Fabrication of Ciprofloxacin-Immobilized Calcium Phosphate Particles for Dental Drug Delivery.

Calcium phosphate (CaP) particles immobilizing antibacterial agents have the potential to be used as dental disinfectants. In this study, we fabricated CaP particles with immobilized ciprofloxacin (CF), a commonly prescribed antibacterial agent, via a coprecipitation process using a supersaturated CaP solution. As the aging time in the coprecipitation process increased from 2 to 24 h, the CaP phase in the resulting particles transformed from amorphous to low-crystalline hydroxyapatite, and their Ca/P elemental ratio, yield, and CF content increased. Despite the higher CF content, the particles aged for 24 h displayed a slower release of CF in a physiological salt solution, most likely owing to their crystallized matrix (less soluble hydroxyapatite), than those aged for 2 h, whose matrix was amorphous CaP. Both particles exhibited antibacterial and antibiofilm activities along with an acid-neutralizing effect against the major oral bacteria, Streptococcus mutans, Porphyromonas gingivalis, and Actinomyces naeslundii, in a dose-dependent manner, although their dose-response relationship was slightly different. The aging time in the coprecipitation process was identified as a governing factor affecting the physicochemical properties of the resulting CF-immobilized CaP particles and their functionality as a dental disinfectant.

Fluoride-Incorporated Apatite Coating on Collagen Sponge as a Carrier for Basic Fibroblast Growth Factor.

Coating layers consisting of a crystalline apatite matrix with immobilized basic fibroblast growth factor (bFGF) can release bFGF, thereby enhancing bone regeneration depending on their bFGF content. We hypothesized that the incorporation of fluoride ions into apatite crystals would enable the tailored release of bFGF from the coating layer depending on the layer's fluoride content. In the present study, coating layers consisting of fluoride-incorporated apatite (FAp) crystals with immobilized bFGF were coated on a porous collagen sponge by a precursor-assisted biomimetic process using supersaturated calcium phosphate solutions with various fluoride concentrations. The fluoride content in the coating layer increased with the increasing fluoride concentration of the supersaturated solution. The increased fluoride content in the coating layer reduced its solubility and suppressed the burst release of bFGF from the coated sponge into a physiological salt solution. The bFGF release was caused by the partial dissolution of the coating layer and, thus, accompanied by the fluoride release. The concentrations of released bFGF and fluoride were controlled within the estimated effective ranges in enhancing bone regeneration. These findings provide useful design guidelines for the construction of a mineralized, bFGF-releasing collagen scaffold that would be beneficial for bone tissue engineering, although further in vitro and in vivo studies are warranted.

Dopamine is involved in reparative dentin formation through odontoblastic differentiation of dental pulp stem cells.

Conventional direct pulp-capping materials induce pulp cells to secrete various biomolecules in pulp tissues that promote reparative dentin formation through induction of odontoblastic differentiation of dental pulp stem cells (DPSCs). However, these biomolecules sometimes induce bone-like dentin with poor sealing properties. Therefore, exploration of biomolecules that allow tight sealing by tubular reparative dentin is required. We recently reported that dopamine (DA) is involved in dentinogenesis. Hence, we investigated the effect of DA on odontoblastic differentiation of DPSCs and reparative dentin formation. Both tyrosine hydroxylase (TH), a DA synthetase, and DA were expressed in odontoblast-like cells in vivo. In vitro, their expression was increased during odontoblastic differentiation of DPSCs. Furthermore, TH-overexpressing DPSCs had promoted odontoblastic differentiation and DA production. Moreover, DA stimulation promoted their differentiation and induced tubular reparative dentin. These results suggest that DA produced by TH is involved in odontoblastic differentiation of DPSCs and has an inductive capacity for reparative dentin formation similar to primary dentin. This study may lead to the development of therapy to preserve vital pulp tissues.

Water-resistant antibacterial properties of a graphene oxide/cetylpyridinium chloride complex formed on medical gauze fibers.

