Development of Antibacterial Resin Composites Incorporating Poly(METAC) Clusters.

This study examined the antibacterial effects and physical properties of a novel resin composite incorporating poly[{2-(methacryloyloxy)ethyl}trimethylammonium chloride] (poly(METAC)), a methacrylate cationic polymer comprising quaternary ammonium compounds (QACs). Resin composites incorporating poly(METAC) were fabricated by adding 6 wt.% METAC aqueous solution to a commercially available resin composite. The FE-SEM/EDS and Raman spec-troscopy analyses showed that METAC was assembled and polymerized in the resin composites after curing. The antibacterial effect was evaluated by inoculating Streptococcus mutans or Strepto-coccus sobrinus suspensions on the surface of cured resin composites, and the experimental resin composites incorporating poly(METAC) clusters exhibited bactericidal effects even after 28 days of ageing. The physical properties of the experimental resin composites were within the ISO-stipulated ranges. Newly fabricated resin composites containing the QAC-based poly(METAC) cluster ex-hibited long-term bactericidal effects against oral bacteria on their surfaces and demonstrated ac-ceptable physical properties for clinical use.

Poly(lactic acid/caprolactone) bilayer membrane achieves bone regeneration through a prolonged barrier function.

Guided bone regeneration (GBR) is a treatment strategy used to recover bone volume. Barrier membranes are a key component of GBR protocols, and their properties can impact treatment outcomes. This study investigated the efficacy of an experimental, slow-degrading, bilayer barrier membrane for application in GBR using in vivo animal models. A synthetic copolymer of poly(lactic acid/caprolactone) (PLCL) was used to prepare a slow-degrading bilayer membrane. The biodegradability of PLCL was evaluated by subcutaneous implantation in a rat model. The barrier function of the PLCL membrane was investigated in a rat calvaria defect model and compared with commercially available membranes composed of type I collagen (Col) and poly(lactic-co-glycolic acid) (PLGA). An alveolar bone defect model in beagle dogs was used to simulate GBR protocols to evaluate the bone regeneration ability of the experimental PLCL membrane. The PLCL membrane showed slow biodegradation, resulting in an efficient and prolonged barrier function compared with commercial materials. In turn, this barrier function enabled the space-making ability of PLCL membrane and facilitated bone regeneration. In the alveolar bone defect model, significantly greater regeneration was achieved by treatment with PLCL membrane compared with Col and PLGA membranes. Additionally, a continuous alveolar ridge contour was observed in PLCL-treated bone defects. In conclusion, the PLCL bilayer membrane is a promising biomaterial for use in GBR given its slow degradation and prolonged barrier function.

Poly(lactic acid/caprolactone) bilayer membrane blocks bacterial penetration.

BACKGROUND AND OBJECTIVE: The clinical outcomes of guided tissue regeneration (GTR) or guided bone regeneration (GBR) procedures can be impaired if a bacterial infection develops at the surgical site. Membrane exposure is one of the causes of the onset of bacterial infection. Previously, we have fabricated a poly(lactic acid/caprolactone) (PLCL) bilayer membrane composed of a porous layer and a compact layer. The compact layer acts as a barrier against connective tissue and epithelial cells, and we hypothesized that it could also be an effective barrier against bacterial cells. The objective of this study was to evaluate the ability of the PLCL bilayer membrane to block bacterial cell penetration, which would be useful for preventing postoperative infections. METHODS: Porphyromonas gingivalis, Streptococcus mutans, and multispecies bacteria collected from human saliva were used in this study. Bacteria were seeded directly on the compact layer of a PLCL bilayer membrane, and bacterial adhesion to the membrane, as well as penetration into the membrane's structure, were assessed. Bacterial adhesion was evaluated by the number of colonies formed at 6, 24, and 72 h, and penetration was observed using a scanning electron microscope at 24 and 72 h. Commercially available membranes, composed of poly(lactic-co-glycolic acid) or type I collagen, were used as controls. RESULTS: P. gingivalis, S. mutans, and the multispecies bacteria obtained from human saliva adhered onto all the membranes after only 6 h of incubation. However, fewer adherent cells were observed for the PLCL bilayer membrane compared with the controls for all experimental periods. The PLCL membrane was capable of blocking bacterial penetration, and no bacterial cells were observed in the structure. In contrast, bacteria penetrated both the control membranes and were observed at depths of up to 80 µm after 72 h of incubation. CONCLUSION: Membrane characteristics may influence how bacterial colonization occurs. The PLCL membrane had reduced bacterial adhesion and blocked bacterial penetration, and these characteristics could contribute to a favorable outcome for regenerative treatments. In the event of membrane exposure at GTR/GBR surgical sites, membranes with an efficient barrier function, such as the PLCL bilayer membrane, could simplify the management of GTR/GBR complications.

