Browsing by Author "Gregory, Richard"
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Item The antibacterial effect of new intracanal medicaments against established mutlispecies biofilm(2017) Troxel, Alex; Spolnik, Kenneth J.; Gregory, Richard; Ehrlich, Ygal; Bringas, Josef; Zunt, Susan L.; Yassen, GhaethWe investigated the antibacterial effect of low concentrations of double antibiotic paste (DAP) loaded into a methylcellulose system against bacterial biofilms obtained from mature and immature teeth with necrotic pulps. Standardized radicular dentin specimens were randomly divided into six experimental groups (n = 20). Group 1: 5mg/mL DAP treatment. Group 2: 1mg/mL DAP treatment. Group 3: Calcium hydroxide (Ca(OH)2) treatment. Group 4: Methylcellulose. Group 5: No treatment. Group 6: No bacteria or treatment. Clinical bacterial isolates were obtained from mature and immature teeth with necrotic pulps indicated for endodontic regeneration or routine endodontic treatment, respectively. Specimens in each group were inoculated with either bacterial isolates (n = 10) and incubated anaerobically for 3 weeks. Specimens were then treated for one week with the assigned group treatment. Treatments were rinsed with sterile saline and biofilms were detached and spiral plated using biofilm disruption assays. Wilcoxon Rank Sum tests followed by pair-wise comparisons were used for statistical analyses. Treatment of infected dentin with 1 mg/ml of DAP, 5 mg/mL of DAP, and Ca(OH)2 demonstrated significant and substantial antibiofilm effects in comparison to untreated control groups or groups treated with placebo paste. Furthermore, 1 mg/mL of DAP caused complete eradication of biofilm obtained from mature tooth with necrotic pulp. However, the same concentration was not able to completely eradicate biofilm obtained from the immature tooth with necrotic pulp. Low concentrations of DAP (1-5 mg/mL) loaded into a biocompatible methylcellulose system demonstrated significant antibacterial effects against biofilm obtained from both mature and immature teeth with necrotic pulps.Item Bactericidal Efficacy of EdgePRO Er,Cr:YSGG Laser-Activated Irrigation Against a Mature Endodontic Multispecies Biofilm Using an in vitro Infected Tooth Model(2024) Patterson, Samuel B.; Spolnik, Kenneth J.; Gregory, Richard; Ehrlich, Ygal; Movila, AlexandruIntroduction: Treatment goals of non-surgical root canal therapy (nsRCT) include the removal of all organic tissue material, bacterial biofilm and their by-products, and debris materials, in order to disinfect the canal system to a level compatible with healing and to further prevent infection. Standard chemo-mechanical protocols have several well-documented shortcomings and subsequent areas for improvement regarding their disinfection abilities. In recent years, emerging laser technology and its application in root canal therapy has been gaining popularity as a safe and promising tool for advancing endodontic treatment. The newest FDA-approved laser for endodontic application is the EdgePRO Erbium,Chromium-doped:Yttrium-Scandium-Gallium-Garnet (Er,Cr:YSGG) infrared laser operating at a 2780 nm wavelength. Previous in vitro studies using Er,Cr:YSGG lasers have demonstrated their ability to enhanced canal debridement, cleaning, smear layer removal, and bacterial disinfection. Additionally, a few in vivo trails have been completed using this laser type as an adjunct in RCT procedures, which have yielded safe and highly successful results in the clinical setting. However, research specifically using the EdgePro device as well as a standardized protocol for optimal clinical usage of the laser is lacking. Objectives: The aim of this study was to evaluate the bactericidal and biofilm dissolution effects of laser-activated irrigation using the EdgePro laser against a mature multispecies biofilm in an infected tooth model and to assess the potential increased disinfection and cleaning ability compared to a standard needle irrigation protocol. Materials and Methods: Single rooted teeth (n=36) were decoronated to a standardized length of 16mm. The root canals were endodontically prepared using a standard irrigation, hand-filing, and rotary protocol to a final size of ISO 25.06 while maintaining a fully patent apical foramen. An irrigation solution reservoir was created in the coronal 4 mm of the canal space. Sterile specimens were inoculated with multispecies bacterial sample containing E. faecalis. The mixed bacteria was grown anaerobically for 10 days to form a mature biofilm using a previously established protocol. The teeth were divided into a negative control group (saline rinse, n=12), positive control