Development of an antimicrobial, 3D printable denture base material with K18 quaternary ammonium silane-functionalized methyl methacrylate and filler.

STATEMENT OF PROBLEM: Denture base materials are highly susceptible to microbial colonization, which can lead to denture stomatitis. In addition, patients who sleep with their dentures have an increased chance of contracting pneumonia. Commercially available antimicrobial denture base materials to prevent or combat microbial colonization are lacking. PURPOSE: The purpose of this in vitro study was to determine the effects of K18 quaternary ammonium methacryloxy silane-functionalized filler (K18-Filler) and methyl methacrylate (K18-MMA) on the polymerization of 3D printed denture base material and its esthetic, mechanical, and antimicrobial properties. MATERIAL AND METHODS: K18-Filler (0%, 10%, 20% w/w) and K18-MMA (0%, 5%, 12.5% w/w) were added to a 3D printable denture base resin (Denture Base Resin, Original Pink; Formlabs Inc) and 3D printed. Specimens were tested by using the Rockwell15T hardness, near infrared FTIR monomer-to-polymer degree of conversion (DoC), transparency parameter (TP), color shift, and 3-point bend and by counting colony forming units against Streptococcus aureus, Streptococcus sanguinis and Candida albicans tests. Data were analyzed using analysis of variance with the Tukey-Kramer HSD post hoc test. RESULTS: Control resins had significantly higher Rockwell15T hardness than most of the K18 groups (P<.05) but had comparable DoC with all K18 groups except one, showing that all groups were well polymerized. Controls had significantly higher TP than most K18 groups, but most K18 groups had ΔE<3.3, so the color shift was not noticeable. However, the 12.5% K18-MMA with 10% and 20% K18-Filler groups, which were also the groups used to test for antimicrobial activity, had ΔE>8. All K18 groups had comparable or greater moduli than the controls, but the controls had significantly higher ultimate transverse strengths than most K18 groups (P<.05). All 12.5% K18-MMA with K18-Filler groups had significant antimicrobial activity against S. aureus, S. sanguinis, and C. albicans. CONCLUSIONS: 12.5% K18-MMA and K18-Filler produced 3D printable denture materials with comparable polymerization properties and significant antimicrobial properties against S. mutans, S. sanguinis, and C. albicans. High K18-MMA and K18-Filler concentrations caused significant color shifts and reductions in ultimate strengths.

Antimicrobial dental composites with K18-methyl methacrylate and K18-filler.

OBJECTIVE: To determine the effects of using K18-methyl methacrylate (K18-MMA) and K18-Filler on composite cure, esthetic, mechanical, polymerization shrinkage, and antimicrobial properties. METHODS: K18-MMA (0-20% w/w) was used to replace TEGDMA in a 70:30 Bis-GMA:TEGDMA composite filled to 70% w/w with barium glass or K18-Filler. Composite degree of cure (Rockwell15T hardness and near Infrared FTIR), hydrophilicity (contact angle measurements), translucency (transparency parameter measurements, TP), mechanical (3-point bend test), polymerization shrinkage (volumetric shrinkage and shrinkage stress), and antimicrobial properties (colony counting assay) against Streptococcus mutans, Streptococcus sanguinis, and Candida albicans were determined. RESULTS: All experimental groups had comparable degrees of cure (near Infrared FTIR and Rockwell15T Hardness), TP, moduli, polymerization volumetric shrinkages and shrinkage stresses to those of controls (Bonferroni corrected p > 0.0018). Only one group (15% K18-MMA+K18-Filler) had significantly different (lower) contact angles as compared to that of controls (Bonferroni corrected p < 0.0018). Most of the K18-Filler-containing composites had significantly lower ultimate transverse strengths (UTS) than controls (Bonferroni corrected p < 0.0018). Controls had significantly greater S mutans colony counts than 15% and 20% w/w K18-MMA+K18-Filler groups, and greater S sanguinis and C albicans colony counts than K18-containing groups. Of the composites with that provided significant antimicrobial properties against S. mutans, S. sanguinis, and C. albicans, only the 20% K18-MMA+K18-Filler group had significantly lower UTS than controls. SIGNIFICANCE: Composites with K18-MMA and K18-Filler with comparable physical properties to control composites and significant antimicrobial properties have been developed. K18-MMA and K18-Filler seem to be suitable for incorporation into commercial dental resins.

Development of a Boron Nitride-Filled Dental Adhesive System.

