Progress in Natural Compounds/siRNA Co-delivery Employing Nanovehicles for Cancer Therapy.

Chemotherapy using natural compounds, such as resveratrol, curcumin, paclitaxel, docetaxel, etoposide, doxorubicin, and camptothecin, is of importance in cancer therapy because of the outstanding therapeutic activity and multitargeting capability of these compounds. However, poor solubility and bioavailability of natural compounds have limited their efficacy in cancer therapy. To circumvent this hurdle, nanocarriers have been designed to improve the antitumor activity of the aforementioned compounds. Nevertheless, cancer treatment is still a challenge, demanding novel strategies. It is well-known that a combination of natural products and gene therapy is advantageous over monotherapy. Delivery of multiple therapeutic agents/small interfering RNA (siRNA) as a potent gene-editing tool in cancer therapy can maximize the synergistic effects against tumor cells. In the present review, co-delivery of natural compounds/siRNA using nanovehicles are highlighted to provide a backdrop for future research.

Quaternary ammonium silane, calcium and phosphorus-loaded PLGA submicron particles against Enterococcus faecalis infection of teeth: An in vitro and in vivo study.

Refractory root canal infection of human teeth is the primary cause of dental treatment failure. Enterococcus faecalis is the major cause of refractory root canal infection. In the present study, poly(D,L-lactic-co-glycolide) (PLGA) submicron particles were used as carriers to deliver an antimicrobial quaternary ammonium silane (code-named K21) as well as calcium and phosphorus elements. The release profiles, antibacterial ability against E. faecalis, extent of infiltration into dentinal tubules, biocompatibility and in vitro mineralization potential of the particles were investigated. In addition, the antimicrobial effects of the particles against E. faecalis infection were evaluated in vivo in the teeth of beagle dogs. The encapsulated components were released from the PLGA particles in a sustained-release manner. The particles also displayed good biocompatibility, in vitro mineralization ability and antibacterial activity against E. faecalis. The particles could be driven into dentinal tubules of dentin slices by ultrasonic activation and inhibited E. faecalis colonization. In the root canals of beagle dogs, PLGA submicron particles loaded with K21, calcium and phosphorus demonstrated strong preventive effects against E. faecalis infection. The system may be developed into a new intracanal disinfectant for root canal treatment.

Comparison of the use of d-enantiomeric and l-enantiomeric antimicrobial peptides incorporated in a calcium-chelating irrigant against Enterococcus faecalis root canal wall biofilms.

OBJECTIVES: To compare the anti-biofilm efficacy of two antimicrobial peptides (AMPs), 1018 and DJK-5, in disrupting canal wall biofilms in the isthmus, canal and dentinal tubules of single-rooted maxillary premolars. METHODS: Enterococcus faecalis single-species biofilms were formed in-situ in the root canal system of the premolars (n = 91). Confocal laser scanning microscopy, bacterial sampling, colony-forming unit counting, XTT assay, lactate dehydrogenase assay and phenol-sulphuric acid method were used to identify the anti-biofilm efficacy of both AMPs and their influence on bacterial metabolic activity. RESULTS: Both AMPs disrupted in-situ E. faecalis biofilms and altered their metabolic activity. At 20 µg/mL, the d-enantiomeric AMP DJK-5 killed 55.5 %, 57.3 % and 55.8 % of biofilm bacteria in the isthmus, canal and dentinal tubules, respectively, in 1 min. In contrast, the l-enantiomeric AMP 1018 only eradicated 25.6 %, 25.5 % and 27.5 % of biofilm bacteria in the isthmus, canal and dentinal tubules, respectively, within the same time. Anti-biofilm efficacy of the root canal irrigants tested were in the order: 6 % NaOCl > 20 µg/mL DJK-5 > 10 µg/mL DJK-5 > 20 µg/mL 1018 > 10 µg/mL 1018 > 0.9 % NaCl. CONCLUSIONS: The present results are confirmatory of previous studies, in that d-enantiomeric AMPs exhibit more potent antibacterial properties than l-enantiomeric AMPs against E. faecalis biofilms within the canal space. Nevertheless, the potency of both AMPs are concentration-dependent. Incorporation of these agents into EDTA, a non-antibacterial calcium-chelating irrigant for removal of the inorganic component of the canal space debris, does not reduce the efficacy of either AMP. CLINICAL SIGNIFICANCE: The present study provides the proof of concept that incorporation of an antimicrobial peptide into a calcium-chelating root canal irrigant enhances the disinfection of intratubular single-species biofilms during smear layer and smear plug removal.

