Hae-Hyoung Lee

Effect of Mineral Trioxide Aggregate on Dentin Bridge Formation and Expression of Dentin Sialoprotein and Heme Oxygenase-1 in Human Dental Pulp

This study was conducted to evaluate the pulpal response to direct capping with either mineral trioxide aggregate (MTA) or calcium hydroxide (CH) cement in humans, with a focus on dentin bridge formation and dentin sialoprotein (DSP) and heme oxygenase-1 (HO-1) expression. Direct pulp capping was performed in 20 cases of caries-free human third molars. The pulps were exposed and capped with either MTA or hard-setting CH. After 2 months, the teeth were extracted, and the specimens were prepared for histologic and immunohistochemical evaluations. Histologically, 100% of the MTA group and 60% of the CH group developed dentin bridges. The mean thickness of the dentin bridges observed in the MTA group was statistically greater than that of CH group. In addition, DSP and HO-1 were expressed in the odontoblast-like cells and pulp fibroblasts beneath the dentin bridge; furthermore, significantly greater immunostaining was observed in the MTA group than in the CH group. Collectively, these results indicate that MTA is superior to CH in terms of inducing the dentinogenic process in human pulp capping.

Apatite-mineralized polycaprolactone nanofibrous web as a bone tissue regeneration substrate

Degradable synthetic polymers with a nanofibrous structure have shown great promise in populating and recruiting cells for the reconstruction of damaged tissues. However, poor cell affinity and lack of bioactivity have limited their potential usefulness in bone regeneration. We produced polymeric nanofiber poly(epsilon-caprolactone) (PCL) with its surface mineralized with bone-like apatite for use as bone regenerative and tissue engineering matrices. PCL was first electrospun into a nanofibrous web, and the surface was further mineralized with apatite following a series of solution treatments. The surface of the mineralized PCL nanofiber was observed to be almost fully covered with nanocrystalline apatites. Through mineralization, the wettability of the nanofiber matrix was greatly improved. Moreover, the murine-derived osteoblastic cells were shown to attach and grow actively on the apatite-mineralized nanofibrous substrate. In particular, the mineralized PCL nanofibrous substrate significantly stimulated the expression of bone-associated genes, including Runx2, collagen type I, alkaline phosphatase, and osteocalcin, when compared with the pure PCL nanofiber substrate without mineralization. The currently developed polymer nanofibrous web with the bioactive mineralized surface is considered to be potentially useful as bone regenerative and tissue engineering matrices.

Bioactivity improvement of poly(ε-caprolactone) membrane with the addition of nanofibrous bioactive glass

Nanofibrous glass with a bioactive composition was added to a degradable polymer poly(epsilon-caprolactone) (PCL) to produce a nanocomposite in thin membrane form ( approximately 260 microm). The bioactivity and osteoblastic responses of the nanocomposite membrane were examined and compared with those of a pure PCL membrane. Glass nanofibers with diameters in the range of hundreds of nanometers were added to a PCL solution at 20 wt.%, and the mixture was stirred vigorously and air dried. The obtained nanocomposite membrane showed that many chopped glass nanofibers formed by the mixing step were embedded uniformly into the PCL matrix. The nanocomposite membrane induced the rapid formation of apatite-like minerals on the surface when immersed in a simulated body fluid. Murine-derived osteoblastic cells (MC3T3-E1) grew actively over the nanocomposite membrane with cell viability significantly improved compared with those on the pure PCL membrane. Moreover, the osteoblastic activity, as assessed by the expression of alkaline phosphatase, was significantly higher on the nanocomposite membrane than on the pure PCL membrane. The currently developed nanocomposite of the bioactive glass-added PCL might find applications in the bone regeneration areas such as the guided bone regeneration (GBR) membrane.