There is a dearth of adhesive systems capable of forming stable bonds between restorative materials and tooth surfaces. To address the concern, this study determined the effects of using methacrylate-functionalized boron nitride nanosheets (BNNSs) in a polymeric dental adhesive system. The bisphenol A glycidyl dimethacrylate (BisGMA):2 hydroxyethyl methacrylate (HEMA) (60:40) adhesive monomer blend with a photoinitiator was filled with 0 wt% (control), 0.1 wt%, and 1 wt% BNNSs and light cured. Fourier transform infrared spectroscopy was performed to determine the conversion degree of monomer double bonds (DoC). Water absorption and solubility were measured. Flexural strength and Youngs's modulus were evaluated to determine the mechanical properties of the composite adhesive system. Finally, dentin bond strength degradation and fracture mode were quantified with a microtensile bond test to confirm the bonding ability of the developed adhesive system. Results showed that the incorporation of BNNSs increased DoC (9.8% and 5.4% for 0.1 and 1 wt%, respectively), but it did not affect water sorption (101.9-119.72 (µg/mm3)), solubility (2.62-5.54 (µg/mm3)), Young's modulus (529.1-1716.1 MPa), or microtensile bond strength (46.66-54.72 MPa). Further studies are needed with varying BNNS loading percentages from 0.1 wt% to 1 wt% in order to more comprehensively determine the effect of BNNSs on dental adhesives.

A low-shrinkage, hydrophobic, degradation-resistant, antimicrobial dental composite using a fluorinated acrylate and an oxirane.

OBJECTIVE: To develop a low shrinkage, hydrophobic, degradation-resistant, antimicrobial dental composite using a fluorinated acrylate, and a difunctional oxirane. METHODS: The effects of a fluorinated acrylate (2-(perfluorooctyl)ethyl acrylate; PFOEA), a difunctional oxirane (EPALLOY™ 5001; EP5001), and a three-component initiator system (camphorquinone/ethyl 4-dimethylaminobenzoate/4-Isopropyl-4'-methyldiphenyl iodonium Tetrakis (pentafluorophenyl) borate; CQ/EDMAB/Borate) on bisphenol A glycidyl dimethacrylate: triethylene glycol dimethacrylate (BisGMA:TEGDMA) composite surface hardness, degree of monomer-to-polymer conversion, hydrophobicity, translucency, mechanical properties, polymerization shrinkage and shrinkage stress, degradation, water imbibition, and antimicrobial properties were determined. RESULTS: Overall the experimental composites had comparable mechanical properties and lower volumetric polymerization shrinkage and shrinkage stress as compared to BisGMA:TEGDMA controls. Addition of PFOEA increased composite hydrophobicity, but it decreased degree of cure, ultimate transverse strength, and translucency. It also decreased polymerization shrinkage and shrinkage stress. The use of the CQ/EDMAB/Borate initiator system was beneficial for the cure and mechanical properties of the 30% w/w PFOEA group. However, it decreased the hydrophobicity and translucency of those composites. The addition of EP5001, at the low concentration used in this work, did not contribute to reduced polymerization volumetric shrinkage or antimicrobial properties, but it did reduce shrinkage stress. CONCLUSIONS: A mechanically viable hydrophobic composite system with reduced polymerization shrinkage and shrinkage stress has been developed by adding PFOEA and EP5001. However, the addition of EP5001 did not render the composite antimicrobial due to the low concentration used. Further research is needed to determine the lowest concentration at which EP5001 provides antimicrobial activity. The composites developed here have the potential to improve longevity of traditional BisGMA:TEGDMA composite systems.

Repair of resin-veneered polyetheretherketone after veneer fracture.

STATEMENT OF PROBLEM: If a composite resin-veneered polyetheretherketone (PEEK) restoration chips or fractures, a repair may be indicated. However, the most appropriate repair protocol for a composite resin-veneered PEEK restoration is unclear. PURPOSE: The purpose of this in vitro study was to determine the efficacy of airborne-particle abrasion and/or a primer in the repair of composite resin-veneered PEEK prostheses. MATERIAL AND METHODS: PEEK specimens (N=80) were airborne-particle abraded with alumina before being conditioned with a methyl methacrylate-based primer. A thin layer of opaquer was applied, and a split mold was then filled with a veneering resin. The specimens underwent 5000 thermocycles, and then shear bond strength (SBS) was determined and used as the positive control group. Specimens that failed in either mixed or adhesive modes were contaminated with saliva and then exposed to 4 different repair treatment methods: no airborne-particle abrasion, bonded without a primer (negative control group); airborne-particle abrasion, bonded without a primer; no airborne-particle abrasion, bonded with a primer; and airborne-particle abrasion, bonded with a primer. All specimens had opaquer applied before being veneered. Repaired specimens then underwent thermocycling before SBS testing. Data were analyzed via ANOVA with a Newman-Keuls post hoc test (α=.05). RESULTS: The SBS values for the negative control were significantly lower than those of all other repair groups and the control group (P<.018). No significant differences in the SBS values were found among these other repair groups or the positive control group (P>.05). CONCLUSIONS: Composite resin-veneered PEEK restorations or prostheses repaired with airborne-particle abrasion and/or primer can provide SBS comparable with that of the initial SBS.

