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Item Effect of nicotine on cariogenic virulence of Streptococcus mutans(Springer, 2016-11) Li, Mingyun; Huang, Ruijie; Zhou, Xuedong; Qiu, Wei; Xu, Xin; Gregory, Richard L.; Department of Biomedical and Applied Sciences, IU School of DentistryNicotine has well-documented effects on the growth and colonization of Streptococcus mutans. This study attempts to investigate the effects of nicotine on pathogenic factors of S. mutans, such as the effect on biofilm formation and viability, expression of pathogenic genes, and metabolites of S. mutans. The results demonstrated that addition of nicotine did not significantly influence the viability of S. mutans cells. The biofilms became increasingly compact as the concentrations of nicotine increased. The expression of virulence genes, such as ldh and phosphotransferase system (PTS)-associated genes, was upregulated, and nlmC was upregulated significantly, while ftf was downregulated. The lactate concentration of S. mutans grown in 1 mg/mL of nicotine was increased up to twofold over either biofilm or planktonic cells grown without nicotine. Changes in the metabolites involved in central carbon metabolism from sucrose indicated that most selected metabolites were detectable and influenced by increased concentrations of nicotine. This study demonstrated that nicotine can influence the pathogenicity of S. mutans and may lead to increased dental caries through the production of more lactate and the upregulation of virulence genes.Item The effects of radicular dentine treated with double antibiotic paste and ethylenediaminetetraacetic acid on the attachment and proliferation of dental pulp stem cells(Wiley, 2015-10) Kim, Ki Wan; Yassen, Ghaeth H.; Ehrlich, Ygal; Spolnik, Kenneth; Platt, Jeffrey A.; Windsor, L. Jack; Department of Biomedical and Applied Sciences, IU School of DentistryAim This study explored the effects of dentine treated with two concentrations of double antibiotic paste (DAP) and ethylenediaminetetraacetic acid (EDTA) on the attachment and proliferation of dental pulp stem cells (DPSCs). Materials and Methods Radicular dentine samples were prepared with identical dimensions and randomized into six groups (n = 4). Four groups were treated with double antibiotic paste (DAP) at concentrations of 500 mg ml−1 or 1 mg ml−1 with or without EDTA. The other two groups were treated with EDTA only or received no treatment. DPSCs were seeded on each dentine sample (10 000 cells per sample). Lactate dehydrogenase activity assays were used to calculate the attached DPSCs after 1 day of incubation. Water soluble tetrazolium assays were performed to investigate DPSCs proliferation on the treated dentine samples after three additional days of incubation. Two-way anova followed by Tukey–Kramer tests was used for statistical analyses (α = 0.05). Results Dentine treated with 1 or 500 mg ml−1 of DAP followed by EDTA caused significant increases in DPSCs attachment compared to the dentine treated with the DAP alone. The 500 mg ml−1 of DAP with or without EDTA caused significant reductions in DPSCs proliferation. However, the treatment of dentine with 1 mg ml−1 of DAP did not have significant negative effects on DPSCs proliferation regardless of the use of EDTA. Conclusion The use of 1 mg ml−1 of DAP followed by 10 min of irrigation with EDTA in endodontic regeneration procedure may have no negative effects on the attachment and proliferation of DPSCs.Item Evaluation of selected properties of a new root repair cement containing surface pre-reacted glass ionomer fillers(Springer, 2016) Yassen, Ghaeth H.; Huang, Ruijie; Al-Zain, Afnan; Yoshida, Takamitsu; Gregory, Richard L.; Platt, Jeffrey A.; Department of Biomedical and Applied Sciences, IU School of DentistryObjective This study evaluated selected properties of a prototype root repair cement containing surface pre-reacted glass ionomer fillers (S-PRG) in comparison to mineral trioxide aggregate (MTA) and intermediate restorative material (IRM). Materials and methods The antibacterial effect of S-PRG, MTA, and IRM cements was tested against Porphyromonas gingivalis and Enterococcus faecalis after 1 and 3 days of aging of the cements. The set cements were immersed in distilled water for 4 h to 28 days, and ion-releasing ability was evaluated. Initial and final setting times of all cements were evaluated using Gilmore needles. The push-out bond strength between radicular dentin and all cements was tested at different levels of the roots. Results S-PRG and IRM cements, but not MTA cement, demonstrated significant antibacterial effect against P. gingivalis. All types of cements exhibited significant antibacterial effect against E. faecalis without being able to eliminate the