High-Fat Diet Influences Dendritic Cells and T-Cell Infiltration in Apical Periodontitis in Mice.

INTRODUCTION: Diet-induced metabolic syndrome may influence the progression and healing of apical periodontitis (AP). The aim of this study was to evaluate the inflammatory immune response of dendritic cells (DCs) and T helper (Th) cells in normal versus obese mice with AP. METHODS: Twenty male C57BL/6 mice were divided into 2 groups: normal chow (NC) and high-fat diet (HFD) for 12 wk. AP was induced in both groups by creating pulp exposure of the right first maxillary molar to the oral environment. Contralateral first molars from each mouse were used as a control. The animal's body mass was recorded on a weekly basis, and they were euthanized after 30 d. The maxillae were removed and processed for micro-computed tomography (micro-CT), histologic analysis, and immunofluorescence staining for DCs (CD11c), Th17 (IL-17A), and T regulatory cells (FOXP3 and IL-10). Different groups were analyzed by Mann-Whitney U test, Student t test, and ordinary 1-way analysis of variance followed by Tukey's multiple comparisons test. The level of significance (α) was set at 0.05. RESULTS: The HFD group showed larger AP lesions than the NC group from micro-CT analysis. For the NC group, induction of AP significantly increased immune cell infiltration when compared with control. HFD showed increased DCs and Th17 infiltration in the control group without AP. In addition, there was no significant change in the amount of DCs and Th17 in the HFD-AP group when compared with the NC-AP and HFD-control groups. CONCLUSIONS: HFD resulted in an increased immune cell infiltration in the periapical area without AP. Despite the larger AP lesion observed in HFD-AP than that of NC-AP, the amount of infiltrated inflammatory cells did not differ significantly. The results of this study suggest that the DCs and Th17 inflammatory pathways are affected by HFD in the periapical region, but their contribution toward AP complicated by metabolic syndrome requires further investigation.

Injectable thermosensitive hydrogel to modulate tolerogenic dendritic cells under hyperglycemic condition.

Hyperglycemia associated with diabetes mellitus is a significant risk factor for periodontitis and it polarizes the immune cells towards an inflammatory state. Specific biomaterials can deliver therapeutic or immunomodulatory agents to regulate the excessive periodontal inflammation. Dendritic cells (DCs) bridge the innate and adaptive immune systems and are crucially involved in periodontitis. Thus, targeting DCs is an attractive treatment option for diabetic periodontitis, which, by modulating the downstream adaptive immune cells could regulate the host immune responses. In this study, a chitosan-based thermosensitive injectable self-assembled hydrogel (TISH) was developed to modulate DCs towards a tolerogenic phenotype, which can induce regulatory T-cells to attenuate inflammation and promote healing. Granulocyte-macrophage colony-stimulating factor (GM-CSF) and resveratrol were loaded into TISH (TISH(G + R)) and were sustainably released. TISH demonstrated good biocompatibility and cell penetration in its porous structure. DCs grown in TISH(G + R) under an in vitro hyperglycemic condition showed reduced maturation and activation markers such as CD80, CD83 and CD86, while simultaneously upregulated tolerogenic genes such as FOXP3, SOCS3, TGFß and IL10. Co-culture of these tolerogenic DCs with naïve T-cells induced regulatory T-cells differentiation, evidenced by elevated gene expressions of FOXP3, TGFß and IL-10. In vivo subcutaneous injection of TISH (G + R) into the mice showed significant infiltration of DCs and regulatory T-cells. In conclusion, TISH was developed and optimized as an injectable hydrogel to modulate DCs towards the tolerogenic phenotype and induce regulatory T-cells under hyperglycemia. TISH has promising potential to improve periodontal parameters in diabetic periodontitis.

PDMS hydrogel-coated tissue culture plates for studying the impact of substrate stiffness on dendritic cell function.

The mechanical properties of polydimethylsiloxane hydrogels can be tuned to mimic physiological tensions, an underappreciated environmental parameter in immunology studies. We describe a workflow to prepare PDMS-coated tissue culture plates with biologically relevant substrate stiffness, and the use of these hydrogel plates to condition isolated primary splenic CD11c+ dendritic cells (DC). Finally, we suggest downstream applications to study the impact of substrate stiffness on DC function and metabolism. The protocol could be adapted to study other mechanosensitive immune cell subsets. For complete details on the use and execution of this protocol, please refer to Chakraborty et al. (2021).

