Oral Biology

High CD3+ lymphocytes, low CD66b+ neutrophils, and scarce tumor budding in the invasive front of lip squamous cell carcinomas Publication date: August 2019 Source: Archives of Oral Biology, Volume 104 Author(s): Karine Duarte da Silva, Patrícia Carlos Caldeira, Alessandro Menna Alves, Ana Carolina Uchoa Vasconcelos, Ana Paula Neutzling Gomes, Maria Cássia Ferreira de Aguiar, Sandra Beatriz Chaves Tarquinio Abstract Objective This study aimed to evaluate tumor budding (TB) and quantify the neutrophilic and lymphocytic infiltration in the invasive front of lip squamous cell carcinomas. In addition, the associations between these features and the histopathological grading in the invasive front were analyzed. Design A total of 43 lip squamous cell carcinoma surgical specimens were included and classified in accordance with a histological invasive front grading system. Immunohistochemistry was performed for CD66b and CD3 for the evaluation of neutrophils and T lymphocytes, respectively, in the invasive front of the tumors. Tumor budding was evidenced by AE1/AE3 staining. Results Thirty-six (83.7%) of the tumors were well-differentiated tumors. Eleven (25.6%) of the cases exhibited high-intensity tumor budding. There were low neutrophil and high T lymphocyte infiltrations in the invasive front, leading to a low neutrophil/T lymphocyte ratio in the same region. Moreover, we found an association between tumor budding and the pattern of invasion, and between the CD3+ cell count and the inflammatory infiltrate (p < 0.05). Conclusions The low neutrophil and high T lymphocyte infiltration in the invasive front, and the few high-intensity tumor budding cases are in accordance with the histopathological features of well-differentiated lip tumors. If these characteristics remain in lip squamous cell carcinomas with more aggressive histopathological features, it deserves to be investigated.

Cannabidiol, cannabinol and their combinations act as peripheral analgesics in a rat model of myofascial pain Publication date: August 2019 Source: Archives of Oral Biology, Volume 104 Author(s): Hayes Wong, Brian E. Cairns Abstract Objective This study investigated whether local intramuscular injection of non-psychoactive cannabinoids, cannabidiol (CBD), cannabinol (CBN), cannabichromene (CBC) and their combinations can decrease nerve growth factor (NGF)-induced masticatory muscle sensitization in female rats. Design In awake rats, changes in mechanical sensitivity induced by intramuscular injection of NGF and cannabinoids were measured by applying an electronic von Frey hair over the masseter muscle to measure the withdrawal response. The effect of CBD (5 mg/ml) and CBN (1 mg/ml) or their combinations CBD/CBN (1:1 mg/ml or 5:1 mg/ml) were assessed. To confirm a peripheral action, electrophysiological experiments were undertaken in anesthetized rats to examine whether intramuscular injections of CBD (5 mg/ml) and CBN (1 mg/ml) altered the mechanical threshold of masticatory muscle mechanoreceptors. Results In behavioral experiments, CBD (5 mg/ml) or CBN (1 mg/ml) decreased NGF-induced mechanical sensitization. Combinations of CBD/CBN induced a longer-lasting reduction of mechanical sensitization than either compound alone. No significant change in mechanical withdrawal threshold was observed in the contralateral masseter muscles and no impairment of motor function was found with the inverted screen test after any of the treatments. Consistent with behavioral results, CBD (5 mg/ml), CBN (1 mg/ml) and the combination of CBD/CBN (1:1 mg/ml) increased the mechanical threshold of masseter muscle mechanoreceptors. However, combining CBD/CBN (5:1 mg/ml) at a higher ratio reduced the duration of this effect. This may indicate an inhibitory effect of higher concentrations of CBD on CBN. Conclusions These results suggest that peripheral application of these non-psychoactive cannabinoids may provide analgesic relief for chronic muscle pain disorders such as temporomandibular disorders and fibromyalgia without central side effects.

