Synthesis, Characterization, and Bioactivity of Mesoporous Bioactive Glass Codoped with Zinc and Silver.

Due to the overconsumption of antimicrobials, antibiotic-resistant bacteria have become a critical health issue worldwide, especially methicillin-resistant S. aureus (MRSA) and vancomycin-resistant E. faecalis (VRE). Recently, many efforts have been made to load metals into bioactive glasses to enhance the multifunctionality of materials, such as antibacterial and osteoinductive functions. Zinc has been documented to stimulate the gene expression of various regulatory factors in bone cells. Meanwhile, previous studies have reported that silver and zinc could be a promising antibacterial combination with synergistic antimicrobial effects. Here, we sought to develop a biomaterial coreleasing zinc and silver, designated 80S-ZnAg, and to evaluate its antibacterial activity and biocompatibility. The textural analyses demonstrated different coreleasing patterns of zinc and silver for the materials. The chemical characterization revealed that the zinc in 80S-ZnAg could be the network modifier when its molar ratio was high, releasing more zinc; zinc could also be the network former when its molar ratio was low, showing an extremely low rate of release. However, the ICP results for 80S-Zn3Ag2 demonstrated up to 7.5 ppm of zinc and 67.6 ppm of silver. Among all the 80S-ZnAg materials, 80S-Zn3Ag2 demonstrated more marked antibacterial activity against MRSA and VRE than the others, with inhibition zones of 11.5 and 13.4 mm, respectively. The cytotoxicity assay exhibited nearly 90% cell viability at 20 mg/mL of 80-Zn3Ag2. Further clinical study is needed to develop an innovative biomaterial to address the issue of antibiotic resistance.

In Vitro Bioactivity and Antibacterial Effects of a Silver-Containing Mesoporous Bioactive Glass Film on the Surface of Titanium Implants.

Peri-implantitis is defined as a bacterial infection-induced inflammation and suppuration of soft and hard tissues surrounding a dental implant. If bacteria further invade the alveolar bone, they can easily cause bone loss and even lead to the early failure of a dental implant surgery. In the present study, an 80SiO2-15CaO-5P2O5 mesoporous bioactive glass film system containing 1, 5, and 10 mol% of silver was prepared on titanium implant discs (MBG-Ag-coated Ti) using sol-gel and spin coating methods. The wettability and adhesion strength of the films were evaluated using contact angle measurements and adhesion strength tests, respectively. The phase composition, chemical bonding, morphology, and oxidation states of the films were analyzed via X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDS), and X-ray photoelectron spectroscopy (XPS). In vitro bioactivity analysis of the films was performed by immersion in a simulated body fluid (SBF) for 24 h. Disk diffusion tests were performed on the early colonizing bacteria Aggregatibacter actinomycetemcomitans and Streptococcus mutans to evaluate the antibacterial ability of the films. A silver-containing mesoporous bioactive glass film with excellent biocompatibility and antibacterial properties was successfully prepared.

Genotyping of intron 22 inversion of factor VIII gene for diagnosis of hemophilia A by inverse-shifting polymerase chain reaction and capillary electrophoresis.

This is the first capillary electrophoresis (CE) analysis for diagnosis of hemophilia A (HA). The intron 22 inversion of factor VIII gene (F8) causes 40-50 % of severe bleeding disorder of HA in all human populations. Consequently, identification of the disease-causing mutations is becoming increasingly important for accurate genetic counseling and prenatal diagnosis. In this study, the key steps of inverse-shifting polymerase chain reaction (IS-PCR) and of short-end injection capillary electrophoresis were used for more specific and rapid genotyping of intron 22 inversion of F8. In IS-PCR, three specific primers were used to amplify 512-bp amplicon for wild type and 584-bp amplicon for patients with intron 22 inversion. The capillary gel electrophoresis (CGE) system was performed using 1× Tris-borate-EDTA (TBE) buffer containing 0.3 % (w/v) polyethylene oxide (PEO). The PCR amplicons were electrokinetically injected at 10 kV for 10 s at a temperature of 25 °C. The optimal short-end injection CGE was applied to detect the F8 gene of HA patients and carriers within 5 min. Intron 22 inversion was indeed found on some HA patients (13/35, 37.1 %). All genotyping results showed good agreement with DNA sequencing method and long-distance polymerase chain reaction (LD-PCR). The IS-PCR combined with short-end injection CGE method was feasible and efficient for intron 22 inversion screening of F8 in the HA populations.

