The Effects of Calcium Hydroxide-loaded Poly(Lactic-co-glycolic Acid) Biodegradable Nanoparticles in the ex vivo External Inflammatory Root Resorption Model.

AIM: To evaluate the calcium ions (Ca2+) diffusion of calcium hydroxide-loaded poly(lactic-co-glycolic acid) biodegradable nanoparticles [Ca(OH)2-loaded PLGA NPs] compared with conventional Ca(OH)2 in a simulated external root resorption ex vivo model using inductively coupled plasma mass spectrometry (ICP-MS). MATERIALS AND METHODS: Thirty human mandibular premolars were prepared by sectioning the root segments to create roots measuring 10 mm from the anatomical apex. The root canals were instrumented and irrigated. The external root surface cavities were created. The specimens were randomly divided into the following three groups: Poly(lactic-co-glycolic acid) (PLGA; control group, n = 10), conventional calcium hydroxide [Ca(OH)2] (Metapaste, n = 10), and Ca(OH)2-loaded PLGA NPs [15% Ca(OH)2, n = 10]. The intracanal materials were placed in the root canals, and the teeth were stored in phosphate-buffered saline at 37°C. The release of Ca2+ was measured at 7, 30, and 60 days using ICP-MS. RESULTS: Both Ca(OH)2-loaded PLGA NPs and Metapaste groups exhibited higher levels of Ca2+ release compared to the PLGA group at all time points. During the initial 7-day period, the Ca(OH)2-loaded PLGA NPs exhibited a significantly greater release of Ca2+ compared to Metapaste. From day 7 to day 30, Metapaste displayed a significantly higher release of Ca2+ than the Ca(OH)2-loaded PLGA NPs, but it experienced a subsequent decline in Ca2+ release after the 30-day period. After the 30-day mark, the Ca(OH)2-loaded PLGA NPs once again exhibited a significantly higher release of Ca2+ compared to Metapaste. CONCLUSION: The Ca(OH)2-loaded PLGA NPs exhibited sustained release of Ca2+ that exceeded conventional Ca(OH)2, particularly during the first week, demonstrating a greater amount of Ca2+ release. CLINICAL SIGNIFICANCE: The utilization of Ca(OH)2-loaded PLGA NPs as an intracanal medication for external inflammatory root resorption provided sustained release and had the potential to enhance the efficacy of inhibiting root resorption more effectively than conventional Ca(OH)2.

Evaluation of the Bond Strength of Epoxy Resin-based Sealer Following Different Calcium Hydroxide Paste Removal Methods in Oval-shaped Root Canal.

AIM: The study aimed to investigate the bond strength of epoxy resin-based sealer following five different calcium hydroxide paste removal methods in oval-shaped root canals. MATERIALS AND METHODS: Sixty single-rooted human mandibular premolars having an oval-shaped root canal as evidenced by radiographs were decoronated and instrumented to size 40/0.04, medicated with calcium hydroxide paste for 7 days, before being randomly assigned to the conventional needle irrigation (CNI), manual dynamic agitation (MDA), sonic irrigation (SI), passive ultrasonic irrigation (PUI), and XP-endo Finisher (XP) groups to be irrigated using different irrigation systems. All specimens were then obturated using an epoxy resin-based root canal sealer and warm vertical compaction obturation technique. After 7 days, each specimen was sectioned into 1 mm root slices at the coronal, middle, and apical third of the root canal and tested for the push-out bond strength using a universal testing machine. RESULTS: The XP group was the only group that had comparable bond strength to the control group at every level of the root canal following removal of calcium hydroxide medicament (p >0.05). The control group had higher bond strength than the CNI group at every level of the root canal (p < 0.05). CONCLUSION: Calcium hydroxide interfered with the bonding of epoxy resin-based sealer to root canal walls. Irrigation with the XP increased the bonding sealer at every level of the root canal. CLINICAL SIGNIFICANCE: The XP was efficacious as a final rinse agitation technique to promote the bonding of the epoxy resin-based sealer at every level of the root canal following calcium hydroxide medication.

Effectiveness of D,L-2-hydroxyisocaproic acid (HICA) and alpha-mangostin against endodontopathogenic microorganisms in a multispecies bacterial-fungal biofilm in an ex vivo tooth model.

