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Item The ability of new intracanal medicaments to prevent the formation of multi-species biofilm on radicular dentin(2017) Jacobs, Jordon C.; Spolnik, Kenneth J.; Ehrlich, Ygal; Gregory, Richard L.; Yassen, Ghaeth; Bringas, JosefThe residual antibacterial effects of antimicrobials used in endodontic regeneration against biofilm bacteria obtained from immature and mature teeth Jordon C. Jacobs DDS, Richard L Gregory PhD, Ygal Ehrlich DMD, Kenneth Spolnik DDS, MS, Josef S. Bringas DMD, DDS, MS, and Ghaeth Yassen BDS, MSD, PhD We explored the residual antibacterial properties of dentin pretreated with low concentrations of double antibiotic paste (DAP) against biofilm bacteria obtained from different clinical sources. Dentin blocks were sterilized and randomized into 4 treatment groups and 2 control groups (n=20). Blocks from treatment groups were pretreated with DAP (1 or 5 mg/ml) loaded into methylcellulose, calcium hydroxide (Ca(OH)2), or methylcellulose paste. After one week, the treatment pastes were removed and all blocks were immersed in PBS. The dentin blocks from treatment groups and one of the control groups were then inoculated with bacterial isolates obtained from immature or mature teeth with pulpal necrosis(n=10). The remaining control group received no bacteria and was used as a sterile control. Blocks were then incubated anaerobically for 3 weeks. Biofilm disruption assays were conducted for all samples. Two-way ANOVA and pair-wise comparisons were used for statistical analyses. The residual antibacterial effect of dentin pretreated with 5 mg/ml of DAP was significantly higher than all other groups regardless of the source of biofilm. Dentin pretreated with 1 mg/ml of DAP demonstrated significantly higher residual antibacterial effects in comparison to dentin pretreated with placebo paste and Ca(OH)2 only in bacterial isolates obtained from mature teeth with pulpal necrosis. Dentin pretreated with Ca(OH)2 did not demonstrate any residual antibacterial effects. Dentin pretreated with 1 or 5 mg/ml of DAP demonstrated significantly better residual antibacterial effects against biofilm bacteria obtained from mature teeth with pulpal necrosis in comparison to bacterial isolates obtained from immature teeth with pulpal necrosis.Item Diluted antibiotics for treating traumatized immature teeth(2014) Sabrah, Ala'a Hussein Aref, 1984-; Platt, Jeffrey A., 1958-; Gregory, Richard L.; Hara, Anderson T.; Spolnik, Kenneth Jacob 1950-; Murray, Peter E.; Goebel, W. ScottEndodontic regeneration (ERP) has been successfully used in the treatment of traumatized immature teeth. The procedure has three essential steps: disinfecting the root canal (i.e. triple antibiotic paste (TAP) or double antibiotic paste (DAP)), provoking bleeding inside the canal to form a scaffold upon which pulp stem cells will be deposited and continue root growth, and creating a good coronal seal. Previous research has reported that antibiotic pastes (TAP and DAP) are cytotoxic to stem cells in the concentrations commonly used in endodontic regeneration (1000 mg/mL). To decrease the adverse effects on stem cells and increase the rate of success of the regeneration, defining appropriate antibiotic concentrations for ERP is critical. In this project, five in-vitro experiments were conducted to determine the breakpoint dilutions of both TAP and DAP medicaments, and to prepare a suitable novel pastes containing diluted TAP or DAP medicaments for ERP. In the first experiment, we compared the antibacterial effect of TAP, and DAP against early biofilm formation of Enterococcus faecalis (E. faecalis) and Porphyromonas gingivalis bacteria. In the second study, we investigated the antibacterial effect of various dilutions of TAP and DAP antibiotic medicaments against established E. faecalis biofilm. In the third experiment, we investigated longitudinally the residual antibacterial activity of human radicular dentin treated with 1000, 1 or 0.5 mg/ml of TAP and DAP. In the fourth study, we investigated the cytotoxic effect of various dilutions of TAP and DAP antibiotic medicaments on the survival of human dental pulp stem cells (DPSC). And in the fifth experiment, we investigated the antibacterial and cytotoxic effect of novel intracanal medicaments consisting of methylcellulose (MC) and/or propylene glycol (PG) mixed with 1mg/ml of TAP or DAP. 1 mg/ml of DAP or TAP medicaments had a significant antibacterial effect against early bacterial biofilm formation, and established bacterial biofilm. Furthermore, 1 mg/ml had a residual antibacterial activity comparable to 1000 mg/ml. The novel intracanal medicaments had comparable antibacterial effect to currently used medicaments (1000 mg/ml). Additionally, the novel intracanal medicaments significantly enhanced