Establishment of novel in vitro culture system with the ability to reproduce oral biofilm formation on dental materials.

Intensive research has been conducted with the aim of developing dental restorative/prosthetic materials with antibacterial and anti-biofilm effects that contribute to controlling bacterial infection in the oral cavity. In situ evaluations were performed to assess the clinical efficacy of these materials by exposing them to oral environments. However, it is difficult to recruit many participants to collect sufficient amount of data for scientific analysis. This study aimed to assemble an original flow-cell type bioreactor equipped with two flow routes and assess its usefulness by evaluating the ability to reproduce in situ oral biofilms formed on restorative materials. A drop of bacterial suspension collected from human saliva and 0.2% sucrose solution was introduced into the assembled bioreactor while maintaining the incubation conditions. The bioreactor was able to mimic the number of bacterial cells, live/dead bacterial volume, and volume fraction of live bacteria in the in situ oral biofilm formed on the surface of restorative materials. The usefulness of the established culture system was further validated by a clear demonstration of the anti-biofilm effects of a glass-ionomer cement incorporating zinc-releasing glasses when evaluated by this system.

Antibacterial effects and physical properties of a glass ionomer cement containing BioUnion filler with acidity-induced ability to release zinc ion.

BioUnion filler is a bioactive glass particle that releases Zn2+ in an acidic environment. In this study, the ion release, antibacterial, and physical properties of a glass ionomer cement (GIC) incorporating BioUnion filler (CA) were assessed in vitro. The concentration of Zn2+ released from CA into acetic acid was higher than that released into water and its minimum inhibitory concentrations against six oral bacterial species. Moreover, the concentration of Zn2+-release was maintained during all the seven times it was exposed to acetic acid. Compared to a conventional cement and resin composite, CA significantly inhibited the growth of oral bacteria and hindered their adhesion on the material surface. Thus, our study outcomes show that the release of Zn2+ from CA in the acidic environment does not affect its compressive strength.

Barrier membranes for tissue regeneration in dentistry.

Background: In dentistry, barrier membranes are used for guided tissue regeneration (GTR) and guided bone regeneration (GBR). Various membranes are commercially available and extensive research and development of novel membranes have been conducted. In general, membranes are required to provide barrier function, biosafety, biocompatibility and appropriate mechanical properties. In addition, membranes are expected to be bioactive to promote tissue regeneration. Objectives: This review aims to organize the fundamental characteristics of the barrier membranes that are available and studied for dentistry, based on their components. Results: The principal components of barrier membranes are divided into nonbiodegradable and biodegradable materials. Nonbiodegradable membranes are manufactured from synthetic polymers, metals or composites of these materials. The first reported barrier membrane was made from expanded polytetrafluoroethylene (e-PTFE). Titanium has also been applied for dental regenerative therapy and shows favorable barrier function. Biodegradable membranes are mainly made from natural and synthetic polymers. Collagens are popular materials that are processed for clinical use by cross-linking. Aliphatic polyesters and their copolymers have been relatively recently introduced into GTR and GBR treatments. In addition, to improve the tissue regenerative function and mechanical strength of biodegradable membranes, inorganic materials such as calcium phosphate and bioactive glass have been incorporated at the research stage. Conclusions: Currently, there are still insufficient guidelines for barrier membrane choice in GTR and GBR, therefore dentists are required to understand the characteristics of barrier membranes.