Silica-based silver nanocomposite 80S/Ag as Aggregatibacter actinomycetemcomitans inhibitor and its in vitro bioactivity.

Background/purpose: As a commonly-found pathogen in periodontal disease, Aggregatibacter actinomycetemcomitans has been reported with several antibiotic resistance. Thus, to develop an alternative and protective therapy for A. actinomycetemcomitans infections is urgently needed in dentistry. In this study, we sought to synthesize a silica-based material to deliver silver nanoparticles for antibacterial purposes. Also, the bioactivities were examined via analyzing the formation of hydroxyapatite. Materials and methods: The 80S/Ag powders were prepared by the evaporation-induced self-assembly method, with Si, Ca, P, and Ag composition ratios of 80, 15, 5, and 1/5/10 (mole percentage), respectively. The nitrogen adsorption-desorption isotherms, transmission electron microscope, selected area electron diffraction, and Fourier transform infrared spectroscopy were conducted for textural analyses. The disk diffusion test was carried out against A. actinomycetemcomitans strain ATCC 29523. In vitro bioactivity assessment involved soaking 80S/Ag membrane powders in acellular simulated body fluid. Results: We successfully developed a material consisting of Si, Ca, P, and Ag, namely the 80S/Ag. In the antibacterial testing, the 80S/Ag demonstrated antibacterial activity against the commonly-found oral pathogen, A. actinomycetemcomitans, with a long-lasting effect for 168h. The formation of hydroxyapatite in simulated body fluid highlighted the characteristic of dentine remineralization for the 80S/Ag. The increased pH values after immersion in simulated body fluid would help neutralize the acidic oral environment. Conclusion: Our results indicate that 80S/Ag possesses remarkable antibacterial properties, hydroxyapatite formation, and increased pH values after immersion in simulated body fluid, supporting the potential therapeutic application of 80S/Ag for treating periodontal disease.

Synergistic effect of drug/antibiotic-impregnated micro/nanohybrid mesoporous bioactive glass/calcium phosphate composite bone cement on antibacterial and osteoconductive activities.

Calcium phosphate bone cements (CPC) can be used in minimally invasive surgery because of their injectability, and they can also be used to repair small and irregular bone defects. This study aimed to release the antibiotic gentamicin sulfate (Genta) to reduce tissue inflammation and prevent infection in the early stages of bone recovery. Subsequently, the sustained release of the bone-promoting drug ferulic acid (FA) mimicked the response of osteoprogenitor D1 cells interaction, thereby accelerating the healing process of the overall bone repair. Accordingly, the different particle properties of micro-nano hybrid mesoporous bioactive glass (MBG), namely, micro-sized MBG (mMBG) and nano-sized MBG (nMBG), were explored separately to generate different dose releases in MBG/CPC composite bone cement. Results show that nMBG had better sustained-release ability than mMBG when impregnated with the same dose. When 10 wt% of mMBG hybrid nMBG and composite CPC were used, the amount of MBG slightly shortened the working/setting time and lowered the strength but did not hinder the biocompatibility, injectability, anti-disintegration, and phase transformation of the composite bone cement. Furthermore, compared with 2.5wt%Genta@mMBG/7.5 wt% FA@nMBG/CPC, 5wt.%Genta@mMBG/5wt.%FA@nMBG/CPC exhibited better antibacterial activity, better compressive strength, stronger mineralization of osteoprogenitor cell, and similar 14-day slow-release trend of FA. The MBG/CPC composite bone cement developed can be used in clinical surgery to achieve the synergistic sustained release of antibacterial and osteoconductive activities.