OBJECTIVES: Graphene oxide (GO) is a nanocarbon material with a high aspect ratio (width:thickness) and abundant anionic functional groups on its surface. In this study, we attached GO to the surface of medical gauze fibers, constructed a complex with a cationic surface active agent (CSAA), and demonstrated that the treated gauze exhibits antibacterial activity even after rinsing with water. METHODS: Medical gauze was immersed in GO dispersion (0.001%, 0.01%, and 0.1%), rinsed with water, dried, and subjected to the Raman spectroscopy analysis. Subsequently, the gauze treated with 0.001% GO dispersion was immersed in 0.1% cetylpyridinium chloride (CPC) solution, immediately rinsed with water, and dried. Untreated, GO-only, and CPC-only gauzes were prepared for comparison. Each gauze was placed in a culture well, seeded with Escherichia coli or Actinomyces naeslundii, and turbidity was measured after 24 h of incubation. RESULTS: The Raman spectroscopy analysis of the gauze after immersion and rinsing showed a G band peak, indicating that GO remained on the surface of the gauze. The turbidity measurements indicated that GO/CPC-treated gauze (GO-treated and rinsed, followed by CPC-treatment and rinsing) significantly decreased turbidity compared to the other gauzes (P < 0.05), suggesting that the GO/CPC complex remained on the gauze fibers even after water rinsing and showed antibacterial activity. CONCLUSIONS: The GO/CPC complex imparts water-resistant antibacterial properties to gauze and has the potential to be widely used for the antimicrobial treatment of clothes.

Periodontal tissue regeneration by recombinant human collagen peptide granules applied with ß-tricalcium phosphate fine particles.

OBJECTIVES: Recombinant human collagen peptide (RCP) is a recombinantly created xeno-free biomaterial enriched in arginine-glycine-aspartic acid sequences with good processability whose use for regenerative medicine applications is under investigation. The biocompatibility and osteogenic ability of RCP granules combined with ß-tricalcium phosphate (TCP) submicron particles (ß-TCP/RCP) were recently demonstrated. In the present study, ß-TCP/RCP was implanted into experimental periodontal tissue defects created in beagles to investigate its regenerative effects. METHODS: An RCP solution was lyophilized, granulated, and thermally cross-linked into particles approximately 1 mm in diameter. ß-TCP dispersion (1 wt%; 500 µL) was added to 100 mg of RCP granules to form ß-TCP/RCP. A three-walled intrabony defect (5 mm × 3 mm × 4 mm) was created on the mesial side of the mandibular first molar and filled with ß-TCP/RCP. RESULTS: A micro-computed tomography image analysis performed at 8 weeks postoperative showed a significantly greater amount of new bone after ß-TCP/RCP grafting (2.2-fold, P < 0.05) than after no grafting. Histological findings showed that the transplanted ß-TCP/RCP induced active bone-like tissue formation including tartaric acid-resistant acid phosphatase- and OCN-positive cells as well as bioabsorbability. Ankylosis did not occur, and periostin-positive periodontal ligament-like tissue formation was observed. Histological measurements performed at 8 weeks postoperative revealed that ß-TCP/RCP implantation formed 1.7-fold more bone-like tissue and 2.1-fold more periodontal ligament-like tissue than the control condition and significantly suppressed gingival recession and epithelial downgrowth (P < 0.05). CONCLUSIONS: ß-TCP/RCP implantation promoted bone-like and periodontal ligament-like tissue formation, suggesting its efficacy as a periodontal tissue regenerative material.

Rose bengal-decorated rice husk-derived silica nanoparticles enhanced singlet oxygen generation for antimicrobial photodynamic inactivation.