The effect of aging methods on the fracture toughness and physical stability of an oxirane/acrylate, ormocer, and Bis-GMA-based resin composites.

PURPOSE: To determine the effect of aging methods on the fracture toughness of a conventional Bis-GMA-based resin composite (Filtek Supreme), an ormocer-based resin composite (Admira), and an experimental hydrophobic oxirane/acrylate interpenetrating network resin system (OASys)-based composite. METHODS: A 25 × 5 × 2.8-mm stainless-steel mold with 2.5 mm single-edge center notch, following ASTM standards [E399-90], was used to fabricate 135 specimens (n = 15) of the composite materials and randomly distributed into groups. For the baseline group, specimens were fabricated and then tested after 24-h storage in water. For the biofilm challenge, specimens were randomly placed in a six-well tissue culture plate and kept at 37 °C with bacterial growth media (Brain Heart Infusion (BHI); Streptococcus mutans) changed daily for 15 days. For the water storage challenge, specimens were kept in 5 ml of deionized distilled autoclaved water for 30 days at 37 °C. µCT evaluation by scanning the specimens was performed before and after the proposed challenge. Fracture toughness (KIc) testing was carried out following the challenges. RESULTS: µCT surface area and volume analyses showed no significant changes regardless of the materials tested or the challenge. Filtek and Admira fracture toughness was significantly lower after the biofilm and water storage challenges. OASys mean fracture toughness values after water aging were significantly higher than that of baseline. Toughness values for OASys composites after biofilm aging were not statistically different when compared to either water or baseline values. CONCLUSION: The fracture toughness of Bis-GMA and ormocer-based dental resin composites significantly decreased under water and bacterial biofilm assault. However, such degradation in fracture toughness was not visible in OASys-based composites. CLINICAL SIGNIFICANCE: Current commercial dental composites are affected by the oral environment, which might contribute to the long-term performance of these materials.

Development of an oxirane/acrylate interpenetrating polymer network (IPN) resin system.

OBJECTIVE: Develop a hydrophobic, degradation-resistant dental restorative based on an Oxirane-Acrylate IPN System (OASys) with low shrinkage-stress to substantially extend clinical lifetime. METHODS: Unfilled OASys blends were prepared using dipenta-erythritol-hexaacrylate (DPHA) and p-cycloaliphatic-diepoxide (EP5000). Varying proportions of camphorquinone/iodonium photoinitiator, with a co-reactant oligomeric-diol, served as the experimental curing system. The effects of oxirane-acrylate ratio on the degree-of-cure (Durometer-D hardness), hydrophobicity (contact angle), mechanical properties (3-point bending), near-infrared FTIR degree-of-conversion (DoC), polymerization shrinkage, and shrinkage stress were determined. 70:30 BisGMA:TEGDMA resin served as control. RESULTS: Oxirane tended to decrease hardness and increase hydrophobicity. 0:100, 25:75, 50:50 EP5000:DPHA are harder after 24h than control. 75:25 and 100:0 EP5000:DPHA increased in hardness over 24h, but were softer than control. All groups increased in contact angle over 24h. After 24h, 50:50, 75:25 and 0:100 EP5000:DPHA were more hydrophobic (∼75-84°) than the control (∼65°). Acrylate DoC was ∼60% across all experimental groups. Initial oxirane conversion varied from ∼42% in 100:0 EP5000:DPHA to ∼82% 75:25 EP5000:DPHA. However, oxirane DoC increased for 100:0 EP5000:DPHA to ∼73° over 24h, demonstrating dark cure. Moduli and ultimate transverse strengths of OASys groups were higher than for 0:100 EP5000:DPHA, with 50:50 EP5000:DPHA having higher modulus than other experimental groups. However, the control had higher modulus and UTS than all experimental groups. Volumetric shrinkage averaged 7% for experimental groups, but stress decreased dramatically with increasing oxirane content. SIGNIFICANCE: Hydrophobic, low shrinkage-stress OASys resins are promising for development of composites that improve longevity and reduce the cost of dental care.