DPSC metabolic activity, compared to currently used medicaments in endodontic regeneration procedures.Item Effect of intracanal medicaments used in endodontic regeneration procedures on microhardness and chemical structure of dentin(Synapse, 2015-05) Hamdon, Ghaeth Yassen; George, Joseph Eckert; Platt, Jeffrey Allen; Department of Restorative Dentistry, School of DentistryOBJECTIVES: This study was performed to investigate the effects of different intracanal medicaments on chemical structure and microhardness of dentin. MATERIALS AND METHODS: Fifty human dentin discs were obtained from intact third molars and randomly assigned into two control groups and three treatment groups. The first control group received no treatment. The second control group (no medicament group) was irrigated with sodium hypochlorite (NaOCl), stored in humid environment for four weeks and then irrigated with ethylenediaminetetraacetic acid (EDTA). The three treatment groups were irrigated with NaOCl, treated for four weeks with either 1 g/mL triple antibiotic paste (TAP), 1 mg/mL methylcellulose-based triple antibiotic paste (DTAP), or calcium hydroxide [Ca(OH)2] and finally irrigated with EDTA. After treatment, one half of each dentin disc was subjected to Vickers microhardness (n = 10 per group) and the other half was used to evaluate the chemical structure (phosphate/amide I ratio) of treated dentin utilizing attenuated total reflection Fourier transform infrared spectroscopy (n = 5 per group). One-way ANOVA followed by Fisher's least significant difference were used for statistical analyses. RESULTS: Dentin discs treated with different intracanal medicaments and those treated with NaOCl + EDTA showed significant reduction in microhardness (p < 0.0001) and phosphate/amide I ratio (p < 0.05) compared to no treatment control dentin. Furthermore, dentin discs treated with TAP had significantly lower microhardness (p < 0.0001) and phosphate/amide I ratio (p < 0.0001) compared to all other groups. CONCLUSIONS: The use of DTAP or Ca(OH)2 medicaments during endodontic regeneration may cause significantly less microhardness reduction and superficial demineralization of dentin compared to the use of TAP.Item Effect of low concentrations of antibiotic intracanal medicaments on crown discoloration and push-out bond strength(2018-07-12) Yaghmoor, Rayan Bahjat M.; Platt, Jeffrey A; Yassen, Ghaeth; Spolnik, Kenneth J.; Chu, Tien Min GabrielIntroduction: Some intracanal medicaments used in regenerative endodontics may compromise the bond strength of root cements and lead to tooth discoloration. Objectives: To evaluate the effects of 1) low concentrations of TAP and DAP (1 mg/mL) on push-out bond strength of various root cements, and 2) low concentrations of TAP and DAP (1 mg/mL and 10 mg/mL) on crown discoloration. Materials and Methods: Single rooted human teeth (n = 144) were horizontally decoronated and instrumented according to standardized protocol. The samples were randomized into six experimental groups (Ca(OH)2, 1000 mg/mL TAP and DAP, 1 mg/mL TAP and DAP, and no medicament control group. After four weeks, the medicaments were removed and each group was divided into three subgroups to receive MTA cement, Biodentine cement, or Endosequence Bioceramic putty cement for two weeks. Then, two root cylinders were obtained from each root and push-out bond strength testing was performed. For the crown discoloration experiment, 160 crowns were obtained from intact human molars and randomized into experimental groups as described earlier with the addition of two groups (10 mg/mL TAP and DAP). The pulp chambers in half of the samples from each group were coated with an adhesive bonding agent before receiving the assigned intracanal medicament. Color changes (ΔE) were detected by spectrophotometer at 1 day, 1 week, and 4 weeks after application, as well as after thermocycling. Results: In the push-out bond strength experiment, 1 mg/mL DAP generally demonstrated significantly higher bond strength of root cements compared with the other treatment groups. For the crown discoloration experiment, when an adhesive bonding agent was used prior to (10 mg/mL or 1000 mg/mL) TAP, the crowns had significantly less discoloration than those without adhesive. DAP 10 mg/mL had the least significant color change at all time points regardless of whether adhesive was used. Conclusion: 1) 1 mg/mL DAP and Ca(OH)2 did not have significant negative effect on the bond strength of calcium-silicate-based cement to radicular dentin. 