Development of endodontic sealers containing antimicrobial-loaded polymer particles with long-term antibacterial effects.

OBJECTIVE: The objective of this study is to prepare new dental resins with a long-lasting antimicrobial activity. Specifically, this study evaluates an approach for controlling infection in root canals using sealers containing polyhydroxyethyl methacrylate trimethylolpropane trimethacrylate (polyHEMA/TMPT) particles loaded with cetylpyridinium chloride (CPC). In addition, the physical properties of sealers containing CPC-loaded polyHEMA/TMPT particles (CLP) are determined. METHODS: PolyHEMA/TMPT particles with 10 (10%-CLP) and 25wt.% CPC (25%-CLP) with different particle sizes were fabricated and incorporated in HEMA-based sealers. CPC-release profiles were evaluated over 14 days of immersion in water, followed by 14 days of storage and 14 days of water immersion. The antibacterial activity of these sealers against Enterococcus faecalis in dentinal tubules was assessed using a root-canal-infection model. Their sealing abilities were evaluated by fluid filtration and physical properties were tested according to the ISO 6876 standard. The long-term antibacterial activity of the cured sealer containing 25%-CLP (∼21µm particle diameter) was re-assessed after 1 year of storage. RESULTS: After 28 days of immersion, 25%-CLP exhibited a higher and sustained CPC release unlike 10%-CLP. Residual bacteria in root dentinal tubules were eradicated by obturation with 25%-CLP-containing sealers. The incorporation of 25%-CLP (∼21µm) had no adverse effects on the sealing ability and physical properties of the sealer and resulted in long-term antibacterial activity. SIGNIFICANCE: The incorporation of CPC-loaded particles in HEMA resins yielded endodontic sealers with long-term bactericidal activity against E. faecalis in root canals. These sealers can potentially be used to prevent recurrent apical periodontitis.

Evaluation of ion release and the recharge ability of glass-ionomer cement containing BioUnion filler using an in vitro saliva-drop setting assembly.

OBJECTIVE: A glass-ionomer cement (GIC) containing BioUnion filler has been reported to release Zn2+ under acidic conditions and to inhibit oral bacteria on its surface. However, previous results are based on in vitro experiments under static conditions. This study aimed to assemble an in vitro saliva-drop setting to simulate in vivo conditions of the oral cavity and to investigate the ion releasing and recharging properties of the GIC containing BioUnion filler. METHODS: The effective concentrations of Zn2+ and F- against Streptococcus mutans and saliva-derived multi-species biofilms were determined. Artificial saliva was dropped on the GIC containing BioUnion filler using the in vitro saliva-drop setting assembly and was periodically replaced with acetic acid. Ion release/recharge properties were investigated by measuring the release concentrations of Zn2+ and F-. RESULTS: The concentration of Zn2+ released from the BioUnion filler-containing GIC during seven days with repeated exposure to acid could be maintained at the level to inhibit S. mutans and saliva-derived multi-species biofilm formation. Moreover, the BioUnion filler-containing GIC could be recharged with Zn2+ and F- by the application of a tooth gel containing Zn2+ and F-. The release concentration of Zn2+ after recharging was significantly higher than the effective concentration of Zn2+ to hinder S. mutans and saliva-derived multi-species biofilm formation on material surfaces. SIGNIFICANCE: The GIC containing BioUnion filler was shown to have the potential to inhibit biofilm formation in the oral cavity. In addition, recharging Zn2+ and F- would further enhance the effect of the GIC containing BioUnion filler.

FGF-2 release and bonding/physical properties of 4-META/MMA-based adhesive resins incorporating small FGF-2-loaded polymer particles.