Hollow mesoporous zirconia delivery system for biomineralization precursors.

Strategies based on the combination of nanocarrier delivery systems and scaffolds provide bone tissue engineering scaffolds with multifunctional capability. Zirconia, a biocompatible ceramic commonly used in orthopedic and dental implants, was used to synthesize hollow mesoporous nanocapsules for loading, storage and sustained release of a novel polyamine-stabilized liquid precursor phase of amorphous calcium phosphate (PAH-ACP) for collagen biomineralization and bone marrow stromal cells osteoinduction. Hollow mesoporous zirconia (hmZrO2) nanocapsules loaded with biomimetic precursors exhibited pH-sensitive release capability and good biocompatibility. The PAH-ACP released from loaded hmZrO2 still retained the ability to infiltrate and mineralize collagen fibrils as well as exhibited osteoinductivity. A collagen scaffold blended with PAH-ACP@hmZrO2 supplement and stem cells may be a promising tool for bone tissue engineering. STATEMENT OF SIGNIFICANCE: The advent of nanotechnology has catalyzed the development of bone tissue engineering strategies based on the combination of nanocarrier delivery systems and scaffolds, which provide distinct advantages, including the possibilities of sustained release and protection of the bioactive agents, site-specific pharmacological effects and reduction of side effects. Herein, hollow mesoporous zirconia (hmZrO2) nanocapsules with pH-sensitive capacity were synthesized for loading, storage and sustained release of a novel polyamine-stabilized liquid precursor phase of ACP (PAH-ACP). The loaded nanocapsules show good biocompatibility and demonstrate bioactivities for collagen biomineralization and bone marrow stromal cells osteoinduction. Our results may offer a promising tool for designing bone tissue engineering "cocktail therapy" involving seeding scaffolds with biomineralization precursors loaded hmZrO2 supplement and stem cells.

Biodegradable mesoporous delivery system for biomineralization precursors.

Scaffold supplements such as nanoparticles, components of the extracellular matrix, or growth factors have been incorporated in conventional scaffold materials to produce smart scaffolds for tissue engineering of damaged hard tissues. Due to increasing concerns on the clinical side effects of using large doses of recombinant bone-morphogenetic protein-2 in bone surgery, it is desirable to develop an alternative nanoscale scaffold supplement that is not only osteoinductive, but is also multifunctional in that it can perform other significant bone regenerative roles apart from stimulation of osteogenic differentiation. Because both amorphous calcium phosphate (ACP) and silica are osteoinductive, a biodegradable, nonfunctionalized, expanded-pore mesoporous silica nanoparticle carrier was developed for loading, storage, and sustained release of a novel, biosilicification-inspired, polyamine-stabilized liquid precursor phase of ACP for collagen biomineralization and for release of orthosilicic acid, both of which are conducive to bone growth. Positively charged poly(allylamine)-stabilized ACP (PAH-ACP) could be effectively loaded and released from nonfunctionalized expanded-pore mesoporous silica nanoparticles (pMSN). The PAH-ACP released from loaded pMSN still retained its ability to infiltrate and mineralize collagen fibrils. Complete degradation of pMSN occurred following unloading of their PAH-ACP cargo. Because PAH-ACP loaded pMSN possesses relatively low cytotoxicity to human bone marrow-derived mesenchymal stem cells, these nanoparticles may be blended with any osteoconductive scaffold with macro- and microporosities as a versatile scaffold supplement to enhance bone regeneration.

Collagen intrafibrillar mineralization as a result of the balance between osmotic equilibrium and electroneutrality.