AIM: To develop a defined multispecies root canal biofilm model ex vivo, and to perform viable compositional analysis following D,L-2-hydroxyisocaproic acid (HICA), alpha-mangostin, Calcicur® , and Odontopaste® exposure. METHODOLOGY: Time-kill assays were conducted in vitro using HICA, alpha-mangostin, Calcicur® , Odontopaste® , and saline solution on the planktonic cultures of C. albicans, E. faecalis, L. rhamnosus, and S. gordonii. Human root dentine blocks were prepared (n = 100) ex vivo, and multispecies suspensions containing each of 1.5 × 108  CFU/mL C. albicans, E. faecalis, L. rhamnosus, and S. gordonii in brain heart infusion (BHI) were incubated within the root canals for 21 days. Canals (n = 20/group) were then exposed to medicaments for 7 days. Samples taken from the inner (first 0.1 mm) and deeper (second 0.1 mm) dentine by drilling with Ash Steel Burs No. 5 and No. 6, and residual roots were cultured in broth for 24 h. Cell growth was detected by spectrophotometry and confirmed by culture on agar. The other set of inner dentine, deeper dentine, and residual root samples were sonicated, and then exposed with 50 µM PMA before DNA was extracted using the QIAamp DNA mini kit. Real-time quantitative PCR was performed to determine the biofilm composition as well as the number of live and total cells remaining in the biofilm following each treatment. The OD data were analysed with Kruskal-Wallis and Friedman with Wilcoxon signed-rank test between and within groups, respectively, agar culture and qPCR data with Pearson chi-square with Mann-Whitney and Cochran with McNemar tests, respectively (p < .0001). RESULTS: Time-kill assays revealed that HICA and Calcicur® killed all planktonic organisms within 24 h, whilst alpha-mangostin killed the organisms within 72 h. However, Odontopaste® was a slow-killing agent: 10 cells of planktonic organisms survived after exposure to the agent for 7 days. The ex vivo tooth model demonstrated that HICA and alpha-mangostin significantly inhibited the cell growth in all sampling depths (p < .0001). All species-specific data revealed the effectiveness of each medicament on the biofilm composition. CONCLUSIONS: D,L-2-hydroxyisocaproic acid and alpha-mangostin had antimicrobial activity against multispecies bacterial-fungal biofilms.

Effects of N-acetyl cysteine on mitochondrial ROS, mitochondrial dynamics, and inflammation on lipopolysaccharide-treated human apical papilla cells.

OBJECTIVES: N-Acetyl cysteine (NAC), a well-known antioxidant molecule, has been used to modulate oxidative stress and inflammation. However, no studies have examined the effect of NAC in regenerative endodontic procedures (REPs). Therefore, the aim of this study was to investigate the effects of NAC on cell survival, mitochondrial reactive oxygen species (mtROS) production, and inflammatory and mitochondria-related gene expression on lipopolysaccharide (LPS)-treated apical papilla cells (APCs). MATERIALS AND METHODS: To assess the NAC concentration, 5 and 10 mM NAC were administered to LPS-treated APCs. Cell proliferation was measured at 24, 48, and 72 h by using AlamarBlue® assay. The 5-mM concentration was further analyzed using different treatment durations: 10 min, 24 h, and the entire study period. The mtROS production was quantified using MitoSOX™ Red and MitoTracker™ Green. RT-PCR was used to detect the expression of IL-6 and TNF-α inflammatory genes and mitochondrial morphology-related genes (Mfn-2/Drp-1 and Bcl-2/Bax) at 6 and 24 h. The statistical significance level was set at 0.05. RESULTS: Five-millimolar NAC promoted the highest LPS-treated APC proliferation. The use of 24-h NAC stimulated cell proliferation, whereas the entire-period NAC application (> 48 h) significantly reduced the cell number. The mtROS levels were slightly altered after NAC induction. Ten-minute NAC treatment downregulated the IL-6 and TNF-α expression, whereas the expression of Bcl-2/Bax and Mfn-2/Drp-1 ratios was upregulated at 6 h. CONCLUSIONS: Under the LPS-induced inflammatory condition, NAC stimulated APC survival and decreased inflammation. Ten-minute NAC treatment was sufficient to reduce the level of inflammation and maintain the mitochondrial dynamics. CLINICAL RELEVANCE: Ten-minute NAC application is sufficient to reduce the level of inflammation and maintain the mitochondrial dynamics. Therefore, NAC may be considered as a potential adjunctive irrigation solution in REPs.

Efficacy of calcium hydroxide-loaded poly(lactic-co-glycolic acid) biodegradable nanoparticles as an intracanal medicament against endodontopathogenic microorganisms in a multi-species biofilm model.

