Differentially Expressed Genes in Dental Pulp Tissues of Individuals With Symptomatic Irreversible Pulpitis With and Without History of COVID-19.

INTRODUCTION: Increased levels of proinflammatory markers have been reported in tissues of individuals with Coronavirus Disease 2019 (COVID-19). We hypothesize that inflamed dental pulp tissues of individuals with previous history of COVID-19 may present a differential inflammatory gene expression profile in comparison with individuals who never had COVID-19. MATERIALS AND METHODS: Dental pulp tissues were collected from 27 individuals referred for endodontic treatment due to symptomatic irreversible pulpitis. Of these, 16 individuals had a history of COVID-19 (6 months to 1 year post infection) and 11 individuals had no previous history of COVID-19 (controls). Total RNA from pulp tissue samples was extracted and subjected to RNA sequencing for comparison of differentially expressed genes (DEGs) among groups. DEGs showing log2(fold change) > 1 or < -1, and P < .05 were considered significantly dysregulated. RESULTS: RNA sequencing identified 1461 genes as differentially expressed among the groups. Of these, 311 were protein coding genes, 252 (81%) that were upregulated and 59 (19%) that were downregulated in the COVID group compared with controls. The top upregulated genes in the COVID group were HSFX1 (4.12-fold change) and LINGO3 (2.06-fold change); significantly downregulated genes were LYZ (-1.52-fold change), CCL15 and IL8 (-1.45-fold change). CONCLUSIONS: Differential gene expression in dental pulp tissues of COVID and non-COVID groups suggests potential contribution of COVID-19 on dysregulating inflammatory gene expression in the inflamed dental pulp.

Evaluation of the Physicochemical and Biological Properties of EndoSequence BC Sealer HiFlow.

INTRODUCTION: Understanding the physicochemical and biological properties of endodontic sealers is important for endodontic treatment planning. This study evaluated the properties of EndoSequence BC Sealer HiFlow (BCH; Brasseler USA, Savannah, GA), EndoSequence BC Sealer (BC, Brasseler USA), and AH Plus (AHP; Dentsply DeTrey, Konstanz, Germany). The effect of temperature on the setting time and flow of these sealers was also evaluated. METHODS: The setting time, flow, radiopacity, pH, solubility, and calcium release were investigated following ISO guidelines. The morphology and chemical composition of the sealers were evaluated by scanning electron microscopy and energy-dispersive spectroscopy. The antibacterial activity of sealers was tested against 2 strains of Enterococcus faecalis. Sealer cytotoxicity and the effects on messenger RNA expression of proinflammatory and mineralization genes were also investigated. Data analysis was performed using analysis of variance, Tukey, Kruskal-Wallis, and Dunn multiple comparison tests. P ≤ .05 was considered statistically significant. RESULTS: The setting time and flow rate of all sealers were affected by heat (P ≤ .05). The setting times and solubility of BCH and BC were significantly higher than AHP (P ≤ .0001). The radiopacity of AHP was higher than BCH and BC (P ≤ .0001). All sealers were alkaline and had antibacterial effects. Cell viability was higher for BCH and BC than AHP (P ≤ .0001). No significant differences in messenger RNA expression of proinflammatory and mineralization genes were observed. CONCLUSIONS: Overall, BCH and BC had similar physicochemical and biological properties. The observed high solubility of BCH and BC as well as the high cytotoxicity of AHP might negatively impact the clinical performance of these materials. The application of heat affected the setting time and flow of all sealers.

Antibiofilm and immunomodulatory resorbable nanofibrous filing for dental pulp regenerative procedures.

Multifunctional scaffolds with host defense peptides designed for regenerative endodontics are desirable nanobiotechnological tools for dentistry. Here, different scaffolds were tested for use during the pulp revascularization process, including poly(vinyl alcohol)-PVA hydrogels or resins, collagen hydrogels and poly(vinyl alcohol) PVA/Chitosan (PVA/CS) nanofibers. Based on time to degradation (21 days), nanofibers were chosen to be incorporated with ciprofloxacin and IDR-1002 (each at 50 mg/g). Nanofibers containing ciprofloxacin and IDR-1002 had anti-biofilm activity against Enterococcus faecalis, Staphylococcus aureus and a multispecies oral biofilm, besides anti-inflammatory activities. The in vivo subcutaneous tissue response to tooth fragments filled with nanofibers demonstrated a pulp-like tissue formation, when compared to empty teeth fragments. Thus, we designed a strong antimicrobial, immunomodulatory and regenerative candidate for pulp revascularization and regeneration procedures.

Evaluation of Passive Ultrasonic Irrigation and GentleWave System as Adjuvants in Endodontic Retreatment.

