The Effect of Calcium Hydroxide Pastes on Isolated Vital Nerve Fibers.

INTRODUCTION: Calcium hydroxide pastes (CHPs), commonly used for disinfecting root canals during endodontic treatment, are generally considered safe. However, accidental extrusions result in minimal injuries and little to no discomfort, except when extruded pastes come into contact with nerve bundles, such as the inferior alveolar nerve. Currently, there is a lack of information about the possible role of specific paste vehicles on the extent of nerve injury. The purpose of this study was to compare the role that paste vehicles, such as water or methylcellulose, may play when nerve fibers are exposed to CHP. METHODS: Isolated sciatic nerves of Sprague-Dawley rats were exposed to either water-based or methylcellulose-based CHP for varying durations of time (30, 60, or 90 minutes). Histopathological changes, including axonal edema, myelin alterations, and loss of cellular outlines, were assessed, and the degrees of changes were compared using chi-square intraclass correlation coefficient tests. RESULTS: Both groups exposed to the pastes demonstrated varying degrees of histopathologic changes, including axonal edema, myelin changes, and loss of cellular outlines, at different exposure times. The water-based calcium hydroxide paste induced these changes more rapidly than the methylcellulose-based paste. Similar patterns were observed in the scanning electron microscopic findings. Exposure time emerged as an important difference in the effects of the 2 pastes. In each of these tests, all observations of water-based paste exposure were rated as moderate to severe, whereas the observed cellular changes (axonal, myelin, and intact cellular outline) were rated as mild to moderate after exposure to methylcellulose-based paste for the same exposure durations. The chi-square tests indicated a statistically significant association between the material and each of the outcomes (axonal changes: χ²15 = 81.0, P < .001; myelin changes: χ²15 = 81.0, P < .001; intact cellular outline, χ²15 = 81.0, P < .001). The intraclass correlation coefficient value was 0.93. CONCLUSIONS: The study demonstrates that axonal and myelin damage increase with longer exposure times, with water-based CHP causing more damage than methylcellulose-based CHP at each time point.

Artificial intelligence-based epigenomic, transcriptomic and histologic signatures of tobacco use in oral squamous cell carcinoma.

Oral squamous cell carcinoma (OSCC) biomarker studies rarely employ multi-omic biomarker strategies and pertinent clinicopathologic characteristics to predict mortality. In this study we determine for the first time a combined epigenetic, gene expression, and histology signature that differentiates between patients with different tobacco use history (heavy tobacco use with ≥10 pack years vs. no tobacco use). Using The Cancer Genome Atlas (TCGA) cohort (n = 257) and an internal cohort (n = 40), we identify 3 epigenetic markers (GPR15, GNG12, GDNF) and 13 expression markers (IGHA2, SCG5, RPL3L, NTRK1, CD96, BMP6, TFPI2, EFEMP2, RYR3, DMTN, GPD2, BAALC, and FMO3), which are dysregulated in OSCC patients who were never smokers vs. those who have a ≥ 10 pack year history. While mortality risk prediction based on smoking status and clinicopathologic covariates alone is inaccurate (c-statistic = 0.57), the combined epigenetic/expression and histologic signature has a c-statistic = 0.9409 in predicting 5-year mortality in OSCC patients.

Oral Squamous Papilloma Mimicking Oral Verrucous Leukoplakia.

The human papillomavirus (HPV) is a non-enveloped DNA virus that causes a variety of skin and mucosal lesions. This report reviews a likely HPV-related lesion of oral squamous cell papilloma clinically mimicking oral verrucous leukoplakia. A 71-year-old white male presented with a raised white lesion on the palatal mucosa. It felt hard on palpation and had a sessile fixed base, and a rough verrucous surface. The lesion was fully excised. Histopathology showed short, thin, fingerlike projections lined by stratified squamous epithelium with thin central connective tissue cores. The epithelial superficial layers demonstrated focal koilocytotic changes suggestive of an HPV infection. High-risk HPV-related lesions have the potential to turn malignant. Early diagnosis and management are critical to preventing serious complications.

Neurotrophin Pathway Receptors NGFR and TrkA Control Perineural Invasion, Metastasis, and Pain in Oral Cancer.

Oral squamous cell carcinoma (OSCC) patients suffer from poor survival due to metastasis or locoregional recurrence, processes that are both facilitated by perineural invasion (PNI). OSCC has higher rates of PNI than other cancer subtypes, with PNI present in 80% of tumors. Despite the impact of PNI on oral cancer prognosis and pain, little is known about the genes that drive PNI, which in turn drive pain, invasion, and metastasis. In this study, clinical data, preclinical, and in vitro models are leveraged to elucidate the role of neurotrophins in OSCC metastasis, PNI, and pain. The expression data in OSCC patients with metastasis, PNI, or pain demonstrate dysregulation of neurotrophin genes. TrkA and nerve growth factor receptor (NGFR) are focused, two receptors that are activated by NGF, a neurotrophin expressed at high levels in OSCC. It is demonstrated that targeted knockdown of these two receptors inhibits proliferation and invasion in an in vitro and preclinical model of OSCC, and metastasis, PNI, and pain. It is further determined that TrkA knockdown alone inhibits thermal hyperalgesia, whereas NGFR knockdown alone inhibits mechanical allodynia. Collectively the results highlight the ability of OSCC to co-opt different components of the neurotrophin pathway in metastasis, PNI, and pain.

