Inhibition of tooth demineralization caused by Streptococcus mutans biofilm via antimicrobial treatment using hydrogen peroxide photolysis.

OBJECTIVES: An antimicrobial technique utilizing hydroxyl radicals generated by the photolysis of 3% H2O2 has been developed recently. The present study aimed to evaluate the effect of H2O2 photolysis treatment on tooth demineralization caused by Streptococcus mutans biofilm. MATERIALS AND METHODS: To induce tooth demineralization, S. mutans biofilm was allowed to form on the maxillary first molars collected from Wistar rats via 24-h culturing. The samples were immersed in 3% H2O2 and irradiated with 365-nm LED (H2O2 photolysis treatment). Viable bacterial counts in the biofilm were evaluated immediately after treatment and after an additional 30-h culturing by colony counting. The acidogenicity of the biofilm, re-established 30 h after treatment, was assessed by measuring the pH. The effect of H2O2 photolysis treatment on tooth demineralization was assessed by measuring the depth of the radiolucent layer in micro-CT images. RESULTS: H2O2 photolysis significantly reduced viable bacterial counts in the biofilm to 3.7 log colony forming units (CFU)/sample, while the untreated group had 7.9 log CFU/sample. The pH of the biofilm re-established after treatment (6.6) was higher than that of the untreated group (5.3). In line with the pH measurement, the treatment group had a significantly lower depth of radiolucent layer in dentin than the untreated group. CONCLUSIONS: H2O2 photolysis treatment was effective not only in killing the biofilm-forming S. mutans but also in lowering the acidogenicity of the biofilm. Thus, this technique could inhibit tooth demineralization. CLINICAL RELEVANCE: H2O2 photolysis can be applicable as a new dental caries treatment.

Introduction of integrated dental training jaw models and rubric criteria.

OBJECTIVE: The objective of the present study was to evaluate the effectiveness of introducing integrated jaw models, rubric criteria and homework tasks to a total clinical simulation training course to improve the clinical competence of preclinical dental students. METHODS: A total simulation training course, which involved six clinical dentistry departments, was held for 110 preclinical students in 2014 and 2015. We prepared integrated jaw models having several morbidities along with corresponding medical information and homework tasks. The students formulated diagnoses and devised treatment plans before performing dental treatment on the mannequin under the direction of instructors from the respective clinical departments. Their performance was assessed by both students and instructors using the rubric criteria. RESULTS: Based on quantitative evaluations, the introduction of integrated jaw models appeared to improve the students' ability to formulate diagnoses and devise dental treatment plans and to understand the respective clinical dentistry disciplines. The rubric criteria provided immediate feedback for the students. Based on a comparison of rubric scores, students tended to significantly underestimate their own performance compared with instructors. Moreover, the introduction of homework tasks improved student seriousness. CONCLUSION: Introducing integrated jaw models, rubric criteria and homework tasks to a total simulation training course may be a good approach for improving student performance in terms of dental diagnoses and treatment.

Gene transfection of human mesenchymal stem cells with a nano-hydroxyapatite-collagen scaffold containing DNA-functionalized calcium phosphate nanoparticles.

This study aimed to fabricate a growth factor-releasing biodegradable scaffold for tissue regeneration. We prepared multishell calcium phosphate (CaP) nanoparticles functionalized with DNA, polyethyleneimine (PEI), protamine and octa-arginine (R8) and compared their respective transfection activity and cell viability measures using human mesenchymal stem cells. DNA-protamine complexes improved the transfection efficiency of CaP nanoparticles with the exception of those functionalized with R8. These complexes also greatly reduced the cytotoxicity of PEI. In addition, we also fabricated DNA-protamine-functionalized CaP nanoparticle-loaded nano-hydroxyapatite-collagen scaffolds and investigated their gene transfection efficiencies. These experiments showed that the scaffolds were associated with moderate hMSC cell viability and were capable of releasing the BMP-2 protein into hMSCs following gene transfection. In particular, the scaffold loaded with protamine-containing CaP nanoparticles showed the highest cell viability and transfection efficiency in hMSCs; thus, it might be suitable to serve as an efficient growth factor-releasing scaffold.

Cutting efficiency of diamond burs operated with electric high-speed dental handpiece on zirconia.