Assessing Macrophage Polarization in Nanoparticle-Guided Wound Repair Using a Lipopolysaccharide Contaminated Intraosseous Model.

INTRODUCTION: Macrophages regulate the processes of inflammation and tissue regeneration/repair through their plasticity and phenotypes of different activation states. Previous studies have shown that disinfection of lipopolysaccharide (LPS)-contaminated dentin with photoactivated rose bengal-functionalized chitosan nanoparticles (CSRBnps) in vivo supported neotissue formation without signs of inflammation and root resorption. The aim of this study was to understand the mechanism underlying CSRBnp-guided attenuation of inflammation in LPS-contaminated dentin using macrophage polarization as an indicator of inflammation and repair. METHODS: To quantify the polarized macrophage populations, M1/M2-specific surface markers CD68, CD80, and CD206 and transcriptional factors signal transducer and activator of transcription (STAT) 1, STAT3, and STAT6 were determined using immunohistochemistry among previously obtained root specimens implanted into mandibles of guinea pigs for 4 weeks. In group 1, the canals were not inoculated; in group 2, the canals were inoculated with Pseudomonas aeruginosa LPS; in group 3, the canals were inoculated and disinfected with sodium hypochlorite; in group 4, the canals were inoculated and disinfected with sodium hypochlorite and calcium hydroxide; and in group 5, the canals were inoculated and disinfected with sodium hypochlorite, and CSRBnps (300 µg/mL) with photoactivation (λ = 540 nm, 40 J/cm2) were analyzed. RESULTS: An increased expression of M2-specific markers was observed in the group treated with CSRBnps compared with the groups treated with either conventional or no root canal disinfection. A statistically significant population of macrophages expressing both M1- and M2-specific markers was observed in all the tested groups. CONCLUSIONS: Disinfection of LPS-contaminated dentin with CSRBnps demonstrated M2-type polarization of macrophages, which corresponded to repair and neotissue formation.

Development of a rat model for type 2 diabetes mellitus peri-implantitis: A preliminary study.

OBJECTIVE: To develop an in vivo model to simulate the complex internal environment of diabetic peri-implantitis (T2DM-PI) model for a better understanding of peri-implantitis in type 2 diabetic patients. MATERIALS AND METHODS: Maxillary first molars were extracted in Sprague-Dawley (SD) rats, and customized cone-shaped titanium implants were installed in the extraction sites. Thereafter, implants were uncovered and customized abutments were screwed into implants. A high-fat diet and a low-dose injection of streptozotocin were utilized to induce T2DM. Finally, LPS was locally injected in implant sulcus to induce peri-implantitis. RESULTS: In the present study, T2DM-PI model has been successfully established. Imaging analysis revealed that abundant inflammatory cells infiltrated in the soft tissue in T2DM-PI group with concomitant excessive secretion of inflammatory cytokines. Moreover, higher expression of MMP and increased number of osteoclasts led to collagen disintegration and bone resorption in T2DM-PI group. CONCLUSIONS: These results describe a novel rat model which stimulate T2DM-PI in vivo, characterized by overwhelming inflammatory response and bone resorption. This model has a potential to be used for investigation of initiation, progression and interventional therapy of T2DM-PI.

Biomaterial Properties Modulating Bone Regeneration.

Biomaterial scaffolds have been gaining momentum in the past several decades for their potential applications in the area of tissue engineering. They function as three-dimensional porous constructs to temporarily support the attachment of cells, subsequently influencing cell behaviors such as proliferation and differentiation to repair or regenerate defective tissues. In addition, scaffolds can also serve as delivery vehicles to achieve sustained release of encapsulated growth factors or therapeutic agents to further modulate the regeneration process. Given the limitations of current bone grafts used clinically in bone repair, alternatives such as biomaterial scaffolds have emerged as potential bone graft substitutes. This review summarizes how physicochemical properties of biomaterial scaffolds can influence cell behavior and its downstream effect, particularly in its application to bone regeneration.

Mechanical Stiffness Controls Dendritic Cell Metabolism and Function.