Brush swab as a noninvasive surrogate for tissue biopsies in epigenomic profiling of oral cancer.

BACKGROUND: Oral squamous cell carcinoma (OSCC) has poor survival rates. There is a pressing need to develop more precise risk assessment methods to tailor clinical treatment. Epigenome-wide association studies in OSCC have not produced a viable biomarker. These studies have relied on methylation array platforms, which are limited in their ability to profile the methylome. In this study, we use MethylCap-Seq (MC-Seq), a comprehensive methylation quantification technique, and brush swab samples, to develop a noninvasive, readily translatable approach to profile the methylome in OSCC patients. METHODS: Three OSCC patients underwent collection of cancer and contralateral normal tissue and brush swab biopsies, totaling 4 samples for each patient. Epigenome-wide DNA methylation quantification was performed using the SureSelectXT Methyl-Seq platform. DNA quality and methylation site resolution were compared between brush swab and tissue samples. Correlation and methylation value difference were determined for brush swabs vs. tissues for each respective patient and site (i.e., cancer or normal). Correlations were calculated between cancer and normal tissues and brush swab samples for each patient to determine the robustness of DNA methylation marks using brush swabs in clinical biomarker studies. RESULTS: There were no significant differences in DNA yield between tissue and brush swab samples. Mapping efficiency exceeded 90% across all samples, with no differences between tissue and brush swabs. The average number of CpG sites with at least 10x depth of coverage was 2,716,674 for brush swabs and 2,903,261 for tissues. Matched tissue and brush swabs had excellent correlation (r = 0.913 for cancer samples and r = 0.951 for normal samples). The methylation profile of the top 1000 CpGs was significantly different between cancer and normal samples (mean p-value = 0.00021) but not different between tissues and brush swabs (mean p-value = 0.11). CONCLUSIONS: Our results demonstrate that MC-Seq is an efficient platform for epigenome profiling in cancer biomarker studies, with broader methylome coverage than array-based platforms. Brush swab biopsy provides adequate DNA yield for MC-Seq, and taken together, our findings set the stage for development of a non-invasive methylome quantification technique for oral cancer with high translational potential.

The REASON score: an epigenetic and clinicopathologic score to predict risk of poor survival in patients with early stage oral squamous cell carcinoma.

BACKGROUND: Oral squamous cell carcinoma (OSCC) is a capricious cancer with poor survival rates, even for early-stage patients. There is a pressing need to develop more precise risk assessment methods to appropriately tailor clinical treatment. Genome-wide association studies have not produced a viable biomarker. However, these studies are limited by using heterogeneous cohorts, not focusing on methylation although OSCC is a heavily epigenetically-regulated cancer, and not combining molecular data with clinicopathologic data for risk prediction. In this study we focused on early-stage (I/II) OSCC and created a risk score called the REASON score, which combines clinicopathologic characteristics with a 12-gene methylation signature, to predict the risk of 5-year mortality. METHODS: We combined data from an internal cohort (n = 515) and The Cancer Genome Atlas (TCGA) cohort (n = 58). We collected clinicopathologic data from both cohorts to derive the non-molecular portion of the REASON score. We then analyzed the TCGA cohort DNA methylation data to derive the molecular portion of the risk score. RESULTS: 5-year disease specific survival was 63% for the internal cohort and 86% for the TCGA cohort. The clinicopathologic features with the highest predictive ability among the two the cohorts were age, race, sex, tobacco use, alcohol use, histologic grade, stage, perineural invasion (PNI), lymphovascular invasion (LVI), and margin status. This panel of 10 non-molecular features predicted 5-year mortality risk with a concordance (c)-index = 0.67. Our molecular panel consisted of a 12-gene methylation signature (i.e., HORMAD2, MYLK, GPR133, SOX8, TRPA1, ABCA2, HGFAC, MCPH1, WDR86, CACNA1H, RNF216, CCNJL), which had the most significant differential methylation between patients who survived vs. died by 5 years. All 12 genes have already been linked to survival in other cancers. Of the genes, only SOX8 was previously associated with OSCC; our study was the first to link the remaining 11 genes to OSCC survival. The combined molecular and non-molecular panel formed the REASON score, which predicted risk of death with a c-index = 0.915. CONCLUSIONS: The REASON score is a promising biomarker to predict risk of mortality in early-stage OSCC patients. Validation of the REASON score in a larger independent cohort is warranted.