Zirconia-based dental restorations are becoming used more commonly. However, limited attention has been given to the difficulties experienced, concerning cutting, in removing the restorations when needed. The aim of the present study was to compare the cutting efficiency of diamond burs, operated using an electric high-speed dental handpiece, on zirconia (Zir) with those on lithium disilicate glass-ceramic (LD) and leucite glass-ceramic (L). In addition, evaluation of the cutting efficiency of diamond burs on Zir of different thicknesses was performed. Specimens of Zir were prepared with thicknesses of 0.5, 1.0, 2.0, and 4.0 mm, and specimens of LD and L were prepared with a thickness of 1.0 mm. Cutting tests were performed using diamond burs with super coarse (SC) and coarse (C) grains. The handpiece was operated at 150,000 rpm with a cutting force of 0.9 N. The results demonstrated that cutting of Zir took about 1.5- and 7-fold longer than cutting of LD and L, respectively. The SC grains showed significantly higher cutting efficiency on Zir than the C grains. However, when the thickness of Zir increased, the cutting depth was significantly decreased. As it is suggested that cutting of zirconia is time consuming, this should be taken into consideration in advance when working with zirconia restorations.

Cementocyte-derived extracellular vesicles regulate osteoclastogenesis and osteoblastogenesis.

Background/purpose: Cementum shares many properties with bone; however, in contrast to bone, it is not innervated or vascularized and has a limited capacity for remodeling. Osteocytes located in the lacunae-canalicular system of bone tissue play a central role in bone remodeling by communicating with osteoblasts and osteoclasts. Although cementocytes are present in cellular cementum and are morphologically similar to osteocytes, it remains unclear whether they are involved in the dynamic functional regulation of metabolism in cementum. The present study focused on the extracellular vesicles (EVs) secreted by cementocytes and examined their effects on osteoclasts and osteoblasts. Materials and methods: EVs were extracted from the mouse cementocyte cell line, IDG-CM6. The effects of EVs on recombinant RANKL-induced osteoclastogenesis and recombinant Bone morphogenetic protein (BMP)-2-mediated osteoblastogenesis were investigated using the mouse osteoclast progenitor cell line, RAW264.7 and mouse pre-osteoblast cell line, MC3T3-E1, respectively. Results: EVs enhanced the formation of tartrate-resistant acid phosphatase activity-positive cells. Real-time PCR revealed that EVs up-regulated the expression of osteoclast-related genes. On the other hand, the cell culture supernatant of cementocytes significantly inhibited the differentiation of osteoclasts. Regarding osteoblastogenesis, EVs suppressed the expression of alkaline phosphatase, bone sialoprotein, and osteocalcin induced by recombinant BMP-2 at the gene and protein levels. Conclusion: A network of cementocytes, osteoblasts, and osteoclasts may exist in cellular cementum, which suggests the involvement of cementocytes in dynamic metabolism of cementum through EVs.

A triple growth factor strategy for optimizing bone augmentation in mice.

With dental implant treatment becoming the gold standard, the need for effective bone augmentation prior to implantation has grown. This study aims to evaluate a bone augmentation strategy integrating three key growth factors: bone morphogenetic protein-2 (BMP-2), insulin-like growth factor 1 (IGF-1), and vascular endothelial growth factor (VEGF). Collagen scaffolds incorporating BMP-2, IGF-1, or VEGF were fabricated and categorized into five groups based on their content: scaffold alone; BMP-2 alone (BMP-2); BMP-2 and IGF-1 (BI); BMP-2, IGF-1, and VEGF (BIV); and BMP-2 and IGF-1 with an earlier release of VEGF (BI + V). The prepared scaffolds were surgically implanted into the calvarias of C57BL/6JJcl mice, and hard tissue formation was assessed after 10 and 28 days through histological, tomographic, and biochemical analyses. The combination of BMP-2 and IGF-1 induced a greater volume of hard tissue augmentation compared with that of BMP-2 alone, regardless of VEGF supplementation, and these groups had increased levels of cartilage compared with others. The volume of hard tissue formation was greatest in the BIV group. In contrast, the BI + V group exhibited a hard tissue volume similar to that of the BI group. While VEGF and CD31 levels were highest in the BIV group at 10 days, there was no correlation at the same time point between hard tissue formation and the quantity of M2 macrophages. In conclusion, the simultaneous release of BMP-2, IGF-1, and VEGF proved to be effective in promoting bone augmentation.