Stiffness in the tissue microenvironment changes in most diseases and immunological conditions, but its direct influence on the immune system is poorly understood. Here, we show that static tension impacts immune cell function, maturation, and metabolism. Bone-marrow-derived and/or splenic dendritic cells (DCs) grown in vitro at physiological resting stiffness have reduced proliferation, activation, and cytokine production compared with cells grown under higher stiffness, mimicking fibro-inflammatory disease. Consistently, DCs grown under higher stiffness show increased activation and flux of major glucose metabolic pathways. In DC models of autoimmune diabetes and tumor immunotherapy, tension primes DCs to elicit an adaptive immune response. Mechanistic workup identifies the Hippo-signaling molecule, TAZ, as well as Ca2+-related ion channels, including potentially PIEZO1, as important effectors impacting DC metabolism and function under tension. Tension also directs the phenotypes of monocyte-derived DCs in humans. Thus, mechanical stiffness is a critical environmental cue of DCs and innate immunity.

Impact of diabetes mellitus simulations on bone cell behavior through in vitro models.

Diabetes mellitus (DM) is related to impaired bone healing and an increased risk of bone fractures. While it is recognized that osteogenic differentiation and the function of osteoblasts are suppressed in DM, the influence of DM on osteoclasts is still unclear. Hyperglycemia and inflammatory environment are the hallmark of DM that causes dysregulation of various pro-inflammatory cytokines and alternated gene expression in periodontal ligament cells, osteoblasts, osteocytes, osteoclasts, and osteoclast precursors. A methodological review on conceptual and practical implications of in vitro study models is used for DM simulation on bone cells. Several major databases were screened to find literature related to the study objective. Published literature within last 20 years that used in vitro DM-simulated models to study how DM affects the cellular behavior of bone cells were selected for this review. Studies utilizing high glucose and serum acquired from diabetic animals are the mainly used methods to simulate the diabetic condition. The combination with various simulating factors such as lipopolysaccharide (LPS), hydrogen peroxide (H2O2), and advanced glycation end products (AGEs) have been reported in diabetic situations in vitro, as well. Through screening procedure, it was evident DM-simulated conditions exerted negative impact on bone-related cells. However, inconsistent results were found among different reported studies, which could be due to variation in culture conditions, concentrations of the stimulating factors and cell lineage, etc. This manuscript has concisely reviewed currently existing DM-simulated in vitro models and provides valuable insights of detailed components in simulating DM conditions in vitro. Studies using DM-simulated microenvironment revealed that in vitro simulation negatively impacted periodontal ligament cells, osteoblasts, osteocytes, osteoclasts, and osteoclast precursors. Contrarily, studies also indicated beneficial influence on bone-related cells when such conditions are reversed.

Effects of Programmed Local Delivery from a Micro/Nano-Hierarchical Surface on Titanium Implant on Infection Clearance and Osteogenic Induction in an Infected Bone Defect.

The two major causes for implant failure are postoperative infection and poor osteogenesis. Initial period of osteointegration is regulated by immunocytes and osteogenic-related cells resulting in inflammatory response and tissue healing. The healing phase can be influenced by various environmental factors and biological cascade effect. To synthetically orchestrate bone-promoting factors on biomaterial surface, built is a dual delivery system coated on a titanium surface (abbreviated as AH-Sr-AgNPs). The results show that this programmed delivery system can release Ag+ and Sr2+ in a temporal-spatial manner to clear pathogens and activate preosteoblast differentiation partially through manipulating the polarization of macrophages. Both in vitro and in vivo assays show that AH-Sr-AgNPs-modified surface renders a microenvironment adverse for bacterial survival and favorable for macrophage polarization (M2), which further promotes the differentiation of preosteoblasts. Infected New Zealand rabbit femoral metaphysis defect model is used to confirm the osteogenic property of AH-Sr-AgNPs implants through micro-CT, histological, and histomorphometric analyses. These findings demonstrate that the programmed surface with dual delivery of Sr2+ and Ag+ has the potential of achieving an enhanced osteogenic outcome through favorable immunoregulation.

Validation of Biofilm Assays to Assess Antibiofilm Efficacy in Instrumented Root Canals after Syringe Irrigation and Sonic Agitation.

