44 mm; Group 2: 0.93 ± 0.45 mm; p = 0.662). At 6 months, no significant differences were observed between groups for keratinized tissue width (Group 1: 4.80 ± 1.48 mm; Group 2: 4.80 ± 1.14 mm; p = 0.999), gingival thickness (Group 1: 1.22 ± 0.25 mm; Group 2: 1.20 ± 0.35 mm; p = 0.650), horizontal bone width at 3 mm from the crest (Group 1: 9.02 ± 2.09 mm; Group 2: 8.41 ± 1.45 mm; p = 0.464), and change in vertical bone at the buccal aspect (Group 1: -0.78 ± 0.59 mm; Group 2: -1.31 ± 1.20 mm; p = 0.244). Ultrasound evaluations were consistent with clinical measurements. Histologic analysis showed similar new bone formation between groups (Group 1: 25.80 ± 15.48%; Group 2: 27.89 ± 18.21%; p = 0.803). Conclusion: The xenogeneic collagen plug alone yielded comparable ARP outcomes to its combination with a collagen membrane, simplifying the treatment process by reducing costs, time, and the need for additional materials.

The role of bacterial plaque and its role as the primary etiology for periodontal disease is well established. The treatment of periodontitis and maintenance of periodontal health is dependent on the elimination of plaque, and calculus, and establishing a stringent dental hygiene maintenance program. The non-surgical treatment of periodontitis relies heavily on scaling and root planning (SRP) therapy to remove subgingival deposits and toxins to allow for the reattachment of periodontal and gingival fibers.

A multitude of sharpening devices are available for maintaining periodontal hand instrument sharpness. However, the impact of these devices on the cutting-edge morphology of periodontal curettes, and subsequently on root surface roughness remains unclear. . Using sharp periodontal hand curettes and scalers is pivotal in the thoroughness of calculus removal, root planing, and achieving a smooth root surface.

This study aims to investigate the effects of sharpening cards, sharpening stones of varying grits, and the Sidekick® on the lateral edge of an unused factory sharpened Gracey 11/12 curette and to evaluate the influence of these sharpened curettes on root surface roughness post-SRP.

The results show that there was a statistically significant difference between the roughness of the medium, fine, and extra-fine cards and between the method of sharpening (horizontal vs vertical) using a stationary-instrument-moving-stone technique. However, no statistically significant difference was observed in the roughness of curette edges sharpened by cards and stones of varying grit sizes, nor in the root surfaces root-planed by these sharpened curettes. This suggests that irrespective of the card or stone utilized for sharpening, the resulting edge of the curette yields relatively similar smooth surfaces. Further studies will need to be conducted to verify this finding.