Folic Acid-Doxorubicin-Double-Functionalized-Lipid-Core Nanocapsules: Synthesis, Chemical Structure Elucidation, and Cytotoxicity Evaluation on Ovarian (OVCAR-3) and Bladder (T24) Cancer Cell Lines.

PURPOSE: Folic acid-doxorubicin-double-functionalized-lipid-core nanocapsules (LNC-CS-L-Zn+2-DOX-FA) were prepared, characterized, and evaluated in vitro against ovarian and bladder cancer cell lines (OVCAR-3 and T24). METHODS: LNC-CS-L-Zn+2-DOX-FA was prepared by self-assembly and interfacial reactions, and characterized using liquid chromatography, particle sizing, transmission electron microscopy, and infrared spectroscopy. Cell viability and cellular uptake were studied using MTT assay and confocal microscopy. RESULTS: The presence of lecithin allows the formation of nanocapsules with a lower tendency of agglomeration, narrower size distributions, and smaller diameters due to an increase in hydrogen bonds at the surface. LNC-L-CS-Zn+2-DOX-FA, containing 98.00 ± 2.34 µg mL-1 of DOX and 105.00 ± 2.05 µg mL-1 of FA, had a mean diameter of 123 ± 4 nm and zeta potential of +12.0 ± 1.3 mV. After treatment with LNC-L-CS-Zn+2-DOX-FA (15 µmol L-1 of DOX), T24 cells had inhibition rates above 80% (24 h) and 90% (48 h), whereas OVCAR-3 cells showed inhibition rates of 68% (24 h) and 93% (48 h), showing higher cytotoxicity than DOX.HCl. The fluorescent-labeled formulation showed a higher capacity of internalization in OVCAR-3 compared to T24 cancer cells. CONCLUSION: Lecithin favored the increase of hydrogen bonds at the surface, leading to a lower tendency of agglomeration for nanocapsules. LNC-CS-L-Zn+2-DOX-FA is a promising therapeutic agent against tumor-overexpressing folate receptors.

Effect on adhesion of a nanocapsules-loaded adhesive system.

This study aimed to evaluate the in situ degree of conversion, contact angle, and immediate and long-term bond strengths of a commercial primer and an experimental adhesive containing indomethacin- and triclosan-loaded nanocapsules (NCs). The indomethacin- and triclosan-loaded NCs, which promote anti-inflammatory and antibacterial effects through controlled release, were incorporated into the primer at a concentration of 2% and in the adhesive at concentrations of 1, 2, 5, and 10%. The in situ degree of conversion (DC, n=3) was evaluated by micro-Raman spectroscopy. The contact angle of the primer and adhesive on the dentin surface (n = 3) was determined by an optical tensiometer. For the microtensile bond strength µTBS test (12 teeth per group), stick-shaped specimens were tested under tensile stress immediately after preparation and after storage in water for 1 year. The data were analyzed using two-way ANOVA, three-way ANOVA and Tukey's post hoc tests with α=0.05. The use of the NC-loaded adhesive resulted in a higher in situ degree of conversion. The DC values varied from 75.07 ± 8.83% to 96.18 ± 0.87%. The use of NCs in only the adhesive up to a concentration of 5% had no influence on the bond strength. The contact angle of the primer remained the same with and without NCs. The use of both the primer and adhesive with NCs (for all concentrations) resulted in a higher contact angle of the adhesive. The longitudinal µTBS was inversely proportional to the concentration of NCs in the adhesive system, exhibiting decreasing values for the groups with primer containing NCs and adhesives with increasing concentrations of NCs. Adhesives containing up to 5% of nanocapsules and primer with no NCs maintained the in situ degree of conversion, contact angle, and immediate and long-term bond strengths. Therefore, the NC-loaded adhesive can be an alternative method for combining the bond performance and therapeutic effects. The use of an adhesive with up to 5% nanocapsules containing indomethacin and triclosan and a primer with no nanocapsules maintained the long-term bond performance.