Antibacterial silver-containing mesoporous bioglass as a dentin remineralization agent in a microorganism-challenged environment.

OBJECTIVES: To provide a suitable material capable of treating dentin hypersensitivity with simultaneous active antibacterial activity. METHODS: We developed silver-containing mesoporous bioglass (MBG-Ag) using the sol-gel technique, which loaded silver nanoparticles as promising bacteriostatic agents. The MBG-Ag with a powder-to-liquid ratio of 0.5 g: 0.01 mL were uniformly mixed with 20 %, 30 %, and 40 % phosphoric acid for 5, 10 and 20 min, respectively. Furthermore, we evaluated the occlusion efficiency, depth of penetration, and antibacterial activity of dentin specimens by simulating a Streptococcus mutans (S. mutans) infection on dentin. Scanning electron microscopy (SEM), X-ray diffraction (XRD), and Fourier transform infrared spectroscopy (FTIR) were used to characterize the powders and assess tubule occlusion. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the MBG-Ag against S. mutans were determined via time-killing curves and colony formation assays. RESULTS: The MIC ranged from 2.5 to 5 mg/mL, and the MBC ranged from 5 to 10 mg/mL. The highest dentinal tubule occlusion efficiency was over 90 %. The colony formation assay confirmed that 5 mg/mL MBG-Ag mixed with phosphoric acid reached the bactericidal concentration. CONCLUSION: The MBG-Ag 40PA achieved a good occlusion efficiency and deep apatite precipitation in a short time, implying its superiority in clinical applications. CLINICAL RELEVANCE: The MBG-Ag formed in this study is a promising candidate for the treatment of demineralized dentin and confers antibacterial effects on the remineralized dentin surface against S. mutans.

The Antibacterial and Remineralization Effect of Silver-Containing Mesoporous Bioactive Glass Sealing and Er-YAG Laser on Dentinal Tubules Treated in a Streptococcus mutans Cultivated Environment.

The aim of this study was to evaluate the remineralization and antibacterial effect of silver-containing mesoporous bioactive glass (MBG-Ag) sealing combined with Er:YAG laser irradiation on human demineralized dentin specimens in a Streptococcus mutans cultivated environment. A total of 48 human dentin specimens were randomly divided into four groups. The characteristics of MBG-Ag and the occlusion efficiency of the dentinal tubules were analyzed using X-ray diffraction patterns, Fourier-transform infrared spectroscopy, scanning electron microscope images and energy dispersive X-ray spectroscopy. Moreover, the antibacterial activity against Streptococcus mutans was evaluated by colony formation assay. The results showed that the dentin specimens with Er:YAG laser irradiation can form a melted occlusion with a size of 3-4 µm. MBG-Ag promoted the deposition of numerous crystal particles on the dentinal surface, reaching the deepest penetration depth of 70 µm. The results suggested that both MBG-Ag and laser have the ability to enhance the remineralization and precipitation of hydroxyapatite crystals. While the results showed that MBG-Ag sealing combined with the thermomechanical subablation mode of Er:YAG laser irradiation-induced dense crystalline deposition, reaching a penetration depth of more than 300 µm, silver nanoparticles without good absorption of the Er:YAG laser resulted in a heterogeneous radiated surface. Er:YAG laser irradiation with a low energy and pulse rate cannot completely inhibit the growth of S. mutans, but MBG-Ag sealing reached the bactericidal concentration. It was concluded that the simultaneous application of MBG-Ag sealing and Er:YAG laser treatment can prevent the drawbacks of their independent uses, resulting in a superior form of treatment for dentin hypersensitivity.

Genotyping of alpha-thalassemia deletions using multiplex polymerase chain reactions and gold nanoparticle-filled capillary electrophoresis.