Activity and distribution pattern of enzymes in the in-situ pellicle of children Publication date: August 2019 Source: Archives of Oral Biology, Volume 104 Author(s): Susann Hertel, Annika Schulz, Roman Lang, Thomas Hofmann, Belinda König, Matthias Hannig, Christian Hannig Abstract Objective This study investigated, for the first time, pellicle enzymes with respect to their activity, distribution and fluorescence pattern in children with different caries experience. Design In-situ pellicles were collected from 41 children (aged 4–6 years) with different caries status; 17 of them were caries-free (dmf = 0), 12 had dental restorations but no current caries (dmf ≥ 2) and 12 had at least two carious lesions (dmf ≥ 2). Bovine enamel samples were fixed on individual upper jaw braces for pellicle formation. After 30 min of intraoral exposure, the pellicle and saliva samples were analysed for the activities of amylase, lysozyme, peroxidase and glucosyltransferase (GTF). The distribution of these enzymes, including GTF-isoforms B, C and D, and the pellicle ultrastructure were examined by gold-immunolabelling and transmission electron microscopy (TEM). Furthermore, interactions between pellicle enzymes and adherent bacteria were visualised using combined fluorescence and immunofluorescence labelling. Results There were no significant differences in the pellicle enzyme activities between the study groups. TEM analysis revealed the absence of GTF C and D in the pellicle of caries-active children. Amylase, peroxidase and GTF-isoforms showed a random distribution within the pellicle layer; lysozyme was found in the form of clusters. A similar ultrastructural pattern was observed for all subjects. Fluorescence labelling technique enabled visualisation of all enzymes, except for GTF B. Conclusion Pellicle enzyme activities and ultrastructure are not associated with children's caries status. Further investigation is needed to assess the influence of individual GTF-isoforms on caries susceptibility in children.

Whole metagenomic shotgun sequencing of the subgingival microbiome of diabetics and non-diabetics with different periodontal conditions Publication date: August 2019 Source: Archives of Oral Biology, Volume 104 Author(s): Roberto Farina, Mattia Severi, Alberto Carrieri, Elena Miotto, Silvia Sabbioni, Leonardo Trombelli, Chiara Scapoli Abstract Objective The aim of this study was to use high-resolution whole metagenomic shotgun sequencing to characterize the subgingival microbiome of patients with/without type 2 Diabetes Mellitus and with/without periodontitis. Design Twelve subjects, falling into one of the four study groups based on the presence/absence of poorly controlled type 2 Diabetes Mellitus and moderate-severe periodontitis, were selected. For each eligible subject, subgingival plaque samples were collected at 4 sites, all representative of the periodontal condition of the individual (i.e., non-bleeding sulci in subjects without a history of periodontitis, bleeding pockets in patients with moderate-severe periodontitis). The subgingival microbiome was evaluated using high-resolution whole metagenomic shotgun sequencing. Results The results showed that: (i) the presence of type 2 Diabetes Mellitus and/or periodontitis were associated with a tendency of the subgingival microbiome to decrease in richness and diversity; (ii) the presence of type 2 Diabetes Mellitus was not associated with significant differences in the relative abundance of one or more species in patients either with or without periodontitis; (iii) the presence of periodontitis was associated with a significantly higher relative abundance of Anaerolineaceae bacterium oral taxon 439 in type 2 Diabetes Mellitus patients. Conclusions Whole metagenomic shotgun sequencing of the subgingival microbiome was extremely effective in the detection of low-abundant taxon. Our results point out a significantly higher relative abundance of Anaerolineaceae bacterium oral taxon 439 in patients with moderate to severe periodontitis vs patients without history of periodontitis, which was maintained when the comparison was restricted to type 2 diabetics.

Applications of the oral microbiome in personalized dentistry Publication date: August 2019 Source: Archives of Oral Biology, Volume 104 Author(s): Georgios N. Belibasakis, Nagihan Bostanci, Philip D. Marsh, Egija Zaura Abstract Objective In the era of personalized medicine, it is imperative that oral health is integrated into this concept. The oral cavity fosters a highly individualized microbiome that has evolved to promote oral health, and which exists in a dynamic balance with the host. Microecological changes to the biology of the mouth [e.g. in the host diet and lifestyle, or status of the immune system] may drive deleterious shifts in the composition or metabolic activity of the oral microbiome ['dysbiosis']. This review aims to explore how knowledge of the oral microbiome may be utilized for personalized dentistry at the point-of-care. Design This is a comprehensive narrative review of the literature, summarizing the perspectives of the authors. Results The huge increase in recent knowledge on the ecology and microbiology of the oral cavity generated by 'OMIC' technologies may indeed be clinically translated to support patient care, in terms of prevention, monitoring, risk classification or early diagnosis. The identified clinical applications may not only include dental caries and periodontal disease, but also dental implants and orthodontics. Population-based applications may include systemic health, pregnancy and elderly populations. Conclusions Applications of selected oral microbiome and host-related biochemical parameters [e.g. the saliva proteome] for personalized dentistry can be customized for different clinical applications or individual populations, at point-of-care hubs.