INTRODUCTION: Different methods to characterize bacterial biofilms have been established, each presenting with distinct advantages and shortcomings. The aim of this study was to validate the ability of microbiological culture, the adenosine-5'-triphosphate (luminescence) assay, molecular, and microscopic methods to assess antibiofilm efficacy. METHODS: Thirty-nine extracted single-rooted teeth were selected. Enterococcus faecalis biofilms were grown for 21 days and randomly distributed into 3 groups. All canals were instrumented (F3 ProTaper Universal; Dentsply Sirona, Johnson City, TN) and irrigated (ProRinse needles, Dentsply Sirona) as follows: group 1, sodium hypochlorite and EDTA irrigation; group 2, supplemented with sonic agitation of NaOCl, and group 3, sterile distilled water irrigation. Bacteriologic samples were collected before (S1) and after canal preparation (S2) and subjected to quantification by culture methods, quantitative reverse transcriptase real-time PCR (qRT-PCR), and luminescence assay. The biofilm structure and bacterial cell viability were evaluated under scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Data were subjected to statistical analysis to determine the statistical significance (P < .05). RESULTS: S1 samples showed approximately 8-log colony-forming-units of bacteria using both culture and qRT-PCR. The reduction in bacterial population and relative luminescence was highly significant in the S2 samples from groups 1 and 2 (P < .001). SEM and CLSM showed well-matured root canal biofilms in the pretreatment samples that were reduced after treatment. Irrigation with NaOCl combined with sonic agitation significantly decreased the percentage of live cells (P < .05) but was not able to eliminate the biofilm structure. CONCLUSIONS: This study highlighted the maximum reduction of microbes after instrumentation-syringe irrigation. Although supplementary sonic agitation reduced the root canal biofilm further, it did not completely eliminate the biofilm from a single root canal model. The merits of combining microbiological and molecular quantification methods with CLSM for the comprehensive assessment of antibiofilm efficacy in root canals were emphasized.

Bioactivity of Photoactivated Functionalized Nanoparticles Assessed in Lipopolysaccharide-contaminated Root Canals In Vivo.

INTRODUCTION: The persistence of dentin-bound lipopolysaccharides (LPS) in disinfected root canals impedes treatment outcomes of endodontic procedures. This study assessed the effects of photoactivated rose bengal-functionalized chitosan nanoparticles (CSRBnps) on LPS-contaminated root dentin in vivo using an intraosseous implantation model and neotissue formation as a marker. METHODS: Fifty human, 3-mm-long root segments with a 1.2-mm canal lumen were divided into 5 groups (n = 10): group 1, canals not contaminated; group 2, canals contaminated with Pseudomonas aeruginosa LPS; group 3, canals contaminated and disinfected with sodium hypochlorite (NaOCl); group 4, canals contaminated and disinfected with NaOCl and calcium hydroxide; and group 5, canals contaminated and disinfected with NaOCl and CSRBnps (300 µg/mL) with photoactivation (λ = 540 nm, 40 J/cm2). Specimens were implanted into mandibles of guinea pigs, block dissected after 4 weeks, and the canal content evaluated histologically and immunohistochemically. The ingrown neotissue interface (50 µm) with dentin was characterized for fibroblasts, osteoclasts, inflammatory markers, dentin resorption, mineralization, and angiogenesis and dichotomized as type 1 (no inflammation and resorption, indicative of LPS inactivation) or type 2 (inflammation and resorption). The frequency of the observed parameters was analyzed using the Fisher exact test. RESULTS: The outcome was categorized as type 1 in groups 1 and 5, type 2 in group 2, and mixed type 1 and 2 in groups 3 and 4. The outcomes in groups 1 and 5 (P > .05) differed significantly (P < .05) from those in groups 2, 3, and 4. CONCLUSIONS: Disinfection of LPS-contaminated root canals with photoactivated CSRBnps in vivo supported ingrowth of neotissue without signs of inflammation or resorption, suggestive of effective inactivation of dentin-bound LPS.

Antibacterial Nanoparticles in Endodontics: A Review.

INTRODUCTION: A major challenge in root canal treatment is the inability of the current cleaning and shaping procedures to eliminate bacterial biofilms surviving within the anatomic complexities and uninstrumented portions of the root canal system. METHODS: Nanoparticles with their enhanced and unique physicochemical properties, such as ultrasmall sizes, large surface area/mass ratio, and increased chemical reactivity, have led research toward new prospects of treating and preventing dental infections. This article presents a comprehensive review on the scientific knowledge that is available on the application of antibacterial nanoparticles in endodontics. RESULTS: The application of nanoparticles in the form of solutions for irrigation, medication, and as an additive within sealers/restorative materials has been evaluated to primarily improve the antibiofilm efficacy in root canal and restorative treatments. In addition, antibiotic or photosensitizer functionalized nanoparticles have been proposed recently to provide more potent antibacterial efficacy. CONCLUSIONS: The increasing interest in this field warrants sound research based on scientific and clinical collaborations to emphasize the near future potential of nanoparticles in clinical endodontics.