Rice husks are well known for their high silica content, and the RH-derived silica nanoparticles (RH NPs) are amorphous and biocompatible; therefore, they are suitable raw materials for biomedical applications. In this study, rose bengal-impregnated rice husk nanoparticles (RB-RH NPs) were prepared for their potential photosensitization and 1O2 generation as antimicrobial photodynamic inactivation. RB is a halogen-xanthene type's photosensitizer showing high singlet oxygen efficiency, and the superior photophysical properties are desirable for RB in the antimicrobial photodynamic inactivation of bacteria. To enhance the binding of anionic RB to RH NPs, we conducted cationization for the RH NPs using polyethyleneimine (PEI). The control of the RB adsorption state on cationic PEI-modified RH NPs was essential for RB RH-NP photosensitizers to obtain efficient 1O2 generation. Minimizing RB aggregation allowed highly efficient 1O2 production from RB-RH NPs at the molar ratio of RB with the PEI, XRB/PEI. = 0.1. The RB-RH NPs have significant antimicrobial activity against Streptococcus mutans compared to free RB after white light irradiation. The RB-RH NP-based antimicrobial photodynamic inactivation can be employed effectively in treating Streptococcus mutans for dental applications. Supplementary Information: The online version contains supplementary material available at 10.1007/s10853-023-08194-z.

Fluoridated Apatite Coating on Human Dentin via Laser-Assisted Pseudo-Biomineralization with the Aid of a Light-Absorbing Molecule.

A simple, area-specific coating technique for fluoridated apatite (FAp) on teeth would be useful in dental applications. Recently, we achieved area-specific FAp coating on a human dentin substrate within 30 min by a laser-assisted biomimetic (LAB) process; pulsed Nd:YAG laser irradiation in a fluoride-containing supersaturated calcium phosphate solution (FCP solution). The LAB-processed, FAp-coated dentin substrate exhibited antibacterial activity against a major oral bacterium, Streptococcus mutans. In the present study, we refined the LAB process with a combination of a dental diode laser and a clinically approved light-absorbing molecule, indocyanine green (ICG). A micron-thick FAp layer was successfully formed on the dentin surface within only 3 min by the refined LAB process, i.e., dental diode laser irradiation in the FCP solution following ICG treatment. The ICG layer precoated on the dentin substrate played a crucial role in inducing rapid pseudo-biomineralization (FAp layer formation) on the dentin surface by absorbing laser light at the solid-liquid interface. In the refined LAB process, the precoated ICG layer was eliminated and replaced with the newly formed FAp layer composed of vertically oriented pillar-like nanocrystals. Cross-sectional ultrastructural analysis revealed a smooth interface between the FAp layer and the dentin substrate. The refined LAB process has potential as a tool for the tooth surface functionalization and hence, is worth further process refinement and in vitro and in vivo studies.

Sustained antibacterial coating with graphene oxide ultrathin film combined with cationic surface-active agents in a wet environment.

Antimicrobial surfactants contained in mouthrinse have excellent efficacy, but are not retained on the tooth surface (are rinsed away) due to their low water resistance and thus do not exhibit sustained antibacterial activity. We have developed a new coating method using graphene oxide (GO) that retains the surfactant on the tooth surface even after rinsing with water, thus providing a sustained antibacterial effect. Ultra-thin films of GO and an antimicrobial agent were prepared by (1) applying GO to the substrate surface, drying, and thoroughly rinsing with water to remove excess GO to form an ultrathin film (almost a monolayer, transparent) on the substrate surface, then (2) applying antimicrobial cationic surface active agents (CSAAs) on the GO film to form a composite coating film (GO/CSAA). GO/CSAA formation was verified by scanning electron microscopy, Raman spectroscopy, X-ray photoelectron spectroscopy, and ζ-potential and contact angle measurements. GO/CSAA was effective at inhibiting the growth of oral pathogens for up to 7 days of storage in water, and antibacterial activity was recovered by reapplication of the CSAA. Antibacterial GO/CSAA films were also formed on a tooth substrate. The results suggest that GO/CSAA coatings are effective in preventing oral infections.