Staircase-pattern neonatal line in human deciduous teeth is associated with tooth type Publication date: August 2019 Source: Archives of Oral Biology, Volume 104 Author(s): Jaana Hurnanen, Matti Sillanpää, Marja-Leena Mattila, Eliisa Löyttyniemi, Carsten Witzel, Jaana Rautava Abstract Objective The staircase (Sc) pattern enamel microstructure is an expression of an impaired ameloblast function. It has been reported to appear in the neonatal line (NNL), the accentuated stria evincing live birth in deciduous tooth enamel. Our objective was to investigate the prevalence of Sc NNL in deciduous tooth types and its possible association with perinatal circumstances. Design Sc in the NNL of 88 teeth, a collection derived from a long-term, prospectively followed population cohort, was recorded with linear polarised transmitted light and analysed for tooth type, duration and mode of delivery, and pain medication used during labour. Results Sc prevalence in the NNL differed highly significantly between tooth types (p < 0.0001). An increase in Sc prevalence was significantly associated with an increased ratio of prenatal to total crown enamel (p < 0.001), when buccal and lingual crown walls were analysed separately. No significant association was found between Sc prevalence and duration or mode of delivery or pain-relieving medication (p = 0.57, p = 0.65, p = 0.58, respectively). Conclusion This research indicates that the NNL location within tooth crown enamel has a strong impact on microstructural changes along the NNL. Considering our results of Sc prevalence, deciduous canines, having the least Sc appearance, could be used in studies that aim to investigate factors associated with NNL width. In addition, Sc prevalence variation in first deciduous molars might enable to investigation of physiological stressors strong enough to cause ameloblast impairment, such as Sc.

Salivary gland metabolism in an animal model of chronic kidney disease Publication date: August 2019 Source: Archives of Oral Biology, Volume 104 Author(s): R.A. Carvalho, A.C. Romero, F.K. Ibuki, F.N. Nogueira Abstract Objective The aim of this study was to determine the effects of experimental CKD into the metabolism of parotid and submandibular glands of rats. CKD was induced by 5/6 nephrectomy. Design Serum analyses of BUN (Blood Urea Nitrogen) and creatinine concentrations were performed. Major salivary glands metabolism was investigated in vivo, both at rest and during salivary stimulation conditions by NMR isotopomer analysis, using [U-13C]glucose as metabolic tracer. Results CKD increases BUN and serum creatinine concentrations (p < 0.001). Multiple metabolic alterations were detected in the parotid glands of this animal model, including decreased concentrations of alanine (p < 0.05) and creatine (p < 0.05) and increased lactate/alanine ratios (p < 0.05). The salivary stimulus fostered accumulations of acetate at both analyzed glands of the CKD model (p < 0.05), indicative of disruption of the oxidative metabolic process. Conclusions Experimental CKD induced by 5/6 nephrectomy altered the parotid salivary gland function, since glucose metabolism is clearly affected after stimulation for salivation in this gland.