group (standard needle irrigation – SNI, n=12), and an experimental group (laser-assisted treatment protocol, n=12). The positive control and experimental laser groups utilized the same irrigation solutions of 2 mL 17% EDTA followed by 5 mL 3% NaOCl using a standard 27-gauge side-vented irrigation needle placed as far apically as possible without binding. The experimental group underwent additional laser activation using laser tip #2 (350 m diameter) and settings of: 15 mJ, 0.75 W, 50 Hz, 0% air, and 0% water spray (Mid-Root Solutions 1 preset). The laser tip was inserted halfway into the irrigation filled canals (8 mm from orifice and apex) and fired upon withdrawal at a speed of 0.8 mm/sec, which comprised a single lasing cycle of 10 seconds. Three lasing cycles were completed with EDTA first followed by NaOCl, for a total of six lasing cycles with 60 seconds of irradiation time per tooth. A final rinse of sterile saline was used in all tooth samples prior to bacterial sample collection via Versa-brushes and sterile paper points. The samples were transferred to a laboratory setting where they underwent ultrasonic agitation, serial dilution, spiral plating on blood-agar, and two days of anaerobic incubation for assessment of bacterial growth. Colony forming units (CFUs/mL) were counted as a means of quantitative analysis. Results: The negative control group yielded the highest level of bacterial growth with an average of 934,771 CFUs/mL. The positive control group displayed a statistically significant lower amount of bacterial growth with an average of 4,698 CFUs/mL and yielded 1 sample with no bacterial growth. The experimental laser group had statistically significant lower bacterial growth present compared to both the positive and negative control groups and produced all negative bacterial samples with none of the 12 agar plates demonstrating CFU growth and averaged 0 CFUs/mL.. Conclusion: Within the scope of this study, laser-activated irrigation (LAI) using the EdgePro Er,Cr:YSGG laser was capable of producing no detectable bacterial samples in an in vitro infected tooth model. EdgePro LAI displayed statistically significant superior cleaning and disinfection of infected canal space compared to teeth treated with standard needle irrigation alone. The EdgePro laser system indeed shows promise as an adjunctive tool in clinical root canal treatment procedures. Further investigation is warranted using similar protocols in teeth with more complicated anatomy and with supplemental methods for analyzing bactericidal potential.Item Combined Effects of Soda Drinks and Nicotine on Streptococcus Mutans Metabolic Activity and Biofilm Activity(2019) Mokeem, Lamia Sami; Gregory, Richard; Cook, Norman Blaine; Windsor, Jack; Eckert, GeorgeItem Differential profiles of soluble and cellular toll like receptor (TLR)-2 and 4 in chronic periodontitis(PLOS, 2018-12-20) AlQallaf, Hawra; Hamada, Yusuke; Blanchard, Steven; Shin, Daniel; Gregory, Richard; Srinivasan, Mythily; Periodontology, School of DentistryChronic periodontitis is a common inflammatory disease initiated by a complex microbial biofilm and mediated by the host response causing destruction of the supporting tissues of the teeth. Host recognition of pathogens is mediated by toll-like receptors (TLRs) that bind conserved molecular patterns shared by large groups of microorganisms. The oral epithelial cells respond to most periodontopathic bacteria via TLR-2 and TLR-4. In addition to the membrane-associated receptors, soluble forms of TLR-2 (sTLR-2) and TLR-4 (sTLR-4) have been identified and are thought to play a regulatory role by binding microbial ligands. sTLR-2 has been shown to arise from ectodomain shedding of the extracellular domain of the membrane receptor and sTLR-4 is thought to be an alternate spliced form. Many studies have previously reported the presence of elevated numbers of viable exfoliated epithelial cells in the saliva of patients with chronic periodontitis. The objective of this study was to investigate the potential value of salivary sTLR-2 and sTLR-4 together with the paired epithelial cell-associated TLR-2/4 mRNA as diagnostic markers for chronic periodontitis. Unstimulated whole saliva was collected after obtaining informed consent from 40 individuals with either periodontitis or gingivitis. The sTLR-2 and sTLR4 in saliva was measured by enzyme-linked immunosorbent assay. The TLR-2 and TLR-4 transcript in the epithelial cells in saliva was measured by real time polymerase chain reaction. While levels of sTLR-2 exhibited an inverse correlation, sTLR-4 positively correlated with clinical parameters in the gingivitis cohort. Interestingly, both