Characterizing the collagen stabilizing effect of crosslinked chitosan nanoparticles against collagenase degradation.

UNLABELLED: Antibacterial and chelating properties of chitosan has been widely studied for various dental applications. OBJECTIVE: To characterize the interaction between chitosan-nanoparticles (CSnp) and collagen, and understand their stabilizing effect against collagenase degradation for dentin matrix stabilization. METHODS: Phase-1: a single Type I collagen-fibril model was used to study the interaction with CSnp along with carbodiimides crosslinking treatment. Degradation of the crosslinked fibrils was studied with bacterial collagenase enzyme and monitored using Fourier Transform Infrared (FTIR) spectroscopy, turbidity measurement (400nm), ninhydrin assay and Atomic Force Microscopy (AFM). Interaction of CSnp with collagenase and Type I collagen, were evaluated using SDS-PAGE, and proteolytic cleavage potential of a synthetic peptide. Phase-2: degradation of dentin collagen crosslinked with/without CSnp was evaluated using FTIR, ninhydrin assay and Scanning Electron Microscopy (SEM). Glutaraldehyde crosslinking was used as a positive control. RESULTS: Both native collagen-fibrils and dentin collagen after crosslinking showed higher resistance to collagenase degradation, as observed in turbidity measurements and FTIR spectra. AFM images showed the interaction of CSnp with single collagen-fibril and crosslinked collagen resisted collagenase degradation up to 54h. The collagen and collagenase both formed complexes with CSnp resulting in thickening of bands and reduction in collagen degradation. CSnp treated collagenase showed significantly reduced cleavage of the fluorescent peptides. Dentin collagen was coated with CSnp following crosslinking with significant increase in resistance to collagenase degradation. SIGNIFICANCE: Crosslinked CSnp on collagen stabilized and enhanced the resistance of dentin matrix against bacterial collagenase degradation due to non-specific interaction with both collagen and collagenase.

Antibacterial Properties Associated with Chitosan Nanoparticle Treatment on Root Dentin and 2 Types of Endodontic Sealers.

INTRODUCTION: The aim of this study was to evaluate the efficacy of carboxymethyl chitosan (CMCS) and chitosan nanoparticles (CNps) to inactivate bacteria and prevent biofilm formation at sealer-dentin interfaces. METHODS: The study was divided into 3 stages: first stage, the experiment was conducted to analyze the antibacterial properties of CMCS in different formulations against biofilms; second stage, direct-contact and membrane-restricted methods were used to evaluate the antibacterial properties of an epoxy resin (ThermaSeal Plus; Dentsply Tulsa Dental, Tulsa, OK) and calcium silicate (MTA Fillapex; Angelus SA, Londrina, PR, Brazil) based-sealers with or without CNps; and third stage, biofilm formation at the sealer dentin interfaces of root dentin treated with CMCS and filled with gutta-percha and CNp incorporated sealer were analyzed after 1- and 4-week aging periods. The samples were treated and filled as follows: (1) distilled water: unaltered sealer (control group), (2) CMCS: sealer+CNps (CMCS group), and (3) CMCS/rose bengal: sealer+CNps (CMCS/RB group). Enterococcus faecalis was used to infect all the samples. Microbiological and microscopic analyses were used to assess the antibacterial characteristics. RESULTS: CMCS-based treatments effectively killed bacteria adherent on root dentin (P < .05). The addition of CNps to ThermaSeal enhanced its antibacterial ability by direct-contact and membrane-restricted tests (P < .05). The CNp incorporation significantly increased the antibacterial efficacy of root canal sealers even after a 4-week aging time (P < .05). CONCLUSIONS: This study highlighted the ability of CMCS to disinfect root canal dentin and inhibit bacterial adhesion. CNps in root canal sealers are capable of maintaining their antibacterial activity even after prolonged aging.

Photoactivated polycationic bioactive chitosan nanoparticles inactivate bacterial endotoxins.

