Melbourne Dental School - Research Publications
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ItemTaxonomy of Oral Bacteria(ELSEVIER ACADEMIC PRESS INC, 2018-01-01)The oral cavity is a collection of diverse microenvironments, each inhabited by a community of microorganisms, the majority of which are bacteria and their phages. Given the appropriate conditions, some of these bacteria can cause destruction of the teeth or their supporting hard and soft tissues. For over 300 years microbiologists have been characterising these microbial communities, in both oral health and disease. In this chapter, we take the reader on a journey through time as we discuss the various methods that have been utilised in the characterisation of the bacteria calling the oral cavity home, and how the use of these methods has informed our understanding of oral bacterial communities and the diversity of their members.
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ItemAdolescent temperament dimensions as stable prospective risk and protective factors for salivary C-reactive protein(WILEY, 2018-02)OBJECTIVE: Temperament has associations with later physical health outcomes, yet there is a dearth of research exploring the connection between temperament and mechanisms that have known associations with these health outcomes. Recent research has delineated a connection between personality and inflammation during adulthood, but this association has not yet been studied in adolescent samples. DESIGN: We investigated whether stable adolescent temperament (averaged over two years), specifically effortful control and negative emotionality, provided a more robust prediction of inflammation as measured by salivary C-reactive protein (sCRP), than depressive symptoms. METHODS: Temperament and depressive symptoms were measured in a sample of sixty-three adolescents (37 males) when they were approximately 12 years old (mean age = 12.30, SD = 0.69) and again when they were approximately 14 years old (mean age = 14.84, SD = 0.49). Levels of sCRP were determined approximately 7 months later (mean = 6.77, SD = 2.99) when participants were approximately 15 years old (mean age = 15.49, SD = 0.49). RESULTS: Regression analyses revealed that effortful control (EC) was significantly associated with lower sCRP levels, while higher negative emotionality (NE) was significantly associated with higher sCRP levels. Furthermore, these associations were larger than those for depressive symptoms and were differentially impacted by the addition of covariates. Implications for the role of stable risk and protective factors in inflammatory processes are discussed. CONCLUSIONS: These findings are the first to show associations between adolescent temperament and inflammation. Furthermore, these findings extend previous personality research to temperamental research in a younger sample of adolescents. Statement of contribution What is already known? There is a large extant literature on the association between depressive symptoms and inflammation. There is a smaller extant literature on the association between personality and inflammation. No studies have examined how adolescent temperament traits may relate to inflammation. What does this study add? Longitudinal data collection over the course of 3 years in an adolescent sample. Addresses the question of whether temperament factors relate to inflammation. Temperament provides a more robust predictor of later inflammation than depressive symptoms.
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ItemA panel of microRNAs can be used to determine oral squamous cell carcinoma(WILEY, 2017-11)BACKGROUND: Subjective histopathology is currently used to diagnose oral squamous cell carcinoma (OSCC). We tested if abundances of a panel of microRNA could be an objective OSCC indicator. METHOD: Literature review enabled identification of 10 microRNAs associated with oral and head and neck malignancies. We extracted RNA from formalin-fixed paraffin-embedded biopsies; 20 each with OSCC, dysplasia, or histologically normal epithelium (HNE) and 10 with oral lichen planus (OLP). Relative abundances of microRNAs in HNE and OSCC were determined using reverse transcription and then real-time PCR with global mean normalization. MicroRNAs differentially expressed (test microRNA, T-miR) and non-differentially expressed (normalization microRNA, N-miR) were identified. The raw microRNA Cq data were incorporated in a developed algorithm that output a T-miR expression value (T-miREV) score. Raw Cq data from HNE, OSCC, dysplasia, and OLP samples were then used to test the algorithm scoring and OSCC classification. RESULTS: Four test and normalization microRNAs were identified. Algorithm output of T-mirEV >1 or <-1 indicated high and low OSCC probability score, respectively, and gave 88.9% sensitivity, 100% specificity, and 93.5% accuracy. Grouping high and intermediate T-mirEV scores (T-miREV ≥-1) resulted in sensitivity of 90%, specificity of 65%, and accuracy of 77.5% in OSCC classification. All 20 dysplasias and eight of 10 OLP had T-miREV ≥-1 indicating intermediate to high probability of malignant changes. CONCLUSION: A microRNA panel combined with our algorithm can identify tissue with probable oncogenic changes. IMPACT: The developed algorithm serves as a baseline for prospective trials, which may result in potential clinical utility.