Osteoclast formation from mouse bone marrow cells on micro/nano-scale patterned surfaces.

OBJECTIVES: Osteoclasts can sense the surface topography of materials. However, it is difficult to identify the structural factors that affect osteoclast formation and its function. Furthermore, we hypothesized that the type of osteoclast precursor cells also affects osteoclastogenesis in the materials. In this study, we investigated the effects of defined micro/nanoscale patterns on osteoclastogenesis from bone marrow cells (BMCs). METHODS: Various cyclo-olefin polymer (COP) patterns were prepared using nanoimprinting. The effects of shape, size, and height of the patterns, and the wettability of the patterned surfaces on osteoclastogenesis from BMCs were evaluated in vitro. RESULTS: Osteoclast formation was promoted on pillars (diameter, 1 µm or 500 nm; height, 500 nm). Notably, osteoclastogenesis from BMCs was better promoted on hydrophobic pillars than on hydrophilic pillars. In contrast, decreased osteoclast formation was observed on the nanopillars (diameter, 100 nm; height, 200 nm). CONCLUSIONS: We demonstrated the promotion of osteoclast formation from BMCs on hydrophobic pillars with diameters of 1 µm and 500 nm. Some cellular behaviors in the patterns were dependent on the type of osteoclast precursor cells. The designed patterns are useful for designing the surface of dental implants or bone replacement materials with a controllable balance between osteoblast and osteoclast activities.

Biological modification of tooth surface by laser-based apatite coating techniques.

BACKGROUND: Development of new clinical regenerative procedures is needed for the reconstruction of the connective tissue attachment lost to periodontal disease. Apatite coating on the affected root surfaces could improve root surface biocompatibility and promote the reestablishment of connective tissue attachment. HIGHLIGHT: We developed two novel techniques that use laser light for coating the tooth surface with apatite. In the laser-assisted biomimetic (LAB) process, a tooth substrate was placed in a supersaturated calcium phosphate solution and irradiated for 30 min with low-energy pulsed laser light. Due to the laser-assisted pseudo-biomineralization, a submicron-thick apatite film was created on the laser-irradiated tooth surface. Furthermore, we created a fluoride-incorporated apatite film on the tooth surface using the LAB process and demonstrated its antibacterial activity against Streptococcus mutans. In the laser-induced forward transfer with optical stamp (LIFTOP) process, a thin apatite film loaded with the cell-adhesion protein, fibronectin, was prepared beforehand as a raw material on the optical stamp (carbon- and polydimethylsiloxane-coated support) by a conventional biomimetic process. After irradiation with a single laser pulse, the film (microchip) was transferred onto a tooth substrate via laser ablation of the carbon sacrificial layer. The LIFTOP process requires only a short processing time and has a minimal heat effect on the film; thus, the film exhibits cell adhesion activity even after the LIFTOP process. CONCLUSION: The LAB and LIFTOP processes have the potential as novel tools for tooth surface modification in the treatment of periodontal disease.

Ultrasonic irrigation of periodontal pocket with surface pre-reacted glass-ionomer (S-PRG) nanofiller dispersion improves periodontal parameters in beagle dogs.