Mineralization of fibrillar collagen with biomimetic process-directing agents has enabled scientists to gain insight into the potential mechanisms involved in intrafibrillar mineralization. Here, by using polycation- and polyanion-directed intrafibrillar mineralization, we challenge the popular paradigm that electrostatic attraction is solely responsible for polyelectrolyte-directed intrafibrillar mineralization. As there is no difference when a polycationic or a polyanionic electrolyte is used to direct collagen mineralization, we argue that additional types of long-range non-electrostatic interaction are responsible for intrafibrillar mineralization. Molecular dynamics simulations of collagen structures in the presence of extrafibrillar polyelectrolytes show that the outward movement of ions and intrafibrillar water through the collagen surface occurs irrespective of the charges of polyelectrolytes, resulting in the experimentally verifiable contraction of the collagen structures. The need to balance electroneutrality and osmotic equilibrium simultaneously to establish Gibbs-Donnan equilibrium in a polyelectrolyte-directed mineralization system establishes a new model for collagen intrafibrillar mineralization that supplements existing collagen mineralization mechanisms.

Effects of different sonic activation protocols on debridement efficacy in teeth with single-rooted canals.

OBJECTIVES: The effects of different EndoActivator® (EA) sonic activation protocols on root canal debridement efficacy were examined. METHODS: Root canals in 48 single-rooted teeth were instrumented, irrigated initially with NaOCl and divided into 6 groups (N=8) based on the application time of QMix (antimicrobial calcium-chelating irrigant), and the time and sequence of EA irrigant activation - Positive Control: 90s QMix; Negative Control: 90s saline; Group 1A: 15s QMix+15s QMix with EA-activation; Group 1B: 30s QMix+30s of QMix with EA-activation; Group 2A: 15s QMix with EA-activation+15s QMix; Group 2B: 30s QMix with EA-activation+30s QMix. Split roots were examined with scanning electron microscopy for assignment of smear and debris scores in locations along the coronal, middle and apical thirds of the canals. The overall cleanliness of pooled canal locations in the Positive Control and the 4 experimental groups were compared with chi-square tests. RESULTS: Significant differences were detected among the 5 groups (P<0.001). Post hoc pairwise comparisons indicated that the overall canal cleanliness was in the order (from best to worst): 1B=2B>2A>1A>Positive Control. Completely clean canals could not be achieved due to the absence of continuous irrigant flow for EA to clear intraradicular debris. CONCLUSIONS: Irrespective of the sonic activation sequence, irrigant activation for 30s during a 60-s period of QMix application appears to maximise the smear layer and debris removal potential of the EndoActivator® system. CLINICAL SIGNIFICANCE: Sonic activation of root canal irrigants produces cleaner root canals and reduces the time required for final delivery of a canal wall smear later-removing irrigant when compared to the use of needle irrigation alone.

Effect of ammonium hexafluorosilicate application for arresting caries treatment on demineralized primary tooth enamel.

Ammonium hexafluorosilicate (AHF) has been applied to arrest caries without discoloration. The purpose of this study was to observe structural and elemental changes of demineralized and AHF applied primary tooth enamel. Enamel from the labial surface of 20 primary canines was divided into an unground side and ground side at the center of the tooth, and demineralized with 35% phosphoric acid for 6 min. The teeth were divided into 4 groups according to a 3-min application of AHF and 1 week of soaking in artificial saliva, as follows: group A (neither AHF nor saliva), group B (only saliva), group C (only AHF), and group D (AHF and saliva), and then subdivided according to whether the enamel was ground or unground. Specimens were analyzed with scanning electron microscopy (SEM) and energy dispersive X-ray spectrometry (EDS). The data were statistically analyzed using ANOVA and Fisher's PLSD test at α = 0.05. In groups A and B, prism structures were seen, however, in groups C and D, enamel surfaces were covered with spherical particles. Ca/P ratio was significantly higher in groups C and D than in groups A and B. There was no significant difference between ground and unground enamel in the content of any element. The values for F, Na, Mg and Si persents and Ca/P ratio were significantly higher for the enamel surface than for points 10-30 µm beneath the surface. Results of this study suggest the possibility that AHF treatment arrests caries, although further study will be required to confirm this result.