A gold nanoparticle-filled capillary electrophoresis method combined with three multiplex polymerase chain reactions (PCRs) was established for simultaneous diagnosis of five common alpha-thalassemia deletions, including the -alpha(3.7) deletion, -alpha(4.2) deletion, Southeast Asian (--(SEA)), Filipino (--(FIL)) and Thai (--(THAI)) deletions. Gold nanoparticles (GNPs) were used as a pseudostationary phase to improve the resolution between DNA fragments in a low-viscosity polymer. To achieve the best CE separation, several parameters were evaluated for optimizing the separation conditions, including the capillary coating, the concentrations of polymer sieving matrix, the sizes and concentrations of GNPs, the buffer concentrations, and the pH. The final CE method for separating a 200-base pair (bp) DNA ladder and alpha-thalassemia deletions used a DB-17 capillary, 0.6% poly(ethylene oxide) (PEO) prepared in a mixture of GNP(32nm) solution and glycine buffer (25mM, pH 9.0) (80:20, v/v) as the sieving matrix with 1microM YO-PRO-1 for fluorescence detection; the applied voltage was -10kV (detector at anode side) and the separation temperature was 25 degrees C. Under these optimal conditions, 15 DNA fragments with sizes ranging from 0.2kb to 3.0kb were resolved within 11.5min. The RSDs of migration times were less than 2.81%. A total of 21 patients with alpha-thalassemia deletions were analyzed using this method, and all results showed good agreement with those obtained by gel electrophoresis.

Thermal cycling effects on adhesion of resin-bovine enamel junction among different composite resins.

Thermal cycling is used to mimic the changes in oral cavity temperature experienced by composite resins when used clinically. The purpose of this study is to assess the thermal cycling effects of in-house produced composite resin on bonding strength. The dicalcium phosphate anhydrous filler surfaces are modified using nanocrystals and silanization (w/NP/Si). The resin is compared with commercially available composite resins Filtek Z250, Z350, and glass ionomer restorative material GIC Fuji-II LC (control). Different composite resins were filled into the dental enamel of bovine teeth. The bond force and resin-enamel junction graphical structures of the samples were determined after thermal cycling between 5 and 55°C in deionized water for 600 cycles. After thermal cycling, the w/NP/Si 30wt%, 50wt% and Filtek Z250, Z350 groups showed higher shear forces than glass ionomer GIC, and w/NP/Si 50wt% had the highest shear force. Through SEM observations, more of the fillings with w/NP/Si 30wt% and w/NP/Si 50wt% groups flowed into the enamel tubule, forming closed tubules with the composite resins. The push-out force is proportional to the resin flow depth and uniformity. The push-out tubule pore and resin shear pattern is the most uniform and consistent in the w/NP/Si 50wt% group. Accordingly, this developed composite resin maintains great mechanical properties after thermal cycling. Thus, it has the potential to be used in a clinical setting when restoring non-carious cervical lesions.

Phosphorus Effects of Mesoporous Bioactive Glass on Occlude Exposed Dentin.

In recent studies, sealing of exposed dentinal tubules is generally considered as one of the most effective strategies to treat dentin hypersensitivity. Mesoporous bioactive glass (MBG) is a potential material for treating dentin hypersensitivity due to its highly specific areas for dissolution and re-precipitated reaction for reduction in dentin permeability. The groups of commercial products of PerioGlas®, synthetic MBG and MBG without phosphorus (MBGNP) were compared. The MBG and MBGNP powders were prepared by the sol-gel method and mixed with different calculated ratios of phosphoric acid (PA) and then was brushed onto dentin surfaces. We used X-ray diffractometer (XRD), scanning electronic microscope (SEM), and Fourier transform infrared spectroscopy (FTIR) to investigate the physiochemistry and the occlusion ability of dentinal tubules. The results showed that MBG paste mixed with PA solution has a better ability for occluding dentinal tubules than MBGNP; it has a short reaction time and good operability. The major crystallite phase of MBG agents was monocalcium phosphate monohydrate [Ca(H2PO4)2·H2O] in the early stages of the reactions. MBG pastes that were mixed with 30% and 40% PA had the ability to create excellent penetration depth greater than 80 µm. These agents have the potential to treat dentin hypersensitivity.

Properties of osteoconductive biomaterials: calcium phosphate cement with different ratios of platelet-rich plasma as identifiers.