2) 1 mg/mL and 10 mg/mL of DAP and Ca(OH)2 had significantly less effect on the color change of the human tooth crown than all intracanal medicaments used in this study.Item The effects of concentration and treatment time on the residual antibacterial properties of DAP(2016) Jenks, Daniel Brent; Spolnik, Kenneth J.; Bringas, Josef; Gregory, Richard L.; Yassen, Ghaeth H.; Ehrlich, Ygal; Warner, Ned A.Introduction: Regenerative endodontic procedures are used to treat immature teeth with pulpal necrosis in order to control infection, enable continued root development and enhance formation of a pulp like tissue in the canal. Canal disinfection is an integral part the regenerative endodontic process. Double antibiotic paste (DAP; i.e., equal parts of ciprofloxacin and metronidazole) has been successfully used for canal disinfection in regenerative endodontics. A comparison of the residual antibacterial effect of dentin treated with various dilutions of DAP pastes on biofilm formation has not yet been investigated thoroughly. Objectives: The aims of this in-vitro study were to investigate how concentration and time of treatment affect the residual antibacterial properties of DAP in preventing E. faecalis biofilm formation on human dentin. Materials and Methods: Extracted human teeth were used to obtain 4x4mm radicular dentin specimens. Each specimen was pretreated for 1 or 4 weeks with the 77 clinically used concentration of DAP (500 mg/mL), low concentrations of DAP (1, 5 or 50 mg/mL) loaded into a methylcellulose system, calcium hydroxide (Ca(OH)2), or placebo paste. After treatment, samples were rinsed and placed in sterile phosphate buffered saline (PBS) for three weeks. Samples were then inoculated with cultured E. faecalis and incubated in anaerobic conditions for three weeks to allow mature biofilm formation. The dentin samples were rinsed and biofilms detached. The detached biofilm cells were then diluted and spirally plated for enumeration on blood agar plates. The plates were then incubated for 24 h and the number of CFUs/mL was determined using an automated colony counter. Data was analyzed using Fisher’s Exact and Wilcoxon rank sum tests were used for statistical comparisons (α=0.05). Results: Dentin pretreatment for 4 weeks with 5, 50 or 500 mg/mL of DAP demonstrated significantly higher residual antibacterial effects and complete eradication of E. faecalis biofilms in comparison to a 1 week pretreatment with similar concentrations. However, dentin pretreated with 1 mg/mL of DAP or Ca(OH)2 did not provide a substantial residual antibacterial effect regardless of the application time. Conclusion: Dentin treated with 500, 50, or 5 mg/mL of DAP for 4 weeks was able to completely prevent the colonization of bacterial biofilm. Four-week treatment of dentin with DAP offers superior residual antibacterial effect in comparison to a one-week treatment. Intracanal application of DAP for 4 weeks during endodontic regeneration may offer an extended residual antibacterial effect.Item The use of traditional and novel techniques to determine the hardness and indentation properties of immature radicular dentin treated with antibiotic medicaments followed by ethylenediaminetetraacetic acid(Wolters Kluwer, 2014-10) Yassen, Ghaeth H.; Al‑Angari, Sarah S.; Platt, Jeffrey; Department of Restorative Dentistry, School of DentistryOBJECTIVE: The objective was to investigate the effect of intracanal antibiotic medicaments followed by ethylenediaminetetraacetic acid (EDTA) on the indentation properties and hardness of radicular dentin using a BioDent reference point indenter and a traditional microhardness technique, respectively. MATERIALS AND METHODS: Specimens with intact root canal dentin surfaces and polished radicular dentin specimens were obtained from immature human premolars. Each type of specimen was randomly assigned (n = 10 per group) and treated with either double antibiotic paste (DAP) for 4-week followed by EDTA for 5 min, triple antibiotic paste (TAP) for 4-week followed by EDTA for 5 min, EDTA for 5 min or Hank's balanced salt solution (control). The BioDent reference point indentor and Vickers microhardness tester were used to measure the indentation properties of root canal surfaces and the hardness of polished dentin specimens, respectively. One-way ANOVA followed by Fisher's protected least significant differences were used for statistical analyses. RESULTS: Both types of radicular dentin treated with antibiotic pastes and/or EDTA had a significant increase in the majority of indentation properties and a significant reduction in hardness compared to the untreated dentin. Furthermore, treatment of dentin with antibiotic pastes and EDTA caused significant increases in indentation properties and a significant reduction in hardness compared to EDTA-treated dentin. However, the RPI technique was not able to significantly differentiate between DAP + EDTA and TAP + EDTA-treated dentin. CONCLUSION: Dentin treated with antibiotic medicaments followed by EDTA had a significant increase the indentation properties and significantly reduction in hardness of radicular dentin.