OBJECTIVES: Surface pre-reacted glass-ionomer (S-PRG) nanofiller, an antibacterial ion-releasing bioactive glass, has been shown to adhere to tooth surfaces and reported to improve inflammatory parameters in experimental periodontitis. In this study, cementum substrate was irrigated ultrasonically with dispersion to examine in-vitro nanofiller adhesion and antibacterial activity. Moreover, periodontal pockets in a beagle dog were ultrasonically irrigated with dispersion to assess periodontal healing. METHODS: The morphology of human cementum irrigated with S-PRG nanofiller dispersion was examined by scanning electron microscopy and energy dispersive X-ray spectrometry. The antibacterial activity of the treated cementum was tested using Actinomyces naeslundii. In addition, experimentally formed periodontal pockets in beagle dog were ultrasonically irrigated with S-PRG nanofiller dispersion. Periodontal parameters (gingival index, bleeding on probing, probing pocket depth, and clinical attachment level) were measured from baseline (0 weeks) through 12 weeks. Moreover, the effects of irrigation with S-PRG nanofiller on changes in periodontal microflora and bone healing were analyzed. RESULTS: After ultrasonic irrigation, S-PRG nanofiller adhered to the cementum and exhibited antibacterial activity. The periodontal parameters were shown to improve following ultrasonic irrigation with S-PRG nanofiller dispersion. Analysis by next-generation sequencing revealed that the ratio of red-complex species decreased in the pockets irrigated with S-PRG nanofiller dispersion. In addition, the S-PRG nanofiller showed the potential to promote bone healing. CONCLUSIONS: Ultrasonic irrigation with S-PRG nanofiller dispersion using an ultrasonic scaler system permitted delivery of the S-PRG nanofiller to the root surface, providing improved parameters in experimental periodontitis and modifying the composition of subgingival periodontal microflora.

Periodontal tissue engineering using an apatite/collagen scaffold obtained by a plasma- and precursor-assisted biomimetic process.

BACKGROUND AND OBJECTIVES: In the treatment of severe periodontal destruction, there is a strong demand for advanced scaffolds that can regenerate periodontal tissues with adequate quality and quantity. Recently, we developed a plasma- and precursor-assisted biomimetic process by which a porous collagen scaffold (CS) could be coated with low-crystalline apatite. The apatite-coated collagen scaffold (Ap-CS) promotes cellular ingrowth within the scaffold compared to CS in rat subcutaneous tissue. In the present study, the osteogenic activity of Ap-CS was characterized by cell culture and rat skull augmentation tests. In addition, the periodontal tissue reconstruction with Ap-CS in a beagle dog was compared to that with CS. METHODS: The plasma- and precursor-assisted biomimetic process was applied to CS to obtain Ap-CS with a low-crystalline apatite coating. The effects of apatite coating on the scaffold characteristics (i.e., surface morphology, water absorption, Ca release, protein adsorption, and enzymatic degradation resistance) were assessed. Cyto-compatibility and the osteogenic properties of Ap-CS and CS were assessed in vitro using preosteoblastic MC3T3-E1 cells. In addition, we performed in vivo studies to evaluate bone augmentation and periodontal tissue reconstruction with Ap-CS and CS in a rat skull and canine furcation lesion, respectively. RESULTS: As previously reported, the plasma- and precursor-assisted biomimetic process generated a low-crystalline apatite layer with a nanoporous structure that uniformly covered the Ap-CS surface. Ap-CS showed significantly higher water absorption, Ca release, lysozyme adsorption, and collagenase resistance than CS. Cell culture experiments revealed that Ap-CS was superior to CS in promoting the osteoblastic differentiation of MC3T3-E1 cells while suppressing their proliferation. Additionally, Ap-CS significantly promoted (compared to CS) the augmentation of the rat skull bone and showed the potential to regenerate alveolar bone in a dog furcation defect. CONCLUSION: Ap-CS fabricated by the plasma- and precursor-assisted biomimetic process provided superior promotion of osteogenic differentiation and bone neoformation compared to CS.

Bone Tissue Regeneration by Collagen Scaffolds with Different Calcium Phosphate Coatings: Amorphous Calcium Phosphate and Low-Crystalline Apatite.