Development of triple-functionalized calcium phosphate nanoparticles as an advanced drug delivery system for bone tissue repair.

Introduction: During tissue repair or regeneration, several bioactive molecules are released and interact with each other and act as complex additives or inhibitors for tissue reconstruction. In this study, the bone-healing effects of the combination treatment with tumor necrosis factor-α (TNF-α) inhibition, vascular endothelial growth factor A (VEGF-A) and bone morphogenetic protein-7 (BMP-7) release by gene silencing, and gene transfection with calcium phosphate nanoparticles (CaP) in the rat femoral head was histologically, morphologically, and biochemically evaluated. Methods: A triple-functionalized paste of CaP carrying plasmid DNA encoding for BMP-7 and for VEGF), and siRNA against TNF-α was developed and denoted as CaP3mix. To compare the effects of 3mixCaP, CaP with plasmid DNA encoding BMP-7, VEGF, or siRNA encoding TNF-α was prepared and denoted as CaP/PEI/pBMP-7/SiO2, CaP/PEI/pVEGF/SiO2, or CaP/PEI/siRNA-TNF-α/SiO2, respectively. The bone healing in bone defects in the rat femoral head was investigated after 10 and 21 days of implantation. Results: The levels of bone formation-related markers OCN, Runx2, and SP7 increased at the protein and gene levels in 3mixCaP after 10 days, and 3mixCaP significantly accelerated bone healing compared with the other treatments after 21 days of implantation. Conclusion: The triple-functionalized CaP paste loading plasmid DNA encoding BMP-7 and VEGF and siRNA encoding TNF-α is a promising bioactive material for bone tissue repair.

Introduction of tenomodulin by gene transfection vectors for rat bone tissue regeneration.

Introduction: Periodontal ligament is regenerated in association with hard tissue regeneration. Tenomodulin (Tnmd) expression has been confirmed in periodontal ligament and it reportedly inhibits angiogenesis or is involved in collagen fibril maturation. The introduction of Tnmd by gene transfection in bone tissue regeneration therapy might inhibit topical hard tissue formation and induce the formation of dense fibrous tissue. Therefore, the effect of Tnmd introduction by gene transfection technique in vitro and in vivo was investigated in this study. Methods: Osteogenesis- and chondrogenesis-related gene expression levels in osteoblastic cells (MC3T3E1) and rat bone marrow derived cells were detected using qPCR three days after gene transfection with plasmid DNA (Tnmd) using non-viral gene transfection vectors: a calcium phosphate-based gene transfection vector (CaP(Tnmd)) or a cationic polymer-based reagent (JetPEI (Tnmd)). Next, an atelocollagen scaffold with or without CaP (Tnmd) or JetPEI (Tnmd) was implanted into a rat calvaria bone defect, and the remaining bone defect volume and the tissue reaction at 28 days after surgery were evaluated. Results: Runx 2 and SP7 mRNA was reduced by JetPEI (Tnmd) in both cells, but not in CaP(Tnmd). The volume of expressed Tnmd was at 9 ng/mL in both gene transfection vector. The remaining bone defect volume of JetPEI (Tnmd) was significantly bigger than that of the other groups and CaP (EGFP), and that of CaP (Tnmd) was significantly bigger than that of CaP (EGFP). Conclusions: Tnmd introduction treatment inhibits bone formation in artificial bone defect, however, the effect of that was dependent on non-viral gene transfection vector.

Gene transfection achieved by utilizing antibacterial calcium phosphate nanoparticles for enhanced regenerative therapy.