bacterium. S-PRG cement provided continuous release of fluoride, strontium, boron, sodium, aluminum, and zinc throughout all tested time points. Both initial and final setting times were significantly shorter for S-PRG and IRM cements in comparison to MTA. The push-out bond strength was significantly lower for S-PRG cement in comparison to MTA and IRM at coronal and middle levels of the roots. Conclusions S-PRG cement demonstrated significant antibacterial effects against endodontic pathogens, multiple ion-releasing ability, relatively short setting time, and low bonding strength. Clinical relevance S-PRG cement can be used as a one-visit root repair material with promising antibacterial properties and ion-releasing capacity.Item Fracture Resistance and Microleakage of Endocrowns Utilizing Three CAD-CAM Blocks(2015-03) El-Damanhoury, Hatem M.; Haj-Ali, Reem N.; Platt, Jeffrey A.; Department of Biomedical and Applied Sciences, IU School of DentistryThis study assessed marginal leakage and fracture resistance of computer-aided design/computer-aided manufacturing (CAD/CAM) fabricated ceramic crowns with intracoronal extensions into the pulp chambers of endodontically treated teeth (endocrowns) using either feldspathic porcelain (CEREC Blocks [CB], Sirona Dental Systems GmbH, Bensheim, Germany), lithium disilicate (e.max [EX], Ivoclar Vivadent, Schaan, Liechtenstein), or resin nanoceramic (Lava Ultimate [LU], 3M ESPE, St Paul, MN, USA).). Thirty extracted human permanent maxillary molars were endodontically treated. Standardized preparations were done with 2-mm intracoronal extensions of the endocrowns into the pulp chamber. Teeth were divided into three groups (n=10); each group was restored with standardized CAD/CAM fabricated endocrowns using one of the three tested materials. After cementation with resin cement, specimens were stored in distilled water at 37°C for one week, subjected to thermocycling, and immersed in a 5% methylene-blue dye solution for 24 hours. A compressive load was applied at 35 degrees to long axis of the teeth using a universal testing machine until failure. Failure load was recorded, and specimens were examined under a stereomicroscope for modes of failure and microleakage. Results were analyzed using one-way analysis of variance and Bonferroni post hoc multiple comparison tests (α=0.05). LU showed significantly (p<0.05) higher fracture resistance and more favorable fracture mode (ie, fracture of the endocrown without fracture of tooth) as well as higher dye penetration than CB and EX. In conclusion, although using resin nanoceramic blocks for fabrication of endocrowns may result in better fracture resistance and a more favorable fracture mode than other investigated ceramic blocks, more microleakage may be expected with this material.Item Green tea catechin inhibits the activity and neutrophil release of Matrix Metalloproteinase-9.(Elsevier, 2016-10) Kim-Park, Wan K.; Allam, Eman S.; Palasuk, Jadesada; Kowolik, Michael; Park, Kichuel K.; Windsor, L. Jack; Department of Biomedical and Applied Sciences, IU School of DentistryGreen tea (Camellia sinensis; 綠茶 lǜ chá) extracts have been shown to possess anti-oxidant and anti-inflammatory effects in various cell types. Green tea extract (GTX) has been shown to significantly inhibit the activity of collagenase-3 (matrix metalloproteinase-13 (MMP-13)) in vitro. MMPs, such asItem In Vitro effects of Plantago major extract, aucubin and baicalein on Candida albicans biofilm formation, metabolic activity and cell surface hydrophobicity(Wiley, 2015) Shirley, Karina Pezo; Windsor, L. Jack; Eckert, George J.; Gregory, Richard L.; Department of Biomedical and Applied Sciences, IU School of DentistryPurpose To determine the in vitro effectiveness of Plantago major extract, along with two of its active components, aucubin and baicalein, on the inhibition of Candida albicans growth, biofilm formation, metabolic activity, and cell surface hydrophobicity. Materials and Methods Twofold dilutions of P. major, aucubin, and baicalein were used to determine the minimum inhibitory concentration (MIC), minimum fungicidal concentration (MFC), and the minimum biofilm inhibitory concentration (MBIC) of each solution. Separately, twofold dilutions of P. major, aucubin, and baicalein were used to determine the metabolic activity of established C. albicans biofilm using a 2,3-bis (2- methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-carboxanilide reduction assay. Twofold dilutions of P. major, aucubin, and baicalein were used to determine the cell surface hydrophobicity of treated C. albicans biofilm by a two-phase assay using hexadecane. The hydrophobicity percentage of the cell surface was then calculated. A mixed-model ANOVA test was used for intergroup comparisons. Results