Reactions: Antibacterial and bioactive dental restorative materials: Do they really work?

PURPOSE: The study and development of antibacterial materials for use in dental applications is growing with the development of novel materials and procedures. Examination of the effects of such antibacterial materials on oral pathogens as well as on stability and longevity of dental restorations is of paramount importance to the field. RESULTS: This review addressed the range of topics covered by the manuscripts presented at the Seoul symposium on antibacterial dental materials. CLINICAL SIGNIFICANCE: Based on the presented works, it seems that the emerging antibacterial and bioactive mate-rials can potentially benefit restorative dentistry; however, like many other subjects in clinical dentistry, good quality evidence on their effectiveness under clinical situations is yet to be accumulated.

A novel antimicrobial orthodontic band cement with in situ-generated silver nanoparticles.

OBJECTIVE: To develop an antimicrobial orthodontic band cement for the prevention of white spot lesions using a novel process that generates silver nanoparticles (AgNP) in situ. MATERIALS AND METHODS: Twenty-seven groups of AgNP-loaded Opal Band Cement (OBC) and two control groups were formulated with varying concentrations of additional benzoyl peroxide (0.5, 1.0, 1.5, or 2.0 wt%) and 2,2-(p-Tolylimino) diethanol (0.5 or 1 wt%). Rockwell15T hardness and near-infrared FTIR were used to assess degree of cure, three-point bending was used to determine modulus and ultimate transverse strength (UTS), and Ag(+) ion release was measured for up to 4 months in vitro using atomic absorption spectroscopy. Antimicrobial activity against Streptococcus mutans and Lactobacillus acidophilus was tested in vitro by counting colony-forming units for up to 28 days. Biocompatibility was evaluated following ISO specifications 7405 (2008), 10993-3 (2003), 10993-5 (2009), and 10993-10 (2010). RESULTS: Most of the experimental groups had hardness, modulus, and UTS values similar to those of the control group. Ag(+) ion release was observed for all AgNP-loaded groups for up to 4 months. Increase in Ag loading increased Ag(+) ion release and in vitro antimicrobial effect. The biocompatibility of the optimal AgNP-loaded OBC was comparable to that of negative controls. CONCLUSION: A novel antimicrobial orthodontic band cement was developed that has comparable mechanical properties to controls, controlled and sustained Ag(+) ion release, significant bacterial inhibition in vitro, and excellent biocompatibility.

Development of a low-color, color stable, dual cure dental resin.

UNLABELLED: Dual-cure (DC) resins are mainly used as cements due to high initial color (generally yellow) and large color shift (ΔE*) after polymerization as compared to light-cured resins. However, even as cements, this color shift is clinically unacceptable, especially when used to cement thin veneers. OBJECTIVE: To develop a novel DC initiator system with both lower initial color (less yellow, i.e., whiter) and smaller ΔE*. METHODS: The effect of using an allyl thiourea (T)/cumene hydroperoxide (CH) self-cure (SC) initiator system in combination with a photo-co-initiator, p-octyloxy-phenyl-phenyl iodonium hexafluoroantimonate (OPPI), in a commercial DC resin cement (PermaFlo DC, Ultradent Products, Inc.) was investigated. Initial color and ΔE* were assessed for 6 weeks in vitro under accelerated aging conditions (75°C water bath). Rockwell15T hardness was used to assess degree of cure (DoC) and the three-point bending test was used to assess mechanical properties. RESULTS: PermaFlo DC (control) was significantly harder than all experimental groups without OPPI but had up to three times higher initial color and four times greater color shift (ΔE*=27 vs. 8). With OPPI, hardness in the experimental groups increased significantly and several were comparable to the controls. Initial color and ΔE* increased slightly (ΔE*=9), but was still 3 times less than that of PermaFlo DC. DC samples containing OPPI had comparable modulus and ultimate transverse strengths to those of the controls. CONCLUSIONS: DC resins that use the T/CH initiator system are weaker but have extremely low color and ΔE*. The addition of OPPI increases DoC and mechanical properties to clinically acceptable levels and maintains extremely low color and ΔE*. SIGNIFICANCE: With this novel initiator system, DC resins potentially can now have comparable color and color stability to light-cure resins and be used in broader esthetic dental applications to improve color stability and reduce shrinkage stress in restorative composites.