Regenerative Endodontic Treatment in Immature Noninfected Ferret Teeth Using Blood Clot or SynOss Putty as Scaffolds.

INTRODUCTION: SynOss Putty (Collagen Matrix, Oakland, NJ) has shown the formation of mineralized tissues when used as a scaffold in regenerative endodontic treatment (RET) in immature human teeth. The aim of this study was to compare the outcome of RET in immature ferret teeth using 2 scaffolds: a blood clot and SynOss Putty. METHODS: Thirty-two immature canine teeth in 8 ferrets (95-105 days old) were divided into 4 groups: group 1, no treatment (positive control, n = 8); group 2, full pulpectomy with no further treatment (negative control, n = 8); group 3, revascularization using a blood clot (n = 8); and group 4, revascularization using a SynOss Putty scaffold (n = 8). After 3 months, the animals were euthanized, and the newly formed tissues were examined histologically. The data were statistically analyzed using chi-square and Fisher exact tests. RESULTS: Normal pulps were found in group 1. No pulp tissue was found in teeth in group 2. In group 3, the pulp tissue and the odontoblastic layer were absent, and the root canal spaces were filled with a hard tissue characterized as bonelike and cementumlike tissues. All teeth except 1 in group 4 showed no hard tissue formation and intracanal/periapical inflammation. SynOss Putty was significantly associated with a lack of tissue formation and intracanal/periapical inflammation (P < .05). CONCLUSIONS: Intracanal hard tissue formation was observed in immature teeth after RET using a blood clot. No tissue regeneration was found in the majority of samples using SynOss Putty as a scaffold.

The haemostatic efficacy and foreign body reaction of epinephrine-Impregnated polyurethane foam in osseous defects.

The objective of this study was to compare the haemostatic efficacy and foreign body reaction of epinephrine-impregnated cotton pellets with those of epinephrine-impregnated polyurethane (PU) foam cubes in osseous defects created in guinea pigs. Initially, these substances were randomly applied to the osseous defects in guinea pigs for 2 min and blood loss was measured. The animals were then sacrificed 7 weeks later and the degree of foreign body reaction was scored. The data were analysed by the independent-samples Kruskal-Wallis test. Epinephrine-impregnated PU foam cubes showed significantly better haemostatic effect compared to epinephrine-impregnated cotton pellets. The PU foam containing epinephrine specimens elicited significantly less foreign body reaction compared to epinephrine cotton pellets (P < 0.05). Based on the results of this study, it is concluded that epinephrine-impregnated PU foam cubes are a good alternative to epinephrine-impregnated cotton pellets as a local haemostatic agent in endodontic surgery.

Effect of Residual Dental Pulp Tissue on Regeneration of Dentin-pulp Complex: An In Vivo Investigation.

INTRODUCTION: Current pulp revascularization procedures in teeth with necrotic pulps and open apices have produced histologic evidence of connective tissue growth, cementum, and bone within the root canals of experimental animals. This study aims to investigate the effect of maintaining uninflamed residual apical pulp tissue on the histologic outcome of pulp-dentin complex regeneration after a revascularization procedure in immature ferret cuspid teeth. METHODS: Twenty-eight cuspid teeth from 7 young male ferrets were used in this experiment. Seven teeth were reserved to serve as positive control samples without any treatment. In another 7 teeth, the pulp was completely extirpated (negative control), whereas the pulp of the remaining 14 teeth were removed to either 1-2 mm short of the apex (7 samples) or 2-4 mm short of the apex (7 samples). Blood clots were covered with mineral trioxide aggregate at the cementoenamel junction level of each tooth. Three months later, block sections were removed for histologic evaluations, and the data were statistically analyzed with the chi-square test (P < .05). RESULTS: All teeth with complete pulp extirpation showed the presence of bone inside the root canal. In contrast, the root canals for most teeth with pulp amputation 1-4 mm from the radiographic apex were filled with normal pulp, which extended coronally to the mineral trioxide aggregate, where hard tissue bridges had formed. CONCLUSIONS: Based on these results, we concluded that regeneration of the pulp-dentin complex is possible when the apical 1-4 mm of the apical pulp remains intact in immature teeth.

Two cases of CD30+, anaplastic lymphoma kinase (ALK)-negative anaplastic large cell lymphoma with oral manifestations.

Anaplastic large cell lymphoma (ALCL), a subgroup of T-cell non-Hodgkin's lymphoma, is an uncommon tumor exhibiting CD30 positivity and a characteristic immunophenotypic profile. Histologically, ALCL is characterized by the proliferation of large, anaplastic lymphoid cells with eccentric horseshoe- or kidney-shaped nuclei and one or more prominent nucleoli. Rare cases have been cited in the literature of ALCL presenting primarily in the oral cavity. The purpose of this article was to present 2 instructive cases of CD30+, anaplastic lymphoma kinase-negative ALCL with oral and systemic involvement.