This study aimed to evaluate the antimicrobial activity of calcium hydroxide-loaded poly(lactic-co-glycolic acid) nanoparticles (CH-loaded PLGA NPs) on multi-species biofilms. Human root blocks were prepared (n = 40), and multi-species suspensions of Candida albicans, Enterococcus faecalis and Streptococcus gordonii were incubated within the root canals for 21 days. Canals (n = 10/group) were then medicated with saline solution (negative control), chlorhexidine (positive control), calcium hydroxide and CH-loaded PLGA NPs for 7 days. Samples taken from the 0.1 mm root canal dentin were collected, and cell growth was detected by culture on BHI agar. The viable cell count of the Ca(OH)2, chlorhexidine gel and CH-loaded PLGA NPs group was significantly lower than the normal saline group (p < 0.001). CH-loaded PLGA NPs demonstrated a significant lower viable cell than Ca(OH)2 (p < 0.001); it has potential as a medicament for endodontic therapy.

The Evaluation of Anti-Osteoclastic Activity of the Novel Calcium Hydroxide Biodegradable Nanoparticles as an Intracanal Medicament.

INTRODUCTION: The aim of this study was to evaluate the anti-osteoclastic activity of calcium hydroxide-loaded poly(lactic-co-glycolic acid) nanoparticles [Ca(OH)2-loaded PLGA NPs] in comparison to calcium hydroxide nanoparticles [Ca(OH)2 NPs]. METHODS: RAW 264.7 cell lines (third-fifth passage) were cultured and incubated with soluble receptor activator of nuclear factor kappa B ligand in triplicate. Subsequently, Ca(OH)2-loaded PLGA NPs and Ca(OH)2 NPs were added for 7 days to evaluate their effects on receptor activator of nuclear factor kappa B ligand-induced osteoclast differentiation of RAW 264.7 cells by tartrate-resistant acid phosphatase activity. Additionally, a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was conducted to confirm the cytotoxicity of treatments to cells. RESULTS: Tartrate-resistant acid phosphatase staining showed a significant reduction in the osteoclast number when treated with Ca(OH)2-loaded PLGA NPs compared with Ca(OH)2 NPs (P < .01). In comparison to the control, the number of osteoclasts significantly reduced upon treatment with Ca(OH)2-loaded PLGA NPs (P < .05), but there was no significant difference in Ca(OH)2 NPs. Furthermore, osteoclast morphology in both treatment groups exhibited smaller sizes than the control group. Neither Ca(OH)2-loaded PLGA NPs nor Ca(OH)2 NPs demonstrated cytotoxic effects on RAW264.7 cells. CONCLUSIONS: Both Ca(OH)2 NPs with and without poly(lactic-co-glycolic acid) have the ability to inhibit osteoclast differentiation. However, Ca(OH)2-loaded PLGA NPs exhibit greater potential than Ca(OH)2 NPs, making them a promising intracanal medicament for cases of root resorption.

The Bond Strength of Root Canal Filling after Calcium Hydroxide Removal with a Simple Apical Negative Pressure Kit in Oval-Shaped Root Canal.

Background: The aim of this study was to evaluate the effect of a simple irrigating device, which produced from the apical negative pressure concept, as a final irrigating method after calcium hydroxide (CH) medication on the bond strength of epoxy resin-based sealer in the oval-shaped root canal. Methods: Forty-eight single-rooted premolars with oval-shaped canals were included in the study. The crown was decoronized and canal instrumented with Mtwo (VDW GmbH, Munich, Germany) up to size 40/04. The specimens were assigned to four groups according to the irrigation protocol after 1 week of CH medication: control group (no medication) and three experimental groups conventional needle irrigation (CNI), passive ultrasonic irrigation (PUI), and simple apical negative pressure kit (sANP). The teeth were obturated with a warm vertical technique using epoxy resin-based sealers. After 1 week, the roots were transversally sectioned at coronal, middle, and apical thirds. A push-out test was performed by a universal testing machine, and statistical analysis was performed using One-Way ANOVA with Duncan's post hoc test. Results: The bond strength in the control group was significantly higher than the CNI group in all root canal thirds (P < 0.05). At coronal third, sANP showed lower bond strength than PUI group (P < 0.05). However, the bond strength of sANP group was comparable to the control group (P > 0.05) which was significantly higher than the PUI group at apical third (P < 0.05). Conclusion: A sANP enhanced the bond strength of epoxy resin-based sealer in the apical third of CH-medicated root canal in the oval-shaped canal.

Efficacy of alpha-mangostin for antimicrobial activity against endodontopathogenic microorganisms in a multi-species bacterial-fungal biofilm model.