OBJECTIVES: Non-biodegradable particles comprising hydroxyethyl methacrylate (HEMA) and trimethylolpropane trimethacrylate (TMPT) have been reported as useful carriers for fibroblast growth factor-2 (FGF-2). They have also been successfully incorporated into the 4-[2-(methacryloyloxy)ethoxycarbonyl]phthalic anhydride/methyl methacrylate-tri-n-butyl borane (4-META/MMA-TBB) resin to promote tissue regeneration. However, smaller particles are required to obtain restorative materials acceptable for clinical use. The aim of this study was to investigate the ability of the 4-META/MMA-TBB resin that comprises small FGF-2-loaded particles to release FGF-2 and promote cell proliferation. In addition, the bonding and physical properties of the experimental resin were evaluated. METHODS: The small particles loaded with FGF-2 were newly fabricated and incorporated into the commercial 4-META/MMA-TBB resin. Release profiles of FGF-2 from the experimental resins were assessed, and the cell proliferation cultured with the eluate was evaluated. The bonding and physical properties of the resins were evaluated using shear bond strength and three-point bending tests, and by measuring the curing time, water absorption, and water dissolution. RESULTS: Sustained release of FGF-2 from the experimental resins for two weeks was observed, and the released FGF-2 was demonstrated to promote cell proliferation. All bonding and physical properties of the 4-META/MMA-TBB resins were found acceptable for clinical use. SIGNIFICANCE: The small FGF-2-loaded particles incorporated into the 4-META/MMA-TBB resin had the same abilities to release FGF-2 and proliferate cells, as those exhibited by the conventionally sized particles. In addition, there were no adverse influences on bonding and physical properties, suggesting that the bioactive adhesive resin was acceptable for clinical use.

Fabrication of novel poly(lactic acid/caprolactone) bilayer membrane for GBR application.

OBJECTIVE: Guided bone regeneration (GBR) often involves the use of membranes as barriers for soft tissues. Commercially available membranes, however, do not possess an adequately low degradation rate, resulting in limited barrier function. The purpose of this study was to develop and assess the physicochemical and biological characteristics of a novel poly(l-lactic acid/caprolactone) (PLCL) bilayer membrane and determine its usefulness for GBR application. METHODS: The experimental bilayer membrane was prepared via a two-step freezing and lyophilization process with a PLCL solution. Next, the PLCL membrane was investigated regarding tensile strength, surface roughness, in vitro degradation and clinical operability. In addition, cell proliferation and differentiation were investigated on each layer of the experimental membrane. For all experiments, a commercially available poly(lactic-co-glycolic) acid membrane was used as a control. RESULTS: In vitro analysis of the PLCL bilayer membrane revealed suitable mechanical strength combined with high breaking strain, which contributed to membrane operability. In addition, the PLCL bilayer membrane had enhanced stability compared to the commercial control due to its slower degradation, and was capable of supporting cell growth and osteogenic differentiation. SIGNIFICANCE: The current study confirmed that the PLCL membrane possessed a high biocompatibility and slow degradation rate that contributes to prolonged barrier function and bone regeneration. Altogether, it was considered that the PLCL bilayer membrane developed in this study was applicable for GBR treatment.

Acidity-induced release of zinc ion from BioUnionTM filler and its inhibitory effects against Streptococcus mutans.

BioUnion filler incorporated into restorative/coating materials is a new bio-functional glass powder. The most unique function of BioUnion filler is its ability to release Zn2+ in acidic environments. In this study, the ion release profile of BioUnion filler under acidic conditions and its antibacterial effects against Streptococcus mutans were evaluated. The concentrations of Zn2+ released from BioUnion fillers into acetic acids were greater than those released into water. S. mutans inhibition by BioUnion fillers was greater with sucrose than without sucrose, reflecting a decrease in suspension pH in response to the addition of sucrose. Exposure to acids increased Zn2+ release from BioUnion fillers, and the fillers after repeated exposure to acids demonstrated inhibitory effects against S. mutans. These findings suggest that BioUnion filler accelerated the release of Zn2+ under acidic conditions, which induced bactericidal/inhibitory effects against S. mutans.

Cutting-edge filler technologies to release bio-active components for restorative and preventive dentistry.

Advancements in materials used for restorative and preventive treatment is being directed toward "bio-active" functionality. Incorporation of filler particles that release active components is a popular method to create bio-active materials, and many approaches are available to develop fillers with the ability to release components that provide "bio-protective" or "bio-promoting" properties; e.g. metal/calcium phosphate nanoparticles, multiple ion-releasing glass fillers, and non-biodegradable polymer particles. In this review paper, recent developments in cutting-edge filler technologies to release bio-active components are addressed and summarized according to their usefulness and functions, including control of bacterial infection, tooth strengthening, and promotion of tissue regeneration.