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ItemRole of microbial biofilms in the maintenance of oral health and in the development of dental caries and periodontal diseases. Consensus report of group 1 of the Joint EFP/ORCA workshop on the boundaries between caries and periodontal disease(WILEY, 2017-03)BACKGROUND AND AIMS: The scope of this working group was to review (1) ecological interactions at the dental biofilm in health and disease, (2) the role of microbial communities in the pathogenesis of periodontitis and caries, and (3) the innate host response in caries and periodontal diseases. RESULTS AND CONCLUSIONS: A health-associated biofilm includes genera such as Neisseria, Streptococcus, Actinomyces, Veillonella and Granulicatella. Microorganisms associated with both caries and periodontal diseases are metabolically highly specialized and organized as multispecies microbial biofilms. Progression of these diseases involves multiple microbial interactions driven by different stressors. In caries, the exposure of dental biofilms to dietary sugars and their fermentation to organic acids results in increasing proportions of acidogenic and aciduric species. In gingivitis, plaque accumulation at the gingival margin leads to inflammation and increasing proportions of proteolytic and often obligately anaerobic species. The natural mucosal barriers and saliva are the main innate defence mechanisms against soft tissue bacterial invasion. Similarly, enamel and dentin are important hard tissue barriers to the caries process. Given that the present state of knowledge suggests that the aetiologies of caries and periodontal diseases are mutually independent, the elements of innate immunity that appear to contribute to resistance to both are somewhat coincidental.
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ItemOral swirl samples - a robust source of microRNA protected by extracellular vesicles(WILEY, 2017-04)BACKGROUND: MicroRNAs are small non-coding RNAs which are dysregulated in disease states, such as oral cancer. Extracellular vesicles, a potential source of microRNA, are found in saliva. OBJECTIVE: To demonstrate that a quantifiable amount of microRNA can be isolated from oral swirl samples. Additionally, we hypothesized that extracellular vesicles may protect contained microRNA from degradation in these samples. METHOD: A polyethylene glycol-based precipitation was used for extracellular vesicle enrichment of oral swirl samples. Comparison was made between samples treated with and without RNase. Further, samples from three subjects were exposed to a range of conditions over 7 days and assessed for presence of microRNA by reverse-transcription quantitative PCR. Extracellular vesicles from samples were identified under transmission electron microscopy. RESULTS: An adequate quantity of microRNA for qPCR analysis was extractable from samples despite exposure to conditions under which degradation of RNA would be expected. CONCLUSION: A technique was developed to isolate an adequate quantity of microRNA for analysis from oral swirl samples. Extracellular vesicle-associated microRNA may be protected from degradation. This technique moves towards chairside application of translational microRNA research in the field of oral cancer prognostics.
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ItemC-Terminal Modification and Multimerization Increase the Efficacy of a Proline-Rich Antimicrobial Peptide(WILEY-V C H VERLAG GMBH, 2017-01-05)Two series of branched tetramers of the proline-rich antimicrobial peptide (PrAMP), Chex1-Arg20, were prepared to improve antibacterial selectivity and potency against a panel of Gram-negative nosocomial pathogens including Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii and Pseudomonas aeruginosa. First, tetramerization was achieved by dithiomaleimide (DTM) conjugation of two C-terminal-cysteine bearing dimers that also incorporated C-terminal peptide chemical modification. DTM-linked tetrameric peptides containing a C-terminal hydrazide moiety on each dimer exhibited highly potent activities in the minimum inhibitory concentration (MIC) range of 0.49-2.33 μm. A second series of tetrameric analogues with C-terminal hydrazide modification was prepared by using alternative conjugation linkers including trans-1,4-dibromo-2-butene, α,α'-dibromo-p-xylene, or 6-bismaleimidohexane to determine the effect of length on activity. Each displayed potent and broadened activity against Gram-negative nosocomial pathogens, particularly the butene-linked tetrameric hydrazide. Remarkably, the greatest MIC activity is against P. aeruginosa (0.77 μm/8 μg mL-1 ) where the monomer is inactive. None of these peptides showed any cytotoxicity to mammalian cells up to 25 times the MIC. A diffusion NMR study of the tetrameric hydrazides showed that the more active antibacterial analogues were those with a more compact structure having smaller hydrodynamic radii. The results show that C-terminal PrAMP hydrazidation together with its rational tetramerization is an effective means for increasing both diversity and potency of PrAMP action.
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ItemCandida virulence and ethanol-derived acetaldehyde production in oral cancer and non-cancer subjects(WILEY, 2016-11)OBJECTIVES: To compare biofilm-forming ability, hydrolytic enzymes and ethanol-derived acetaldehyde production of oral Candida isolated from the patients with oral cancer and matched non-oral cancer. MATERIAL AND METHODS: Fungal biofilms were grown in RPMI-1640 medium, and biofilm mass and biofilm activity were assessed using crystal violet staining and XTT salt reduction assays, respectively. Phospholipase, proteinase, and esterase production were measured using agar plate method, while fungal acetaldehyde production was assessed via gas chromatography. RESULTS: Candida isolated from patients with oral cancer demonstrated significantly higher biofilm mass (P = 0.031), biofilm metabolic activity (P < 0.001), phospholipase (P = 0.002), and proteinase (P = 0.0159) activity than isolates from patients with non-oral cancer. High ethanol-derived acetaldehyde-producing Candida were more prevalent in patients with oral cancer than non-oral cancer (P = 0.01). In univariate regression analysis, high biofilm mass (P = 0.03) and biofilm metabolic activity (P < 0.001), high phospholipase (P = 0.003), and acetaldehyde production ability (0.01) were significant risk factors for oral cancer; while in the multivariate regression analysis, high biofilm activity (0.01) and phospholipase (P = 0.01) were significantly positive influencing factors on oral cancer. CONCLUSION: These data suggest a significant positive association between the ability of Candida isolates to form biofilms, to produce hydrolytic enzymes, and to metabolize alcohol to acetaldehyde with their ability to promote oral cancer development.