Chitosan–nanobioactive glass electrophoretic coatings with bone regenerative and drug delivering potential

Nanocomposites with bone-bioactivity and drug eluting capacity are considered as potentially valuable coating materials for metallic bone implants. Here, we developed composite coatings of chitosan (CH)–bioactive glass nanoparticles (BGn) via cathodic electrophoretic deposition (EPD). BGn 50–100 nm in size with aminated surface were suspended with CH molecules at different ratios (5–20 wt% BGn) in aqueous medium, and EPD was performed. Uniform coatings with thicknesses of a few to tens of micrometers were produced, which was controllable by the EPD parameters (voltage, pH and time). Thermogravimetric analysis revealed the quantity of BGn within the coatings that well corresponded to that initially incorporated. Apatite forming ability of the coatings, performed in simulated body fluid, was significantly improved by the addition of BGn. Degradation of the coatings increased with increasing BGn addition. Of note, the degradation profile was almost linear with time; degradation of 5–13 wt% during 1 week became 30–40 wt% after 7 weeks at almost a constant rate. The CH–BGn coatings showed favorable cell adhesion and growth, and stimulated osteogenic differentiation. Drug loading and release capacity of the CH–BGn coatings were performed using the ampicillin antibiotic as a model drug. Ampicillin, initially incorporated within the CH–BGn suspension, was eluted from the coatings continuously over 10–11 weeks, confirming long-term drug delivering capacity. Antibacterial tests also confirmed the effects of released ampicillin using agar diffusion assay against Streptococcus mutants. The CH–BGn may be potentially useful as a coating composition for metallic implants due to the excellent bone bioactivity and cell responses, as well as the capacity for long-term drug delivery.

Biomedical nanocomposites of poly(lactic acid) and calcium phosphate hybridized with modified carbon nanotubes for hard tissue implants

Degradable polymer-based materials are attractive in orthopedics and dentistry as an alternative to metallic implants for use as bone fixatives. Herein, a degradable polymer poly(lactic acid) (PLA) was combined with novel hybrid nanopowder of carbon nanotubes (CNTs)-calcium phosphate (CP) for this application. In particular, CNTs-CP hybrid nanopowders (0.1 and 0.25% CNTs) were prepared from the solution of ionically modified CNTs (mCNTs), which was specifically synthesized to be well-dispersed and thus to effectively adsorb onto the CP nanoparticles. The mCNTs-CP hybrid nanopowders were then mixed with PLA (up to 50%) to produce mCNTs-CP-PLA nanocomposites. The mechanical tensile strength of the nanocomposites was significantly improved by the addition of mCNTs-CP hybrid nanopowders. Moreover, nanocomposites containing low concentration of mCNTs (0.1%) showed significantly stimulated biological responses including cell proliferation and osteoblastic differentiation in terms of gene and protein expressions. Based on this study, the addition of novel mCNT-CP hybrid nanopowders to PLA biopolymer may be considered a new material choice for developing hard tissue implants.

Bone regeneration by bioactive hybrid membrane containing FGF2 within rat calvarium

This study examined the bone regeneration potential of a novel hybrid membrane consisting of collagen and nano-bioactive glass (nBG) incorporating basic fibroblast growth factor (FGF2) for use in guided bone regeneration. nBG was added to a reconstitution of collagen at a concentration of 30%, and the hybrid was formulated into a thin membrane. FGF2 (50 microg/ml) was adsorbed to the hybrid membrane. This level of FGF2 was found to be the optimal concentration to stimulate osteoblastic differentiation in vitro. Three membrane groups, including pure collagen, collagen-nBG hybrid and its combination with FGF2 were implanted within a rat calvarium defect (phi = 5 mm) for a period of 3 weeks. Histomorphometric analysis was carried out to evaluate the bone regeneration within the defect. The results showed that the defect in the collagen-nBG-FGF2 membrane was recovered almost completely, while partial recovery was observed in the other membrane groups (collagen and collagen-BG). However, there was little defect recovery in the blank control. The new bone formation was as high as approximately 60, approximately 45, and approximately 30% of the defect treated with the collagen-nBG-FGF2, collagen-BG, and collagen, respectively, whilst only 4% of new bone was observed in the blank control. Overall, the nBG was shown to stimulate bone formation of the collagen membrane, and FGF2 synergistically accelerated the bone regeneration within a rat calvarium defect.