Freeze-dry processing of three-dimensional cell constructs for bone graft materials.

Freeze-dry processing improves the operability and stability of cell-based biomaterials and facilitates sterilization for clinical application. However, there is no established freeze-drying protocol for engineered tissues. Recently, we reported that biomimetic bone tissues can be fabricated using scaffold-free three-dimensional (3D) cell constructs with potential applications as bone graft materials. The purpose of this study was to assess the influence of freeze drying on the morphology and components of 3D cell constructs. Cell constructs freeze dried in phosphate buffered saline (PBS) maintained organic and inorganic components; whereas sodium citrate buffer (SCB)-treated constructs had significantly lower amounts of calcium and bone-related proteins. Alkaline phosphatase (ALP) activity in cell constructs was maintained by freeze drying in 10% sucrose-containing PBS, whereas cell constructs treated with PBS without sucrose or with sucrose-containing SCB showed significant reductions of ALP activity. In this study, we found that sucrose-containing phosphate buffer was suitable for freeze drying to maintain minerals and protein functions within 3D cell constructs, whereas citrate buffer was inappropriate. The insights gained by this study may facilitate the development of novel cell-based biomaterials fabricated by tissue engineering approaches and bone graft biomaterials.

Development of a novel dental resin cement incorporating FGF-2-loaded polymer particles with the ability to promote tissue regeneration.

OBJECTIVE: Aiming to achieve bioactive dental resins that promote healing of surrounding tissues, we developed novel poly(2-hydroxyethyl methacrylate/trimethylolpropane trimethacrylate) (polyHEMA/TMPT) particles. These particles have been reported to be useful as a non-biodegradable carrier for fibroblast growth factor-2 (FGF-2) in vitro. The aim of this study was to evaluate the ability of an adhesive resin incorporating FGF-2-loaded polymer particles to promote tissue regeneration in vitro and in vivo. METHODS: Experimental adhesive resins were prepared by incorporating FGF-2-loaded polyHEMA/TMPT particles into a 4-META/MMA-based adhesive resin, and the release profiles of FGF-2 were evaluated. The proliferation of osteoblast-like cells in the eluate from cured experimental resin was assessed. When the experimental resin was implanted into rat calvaria defects, bone regeneration was evaluated by microcomputed tomography and histological observations. RESULTS: Sustained release of FGF-2 from the experimental resin was observed for 14 days. Eluate from the cured experimental resin significantly promoted the proliferation of osteoblast-like cells. Significantly greater bone regeneration was observed using the experimental resin compared with the control resin without FGF-2. SIGNIFICANCE: 4-META/MMA-based adhesive resin incorporating FGF-2-loaded polymer particles is useful to promote tissue regeneration, suggesting that its application would be beneficial for root-end filling or the repair of fractured roots in cases with severely damaged periodontal tissue.

Development of layered PLGA membranes for periodontal tissue regeneration.

OBJECTIVE: Various commercial products are available for guided tissue regeneration (GTR) therapy; however, they do not combine biosafety with the ability to control cell function. The purpose of this study was to evaluate the physicochemical and biological characteristics of the novel bilayer biodegradable poly(lactic-co-glycolic acid) (PLGA) membrane, and to assess whether the bilayer PLGA membrane could be used for periodontal tissue regeneration. METHODS: Bilayer biodegradable membrane was fabricated thorough a two-step freezing and lyophilization process using PLGA solution. The characteristics of bilayer membranes were evaluated with respect to surface morphology, stability, mechanical strength, and operability for clinical use. Cell proliferation and osteogenic differentiation were investigated on the each surface of bilayer membrane. Then, these membranes were implanted to the rat calvaria bone defect models and evaluated their capability for tissue regeneration. RESULTS: Biodegradable membranes composed of the solid and porous layer were successfully prepared and the surface morphologies analyzed. Physicochemical analyses revealed that the membranes possessed enough stability and mechanical properties for clinical use. It was also confirmed that the solid layer inhibited cell proliferation and subsequent connective tissue invasion, while the inner layer promoted proliferation and osteogenic differentiation, thus resulting in bone regeneration in vivo. SIGNIFICANCE: The layering technology used to fabricate the bilayer polymer membrane could be applied in the developing of other novel biomaterials. The present study demonstrates that the bilayer biodegradable polymer membranes facilitate tissue regeneration in vivo, and therefore represent a prospective biomaterial for GTR therapy.