Effect on adhesion of a nanocapsules-loaded adhesive system

Abstract This study aimed to evaluate the in situ degree of conversion, contact angle, and immediate and long-term bond strengths of a commercial primer and an experimental adhesive containing indomethacin- and triclosan-loaded nanocapsules (NCs). The indomethacin- and triclosan-loaded NCs, which promote anti-inflammatory and antibacterial effects through controlled release, were incorporated into the primer at a concentration of 2% and in the adhesive at concentrations of 1, 2, 5, and 10%. The in situ degree of conversion (DC, n=3) was evaluated by micro-Raman spectroscopy. The contact angle of the primer and adhesive on the dentin surface (n = 3) was determined by an optical tensiometer. For the microtensile bond strength µTBS test (12 teeth per group), stick-shaped specimens were tested under tensile stress immediately after preparation and after storage in water for 1 year. The data were analyzed using two-way ANOVA, three-way ANOVA and Tukey's post hoc tests with α=0.05. The use of the NC-loaded adhesive resulted in a higher in situ degree of conversion. The DC values varied from 75.07 ± 8.83% to 96.18 ± 0.87%. The use of NCs in only the adhesive up to a concentration of 5% had no influence on the bond strength. The contact angle of the primer remained the same with and without NCs. The use of both the primer and adhesive with NCs (for all concentrations) resulted in a higher contact angle of the adhesive. The longitudinal μTBS was inversely proportional to the concentration of NCs in the adhesive system, exhibiting decreasing values for the groups with primer containing NCs and adhesives with increasing concentrations of NCs. Adhesives containing up to 5% of nanocapsules and primer with no NCs maintained the in situ degree of conversion, contact angle, and immediate and long-term bond strengths. Therefore, the NC-loaded adhesive can be an alternative method for combining the bond performance and therapeutic effects. The use of an adhesive with up to 5% nanocapsules containing indomethacin and triclosan and a primer with no nanocapsules maintained the long-term bond performance.

Antimicrobial effect and physicochemical properties of an adhesive system containing nanocapsules.

OBJECTIVE: To incorporate indomethacin and triclosan-loaded nanocapsules into primer and adhesive, and evaluate its properties. METHODS: Indomethacin and triclosan were encapsulated by deposition of preformed polymer and subsequently characterized regarding morphology, particle size, drug content and cytotoxicity. Nanocapsules (NCs) were incorporated into primer at 2% and into adhesive at 1, 2, 5, and 10% concentrations. Degree of conversion (DC) and softening in ethanol of the adhesive were evaluated. Drug release and drug diffusion through dentin was quantified by high performance liquid chromatography. Antimicrobial test was performed until 96h. RESULTS: Spherical and biocompatible NCs presented mean size of 159nm. Drugs content was 3mg indomethacin/g powder and 2mg triclosan/g powder. Incorporating NCs in adhesive showed no influence in DC (p=0.335). The addition of 2% of NCs showed no influence in softening in ethanol (p>0.05). After 120h, 93% of indomethacin and 80% of triclosan were released from primer, 20% of indomethacin and 17% of triclosan were released from adhesive with 10% of NCs. Indomethacin showed diffusion through dentin. In 24h, adhesive containing 2 and 5% of NCs using primer with NCs showed antimicrobial effect. In 96h, adhesives containing different concentration of NCs promoted antimicrobial effect. CONCLUSIONS: Indomethacin and triclosan-loaded nanocapsules were successfully incorporated into primer and adhesive, promoting controlled drugs release, indomethacin diffusion through dentin and antimicrobial effect without compromising its physicochemical properties. SIGNIFICANCE: Indomethacin and triclosan-loaded nanocapsules have potential to prevent recurrent caries and to be used in deep cavities controlling pulpar inflammatory process.

Effect of indomethacin-loaded nanocapsules incorporation in a dentin adhesive resin.

OBJECTIVES: The aim of this study was to produce indomethacin-loaded nanocapsules (IndOH-NCs) and evaluate the influence of their incorporation into an adhesive resin. MATERIALS AND METHODS: Indomethacin was encapsulated by the deposition of preformed polymer. IndOH-NCs were characterized by laser diffractometry, Fourier transformed infrared spectrometry, transmission electron microscopy (TEM), scanning electron microscopy, high-performance liquid chromatography (HPLC), and MTT assay. Nanocapsules (NCs) were incorporated into an adhesive in concentrations of 1, 2, 5, and 10 %. The addition was visualized by TEM and drug release was evaluated by HPLC until 120 h of immersion in simulated body fluid (SBF). Drug diffusion through dentin was tested using a Franz diffusion cell apparatus and quantified by HPLC. The degree of conversion (DC), softening in ethanol, and microtensile bond strength (µTBS) were evaluated to determine whether the nanocapsules influenced the adhesive. Data were analyzed using one-way ANOVA and Tukey's post hoc test for DC, softening in ethanol, µTBS, and cytotoxicity, and paired t test for comparison between the initial and final Knoop microhardness. RESULTS: IndOH-NCs, with a spherical shape and a mean diameter of 165 nm, were incorporated into an adhesive. Indomethacin content was 7 mg drug/g powder. IndOH-NCs maintained high cell viability. At 120 h, an amount of 13.83 % of indomethacin was released, and after 7 days, 7.07 % of this drug was diffused through dentin for an adhesive containing 10 % of nanocapsules. No alteration in the DC, softening in ethanol, and µTBS resulted from NC addition. CONCLUSIONS: IndOH-NCs may be incorporated into adhesive systems, without compromising properties, to add an anti-inflammatory drug controlled release for restorative procedures in deep cavities. CLINICAL SIGNIFICANCE: Here is the first step toward the goal of providing agents to act at an inflammatory process of pulp tissue through dental adhesives via encapsulation of drug.