Protamine-coated multi-shell calcium phosphate (CaP) was developed as a non-viral vector for tissue regeneration therapy. CaP nanoparticles loaded with different amounts of plasmid DNA encoding bone morphogenetic protein 2 (BMP-2) and insulin-like growth factor 1 (IGF-1) were used to treat MC3T3E1 cells, and the yield of the released BMP-2 or IGF-1 was measured using ELISA 3 days later. Collagen scaffolds containing CaP nanoparticles were implanted into rat cranial bone defects, and BMP-2 and IGF-1 yields, bone formation, and bone mineral density enhancement were evaluated 28 days after gene transfer. The antibacterial effects of CaP nanoparticles against Streptococcus mutans and Aggregatibacter actinomycetemcomitans increased with an increase in the protamine dose, while they were lower for Staphylococcus aureus and Porphyromonas gingivalis. In the combination treatment with BMP-2 and IGF-1, the concentration ratio of BMP-2 and IGF-1 is an important factor affecting bone formation activity. The calcification activity and OCN mRNA of MC3T3E1 cells subjected to a BMP-2:IGF-1 concentration ratio of 1:4 was higher at 14 days. During gene transfection treatment, BMP-2 and IGF-1 were released simultaneously after gene transfer; the loaded dose of the plasmid DNA encoding IGF-1 did not impact the BMP-2 or IGF-1 yield or new bone formation ratio in vitro and in vivo. In conclusion, two growth factor-releasing systems were developed using an antibacterial gene transfer vector, and the relationship between the loaded plasmid DNA dose and resultant growth factor yield was determined in vitro and in vivo.

Histomorphometric assessments of peri-implant bone around Ti-Nb-Sn alloy implants with low Young's modulus.

Many ß-Ti alloys have been developed for, and used in, medical devices because of the corrosion resistance, biocompatibility, and exceptionally low Young's modulus. The aim of the present study was to investigate the histomorphometric aspects of peri-implant bone around Ti-Nb-Sn alloy implants and compare them with those in the case of commercially pure Ti (Ti). Fluorescent morphological observations of ST-2 cells on the substrate were performed and bone morphogenesis around implants in rat femur was evaluated. There was no difference between the cell morphology on Ti and those on the Ti-Nb-Sn alloy during observation for 24 h. A comparison of the Ti-Nb-Sn alloy implant and the Ti implant showed no significant differences between the bone-to-implant contact ratios or the bone fractions. These results suggest that the biological adaptations with Ti-Nb-Sn implants during a healing period are similar to those with Ti. Ti-Nb-Sn is therefore suitable for use in dental implants.

Proanthocyanidin-rich grape seed extract improves bone loss, bone healing, and implant osseointegration in ovariectomized animals.

The purpose of the present study was to confirm if proanthocyanidin-rich grape seed extract (GSE) had the ability to improve bone health such as bone loss, bone healing, and implant osseointegration (defined as the direct connection between bone tissue and an implant) in ovariectomized (OVX) animals. We demonstrated that daily oral administration of GSE prevented bone loss in the lumbar vertebrae and femur in OVX mice. In addition, osteoclastogenesis in the lumbar spine bone of OVX mice, as assessed by histological and histomorphometric analyses, was accelerated but GSE prevented this dynamization, suggesting that GSE could counteract OVX-induced accelerated osteoclastogenic activity. In rats, OVX clearly impaired the healing of defects created on the calvaria, and GSE overcame this OVX-impaired healing. In the same way, osseointegration of a tibial implant in rats was retarded by OVX, and GSE counteracted the OVX-induced poor osseointegration, likely promoting bone healing by preventing imbalanced bone turnover. These results suggest that orally administered GSE improved implant osseointegration by mitigating the impaired bone health induced by OVX as a model of estrogen deficiency.

Bactericidal activity and recovery effect of hydroxyl radicals generated by ultraviolet irradiation and silver ion application on an infected titanium surface.

This study investigated the bactericidal effect, the underlying mechanisms of treatment, and recovery of biocompatibility of the infected titanium surface using a combination treatment of silver ion application and ultraviolet-A (UV-A) light irradiation. Streptococcus mutans and Aggregatibacter actinomycetemcomitans were used in suspension and as a biofilm on a titanium surface to test for the bactericidal effect. The bactericidal effect of the combination treatment was significantly higher than that of silver ion application or UV-A light irradiation alone. The bactericidal effect of the combination treatment was attributable to hydroxyl radicals, which generated from the bacterial cell wall and whose yield increased with the silver concentration. To assess the biocompatibility, proliferation and calcification of MC3T3E1 cells were evaluated on the treated titanium surface. The treated titanium screws were implanted into rat tibias and the removal torques were measured 28 days post-surgery. The titanium surface that underwent the combination treatment exhibited recovery of biocompatibility by allowing cellular proliferation or calcification at levels observed in the non-infected titanium surfaces. The removal torque 28 days after surgery was also comparable to the control values. This approach is a novel treatment option for peri-implantitis.