Surface-mineralized collagen sponges have attracted much attention as scaffolds for bone tissue engineering. Recently, we developed amorphous calcium phosphate (ACP) and low-crystalline apatite coating processes on collagen sponges. In the present study, we applied these coating processes to granular collagen sponges (referred to as Col) to compare the bone tissue regeneration capabilities of ACP-coated and apatite-coated Col (referred to as Col-ACP and Col-Ap, respectively) using a rat cranial bone defect model. According to micro-CT and histological analyses, Col-Ap enhanced bone tissue regeneration compared to Col, whereas Col-ACP did not. These results not only demonstrated the superior bone tissue regeneration capability of Col-Ap, but also indicated limitations of the in vitro simulated body fluid (SBF) test used in our previous study. Despite the apatite-forming ability of Col-ACP in SBF, it was ineffective in improving bone tissue regeneration in vivo, unlike Col-Ap, most likely due to the quick resorption of the ACP coating in the defect site. The present results clarified the importance of the coating stability in vivo and revealed that the low-crystalline apatite coating was more beneficial than the ACP coating in the fabrication of surface-mineralized collagen sponges for use as bone tissue engineering scaffolds.

Antibacterial and Antibiofilm Photodynamic Activities of Lysozyme-Au Nanoclusters/Rose Bengal Conjugates.

Antibacterial photodynamic therapy (aPDT) utilizes reactive oxygen species such as singlet oxygen (1O2) and free radicals via photosensitizers, which are light and light-sensitive agents, to reduce bacterial infections. It has been utilized as a treatment for dental diseases in place of antibiotic therapies. However, aPDT does not always cause the desired therapeutic effect due to the instability of organic photosensitizers and the formation of bacterial biofilms. To promote the antibacterial and antibiofilm effects of aPDT, we have proposed a lysozyme (Lys)-gold nanoclusters (Au NCs)/rose bengal (Lys-Au NCs/RB) conjugate as a novel photosensitizer. This conjugate was found to effectively impede the growth of both gram-positive and gram-negative bacteria when exposed to white light-emitting diode (LED) irradiation. The photoexcited Lys-Au NCs/RB showed significantly higher antibacterial activity than photoexcited Lys-Au NCs or RB alone. The synergistic effect is a result of the combination of Lys (an antibacterial protein) and enhanced 1O2 generation related to resonance energy transfer (RET) in the Au NCs/RB conjugate. Photoexcited Lys-Au NCs/RB increased the effects of aPDT in a dose- and time-dependent manner. Furthermore, the photoexcited Lys-Au NCs/RB successfully decreased Streptococcus mutans biofilm formation. However, in contrast, it did not have a negative effect on the proliferation, adhesion, or spread of mammalian cells, indicating low cytotoxicity. Lys-Au NCs/RB is a novel photosensitizer with low cytotoxicity that is capable of bacterial inactivation and the suppression of biofilm formation, and could help to improve dental treatments in the future.

Antibacterial coating of tooth surface with ion-releasing pre-reacted glass-ionomer (S-PRG) nanofillers.

OBJECTIVES: Surface pre-reacted glass-ionomer (S-PRG) fillers release antibacterial borate and fluoride ions. We fabricated nanoscale S-PRG fillers (S-PRG nanofillers) for antibacterial coating of tooth surfaces and assessed the antibacterial effects of this coating in vitro. In addition, we creating a canine model of periodontitis to evaluate the effectiveness of S-PRG nanofiller application on tooth roots and improvement of periodontal parameters. METHODS: Human dentin blocks were coated with S-PRG nanofiller (average particle size: 0.48 µm) and then characterized by scanning electron microscopy (SEM), energy dispersive X-ray spectrometer (EDX), and ion-releasing test. Antibacterial effects of dentin blocks coated with S-PRG nanofiller were examined using bacterial strains, Streptococcus mutans and Actinomyces naeslundii. Next, we created an experimental model of periodontitis in furcation of premolars of beagle dogs. Then, S-PRG nanofiller coating was applied onto exposed tooth root surfaces. Periodontal parameters, gingival index (GI), bleeding on probing (BOP), probing pocket depth (PPD), and clinical attachment level (CAL), were measured from baseline until 4 weeks. In addition, bone healing was radiographically and histologically examined. RESULTS: SEM and EDX revealed that S-PRG nanofillers uniformly covered the dentin surface after coating. Dentin blocks coated with S-PRG nanofiller showed ion-releasing property, bacterial growth inhibition, and sterilization effects. In the experimental periodontitis model, S-PRG nanofiller coating significantly reduced clinical inflammatory parameters, such as GI (P < 0.01) and BOP (P < 0.05), compared to uncoated samples. In addition, PPD and CAL significantly decreased by S-PRG nanofiller coating (2 weeks: P < 0.05; 3 and 4 weeks: P < 0.01), suggesting the improvement of periodontitis. Micro-CT and histology revealed that bone healing of furcation defects was enhanced by S-PRG nanofiller coating. CONCLUSION: S-PRG nanofiller coating provides antibacterial effects to tooth surfaces and improves clinical parameters of periodontitis.