Degradable segmented polyurethane elastomers for bone tissue engineering: effect of polycaprolactone content.

Segmented polyurethanes (PURs), consisting of degradable poly(a-hydroxy ester) soft segments and aminoacid-derived chain extenders, are biocompatible elastomers with tunable mechanical and degradative properties suitable for a variety of tissue-engineering applications. In this study, a family of linear PURs synthesized from poly(ϵ-caprolactone) (PCL) diol, 1,4-diisocyanobutane and tyramine with theoretical PCL contents of 65-80 wt% were processed into porous foam scaffolds and evaluated for their ability to support osteoblastic differentiation in vitro. Differential scanning calorimetry and mechanical testing of the foams indicated increasing polymer crystallinity and compressive modulus with increasing PCL content. Next, bone marrow stromal cells (BMSCs) were seeded into PUR scaffolds, as well as poly(lactic-co-glycolic acid) (PLGA) scaffolds, and maintained under osteogenic conditions for 14 and 21 days. Analysis of cell number indicated a systematic decrease in cell density with increasing PUR stiffness at both 14 and 21 days in culture. However, at these same time points the relative mRNA expression for the bone-specific proteins osteocalcin and the growth factors bone morphogenetic protein-2 and vascular endothelial growth factor gene expression were similar among the PURs. Finally, prostaglandin E2 production, alkaline phosphatase activity and osteopontin mRNA expression were highly elevated on the most-crystalline PUR scaffold as compared to the PLGA and PUR scaffolds. These results suggest that both the modulus and crystallinity of the PUR scaffolds influence cell proliferation and the expression of osteoblastic proteins.

Effects of the analgesic acetaminophen (Paracetamol) and its para-aminophenol metabolite on viability of mouse-cultured cortical neurons.

Acetaminophen has been used as an analgesic for more than a hundred years, but its mechanism of action has remained elusive. Recently, it has been shown that acetaminophen produces analgesia by the activation of the brain endocannabinoid receptor CB1 through its para-aminophenol (p-aminophenol) metabolite. The objective of this study was to determine whether p-aminophenol could be toxic for in vitro developing mouse cortical neurons as a first step in establishing a link between acetaminophen use and neuronal apoptosis. We exposed developing mouse cortical neurons to various concentrations of drugs for 24 hr in vitro. Acetaminophen itself was not toxic to developing mouse cortical neurons at therapeutic concentrations of 10-250 µg/ml. However, concentrations of p-aminophenol from 1 to 100 µg/ml produced significant (p < 0.05) loss of mouse cortical neuron viability at 24 hr compared to the controls. The naturally occurring endocannabinoid anandamide also caused similar 24-hr loss of cell viability in developing mouse cortical neurons at concentrations from 1 to 100 µg/ml, which indicates the mechanism of cell death could be through the cannabinoid receptors. The results of our experiments have shown a detrimental effect of the acetaminophen metabolite p-aminophenol on in vitro developing cortical neuron viability which could act through CB1 receptors of the endocannabinoid system. These results could be especially important in recommending an analgesic for children or individuals with traumatic brain injury who have developing cortical neurons.

Antimicrobial acrylic materials with in situ generated silver nanoparticles.

UNLABELLED: Polymethyl methacrylate (PMMA) is widely used to treat traumatic head injuries (cranioplasty) and orthopedic injuries (bone cement), but there is a problem with implant-centered infections. With organisms such as Acinetobacter baumannii and methicillin-resistant staphylococcus aureus developing resistance to antibiotics, there is a need for novel antimicrobial delivery mechanisms without risk of developing resistant organisms. OBJECTIVES: To develop a novel antimicrobial implant material by generating silver nanoparticles (AgNP) in situ in PMMA. RESULTS: All PMMA samples with AgNP's (AgNP-PMMA) released Ag(+) ions in vitro for over 28 days. In vitro antimicrobial assays revealed that these samples (even samples with the slowest release rate) inhibited 99.9% of bacteria against four different strains of bacteria. Long-term antimicrobial assay showed a continued antibacterial effect past 28 days. Some AgNP-loaded PMMA groups had comparable Durometer-D hardness (a measure of degree of cure) and modulus to control PMMA, but all experimental groups had slightly lower ultimate transverse strengths. CONCLUSIONS: AgNP-PMMA demonstrated a tremendously broad-spectrum and long-intermediate-term antimicrobial effect with comparable mechanical properties to control PMMA. Current efforts are focused on further improving mechanical properties by reducing AgNP loading and assessing fatigue properties.