Non-biodegradable polymer particles for drug delivery: A new technology for "bio-active" restorative materials.

To develop dental restorative materials with "bio-active" functions, addition of the capability to release active agents is an effective approach. However, such functionality needs to be attained without compromising the basic properties of the restorative materials. We have developed novel non-biodegradable polymer particles for drug delivery, aimed for application in dental resins. The particles are made using 2-hydroxyethyl methacrylate (HEMA) and a cross-linking monomer trimethylolpropane trimethacrylate (TMPT), with a hydrophilic nature to adsorb proteins or water-soluble antimicrobials. The polyHEMA/TMPT particles work as a reservoir to release fibroblast growth factor-2 (FGF-2) or cetylpyridinium chloride (CPC) in an effective manner. Application of the polyHEMA/TMPT particles loaded with FGF-2 to adhesives, or those loaded with CPC to resin-based endodontic sealers or denture bases/crowns is a promising approach to increase the success of the treatments by conferring "bio-active" properties to these materials to induce tissue regeneration or to inhibit bacterial infection.

Influence of polymerization properties of 4-META/MMA-based resin on the activity of fibroblast growth factor-2.

Dental adhesive resins based on 4-methacryloxyethyl trimellitate anhydride (4-META)/methyl methacrylate (MMA) have been utilized for root-end filling and the bonding of fractured roots. To increase the success rate of these treatments, it would be beneficial to promote the healing of surrounding tissue by applying growth factors. In this study, the influences of the polymerization properties of 4-META/MMA-based resins on the activity of fibroblast growth factor-2 (FGF-2) were evaluated in vitro. The temperature increase caused by the heat generation during polymerization of the 4-META/MMA-based resin was insufficient to change the structure and function of FGF-2. Unpolymerized monomers released from the cured 4-META/MMA-based resin had no negative influences on the ability of FGF-2 to promote the proliferation of osteoblast-like cells. These findings suggest that it is possible to use FGF-2 in combination with 4-META/MMA-based resins.

Effectiveness of non-biodegradable poly(2-hydroxyethyl methacrylate)-based hydrogel particles as a fibroblast growth factor-2 releasing carrier.

OBJECTIVES: Dental resin-based restorative materials are used in a variety of dental treatment modalities such as root-end filling, perforation sealing, and adhesion of fractured roots. However, the prognosis after such treatments is not necessarily favorable because they fail to promote healing of the surrounding alveolar tissue. In the present study, non-biodegradable poly-2-hydroxyethyl methacrylate (polyHEMA)-based hydrogel particles were fabricated as a carrier vehicle for drug delivery that is applied to dental resins. METHODS: The loading and release characteristics of bovine serum albumin (BSA) and fibroblast growth factor-2 (FGF-2) from the polyHEMA-based hydrogel particles were evaluated over time in culture. The hydrogel particles were immersed into an aqueous FITC-labeled BSA solution and were observed using confocal laser scanning microscopy (CLSM). To determine the activity of the FGF-2 released from the particles, the proliferation of osteoblast-like cells cultured with eluates collected from the particles for up to 14 days was determined. RESULTS: CLSM revealed that BSA was adsorbed to the surface of the hydrogel particles. A sustained release of BSA and FGF-2 from the particles was detected for up to 14 days. The eluates from the FGF-2-loaded particles increased the proliferation of the osteoblast-like cells, suggesting that the activity of FGF-2 was maintained for at least 2 weeks within the particles. SIGNIFICANCE: These polyHEMA-based non-degradable hydrogel particles may be useful tools that can be applied to dental restorative materials to achieve sustained delivery of drugs that promote tissue regeneration.