correlations were lost in the periodontitis cohort indicating a dysregulated host response. On the other hand, while the sTLR-2 and the paired epithelial cell associated TLR-2 mRNA exhibited a direct correlation (r2 = 0.62), that of sTLR4 and TLR-4 mRNA exhibited an inverse correlation (r2 = 0.53) in the periodontitis cohort. Collectively, assessments of salivary sTLR2 and sTLR4 together with the respective transcripts in the epithelial cells could provide clinically relevant markers of disease progression from gingivitis to periodontitis.Item Differential profiles of soluble and cellular toll like receptor (TLR)-2 and 4 in chronic periodontitis(PLOS, 2018-12-20) AlQallaf, Hawra; Hamada, Yusuke; Blanchard, Steven; Shin, Daniel; Gregory, Richard; Srinivasan, Mythily; Periodontology, School of DentistryChronic periodontitis is a common inflammatory disease initiated by a complex microbial biofilm and mediated by the host response causing destruction of the supporting tissues of the teeth. Host recognition of pathogens is mediated by toll-like receptors (TLRs) that bind conserved molecular patterns shared by large groups of microorganisms. The oral epithelial cells respond to most periodontopathic bacteria via TLR-2 and TLR-4. In addition to the membrane-associated receptors, soluble forms of TLR-2 (sTLR-2) and TLR-4 (sTLR-4) have been identified and are thought to play a regulatory role by binding microbial ligands. sTLR-2 has been shown to arise from ectodomain shedding of the extracellular domain of the membrane receptor and sTLR-4 is thought to be an alternate spliced form. Many studies have previously reported the presence of elevated numbers of viable exfoliated epithelial cells in the saliva of patients with chronic periodontitis. The objective of this study was to investigate the potential value of salivary sTLR-2 and sTLR-4 together with the paired epithelial cell-associated TLR-2/4 mRNA as diagnostic markers for chronic periodontitis. Unstimulated whole saliva was collected after obtaining informed consent from 40 individuals with either periodontitis or gingivitis. The sTLR-2 and sTLR4 in saliva was measured by enzyme-linked immunosorbent assay. The TLR-2 and TLR-4 transcript in the epithelial cells in saliva was measured by real time polymerase chain reaction. While levels of sTLR-2 exhibited an inverse correlation, sTLR-4 positively correlated with clinical parameters in the gingivitis cohort. Interestingly, both correlations were lost in the periodontitis cohort indicating a dysregulated host response. On the other hand, while the sTLR-2 and the paired epithelial cell associated TLR-2 mRNA exhibited a direct correlation (r2 = 0.62), that of sTLR4 and TLR-4 mRNA exhibited an inverse correlation (r2 = 0.53) in the periodontitis cohort. Collectively, assessments of salivary sTLR2 and sTLR4 together with the respective transcripts in the epithelial cells could provide clinically relevant markers of disease progression from gingivitis to periodontitis.Item Effect of Gap Geometry on Secondary Caries in Vitro(2009) Nassar, Hani M.; Cabezas, Carlos Gonzales, 1966-; Chu, Tien-Min Gabriel; Fontana, Margherita Ruth, 1966-; Gregory, Richard; Matis, Bruce; Cochran, MichaelObjective: To investigate the effect of the size of the space between the restoration and the dentinal wall of the tooth (i.e. the dentinal portion of the gap) on the development of secondary caries. Methods: Tooth-resin-matrix composite specimens were mounted on custom-made gap-model stages. Specimens were divided into four groups (n=10). Group 1 had a uniform gap size of 30μm throughout both enamel and dentin. Group 2 had a 30μm enamel gap size with a 530μm dentinal gap. Group 3 had 525μm gaps in both enamel and dentin. Group 4 had 525μm and 1025μm gaps in enamel and dentin, respectively. Specimens were attached to plastic Petri plates, gas-sterilized and then incubated in a microbial caries model with S. mutans TH16 in (1% sucrose tryptic soy broth for 1 h, 4 times/day, and with a buffer solution for the rest of the day). After 8 days of incubation, tooth specimens were sectioned and stained with a rhodamine B solution. Digital images were taken under a confocal microscope and analyzed for lesion size at the enamel outer lesion (EOL), enamel wall lesion (EWL), dentin wall lesion next to the DEJ (DWL-A) and dentin wall lesion at 750µm from the DEJ (DWL-B). Results: No difference in EOL size was found between the groups. DWL-A and -B were larger in group 3 than groups 1and 2. Larger DWL-B was found in group 3 than group 4. Group 4 had marginally significant larger EWL than groups 1 and 2 (p=0.0652 and p=0.0648, respectively). Also, group 4 had marginally