Reduction of inflammation in a chronic periodontitis model in rats by TNF-α gene silencing with a topically applied siRNA-loaded calcium phosphate paste.

We developed a calcium phosphate-based paste containing siRNA against TNF-α and investigated its anti-inflammatory and bone-healing effects in vitro and in vivo in a rat periodontitis model. The bioactive spherical CaP/PEI/siRNA/SiO2 nanoparticles had a core diameter of 40-90 nm and a positive charge (+23 mV) that facilitated cellular uptake. The TNF- α gene silencing efficiency of the nanoparticles in J774.2 monocytes, gingival-derived cells, and bone marrow-derived cells was 12 ± 2%, 36 ± 8%, and 35 ± 22%, respectively. CaP/PEI/siRNA/SiO2 nanoparticles cancelled the suppression of alkaline phosphatase (ALP) activity in LPS-stimulated bone marrow-derived cells. In vivo, ALP mRNA was up-regulated, TNF-α mRNA was down-regulated, and the amount of released TNF-α was significantly reduced after topical application of the calcium phosphate-based paste containing siRNA-loaded nanoparticles. The number of TNF-α-positive cells in response to CaP/PEI/siRNA/SiO2 nanoparticle application was lower than that observed in the absence of siRNA. Elevated ALP activity and numerous TRAP-positive cells (osteoclasts) were observed in response to the application of all calcium phosphate pastes. These results demonstrate that local application of a paste consisting of siRNA-loaded calcium phosphate nanoparticles successfully induces TNF-α silencing in vitro and in vivo and removes the suppression of ALP activity stimulated by inflammation. STATEMENT OF SIGNIFICANCE: We developed a calcium phosphate-based paste containing nanoparticles loaded with siRNA against TNF-α. The nanoparticles had a core diameter of 40-90 nm and positive charge (+23 mV). The anti-inflammatory and osteoinductive effects of the paste were investigated in vitro and in vivo in a rat periodontitis model. In vitro, the TNF-α gene silencing efficiency of the nanoparticles in J774.2 monocytes, gingival-derived cells, and bone marrow-derived cells was 12 ± 2%, 36 ± 8%, and 35 ± 22%, respectively. The ALP activity of bone marrow-derived cells was recovered. In vivo, TNF-α mRNA was down-regulated and the amount of released TNF-α was significantly reduced, whereas the ALP mRNA was up-regulated. Elevated ALP activity and TRAP-positive cells were observed by immunohistochemistry.

Hydroxyl radicals generated by hydrogen peroxide photolysis recondition biofilm-contaminated titanium surfaces for subsequent osteoblastic cell proliferation.

Titanium dental implants have been successfully used for decades; however, some implants are affected by peri-implantitis due to bacterial infection, resulting in loss of supporting bone. This study aimed to evaluate the effect of an antimicrobial chemotherapy employing H2O2 photolysis-developed to treat peri-implantitis-on biofilm-contaminated titanium surfaces in association with osteoblastic cell proliferation on the treated surface. Titanium discs were sandblasted and acid-etched, followed by contamination with a three-species biofilm composed of Porphyromonas gingivalis, Fusobacterium nucleatum, and Streptococcus mitis. This biofilm model was used as a simplified model of clinical peri-implantitis biofilm. The discs were subjected to ultrasound scaling, followed by H2O2 photolysis, wherein 365-nm LED irradiation of the disc immersed in 3% H2O2 was performed for 5 min. We analysed proliferation of mouse osteoblastic cells (MC3T3-E1) cultured on the treated discs. Compared with intact discs, biofilm contamination lowered cell proliferation on the specimen surface, whereas H2O2 photolysis recovered cell proliferation. Thus, H2O2 photolysis can recover the degraded biocompatibility of biofilm-contaminated titanium surfaces and can potentially be utilised for peri-implantitis treatment. However, to verify the findings of this study in relation to clinical settings, assessment using a more clinically relevant multi-species biofilm model is necessary.

Oral mucosal irritation potential of antimicrobial chemotherapy involving hydrogen peroxide photolysis with high-power laser irradiation for the treatment of periodontitis.