INTRODUCTION: Nonsurgical endodontic retreatment continues to be a challenge in endodontics, particularly when dealing with a complex tooth anatomy. This study evaluated the efficacy of passive ultrasonic irrigation (PUI) and the GentleWave system as supplementary techniques to remove remaining filling materials from oval-shaped root canals. METHODS: Twenty distal roots of human mandibular molars with single and oval-shaped canals were shaped with R40 (40.06) instrument and filled with gutta-percha and AH Plus sealer using warm vertical obturation. Initial filling material removal was performed with R50 (50.05) instrument, followed by the use of PUI (n = 10) or GentleWave system (n = 10). Micro-computed tomographic images were obtained after obturation, initial material removal, and after the use of PUI and GentleWave. The volume of remaining filling material was calculated for the entire canal as well as for the coronal, middle, and apical thirds. Statistical analyses were performed by using analysis of variance, Kruskal-Wallis and Mann-Whitney tests. P ≤ .05 was considered significant. RESULTS: The use of PUI and GentleWave as supplementary techniques significantly reduced the volume of remaining filling material after initial instrumentation (P < .05). However, none of these techniques was able to render canals free from filling materials. PUI showed better performance by removing 18% of the remaining filling material, whereas the GentleWave system was able to remove approximately 10% (P = .02). CONCLUSIONS: The use of supplementary techniques optimized filling material removal after initial instrumentation. PUI enhanced the overall cleaning of the root canal system during endodontic retreatment in oval-shaped canals.

Analysis of multiple cytokine polymorphisms in individuals with untreated deep carious lesions reveals IL1B (rs1143643) as a susceptibility factor for periapical lesion development.

INTRODUCTION: It has been proposed that individual genetic predisposition may contribute to persistent apical periodontitis. Cytokines are associated with levels of inflammation and are involved in caries, pulpal, and periapical tissue destruction. We hypothesized that polymorphisms in cytokine genes may contribute to an individual's increased susceptibility to apical tissue destruction in response to deep carious lesions. METHODS: Subjects with deep carious lesions with or without periapical lesions (≥3 mm) were recruited at the University of Pittsburgh, Pittsburgh, PA, and the University of Texas at Houston, Houston, TX. Genomic DNA samples of 316 patients were sorted into 2 groups: 136 cases with deep carious lesions and periapical lesions (cases) and 180 cases with deep carious lesions but no periapical lesions (controls). Nine single-nucleotide polymorphisms in IL1B, IL6, TNF, RANK, RANKL, and OPG genes were selected for genotyping. Genotypes were generated by end point analysis using TaqMan chemistry (Invitrogen, Carlsbad, CA) in a real-time polymerase chain reaction instrument. Allele and genotype frequencies were compared among cases and controls using the PLINK program (http://pngu.mgh.harvard.edu/purcell/plink/). Ninety-three human periapical granulomas and 24 healthy periodontal ligament tissues collected postoperatively were used for messenger RNA expression analyses of IL1B. RESULTS: A single-nucleotide polymorphism in IL1B (rs1143643) showed allelic (P = .02) and genotypic (P = .004) association with cases of deep caries and periapical lesions. We also observed altered transmission of IL1B marker haplotypes (P = .02) in these individuals. IL1B was highly expressed in granulomas (P < .001). CONCLUSIONS: Variations in IL1B may be associated with periapical lesion formation in individuals with untreated deep carious lesions. Future studies could help predict host susceptibility to developing periapical lesions.

Expression of heat shock proteins in periapical granulomas.

INTRODUCTION: Cells from virtually all organisms respond to a variety of stresses by the rapid synthesis of a highly conserved set of polypeptides termed heat shock proteins (HSPs). HSPs protect cells under adverse conditions such as infection, inflammation, and disease. We hypothesize that endodontic infection might result in an imbalance in the expression of heat shock genes, accounting for different clinical outcomes in periapical lesions. METHODS: We analyzed the expression of 44 HSPs genes using a pathway-specific real-time polymerase chain reaction array in 93 human periapical granulomas and 24 healthy periodontal ligament tissues collected postoperatively. Observed variations in the expression of HSP genes were also analyzed based on the classification of periapical granulomas as active or inactive. In addition, U937 cells were differentiated into macrophages, infected with different concentrations of purified Escherichia coli lipopolysaccharide (LPS), and used as templates for the HSP gene array. Protein expression was assessed by immunohistochemistry. RESULTS: The expression of HSP genes was significantly increased in granulomas compared with healthy periodontal ligament (P < .00001). Among the 44 HSP genes, DNAJC3, HSPA4, HSPA6, and HSPB1 showed the highest expression levels in both granulomas and LPS-treated macrophages. DNAJC3, HSPA6, and HSPB1 were highly expressed in active lesions, whereas HSPA4 expression was higher in inactive lesions (P < .005). Higher concentrations of LPS led to increased HSP expression in macrophages (P < .0001). Immunocytochemistry confirmed the expression and colocalization of HSPB1 and HSPA6 proteins in the cytoplasm of LPS-infected macrophages. CONCLUSIONS: The observed differential expression patterns of HSPs in periapical granulomas and LPS-infected macrophages suggest that HSP genes and proteins are involved in periapical lesion development and may account for different clinical outcomes. Understanding the role of the heat shock response might provide additional insights into the process of periapical lesion development.