significant (p=0.0607) larger DWL-B than group 1. Conclusions: Based on the results of this study, it can be concluded that the presence of additional space at the dentinal wall area did not affect secondary caries development as long as the enamel gap was small. However, with enamel gaps of ≈500 µm, the presence of the additional gap space at the dentinal wall led to the development of smaller dentinal wall lesions at the deeper parts of the simulated cavity. Also, in uniform gaps, the size of the interface was positively correlated with size of the dentinal wall lesions.Item Extrinsic Characterization Sustainability in Zirconia Reinforced Lithium Silicate Ceramics(2021-04-08) Gadah, Thrya; May, Jaren; Levon, John; Chu, Tien-Min G; Gregory, Richard; Wei Shao, Lin; Feitosa, SabrinaABSTRACT OBJECTIVE. To investigate the effect of aging on the surface roughness and the color sustainability of externally characterized zirconia reinforced lithium silicate glass-ceramics treated with different surface protocols. METHODS. Sixty blocks (12-mm X 14-mm; 1.5-mm) of pre-crystalized zirconia reinforced-lithium silicate glass-ceramic (Vita Suprinity, Vita Zahnfabrick, Germany) CAD/CAM were crystalized and treated with different surface protocols, as extrinsic characterization (EC), mechanical polishing (MP), glaze layer (GL), surface adjustment (SA) and no treatment – control group (CG). Experimental groups (n=10) were divided as follow: CG; EC-MP-GZ; EC-GZ; EC-MP; EC-GZ-SA-GZ; EC-GZ-SA-MP and submitted to thermocycling (5,000 cycles, 5-55C) and toothbrushing simulation (5,000 cycles). Surface roughness (Ra and Rq), color change (CIED2000) and biofilm growth were evaluated. Statistical analysis was performed with a two-sided 5% significance level for all tests. RESULTS. For the parameter RaX, the control differed from EC-GZ and EC-MP (p = 0.04). For the parameter RqX, EC-GZ presented higher surface roughness compared than EC-MP-GZ and the group EC-GZ-SA-MP (p=0.02). EC-MP (p<0.01) and the EC-GZ-SA-MP (p<0.01) showed higher color change E00 after aging, while CG and EC-MP-GZ the least. For the biofilm growth, no significant group effect on bacteria counts was found (p=0.089). CONCLUSION. The aging protocol affected the surface roughness, and color of externally characterized zirconia reinforced lithium silicate glass-ceramics submitted to different surface treatment protocols. In the present study, when the mechanical polishing was performed before glaze application, the slightest color change and surface roughness were observed compared to the other surface treatments. Bacteria were not able to grow in the material surface, under the conditions tested in the present study.Item The impact of hydroxyapatite on alkaline phosphatase activity and mineral deposition of dental pulp stem cells using a double antibiotic paste loaded methylcellulose carrier(2020) Fischer, Benjamin I.; Bruzzaniti, Angela; Spolnik, Kenneth; Ehrlich, Ygal; Bringas, Josef; Gregory, RichardIntroduction: Regenerative endodontic procedures (REPs) are a type of endodontic treatment aimed at replacing damaged tooth structures, including dentin and root structures, as well as cells of the pulp-dentin complex. Double antibiotic paste (DAP) has been shown to be efficacious in achieving disinfection of the root canal system while minimizing cytotoxicity to dental pulp stem cells (DPSCs). Hydroxyapatite (HA) is an extracellular, mineralized component of bone that has shown much promise as a scaffold in the field of regenerative medicine. Objective: The objective of this study was to evaluate the effects of HA in a DAP loaded methylcellulose (MC) carrier on the differentiation and mineral deposition of DPSC over time. Materials and Methods: DPSCs were plated in 24-well plates with culture media. The following day, semi-permeable 0.1 m chambers were inserted into the wells to separate the reservoirs and permit delivery of medicaments. 100 L treatment paste composed of MC with 1% DAP and either 0.5% or 1.0% nano-HA was added, followed by additional culture media. After 3 days of treatment, medicaments were removed and DPSCs were cultured for an additional 9 days with replacement of media every 3-4 days. At Day 12, DPSCs were evaluated for alkaline phosphatase (ALP) activity using a biochemical assay and mineral deposition using an Alizarin Red S Ca2+ staining assay (4 wells/group). Comparisons between groups were performed using one-way analysis of variance (ANOVA) with a 5% significance level used for all tests. Results: A trend towards increased ALP and mineral deposition activity was noted among the groups with HA added to DAP with MC. Although these trends were not