INTRODUCTION: The current root canal disinfection protocols fail to markedly inactivate bacterial endotoxins from infected root dentin. This study aimed to evaluate the ability of antibacterial photodynamic therapy with chitosan-conjugated rose bengal nanoparticles (CSRBnps) to selectively inactivate endotoxins/lipopolysaccharides (LPSs). METHODS: Antimicrobial agents such as calcium hydroxide (Ca[OH]2), chitosan nanoparticles (CSnps), CSRBnps, and methylene blue (MB) were assessed for their ability to neutralize LPSs obtained from Pseudomonas aeruginosa in a time-dependent interaction with/without photoactivation (20 and 40 J/cm(2)). The inflammatory potential of the treated/untreated LPSs was assessed on macrophage cells (RAW 267.4) using nitric oxide- and enzyme-linked immunosorbent assay (tumor necrosis factor α and interleukin-6 expression)-based analysis. These antimicrobials were tested directly on macrophage cells for cytotoxicity using the mitochondrial activity assay and light microscopy. The data were analyzed using 1-way analysis of variance and the Tukey test. RESULTS: CSnps were least effective in LPS inactivation. Interluekin-6 expression was reduced only with CSRBnp treatment. CSnps and CSRBnps were completely nontoxic, and MB showed slight toxicity to macrophage cells. Ca(OH)2 was highly cytotoxic (P < .005) even at 30 minutes of exposure. CSRBnps and MB with/without photoactivation significantly inactivated LPSs with reduced nitric oxide and tumor necrosis factor α expression (P < .05). Cell death and detachment after Ca(OH)2 treatment resulted in complete absence of all 3 inflammatory markers. CONCLUSIONS: Photodynamically activated CSRBnps caused significant inactivation of endotoxins and the subsequent reduction of all tested inflammatory markers from activated macrophages. Antimicrobial CSRBnps in combination with photodynamic therapy showed the potential to effectively inactivate bacterial endotoxins.

Antibiofilm efficacy of photosensitizer-functionalized bioactive nanoparticles on multispecies biofilm.

INTRODUCTION: Newer disinfection strategies based on antibacterial nanoparticles and photodynamic therapy (PDT) aim to eliminate residual biofilm bacteria during root canal treatment. The aim of the current study was to test the newly developed rose bengal-functionalized chitosan nanoparticles (CSRBnps) for their interaction/uptake with monospecies bacteria/biofilm and assess their antibiofilm efficacy on a multispecies biofilm model in vitro. METHODS: The interaction of CSRBnps with bacterial cells was conducted using atomic force microscopy. Their membrane-damaging effect was determined by measuring the absorbance at 260 nm (OD260nm) using Enterococcus faecalis. The penetration of CSRBnps into E. faecalis biofilms was evaluated using confocal laser scanning microscopy (CLSM). Multispecies biofilms of Streptococcus oralis, Prevotella intermedia, and Actinomyces naeslundii were grown on dentin sections for 21 days to assess the antibiofilm efficacy. The biofilms were subjected to PDT (60 J/cm(2)) using CSRBnps and rose bengal. The treated/untreated biofilms were examined under scanning electron microscopy and CLSM. RESULTS: The CSRBnps synthesized were 60 ± 20 nm and showed absorption spectra similar to rose bengal. Atomic force microscopy showed adherence of CSRBnps to bacteria, roughening of cell surface, and cell disruption after PDT. CSRBnp treatment resulted in significantly increased bacterial membrane damage (P < .05). CSRBnps exhibited deeper penetration into the biofilm structure. Scanning electron microscopy and CLSM confirmed the complete disruption of multispecies biofilm with a reduction in viable bacteria and biofilm thickness (P < .05). CONCLUSIONS: These novel photosensitizer functionalized bioactive nanoparticles with increased affinity to bacterial cell membrane, higher penetration into biofilm structure, and enhanced ability to eliminate clinically relevant multispecies bacterial biofilm present a potential antibiofilm agent for root canal disinfection.

Antibacterial efficacy of photosensitizer functionalized biopolymeric nanoparticles in the presence of tissue inhibitors in root canal.