Development of an antimicrobial resin--a pilot study.

OBJECTIVES: To demonstrate that silver nanoparticles (AgNPs) could be synthesized in situ in acrylic dental resins. METHODS: Light-cure (LC; bisphenol A glycidyl methacrylate, tetraethyleneglycol dimethacrylate, bisphenol A ethoxylate dimethacrylate blend) and chemical-cure systems (CC; orthodontic denture resin) were used to synthesize AgNPs using different concentrations of Ag benzoate (AgBz). RESULTS: Rockwell hardness for LC resins showed that resins could be cured with up to 0.15% AgBz, while the hardness of CC resins were unaffected in the concentrations tested. UV-Vis spectroscopy and transmission electron microscopy confirmed the presence of AgNPs in both LC and CC resins. Generally, CC resins had better distribution of and much smaller AgNPs as compared to LC resins overall. In some samples, especially in LC resins, nanoclusters were visible. An in vitro release study over four-weeks showed that CC resins released the most Ag(+) ions, with release detected in all samples. However, LC resins only released Ag(+) ions when AgBz concentration was greater than 0.1% (w/w). AgNP-loaded CC resins made with 0.2 and 0.5% (w/w) AgBz were tested for antibacterial activity in vitro against Streptococcus mutans, and results showed 52.4% and a 97.5% bacterial inhibition, respectively. Further work is now warranted to test mechanical properties and to optimize the initiator system to produce commercially useful dental and medical resins. SIGNIFICANCE: Success in this work could lead to a series of antimicrobial medical and dental biomaterials that can prevent secondary caries and infection of implants.

A novel osteotropic biomaterial OG-PLG: Synthesis and in vitro release.

Statins (e.g., simvastatin) have shown to induce expression of the bone morphogenic protein-2 gene in bone cells, but they are not used clinically because of a lack of a suitable delivery device. The overall objective is to develop optimized statin delivery devices for bone regeneration. The specific objective was to determine the effect of grafting statins to biodegradable poly[lactide-co-glycolide] (PLG) on release kinetics. Simvastatin was grafted to PLG (OG-PLG) and characterized using contact-angle measurements, attenuated total reflectance-Fourier transform infrared, and ultraviolet-visible spectroscopy to determine success of the synthesis. An ultraviolet-visible assay for measuring release of statins and degraded OG-PLG in media was also developed. In vitro release studies using films and scaffolds made with PLG, PLG blended with simvastatin (PLG + Sim), and OG-PLG (simvastatin grafted to PLG) blended into PLG at different concentrations showed that release rate of OG-PLG from films was significantly greater than that of PLG + Sim. However, release rate from scaffolds showed PLG + Sim to be significantly higher than that of OG-PLG. The diffusion-controlled release kinetics of simvastatin from PLG + Sim seems to be more heavily affected by device morphology, whereas the degradation-controlled release kinetics seem to be less affected. In short, release kinetics can be modulated by grafting statins to PLG.

A novel osteotropic biomaterial OG-PLG: in vitro efficacy.

Previously, a novel osteotropic biomaterial, OG-PLG [simvastatin grafted to poly(lactide-co-glycolide), PLG], was synthesized and shown to have degradation-controlled release kinetics. The objective here was to determine the effect of grafting statins to PLG on bone regeneration in vitro. Rat bone marrow cells were stimulated in vitro with simvastatin dissolved in media, saponified simvastatin dissolved in media, simvastatin released through diffusion from emulsion freeze-dried scaffolds, and OG-PLG. Unstimulated cultures and cultures stimulated with dexamethasone were used as negative and positive controls, respectively. In vitro bone formation was assessed using the alkaline phosphatase (ALP) and von Kossa assays at different times up to 16 days. ALP analysis revealed that saponified simvastatin at 10(-7)M and OG-PLG significantly increased ALP expression at various time points. von Kossa assay showed that simvastatin, saponified simvastatin, and OG-PLG significantly enhanced mineralization, with the effect from OG-PLG being the most significant. In short, OG-PLG significantly enhanced in vitro bone cell mineralization beyond the effect of simvastatin or saponified simvastatin dissolved in media and simvastatin released via diffusion from scaffolds.