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ItemThe assessment of the robustness of microRNAs from oral cytological scrapings(WILEY, 2017-05)BACKGROUND: Sampling of suspect oral lesions in the general dental clinic may increase early carcinoma detection thus oral cancer survival rates. One means of lesion sampling that is an alternative to incisional biopsy is cytological scraping. MicroRNA alterations are also being explored as a means of diagnosing carcinoma as an alternative to histopathology. METHODS: We obtained cytological scrapings using 10 strokes ('light') or 40 strokes ('heavy') from the buccal mucosa of one healthy subject using a dermatological curette. MicroRNA was isolated from oral cytological scrapings immediately, or the scrapings were stored in buffer or RNA later, at 4°C, room temperature or 36°C, from 1 to 7 days prior to RNA isolation. All scrape comparisons and test conditions were conducted in triplicate. MicroRNAs were measured using qRT-PCR. RESULTS: MicroRNAs can be obtained from cytological scrapings independent of the number of strokes and can be measured using qRT-PCR after storage under all conditions tested. CONCLUSION: MicroRNAs are robust to a wide range of storage conditions that bodes well for use of cytological scrapings to be of use in a clinical setting as a chair side sampling method for suspect oral lesions.
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ItemImportance of bioavailable calcium in fluoride dentifrices for enamel remineralization(ELSEVIER SCI LTD, 2018-11)OBJECTIVES: To compare remineralization of enamel subsurface lesions by fluoride dentifrices with added calcium in a double-blind, randomized, cross-over, in situ study. METHODS: Human enamel with subsurface lesions were prepared and inserted into intra-oral appliances worn by volunteers. A slurry (1 g toothpaste/4 ml H2O) was rinsed for 60 s, 4 times per day for 14 days. Seven toothpastes were tested: (i) 1450 ppm F (NaF), (ii) 5000 ppm F (NaF), (iii) 1450 ppm F (MFP) with calcium sodium phosphosilicate (CSP), (iv) 1450 ppm F (MFP) with CaCO3/Arg, (v) 1150 ppm F (SnF2) with amorphous calcium phosphate (ACP), (vi) 1100 ppm F (NaF) with casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) and (vii) 5000 ppm F (NaF) with functionalized tri-calcium phosphate (TCP). Total (acid soluble) and bioavailable (water soluble) calcium, inorganic phosphate and fluoride levels of the dentifrices were measured using ion chromatography (F/MFP) and spectrophotometry (Ca and inorganic phosphate). Enamel lesion mineral content was measured using transverse microradiography. Data were statistically analysed using a linear mixed model. RESULTS: All calcium and fluoride containing toothpastes released > 90% of bioavailable fluoride and were superior to the respective fluoride alone toothpastes in remineralization of enamel subsurface lesions. The level of remineralization followed the order: CPP-ACP/1l00 ppm F > ACP/1150 ppm F = TCP/5000 ppm F > 5000 ppm F = CaCO3/Arg/1450 ppm F = CSP/1450 ppm F > 1450 ppm F. Bioavailable calcium levels significantly correlated with enhanced remineralization of enamel subsurface lesions. CONCLUSIONS: Bioavailable calcium in fluoride dentifrices enhanced remineralization of enamel subsurface lesions.
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ItemArchitectural Effects of Star-Shaped "Structurally Nanoengineered Antimicrobial Peptide Polymers" (SNAPPs) on Their Biological Activity(WILEY, 2018-11)In this work, the effect of two key structural parameters, number of arms and arm length, of star-shaped "structurally nanoengineered antimicrobial peptide polymers" (SNAPPs) on their antimicrobial activity and biocompatibility, is investigated. A library of star-shaped SNAPPs is prepared, containing varying arm numbers and arm lengths. Antimicrobial assays are then performed to assess the capacity of the SNAPPs to disrupt the membrane, inhibit the growth, and kill pathogenic bacteria. A major finding of the study is that increasing arm number and length of SNAPPs enhanced antimicrobial activity, which can be respectively attributed to the higher local concentrations of polypeptide arms and increased α-helical content. SNAPP architecture is shown to affect the bacteria membrane state and therefore mechanism of killing. Two more potent structures with up to twice the antimicrobial activity of the previously reported SNAPP are discovered in this process. Toxicities of the SNAPPs also increase with arm number and arm length, however therapeutic index calculations identified a 16-arm SNAPP and an easier to prepare 4-arm SNAPP as the best therapeutic agents. The biocompatibility of the SNAPP with the best biological activity is also evaluated in vivo, showing no markers of systemic damage in mice.