Bone Formation on the Apatite-coated Zirconia Porous Scaffolds within a Rabbit Calvarial Defect:

Previously, a strong and bioactive ceramic scaffold consisting of a porous zirconia body coated with apatite double layers (fluorapatite (FA) as an inner layer and hydroxyapatite (HA) as an outer layer) was successfully fabricated. In this contribution, the authors investigate the in vivo performance of the engineered bioceramic scaffolds using a rabbit calvarial defect model. In particular, the porosity and pore size of the scaffolds are varied in order to observe the geometrical effects of the scaffolds on their bone formation behaviors. The scaffolds supported on a zirconia framework can be produced with an extremely high porosity (~84—87%), while retaining excellent compressive strength (~7—8 MPa), which has been unachievable in the case of pure apatite scaffolds (~74% porosity with ~2MPa strength). The experimental groups used in this study include three types of zirconia scaffolds coated with apatite; high porosity (~87%) with large pore size (~500— 700 μm): AZ-HL, high porosity (~84%) with small pore size (~150—200 μm): AZ-HS, and low porosity (~75%) with large pore size (~500—700 μm): AZ-LL, as well as one type of HA porous scaffold: low porosity (~74%) with a large pore size (~500—700 μm) for the purpose of comparison. The scaffolds prepared with dimensions of ~ 10 mm (diameter) × 1.2 mm (thickness) are grafted in rabbit calvaria defects. The histological sections are made at 4 and 12 weeks after surgery and immunohistochemical analyses are performed on the samples. All of the specimens show a good healing response without adverse tissue reactions. Good healing is shown at 4 weeks post-surgery with the ingrowth of new bone into the macropore-channels of the scaffolds. The newly formed bone amounts to ~19.9—24.2% of the initial defect area, depending on the scaffold type, but there is no statistical significance between the scaffold groups. However, the defects without the scaffolds (control group) show a significantly lower bone formation ratio (~4.3%). At twelve weeks after surgery, the extent of new bone formation is more pronounced in all of the scaffold groups. All of the scaffold groups show significantly higher bone formation ratios (26.7—46.9%) with respect to the control without the graft. In the comparison between the scaffold groups, those with high porosities (AZ-HL and AZ-HS) exhibit significantly higher bone formation as compared to the scaffold with low porosity (AZ-LL). Based on the present in vivo test performed within a rabbit calvaria defect model, it is concluded that the apatite-coated zirconia scaffolds show good bone forming ability and are considered to be a promising scaffolding material for bone regeneration since they possess a high level of both mechanical and biological properties.

Development of long-term antimicrobial poly(methyl methacrylate) by incorporating mesoporous silica nanocarriers.

OBJECTIVE Poly(methyl methacrylate) (PMMA) used as removable denture bases or orthodontic appliances has relatively poor antimicrobial properties, which accelerate oral infection and induce unfavorable odors. Mesoporous silica nanoparticles (MSNs) have been highlighted as a potential additive to overcome this issue because of their drug-loading capacity. Here, we present the long-term antimicrobial effect of MSN-incorporated PMMA with drug-loading capacity. METHODS After the MSNs were characterized, MSN incorporation into chemically activated PMMA (0.5, 1, 2.5 or 5wt%) relative to the methyl methacrylate powder by mass was fabricated into a rectangular specimen (1.4×3.0×19.0mm) for a 3-point flexural test at a speed of 1mm/min or a disk (∅=11.5mm and d=1.5mm) for investigation of its antimicrobial effects. RESULTS A typical spherical morphology with a well-ordered mesoporous structure of the MSNs was visualized and is beneficial for loading drugs and combining in matrixes. Among the tested levels of MSN incorporation in PMMA (0.5, 1, 2.5 or 5wt%), only 5wt% decreased the flexural strength (p<0.05), whereas the flexural modulus was not significantly decreased (p>0.05). The surface roughness and surface energy were increased with 2.5wt% or 5wt% incorporation. An anti-adherent effect against Candida albicans and Streptococcus oralis after 1h of attachment was only observed with 2.5 and 5wt% incorporation compared to a lack of MSNs (p<0.05). A long-term antimicrobial effect was observed for 2 weeks with 2.5wt% MSN-incorporated PMMA when amphotericin B was loaded into the MSNs on the PMMA surface. SIGNIFICANCE The long-term antimicrobial performance after loading amphotericin B into the MSN-incorporated PMMA suggests the potential clinical usefulness of MSN-incorporated PMMA resin.