OBJECTIVE: To determine the activity of alpha-mangostin on preformed bacterial-fungal multi-species biofilms in vitro, and to ascertain the impact on metabolic activity, biofilm structure and viability. DESIGN: Inhibitory concentrations (ICs) for alpha-mangostin against planktonic cultures of Candida albicans, Enterococcus faecalis, Lactobacillus rhamnosus, and Streptococcus gordonii were determined using a standard broth microdilution method. Single and multi-species (all species 1:1:1:1) biofilms were grown on polystyrene coverslips in Roswell Park Memorial Institute Medium for 48 h. The biofilms were then exposed to 0.2% (w/v) alpha-mangostin for 24 h. These concentrations were selected based on pilot experiments and the solubility of these compounds. 2% (v/v) chlorhexidine was used as a positive control and Roswell Park Memorial Institute Medium as a negative control. The metabolic activity of the biofilms after exposure was measured using metabolic (XTT) assays. Biofilms were visualised and quantified using fluorescent BacLight™ LIVE/DEAD staining. The biofilms were assessed for cell viability by culture and colony counting (CFU/mL). RESULTS: 8 mg/L of alpha-mangostin was cidal against planktonic bacteria and 1000 mg/L for Candida. Alpha-mangostin was most active against L. rhamonosus biofilms and least active against C. albicans biofilm (metabolism inhibited by 99% and 78%, respectively). Alpha-mangostin exposure reduced the number of viable cells in the biofilms. CONCLUSION: Alpha-mangostin inhibited the metabolic activity of bacterial-fungal biofilms effectively. The anti-biofilm activity of alpha-mangostin was comparable to chlorhexidine and thus has potential as a novel agent for endodontic therapy.

Ameliorative Effect of Bouea macrophylla Griffth Seed Extract Against Bacteria-Induced Acne Inflammation: in vitro study.

Currently, bioactive compounds derived from nature have been thought to be promising anti-acne substances owing to the variety of potential biological effects. This study aimed to evaluate the ameliorative effect of Bouea macrophylla Griffth seed extract against bacteria-induced acne inflammation for the first time in terms of antibacterial effects against acne-inducing bacteria, anti-inflammatory, and antioxidant properties. Initially, extracting procedures were optimized and five different extracts were obtained. Considering their antibacterial activities against Cutibacterium acnes, Staphylococcus aureus, and Staphylococcus epidermidis, ethanolic and ethyl acetate fractions exerted a notable effect which were highly superior above those of polyphenol standards. Additionally, these two extracts presented outstanding antioxidant capacities in terms of DPPH and ABTS radicals scavenging effects, reducing power, and inhibitory effect on lipid peroxidation which also play a role in the exacerbation of acne inflammation. Besides, inhibition on lipid peroxidation and reducing power of ethanolic fraction were significantly (p<0.05) better than those of ethyl acetate fraction which was corresponding to their phenolic and ellagic acid contents. However, flavonoids found in ethyl acetate fraction might play an important role in its potentials. After that, the anti-inflammatory effects of the extracts were elucidated by means of inhibition on nitric oxide production from LPS-induced RAW 264.7 cell lines at which the effects of both extracts were dosedependency. Taken together, our findings have apparently proven that B. macrophylla seed extracts exerted a variety of potential properties including antioxidation, anti-acne-inducing bacteria, and anti-inflammatory effects which could serve as a promising anti-acne agent for cosmeceutical applications.

Potential of Bouea macrophylla kernel extract as an intracanal medicament against mixed-species bacterial-fungal biofilm. An in vitro and ex vivo study.

OBJECTIVE: To investigate the antimicrobial activity of B. macrophylla kernel extract against mixed-species biofilms of E. faecalis, S. gordonii and C. albicans in vitro. To evaluate the efficacy of the extract as an intracanal medicament compared with Ca(OH)2 and chlorhexidine in ex vivo tooth model. METHODS: The antibiofilm effect of B. macrophylla kernel extract was determined by AlamarBlue™ assay and the effect on biofilms was visualized by LIVE/DEAD® BacLight™ viability test. Mixed-species biofilms were incubated into the tooth model (N = 42) for 21 days. The teeth were randomly divided into 4 medicament groups for 7 days: (i) normal saline, (ii) calcium hydroxide (Ca(OH)2), (iii) chlorhexidine gel, (iv) B. macrophylla kernel extract. Dentine samples were collected, qPCR with PMA was used to quantify the viability and species composition of each sample. SEM was used to visualize the effect of medicament on biofilm structure. RESULTS: The MBIC was 6.25 mg/mL and the MBEC was 50 mg/mL. The integrity of microbial cells was progressively compromised as concentration increased, resulting in greater cell death. Ex vivo tooth model revealed that biofilm treated with 50 mg/mL of the B. macrophylla extract demonstrated a significantly higher proportions of dead cells than in Ca(OH)2, chlorhexidine and normal saline groups (p < 0.01). Disruption of biofilm structure and enlargement of dentinal tubules was observed in B. macrophylla group on SEM. CONCLUSION: The extract of B. macrophylla kernel exhibited significant antibiofilm effect against the mixed-species biofilms of E. faecalis, S. gordonii and C. albicans.