In the present study, we assessed the oral mucosal irritation potential of antimicrobial chemotherapy involving hydrogen peroxide (H2O2) photolysis with a 405-nm laser device at an output power of ≥100 mW in hamsters. Twenty-four cheek pouches from 12 male Syrian hamsters received 7-min treatment with pure water (PW), 3% H2O2, laser irradiation of PW at 100 mW, laser irradiation of 3% H2O2 at 100 mW, laser irradiation of PW at 200 mW, or laser irradiation of 3% H2O2 at 200 mW (n = 4 each). The diameter of the irradiation area was set at 3 mm; accordingly, the calculated irradiances (optical power densities) of the 100- and 200-mW laser lights were approximately 1400 and 2800 mW/cm2, respectively. In addition, 12 cheek pouches from six animals received laser irradiation of 3% H2O2 at 100 mW for 1, 3, or 5 min (n = 4 each). Each treatment was repeated three times at 1-h intervals. Macroscopic and histological changes were evaluated 24 h after the last treatment. In addition, in vitro bactericidal activity of the treatment against periodontal pathogens was evaluated. We found that 405-nm laser irradiation of 3% H2O2 caused moderate to severe oral mucosal irritation when performed at powers of 100 and 200 mW for ≥3 min, while the same treatment performed at 100 mW for 1 min resulted in mild irritation. Moreover, 1-min H2O2 photolysis at 100 mW caused a >4-log decrease in viable bacterial counts. These findings suggest that 1-min H2O2 photolysis, which can effectively kill periodontal pathogens, may be acceptable when a 405-nm laser device is used at 100 mW. However, use of the laser at a lower power would be preferable for the prevention of unnecessary oral mucosal irritation.

Potential adverse effects of antimicrobial chemotherapy based on ultraviolet-A irradiation of polyphenols against the oral mucosa in hamsters and wounded skin in rats.

Antimicrobial chemotherapy based on ultraviolet-A (UVA) irradiation of polyphenol solution has been proposed as an adjunctive treatment for dental caries. However, the safety of this treatment has not been thoroughly evaluated. Therefore, the aim of the present study was to assess the influence of this treatment on the oral mucosa in hamsters and wounded skin in rats. An oral mucosal irritation test was performed in hamsters. The cheek pouch was everted and treated with UVA irradiation (wavelength: 365 nm) of pure water, 1 mg/mL of caffeic acid, or 1 mg/mL of grape seed extract using a light-emitting diode at an irradiance of 275 mW/cm2. Each treatment was performed for 2 or 5 min and repeated three times. Macroscopic and histological evaluations were performed 24 h after the last treatment. We also examined the effects of the treatment on the healing process of skin wounds in rats. Full-thickness skin wounds created on the back of the rats were treated for 2 min as described above, but only once. The wound area was then assessed daily for 9 days. The results demonstrated that the treatment induced oral mucosal irritation depending on the irradiation time; however, it did not influence the wound healing process. The oral mucosal irritation potential of three treatment sessions performed for 2 and 5 min was minimal and mild to moderate, respectively, according to histological analysis. These findings suggest that the duration of treatment based on UVA irradiation of polyphenols in the oral cavity should be as short as possible, considering the clinical efficacy of the antimicrobial effects and the irritation potential.

The effect of low-magnitude high-frequency loading on peri-implant bone healing and implant osseointegration in Beagle dogs.

PURPOSE: Low-magnitude, high-frequency (LMHF) loading plays an important role in bone healing. The present study aimed to evaluate the effect of LMHF loading applied directly to titanium dental implants on peri-implant bone healing and implant osseointegration. METHODS: The mandibular premolars and molars were extracted from six male Beagle dogs. Three months post-extraction, each of the six dogs had three titanium implants (Aadva Standard Implant Narrow, Φ3.3×8mm) inserted into the mandibular premolar and molar area (three implants per side). In each animal, one side was randomly selected to undergo daily LMHF loading (treatment group), while the other side had no further intervention (control). The loading was applied directly to the implant abutment using an individual jig and a custom-made loading device (8µm, 100Hz). The implant stability quotient (ISQ) was tested every week. Three dogs were euthanized after 2 weeks, and three were euthanized after 8 weeks. Tissue samples were fixed and stained for micro-computed tomography (micro-CT) and histomorphometric analyses. Data were analyzed statistically, with significance set at p<0.05. RESULTS: The treatment group had significantly increased peri-implant bone volume relative to tissue volume in region of interest 2 (100-500µm) compared with the control group after 2 weeks of loading (p<0.05); however, there was no significant difference between groups after 8 weeks. The ISQ value and the micro-CT results did not differ between groups during the study period. CONCLUSIONS: LMHF loading positively influenced peri-implant bone healing in the early healing period.