statistically significant, a trend towards increased ALP and mineral deposition was observed after 3-day medicament exposure. The results were similar to previous findings using 7-day medicament treatments. Conclusion: The addition of HA showed a trend towards improved differentiation and mineral deposition of DPSCs compared to DAP with MC. Although additional studies are required, these results showed suggest that even with a shortened treatment time, increased differentiation and mineral deposition of DPSCs may be possible. This study provides additional support that low concentration DAP in a MC carrier has potential application in regenerative endodontic procedures. The novel addition of HA may provide additional osteogenic potential.Item INTERACTIONS OF HUMAN GINGIVAL FIBROBLASTS WITH TOBACCO TREATED PORPHYROMONAS GINGIVALIS(Office of the Vice Chancellor for Research, 2012-04-13) Lanier, Branden; Al-Shibani, Nouf Khider; Windsor, L. Jack; Gregory, RichardPorphyromonas gingivalis (P. gingivalis) and tobacco are risk factors for periodontal disease. The objective of this study was to determine the effects that tobacco treated P. gingivalis cells have on human gingival fibroblasts (HGFs). The study was conducted to examine the effects that cigarette smoke condensate (CSC), nicotine, and dissolvable smokeless tobacco (DST) strips treated P. gingivalis has on cell cytotoxicity and the expression of cy-tokines and growth factors from HGFs. The P. gingivalis was grown at 37°C and then the cells and supernatant were separated. P. gingivalis cells were then washed and killed. The concentration of protein in the cell pellet and supernatant were determined by protein assay using the Bradford method. The lowest non-toxic levels of the cell pellet and supernatant will be used to treat the HGFs for 72 hours and then cytotoxicity was determined by lactate dehydrogenase (LDH) assays. The cytokine/growth factor expression will be determined by antibody protein arrays. The protein assays showed that the tobacco products reduced the protein amounts as compared to untreated bacteria. The results should show an increase in cytotoxicity with increasing protein concentrations, along with increased pro-inflammatory cyto-kine/growth factors expression by the HGFs treated with tobacco treated P. gingivalis compared to P. gingivalis that was not treated with tobacco prod-ucts. A better understanding of the detrimental effects that tobacco has on the underlining causes of periodontal disease can advance the quest of con-trolling the disease. This study was funded by the Indiana University–Purdue University Indianapolis Multidisci-plinary Undergraduate Research Institute (MURI).Item Primary Coronal Caries Prevention with Silver Diamine Fluoride – Investigations into Efficacy and Mode of Action(2021-01) Sorkhdini, Parand; Lippert, Frank; Gregory, Richard; Martinez Mier, E. Angeles; Crystal, Yasmi O.; Stelzner, SarahDental caries continues to be one of the most prevalent preventable diseases worldwide. Silver diamine fluoride (SDF) is a topical solution comprised of silver, ammonia and fluoride. It is a safe, effective, efficient, noninvasive and cost-effective method in caries management. However, there is little clinical evidence supporting the use of SDF (or SDF followed by application of potassium iodide[KI] to mitigate staining) as anti-caries agents on sound enamel and early enamel carious lesions. In this dissertation, I studied the mechanism behind SDF’s ability to prevent coronal caries which has not been studied yet. In the first and second aims, I investigated the effectiveness of SDF, SDF+KI, fluoride (potassium fluoride [KF]) and silver (silver nitrate [AgNO3]) controls to SDF and deionized water (DIW) in preventing enamel demineralization and enhancing remineralization using chemical, biofilm and pH-cycling models. In both chemical demineralization and pH-cycling models there were no statistically significant differences between SDF and SDF+KI in preventing coronal caries. In the biofilm model, however, SDF+KI was significantly less effective in preventing demineralization than SDF. In the third aim, I investigated the efficacy of SDF, SDF+KI, KF, AgNO3, and DIW on the remineralization of active subclinical enamel carious lesions. Here, SDF+KI was significantly more effective in promoting remineralization than SDF. I calculated changes in color, and the results show applying KI after SDF significantly reduced the dark staining caused by SDF. In conclusion: SDF and SDF+KI appear to be effective options in preventing and in the treatment of primary coronal caries. Further clinical research is required to confirm the present findings.