Evaluation of antibacterial and cytocompatible properties of multiple-ion releasing zinc-fluoride glass nanoparticles.

Zinc-fluoride glass nanoparticles (Zinc-F) release several ions, such as fluoride, zinc and calcium ions, through acid-base reactions. The aim of this study was to evaluate the antibacterial and cytotoxic properties of Zinc-F. Antibacterial tests showed that a Zinc-F eluting solution significantly reduced the turbidity and colony-forming units of Streptococcus mutans and Actinomyces naeslundii, compared to that of calcium-fluoroaluminosilicate glass nanoparticles without zinc ions. In live/dead staining, Zinc-F eluate significantly decreased green-stained bacterial cells, indicating live cells, compared with the control (no application). Human dentin coated with Zinc-F showed suppressed S. mutans and A. naeslundii biofilm formation. Additionally, Zinc-F eluate showed low cytotoxic effects in osteoblastic and fibroblastic cells. Therefore, our findings suggested that Zinc-F exhibits antibacterial and biocompatible properties through multiple-ion release.

Antibacterial tooth surface created by laser-assisted pseudo-biomineralization in a supersaturated solution.

A technique for implementing biocompatible and antibacterial functions to a targeted region on tooth surfaces has potential in dental treatments. We have recently demonstrated pseudo-biomineralization, i.e., the growth of an apatite layer on a human dentin substrate by a laser-assisted biomimetic (LAB) process, based on pulsed laser irradiation in a supersaturated CaP solution. In this study, pseudo-biomineralization was induced in the presence of fluoride ions using the LAB process in order to fabricate an antibacterial fluoride-incorporated apatite (FAp) layer on the dentin surface. After processing for 30 min, a micron-thick FAp layer was formed heterogeneously at the laser-irradiated solid-liquid interface via pseudo-biomineralization. A time-course study revealed that the LAB process first eliminated the pre-existing organic layer, while allowing fluoride incorporation into the dentin surface within 1 min. Within 5 min, FAp nanocrystals precipitated on the dentin surface. Within 30 min, these nanocrystals acquired a pillar-like structure that was weakly oriented in the direction normal to the substrate surface to form a dense micron-thick layer. This layer was integrated seamlessly with the underlying dentin without any apparent gaps. The FAp layer exhibited antibacterial activity against a major oral bacterium, Streptococcus mutans. The proposed LAB process is expected to be a useful new tool for tooth surface functionalization via facile and area-specific pseudo-biomineralization.

In situ precipitation of amorphous calcium phosphate nanoparticles within 3D porous collagen sponges for bone tissue engineering.