INTRODUCTION: Application of antibacterial nanoparticles to improve root canal disinfection has received strong interest recently. The current study aims to assess the antibacterial effect of a novel photosensitizer (rose bengal functionalized chitosan nanoparticles [CSRBnp]) to eliminate bacteria in the presence of various root canal constituents that are known to inhibit the antibacterial efficacy of root canal disinfectants. METHODS: The synthesized CSRBnp were evaluated for size, charge, and singlet oxygen release. The antibacterial effect of CSRBnp was tested on planktonic Enterococcus faecalis with or without pretreatment by using different inhibiting agents such as dentin, dentin-matrix, pulp tissue, bacterial lipopolysaccharides, and bovine serum albumin (BSA). Bacterial survival was assessed in a time-dependent manner. The antibacterial effects after photodynamic activation on CSRBnp, a cationic photosensitizer (methylene blue), and an anionic photosensitizer (rose bengal [RB]) in the presence of inhibitors were also evaluated. RESULTS: CSRBnp were 60 ± 20 nm in size and showed reduced rate of singlet oxygen release as compared with methylene blue and RB. Pulp and BSA inhibited the antibacterial effect of CSRBnp (without photoactivation) significantly (P < .05) even after 24 hours of interaction. In case of photodynamic therapy, the pulp and BSA significantly inhibited the antibacterial activity of all 3 photosensitizers. CSRBnp showed residual effect and completely eliminated the bacteria after 24 hours of interaction after photodynamic therapy. CONCLUSIONS: The inherent antibacterial activity of polycationic chitosan nanoparticles and the singlet oxygen released after photoactivation of RB synergistically provided CSRBnp the potential to achieve significant antibacterial efficacy even in the presence of tissue inhibitors within root canals.

Photoactivated rose bengal functionalized chitosan nanoparticles produce antibacterial/biofilm activity and stabilize dentin-collagen.

Treatment of infected teeth presents two major challenges: persistence of the bacterial-biofilm within root canals after treatment and compromised structural integrity of the dentin hard-tissue. In this study bioactive polymeric chitosan nanoparticles functionalized with rose-bengal, CSRBnp were developed to produce antibiofilm effects as well as stabilize structural-integrity by photocrosslinking dentin-collagen. CSRBnp were less toxic to fibroblasts and had significant antibacterial activity even in the presence of bovine serum albumin. CSRBnp exerted antibacterial mechanism by adhering to bacterial cell surface, permeabilizing the membrane and lysing the cells subsequent to photodynamic treatment. Photoactivated CSRBnp resulted in reduced viability of Enterococcus faecalis biofilms and disruption of biofilm structure. Incorporation of CSRBnp and photocrosslinking significantly improved resistance to degradation and mechanical strength of dentin-collagen (P<0.05). The functionalized chitosan nanoparticles provided a single-step treatment of infected root dentin by combining the properties of chitosan and that of photosensitizer to eliminate bacterial-biofilms and stabilize dentin-matrix. FROM THE CLINICAL EDITOR: In this study, bioactive polymeric chitosan nanoparticles functionalized with rose-bengal (a photosensitizer), CSRBnp were developed to produce antibiofilm effects as well as stabilize structural-integrity of dental root dentin by photocrosslinking dentin-collagen, leading to efficient elimination of bacterial-biofilms and stabilization of dentin-matrix.

The effect of tissue inhibitors on the antibacterial activity of chitosan nanoparticles and photodynamic therapy.

INTRODUCTION: Newer antibacterial alternatives such as chitosan nanoparticles (CSnps) and photodynamic therapy (PDT) have been investigated to achieve effective root canal disinfection. The current study aims to assess the effect of various tissue inhibitors such as dentin, dentin matrix, pulp tissue, bacterial lipopolysaccharides (LPSs), and bovine serum albumin (BSA) on the antibacterial activity of CSnps and PDT. METHODS: The antibacterial effect of CSnps and PDT using photosensitizers, rose bengal (RB), and methylene blue (MB) were tested on planktonic Enterococcus faecalis American Type Culture Collection 29212 with or without pretreatment using different tissue inhibitors for an hour. Bacterial survival was assessed after 1, 8, and 24 hours of incubation with CSnps and after PDT using RB and MB. RESULTS: Pulp and BSA inhibited the antibacterial effect of CSnps significantly (P < .05). The antibacterial effect of CSnps was not affected by dentin, dentin matrix, or LPSs. The antibacterial activity of PDT using MB and RB was inhibited in a decreasing order by dentin matrix, BSA, pulp, dentin, and LPSs (P < .05). The effect of tissue inhibitors was higher in the case of PDT with RB. Depending on the antibacterial mechanism of CSnps and PDT, different inhibitory patterns were observed with different tissue inhibitors. CONCLUSIONS: The tissue inhibitors existing within the root canal affected the antibacterial activity of CSnps and PDT at varying degrees. Further research is required to enhance their antimicrobial efficacy in an endodontic environment.