The MICs of P. major extract (diluted 1:2 to 1:8), aucubin (61 to 244 μg/ml), and baicalein (0.0063 to 100 μg/ml) on the total growth of C. albicans were noticeable at their highest concentrations, and the inhibition was dose dependent. The MFC was evaluated after 48 hours of incubation, and aucubin (244 μg/ml) exhibited a strong fungicidal activity at its highest concentration against C. albicans growth. The MBIC indicated no growth or reduced growth of C. albicans biofilm at the highest concentrations of aucubin (61 to 244 μg/ml) and baicalein (25 to 100 μg/ml). Similarly, the effects of these reagents on C. albicans biofilm metabolic activity and hydrophobicity demonstrated high effectiveness at their highest concentrations. Conclusion P. major extract, aucubin, and baicalein caused a dose-dependent reduction on the total growth, biofilm formation, metabolic activity, and cell surface hydrophobicity of C. albicans. This demonstrates their effectiveness as antifungals and suggests their promising potential use as solutions for C. albicans biofilm-related infections.Item Inhibitory effect of gels loaded with a low concentration of antibiotics against biofilm formation by Enterococcus faecalis and Porphyromonas gingivalis(2015-09) Algarni, Amnah A.; Yassen, Ghaeth H.; Gregory, Richard L.; Department of Biomedical and Applied Sciences, IU School of DentistryWe explored longitudinally the inhibitory effect of gels loaded with 1 mg/mL modified triple antibiotic paste (MTAP) or double antibiotic paste (DAP) against biofilm formation by Enterococcus faecalis and Porphyromonas gingivalis. Methylcellulose-based antibiotic gels of MTAP (ciprofloxacin, metronidazole and clindamycin) and DAP (ciprofloxacin and metronidazole) were prepared at a concentration of 1 mg/mL. Individually cultured E. faecalis and P. gingivalis bacterial suspensions were treated with MTAP, DAP, or placebo (vehicle only) gels at different dilutions and allowed to grow in 96-well microtiter plates. Untreated bacterial suspensions served as a negative control. Crystal violet assays were used to evaluate biofilm formation after 48 h. The ability of the gels to inhibit biofilm formation was determined immediately, and at 1 month and 3 months after the gels had been prepared. Data were analyzed using a mixed-model ANOVA. The MTAP and DAP gels significantly reduced biofilm formation by both bacterial species at all time points, regardless of the tested dilution. No-significant differences in biofilm-inhibitory effects between MTAP and DAP gels were observed at the majority of the tested dilutions through various time points. Gels loaded with 1 mg/mL MTAP and DAP demonstrated a significant antibiofilm effect against E.faecalis and P. gingivalis.Item Limitations of a morphological criterion of adaptive inference in the fossil record(Wiley, 2015) Ravosa, Matthew J.; Menegaz, Rachel A.; Scott, Jeremiah E.; Daegling, David J.; McAbee, Kevin R.; Department of Biomedical and Applied Sciences, IU School of DentistryExperimental analyses directly inform how an anatomical feature or complex functions during an organism's lifetime, which serves to increase the efficacy of comparative studies of living and fossil taxa. In the mammalian skull, food material properties and feeding behaviour have a pronounced influence on the development of the masticatory apparatus. Diet-related variation in loading magnitude and frequency induce a cascade of changes at the gross, tissue, cellular, protein and genetic levels, with such modelling and remodelling maintaining the integrity of oral structures vis-à-vis routine masticatory stresses. Ongoing integrative research using rabbit and rat models of long-term masticatory plasticity offers unique insight into the limitations of functional interpretations of fossilised remains. Given the general restriction of the palaeontological record to bony elements, we argue that failure to account for the disparity in the hierarchical network of responses of hard versus soft tissues may overestimate the magnitude of the adaptive divergence that is inferred from phenotypic differences. Second, we note that the developmental onset and duration of a loading stimulus associated with a given feeding behaviour can impart large effects on patterns of intraspecific variation that can mirror differences observed among taxa. Indeed, plasticity data are relevant to understanding evolutionary transformations because rabbits raised on different diets exhibit levels of morphological disparity comparable to those found between closely related primate species that vary in diet. Lastly, pronounced variation in joint form, and even joint function, can also characterise adult conspecifics that differ solely in age. In sum, our analyses emphasise the