Effects of Candida albicans infection on defense effector secretion by human oral mucosal epithelial cells Publication date: July 2019 Source: Archives of Oral Biology, Volume 103 Author(s): Fan Huang, Yuefeng Song, Wei Chen, Qin Liu, Qiong Wang, Weida Liu, Xiang Wang, Wenmei Wang Abstract Objective The aim of this study was to investigate the effects of Candida albicans on the production of defense effector molecules by human oral mucosal epithelial cells in vitro. Design Immortalized human oral mucosal epithelial (Leuk-1) cells and C. albicans strain 5314 were cocultured at different cell-to-C. albicans ratios. The viability of Leuk-1 cells was determined by MTT and RTCA measurements. The secretory levels of multiple defense effector molecules were determined by Enzyme-linked immunosorbent assay (ELISA). Results Our results indicated that C. albicans significantly decreased the secretion of IgG, cystatin C, lactoferrin, and TGF-β1 in a dose-dependent manner and remarkably reduced the production of IgA independent of the cell-to-C. albicans ratio. However, C. albicans clearly enhanced the secretion of IgM, galectin-3, P-selectin, granzyme B and perforin. Conclusion These results suggest that C. albicans may exert a regulatory role in the defense response of oral mucosal epithelial cells by altering secretory levels of defense effector molecules.

Liposome-coated nano doxorubicin induces apoptosis on oral squamous cell carcinoma CAL-27 cells Publication date: July 2019 Source: Archives of Oral Biology, Volume 103 Author(s): Ehab S. Abd El-Hamid, Amira M. Gamal-Eldeen, Abdelrahman M. Sharaf Eldeen Abstract Objective This study aims to investigate the apoptotic effect of Doxorubicin and its nano-formulated form (Doxil) on oral squamous cell carcinoma CAL-27 cells. Design Cell viability using MTT assay, mode of cell death using fluorescence analysis, expression of the apoptotic marker caspase-3 using indirect ELISA technique and expression of C-Myc gene using reverse transcriptase and real time PCR. Results Doxil treatment resulted in a higher percentage of apoptotic cells than doxorubicin treatment, while doxorubicin treatment resulted in a higher percentage of necrotic cells than Doxil treatment. Doxil-treated cells exhibited 3.38-fold higher caspase-3 levels than control cells, while doxorubicin significantly increased caspase-3 levels by 2.72-fold. The percentage of C-Myc mRNA inhibition was 27% in doxorubicin-treated cells and 41% in Doxil-treated cells. Conclusions Doxil exerted a higher apoptotic effect on CAL-27 cells compared to doxorubicin. It showed a higher increase in capase-3 level than doxorubicin and also exerted a more percentage of C-Myc mRNA inhibition.

BMP4 mutations in tooth agenesis and low bone mass Publication date: July 2019 Source: Archives of Oral Biology, Volume 103 Author(s): Miao Yu, Hao Wang, Zhuangzhuang Fan, Chencheng Xie, Haochen Liu, Yang Liu, Dong Han, Sing-Wai Wong, Hailan Feng Abstract Objective To identify an uncommon genetic cause of tooth agenesis (TA) by utilizing whole exome sequencing (WES) and targeted Sanger sequencing in a cohort of 120 patients with isolated TA. Design One deleterious mutation in the gene encoding bone morphogenetic protein 4 (BMP4) was identified in 6 unrelated patients with TA by WES. After that, the coding exons of BMP4 were examined in 114 TA patients using Sanger sequencing. Dual-energy X-ray absorptiometry (DEXA) was used to measure the bone mineral density of patients who carried a BMP4 mutation. Finally, preliminary functional studies of two BMP4 mutants were performed. Results We detected 3 novel missense mutations (c.58 G > A: p.Gly20Ser, c.326 G > T: p.Arg109Leu and c.614 T > C: p.Val205Ala) and 1 reported mutation in the BMP4 gene among 120 TA probands. The previously reported BMP4 mutation (c.751C > T: p.His251Tyr) was associated with urethra and eye anomalies. By extending the pedigrees, we determined that the tooth phenotypes had an autosomal dominant inheritance pattern, as individuals carrying a BMP4 mutation exhibit different types of dental anomalies. Interestingly, we observed that patients harboring a BMP4 mutation manifested early onset osteopenia or osteoporosis. Further in vitrofunctional assays demonstrated that two BMP4 mutants resulted in a decreased activation of Smad signaling. Therefore, a loss-of-function in BMP4 may contribute to the clinical phenotypes seen in this study. Conclusions We identified 4 mutations in the BMP4 gene in 120 TA patients. To our knowledge, this is the first study to describe human skeletal diseases associated with BMP4 mutations.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
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6948891480

alsfakia@gmail.com