Amorphous calcium phosphate (ACP) plays an important role in biomineralization within the three-dimensional (3D) collagen network in human hard tissues, and exhibits osteoconductivity. Porous collagen sponges coated with ACP nanoparticles could be considered as potential scaffolds for use in bone tissue engineering. In this study, such composite materials were fabricated via homogeneous ACP precipitation using a supersaturated calcium phosphate (CaP) solution. Homogeneous ACP precipitation was induced in situ within the sponges by a temperature-controlled coating process composed of two steps. In the first step, the CaP solution was cooled to 4 °C to suppress precipitation until the solution penetrated fully into the sponge's internal pores. In the second step, the CaP solution was warmed up to 25 °C with continuous shaking to induce ACP precipitation within the sponges. The resulting sponges were therefore coated with ACP nanoparticles on their inner and outer surfaces. A simulated body fluid (SBF) test indicated osteoconductivity of the collagen sponges coated with ACP nanoparticles. Further, ACP-coated collagen sponges immobilizing basic fibroblast growth factor (bFGF) were fabricated using the CaP solution supplemented with bFGF. The fabricated sponges allowed the sustained release of bFGF in a culture medium and enhanced proliferation of osteoblastic MC3T3-E1 cells. Such ACP-coated collagen sponges have the potential to be used as scaffolds in bone tissue engineering if pursued for further in vitro and in vivo studies.

Technique for simple apatite coating on a dental resin composite with light-curing through a micro-rough apatite layer.

Tooth root surfaces restored with dental resin composites exhibit inferior biocompatibility. The objective of this study was to develop a simple technique for coating apatite onto a resin composite to improve its surface biocompatibility. First, we fabricated a polymer film coated with a micro-rough apatite layer and pressed it (coating-side down) onto a viscous resin composite precursor. As a result of light-induced curing of the precursor through the overlaid film, the micro-rough apatite layer was integrated with the resin composite and, thus, transferred from the polymer film surface to the cured resin composite surface as a result of the difference in interfacial adhesion strength. The transferred apatite layer attached directly to the cured resin composite without any gaps at the microscopic level. The adhesion between the apatite layer and the cured resin composite was so strong that the layer was not peeled off even by a tape-detaching test. The flexural strength of the resulting apatite-coated resin composite was comparable to that of the clinically used resin composite while satisfying the ISO requirement for dental polymer-based restorative materials. Furthermore, the apatite-coated resin composite showed better cell compatibility than the uncoated resin composite. The present apatite coating technique is well suited for dental treatment because the coating is applied during a conventional light curing procedure through simple utilization of the apatite-coated polymer film in place of an uncoated film. The proposed technique represents a practical evolution in dental treatment using light-curing resin composites, although further in vitro and in vivo studies are needed.

Bone forming ability of recombinant human collagen peptide granules applied with ß-tricalcium phosphate fine particles.

Recombinant human collagen peptide, developed based on human collagen type I, contains an arginyl-glycyl-aspartic acid (RGD)-rich motif to enhance cell behavior and is anticipated as a xeno-free polymer material for use in tissue engineering. We fabricated granules containing recombinant human collagen peptide (RCP) applied with beta-tricalcium phosphate fine particles (RCP/ß-TCP) as bone filling scaffold material and assessed the bone forming ability of RCP/ß-TCP. Recombinant peptide was thermal crosslinked and freeze-dried to prepare RCP. An aqueous dispersion of ß-TCP fine particles was added to RCP to obtain RCP/ß-TCP. Subsequently, RCP/ß-TCP were characterized using scanning electron microscopy (SEM), energy dispersive X-ray spectrometry (EDX), and cell culture assessments. Furthermore, RCP/ß-TCP were implanted into rat cranial bone defects for radiographic and histological evaluations. In SEM and EDX analyses of RCP/ß-TCP, ß-TCP particles dose-dependently covered the surface of RCP. Cell culture tests showed that RCP/ß-TCP remarkably promoted proliferation and mRNA expression of various genes, such as integrin ß1 and osteogenic markers, of osteoblastic MC3T3-E1 cells. Histomorphometric assessment at 4 weeks showed that RCP/ß-TCP significantly promoted new skull bone formation compared to RCP (p < 0.05) and control (no application) (p < 0.01). Accordingly, these findings suggest RCP/ß-TCP possess bone forming capability and would be beneficial for bone tissue engineering therapy.