importance of a multi-site and hierarchical approach to understanding determinants of morphological variation, one which incorporates critical data on performance.Item Nicotine promotes Streptococcus mutans extracellular polysaccharide synthesis, cell aggregation and overall lactate dehydrogenase activity(Elsevier, 2015-08) Huang, R.; Li, M.; Gregory, Richard L.; Department of Biomedical and Applied Sciences, IU School of DentistrySeveral epidemiology studies have reported a positive relationship between smoking and dental caries. Nicotine, an alkaloid component of tobacco, has been demonstrated to stimulate biofilm formation and metabolic activity of Streptococcus mutans, one of the most important pathogens of dental caries. The first aim of the present study was to explore the possible mechanisms leading to increased biofilm by nicotine treatment from three aspects, extracellular polysaccharides (EPS) synthesis, glucosyltransferase (Gtf) synthesis and glucan-binding protein (Gbp) synthesis at the mRNA and protein levels. The second aim was to investigate how nicotine affects S. mutans virulence, particular in lactate dehydrogenase (LDH) activity. Confocal laser scanning microscopy results demonstrated that both biofilm bacterial cell numbers and EPS were increased by nicotine. Gtf and GbpA protein expression of S. mutans planktonic cells were upregulated while GbpB protein expression of biofilm cells were downregulated by nicotine. The mRNA expression trends of those genes were mostly consistent with results on protein level but not statistically significant, and gtfD and gbpD of biofilm cells were inhibited. Nicotine was not directly involved in S. mutans LDH activity. However, since it increases the total number of bacterial cells in biofilm, the overall LDH activity of S. mutans biofilm is increased. In conclusion, nicotine stimulates S. mutans planktonic cell Gtf and Gbp expression. This leads to more planktonic cells attaching to the dental biofilm. Increased cell numbers within biofilm results in higher overall LDH activity. This contributes to caries development in smokers.Item Ontogenetic and functional modularity in the rodent mandible(Elsevier, 2017) Menegaz, Rachel A.; Ravosa, Matthew J.; Department of Biomedical and Applied Sciences, IU School of DentistryThe material properties of diets consumed by juvenile individuals are known to affect adult morphological outcomes. However, much of the current experimental knowledge regarding dietary effects on masticatory form is derived from studies in which individuals are fed a non-variable diet for the duration of their postweaning growth period. Thus, it remains unclear how intra-individual variation in diet, due to ontogenetic variation in feeding behaviors or seasonal resource fluctuations, affects the resulting adult morphology. Furthermore, the mandible is composed of multiple developmental and functional subunits, and the extent to which growth and plasticity among these modules is correlated may be misestimated by studies that examine non-variable masticatory function in adults only. To address these gaps in our current knowledge, this study raised Sprague Dawley rats (n = 42) in four dietary cohorts from weaning to skeletal maturity. Two cohorts were fed a stable (“annual”) diet of either solid or powdered pellets. The other two cohorts were fed a variable (“seasonal”) diet consisting of solid/powdered pellets for the first half of the study, followed by a shift to the opposite diet. Results of longitudinal morphometric analyses indicate that variation in the mandibular corpus is more prominent at immature ontogenetic stages, likely due to processes of dental eruption and the growth of tooth roots. Furthermore, adult morphology is influenced by both masticatory function and the ontogenetic timing of this function, e.g., the consumption of a mechanically resistant diet. The morphology of the coronoid process was found to separate cohorts on the basis of their early weanling diet, suggesting that the coronoid process/temporalis muscle module may have an early plasticity window related to high growth rates during this life stage. Conversely, the morphology of the angular process was found to be influenced by the consumption of a mechanically resistant diet at any point during the growth period, but with a tendency to reflect the most recent diet. The prolonged plasticity window of the angular process/pterygomasseteric muscle module may be related to delayed ossification and muscular maturation within this module. The research presented here highlights the importance of more naturalistic models of mammalian feeding, and underscores the need for a better understanding of the processes of both morphological and behavioral maturation that follow weaning.