This study aims to evaluate further the performance of a platelet-rich plasma (PRP) additive incorporated with calcium phosphate bone cement (CPC) in vitro to prove its efficiency as bone graft substitutes and its compatibility to be incorporated into the CPC with other techniques in clinical restoration in vivo. The growth factor release ability and the osteogenic evaluation of PRP, CPC, and PRP/CPC testing groups with 5, 10, and 15 wt.% PRP were compared in vitro. Four groups were measured using non-decalcified staining methods in vivo, which include the testing group of 10 wt.% PRP/CPC selected from the evaluation in vitro, by using both the autograft with rabbit trabecular and CPC-only as comparison groups and the group without grafting material as the control sample. The results obtained through specimen immersion show that growth factor release and alkaline phosphatase activities after osteoprogenitor cell culture had a significantly better effect on 10 and 15 wt.% PRP/CPC than on the other groups in vitro. Analysis results suggest that PRP was still retained in the CPC matrix even after 32 days of immersion. The results in vivo show that the histology of the autograft bone and the control group without grafting material exhibited fibrous connective and adipose tissues, which obviously filled the created cavity even at nine weeks after the operation. Osteoregeneration was more successful in the PRP-additive group, which accumulated bone remodeling than in the other groups. In conclusion, CPC could be a potential carrier with adequate PRP additives that bear a therapeutic potential for enhanced bone tissue regeneration.

Calcium phosphate bone cement with 10 wt% platelet-rich plasma in vitro and in vivo.

OBJECTIVES: The aim of this study was to evaluate the performance of a 10 wt% platelet-rich plasma (PRP) additive composite with calcium phosphate cement (CPC) in vitro and in vivo. METHODS: The in vitro testing of modulus, the apatite conversion rate, morphology, cell and alkaline phosphatase (ALP) activities, and in vivo testing of histological examinations between two groups of 10 wt% PRP/CPC and CPC were characterised and compared. RESULTS: Although the crystallite morphologies showed a retarded effect in the PRP/CPC group in vitro, the modulus results showed that the 10 wt% PRP/CPC group had a significant reduction in strength but had no significant changes in the relative conversion ratio of the apatite phase with CPC only. The osteogenic evaluation of ALP expression was significantly increased by the PRP additives group with stem cells (D1) cultured for different periods (2-32 days). Our histological examinations showed that greater remodelling and the phenomenon of isolated/detached CPC particles were significantly observed at 9 weeks after implantation when the 10 wt% PRP/CPC composite was used. CONCLUSION: The results demonstrate that CPC may be a potential candidate as a carrier with PRP additives for bone regeneration.

Single nucleotide polymorphism detection in the hMSH2 gene using conformation-sensitive CE.

CE allows for highly reproducible analysis of DNA fragments which can be used to detect DNA mutations including SNPs. We have utilized a simple and direct CE analysis method for SNP analysis called conformation-sensitive CE (CSCE), based on the principle of single nucleotide different to produce conformational changes in the mildly denaturing solvent system. This method was applied to analysis of a mutation in the promoter region of the hMSH2 gene. This gene belongs to the human DNA mismatch repair system, which is responsible for recognizing and repairing mispaired nucleotides, and mutations in the hMSH2 gene are known to cause hereditary nonpolyposis colorectal cancer (HNPCC). PCR fragments generated from the promoter region of the hMSH2 gene, displaying either a C/C homozygote, C/T heterozygote, or T/T homozygote genotype, did not require further pretreatment before electrokinetic injection. The CE separation, using a 1xTris-borate-EDTA (TBE) buffer containing 3% w/v hydroxylethyl cellulose (HEC) and 6 M urea, was performed under reverse polarity with a separation temperature of 15 degrees C. The genotypes of 204 healthy volunteers and 13 colorectal cancer patients were determined using CSCE, and the results confirmed by DNA sequencing. While the CSCE separations were shown to be highly reproducible and sensitive for screening large populations, no correlation was observed between cancer patients and this hMSH2 gene polymorphism.