Effects of Two Types of Melatonin-Loaded Nanocapsules with Distinct Supramolecular Structures: Polymeric (NC) and Lipid-Core Nanocapsules (LNC) on Bovine Embryo Culture Model.

Melatonin has been used as a supplement in culture medium to improve the efficiency of in vitro produced mammalian embryos. Through its ability to scavenge toxic oxygen derivatives and regulate cellular mRNA levels for antioxidant enzymes, this molecule has been shown to play a protective role against damage by free radicals, to which in vitro cultured embryos are exposed during early development. In vivo and in vitro studies have been performed showing that the use of nanocapsules as active substances carriers increases stability, bioavailability and biodistribution of drugs, such as melatonin, to the cells and tissues, improving their antioxidant properties. These properties can be modulated through the manipulation of formula composition, especially in relation to the supramolecular structures of the nanocapsule core and the surface area that greatly influences drug release mechanisms in biological environments. This study aimed to evaluate the effects of two types of melatonin-loaded nanocapsules with distinct supramolecular structures, polymeric (NC) and lipid-core (LNC) nanocapsules, on in vitro cultured bovine embryos. Embryonic development, apoptosis, reactive oxygen species (ROS) production, and mRNA levels of genes involved in cell apoptosis, ROS and cell pluripotency were evaluated after supplementation of culture medium with non-encapsulated melatonin (Mel), melatonin-loaded polymeric nanocapsules (Mel-NC) and melatonin-loaded lipid-core nanocapsules (Mel-LNC) at 10-6, 10-9, and 10-12 M drug concentrations. The highest hatching rate was observed in embryos treated with 10-9 M Mel-LNC. When compared to Mel and Mel-NC treatments at the same concentration (10-9 M), Mel-LNC increased embryo cell number, decreased cell apoptosis and ROS levels, down-regulated mRNA levels of BAX, CASP3, and SHC1 genes, and up-regulated mRNA levels of CAT and SOD2 genes. These findings indicate that nanoencapsulation with LNC increases the protective effects of melatonin against oxidative stress and cell apoptosis during in vitro embryo culture in bovine species.

Melatonin delivery by nanocapsules during in vitro bovine oocyte maturation decreased the reactive oxygen species of oocytes and embryos.

In this work, a promising approach to increase the advantageous properties of melatonin through its encapsulation into lipid-core nanocapsules (LNC) was examined. Oocytes were treated during in vitro maturation with non-encapsulated melatonin (Mel), melatonin-loaded lipid-core nanocapsules (Mel-LNC), and unloaded LNC. Cytotoxicity, meiotic maturation rate, development to the blastocyst stage, reactive oxygen species (ROS) and glutathione levels, mean cell number and apoptotic cell/blastocyst, and mRNA quantification were evaluated. Both Mel and Mel-LNC enhanced in vitro embryo production, however, Mel-LNC proved to be more effective at decreasing ROS levels and the apoptotic cell number/blastocyst, increasing the cleavage and blastocyst rates, up-regulating the GPX1 and SOD2 genes, and down-regulating the CASP3 and BAX genes. Mel-LNC could penetrate into oocytes and remain inside the cells until they reach the blastocyst stage. In conclusion, when melatonin was encapsulated in LNC and applied during in vitro oocyte maturation, some quality aspects of the blastocysts were improved.

Development of Novel Chitosan Microcapsules for Pulmonary Delivery of Dapsone: Characterization, Aerosol Performance, and In Vivo Toxicity Evaluation.

Pneumocystis carinii pneumonia (PCP) is a major opportunistic infection that affects patients with human immunodeficiency virus. Although orally administered dapsone leads to high hepatic metabolism, decreasing the therapeutic index and causing severe side effects, this drug is an effective alternative for the treatment of PCP. In this context, microencapsulation for pulmonary administration can offer an alternative to increase the bioavailability of dapsone, reducing its adverse effects. The aim of this work was to develop novel dapsone-loaded chitosan microcapsules intended for deep-lung aerosolized drug delivery. The geometric particle size (D 4,3) was approximately 7 µm, the calculated aerodynamic diameter (d aero) was approximately 4.5 µm, and the mass median aerodynamic diameter from an Andersen cascade impactor was 4.7 µm. The in vitro dissolution profile showed an efficient dapsone encapsulation, demonstrating the sustained release of the drug. The in vitro deposition (measured by the Andersen cascade impactor) showed an adequate distribution and a high fine particles fraction (FPF = 50%). Scanning electron microscopy of the pulmonary tissues demonstrated an adequate deposition of these particles in the deepest part of the lung. An in vivo toxicity experiment showed the low toxicity of the drug-loaded microcapsules, indicating a protective effect of the microencapsulation process when the particles are microencapsulated. In conclusion, the pulmonary administration of the novel dapsone-loaded microcapsules could be a promising alternative for PCP treatment.