Cytotoxicity and anti-inflammatory effects of zinc ions and eugenol during setting of ZOE in immortalized human oral keratinocytes grown as three-dimensional spheroids

OBJECTIVES The objective of this study is to assess the cytotoxic and anti-inflammatory effects of ZOE cement during setting in two-dimensional (2D) or three-dimensional (3D) cultures of immortalized human oral keratinocytes (IHOKs) with determining the extract components responsible for these effects. METHODS Extracts of mixed ZOE at different stages of setting were analyzed by a digital pH meter, ICP-MS, and GC-MS. Serial concentrations of extract and their mixture of ZnCl2, ZnSO4·H2O, and eugenol liquid were added to the 2D and 3D IHOK cultures to determine the half maximal effective concentration in investigating the cause of cytotoxicity by means of WST assay and to investigate mRNA expression levels of inflammatory cytokines by RT-PCR. RESULTS Zn(2+) and eugenol (4-19 ppm) were detected in the extracts. In the early setting stage, significant cytotoxicity was observed in the 2D and 3D IHOK cultures (P<0.05). The EC50 of Zn(2+) from ZnCl2 was 5-44 ppm in both cultures, whereas the EC50 of eugenol was not detectable under 100 ppm. Along with the lower levels of inflammatory cytokine gene expressions in the extract, treatment of the 2D IHOKs with Zn(2+) alone and treatment of the 3D IHOKs with Zn(2+) plus eugenol resulted in significantly lower expression levels of IL-1β, IL-6, and IL-8 (P<0.05). SIGNIFICANCE The cytotoxic effect of ZOE on IHOKs was greater during the setting stage owing to the presence of Zn(2+). The anti-inflammatory response to ZOE was induced by a combination of Zn(2+) and eugenol. Cytotoxic and anti-inflammatory effects differed between the 2D and 3D IHOK cultures.

Development of a novel aluminum-free glass ionomer cement based on magnesium/strontium-silicate glasses.

The effects of strontium substitution for magnesium in a novel aluminum-free multicomponent glass composition for glass ionomer cements (GICs) were investigated. A series of glass compositions were prepared based on SiO2-P2O5-CaO-ZnO-MgO(1-X)-SrOX-CaF2 (X=0, 0.25, 0.5 and 0.75). The mechanical properties of GICs prepared were characterized by compressive strength, flexural strength, flexural modules, and microhardness. Cell proliferation was evaluated indirectly by CCK-8 assay using various dilutions of the cement and rat mesenchyme stem cells. Incorporation of strontium instead of magnesium in the glasses has a significant influence on setting time of the cements and the properties. All mechanical properties of the GICs with SrO substitution at X=0.25 were significantly increased, then gradually decreased with further increase of the amount of strontium substitution in the glass. The GIC at X=0.25, also, showed an improved cell viability at low doses of the cement extracts in comparison with other groups or control without extracts. The results of this study demonstrate that the glass compositions with strontium substitution at low levels can be successfully used to prepare aluminum-free glass ionomer cements for repair and regeneration of hard tissues.