Prolonged release of bone morphogenetic protein-2 in vivo by gene transfection with DNA-functionalized calcium phosphate nanoparticle-loaded collagen scaffolds.

In the combination of scaffolds immersed in growth factor solutions, the release of growth factors mainly depends on scaffold degradation. However, the release of bone morphogenetic protein (BMP)-2 at an appropriate concentration during the stage of tissue regeneration would enhance bone regeneration. To achieve this condition, the present study was performed to investigate the effects of scaffolds combined with gene transfection using non-viral vectors. Nanohydroxyapatite-collagen (nHAC) scaffolds cross-linked with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) or ascorbic acid/copper chloride, and a collagen scaffold (Terdermis®) were prepared, loaded with BMP-2-encoding plasmid DNA-functionalized calcium phosphate nanoparticles (CaP), naked plasmid DNA, or BMP-2 solution, and implanted in rats. The yield of released BMP-2 and its releasing period, respectively, were larger and longer from the scaffolds loaded with CaP than from those incubated with BMP-2 solution. In addition, the alkaline phosphatase activity induced by the CaP-loaded scaffolds was higher. Histological analysis showed that released BMP-2 could be observed on the macrophages or multinuclear giant cells surrounding the nHAC fragments or collagen fibres. TRAP-positive or OCN-positive sites were observed in all groups and a mineralization area was observed in the Terdermis®/CaP sample. The present study demonstrates that gene transfection by scaffold loaded with CaP gene transfer vectors induces a larger yield of BMP-2 for a longer period than by scaffolds loaded with BMP-2 solution or naked plasmid.

Amiloride-enhanced gene transfection of octa-arginine functionalized calcium phosphate nanoparticles.

Nanoparticles represent promising gene delivery systems in biomedicine to facilitate prolonged gene expression with low toxicity compared to viral vectors. Specifically, nanoparticles of calcium phosphate (nCaP), the main inorganic component of human bone, exhibit high biocompatibility and good biodegradability and have been reported to have high affinity for protein or DNA, having thus been used as gene transfer vectors. On the other hand, Octa-arginine (R8), which has a high permeability to cell membrane, has been reported to improve intracellular delivery systems. Here, we present an optimized method for nCaP-mediated gene delivery using an octa-arginine (R8)-functionalized nCaP vector containing a marker or functional gene construct. nCaP particle size was between 220-580 nm in diameter and all R8-functionalized nCaPs carried a positive charge. R8 concentration significantly improved nCaP transfection efficiency with high cell compatibility in human mesenchymal stem cells (hMSC) and human osteoblasts (hOB) in particular, suggesting nCaPs as a good option for non-viral vector gene delivery. Furthermore, pre-treatment with different endocytosis inhibitors identified that the endocytic pathway differed among cell lines and functionalized nanoparticles, with amiloride increasing transfection efficiency of R8-functionalized nCaPs in hMSC and hOB.

Electrochemical evaluation of the corrosion resistance of cup-yoke-type dental magnetic attachments.

The corrosion resistance of different magnetic assemblies­Magfit DX800 (Aichi Steel), Gigauss D800 (GC), Hyper Slim 4013, and Hicorex Slim 4013 (Hitachi Metals)­were electrochemically evaluated using anodic polarization curves obtained in 0.9% NaCl solution at 37°C. Stainless steels (444, XM27, 447J1, and 316L) composing the magnetic assemblies were also examined as controls. This revealed that all of the magnetic assemblies break down at 0.6-1.1 V; however, their breakdown potentials were all still significantly higher (p<0.05) than that of 316L. The distribution of elements in the laser welding zone between the yoke and shield ring was analyzed using EPMA; except with Magfit DX800, where the Cr content of the shield ring weld was greater than that of 316L. These magnetic assemblies are expected to have good corrosion resistance in the oral cavity, as their breakdown potentials are sufficiently higher than the 316L commonly used as a surgical implant material.