New trends in dentin bonding: treatment with Chlorhexidine, Hyaluronic acid, vitamin C and green tea / Novas tendências em adesão dentinária: tratamento com clorexidina, ácido hialurônico, vitamina C e chá verde

Objetivo: O objetivo deste estudo foi avaliar a resistência de união dentinária tratada com clorexidina, ácido hialurônico, vitamina C e chá verde. Material e Métodos: As raízes de 50 dentes bovinos foram removidas e a superfície de dentina vestibular exposta. Após o condicionamento ácido, os espécimes foram divididos em 5 grupos (n = 10), de acordo com as estratégias adesivas na dentina: CO - dentina não tratada; CHX – tratamento com clorexidina 2% por 30 s; HA - tratamento com ácido hialurônico 1% por 30 s; VC - tratamento com vitamina C 10% por 30 s; GT – tratamento com extrato de chá verde 1% por 30 s. Sistema adesivo Adper Single Bond II foi aplicado segundo recomendações do fabricante. Os espécimes foram restaurados com uma camada de 4 mm de espessura de resina composta, que foi polimerizada por 40 s. As amostras foram armazenadas em água destilada a 37 ° C durante 24 h, e seccionada em palitos de 1 x 1 mm2 contendo a interface adesiva. Teste de microtração foi realizado com uma máquina de ensaios universal a uma velocidade de 0,5 mm/min. Resultados: Os resultados foram analisados com teste de variância ANOVA um-fator e testes de múltipla comparação de Tukey (p < 0,05). Grupo GT apresentou os maiores valores de resistência de união (29,4 ± 3,1) a, mas sem diferença significativa em relação aos demais grupos experimentais HA (26,7 ± 3,1) ab, CHX (25,4 ± 2,6) ab e VC (22,4 ± 6,0) b. Resistência de união dos grupos experimentais não foi significativamente diferente do grupo CO. Conclusão: a resistência de união imediata foi mantida mesmo após os diferentes tratamentos dentinários.

Objective: The aim of this study was to evaluate the bond strength of dentin treated with chlorhexidine, hyaluronic acid, vitamin C and green tea. Material and Methods: The roots of 50 bovine teeth were removed and buccal coronal dentin was exposed. After acidetching, the specimens were divided into 5 groups (n= 10), according to the dentin treatment strategy: CO -untreated dentin; CHX - treated with 2 wt% chlorhexidine for 30 s; HA - treated with 1 wt% hyaluronic acid for 30s; VC - treated with 10 wt% vitamin C for 30 s; GT - treated a 1% green tea extract for 30 s. Adper Single Bond was then applied to the treated according to the manufacturer’s recommendations. The specimens were restored with a 4 mm thick layer of the resin composite, which was polymerized for 40 s. The specimens were stored in distilled water at 37°C for 24 h and sectioned into 1 x 1 mm2 sticks containing the adhesive interface. Microtensile bond strength testing was performed with a universal testing machine at a cross-head speed of 1.0 mm/min. Results: The results were analyzed with onefactor ANOVA and Tukey’s multiple comparison tests. GT group presented the highest values bond strength (29.4 ± 3.1) a, but no significant difference compared to the other experimental groups HA (26.7 ± 3.1) ab, CHX (25.4 ±2.6) ab and VC (22.4 ± 6.0) b. Bond strengths of experimental groups were not significantly different from the CO. Conclusion: Immediate bond strength was preserved after acid-etched dentin was treated.

A chemical phosphorylation-inspired design for Type I collagen biomimetic remineralization.

OBJECTIVES: Type I collagen alone cannot initiate tissue mineralization. Sodium trimetaphosphate (STMP) is frequently employed as a chemical phosphorylating reagent in the food industry. This study examined the feasibility of using STMP as a functional analog of matrix phosphoproteins for biomimetic remineralization of resin-bonded dentin. METHODS: Equilibrium adsorption and desorption studies of STMP were performed using demineralized dentin powder (DDP). Interaction between STMP and DDP was examined using Fourier transform-infrared spectroscopy. Based on those results, a bio-inspired mineralization scheme was developed for chemical phosphorylation of acid-etched dentin with STMP, followed by infiltration of the STMP-treated collagen matrix with two etch-and-rinse adhesives. Resin-dentin interfaces were remineralized in a Portland cement-simulated body fluid system, with or without the use of polyacrylic acid (PAA) as a dual biomimetic analog. Remineralized resin-dentin interfaces were examined unstained using transmission electron microscopy. RESULTS: Analysis of saturation binding curves revealed the presence of irreversible phosphate group binding sites on the surface of the DDP. FT-IR provided additional evidence of chemical interaction between STMP and DDP, with increased in the peak intensities of the PO and P-O-C stretching modes. Those peaks returned to their original intensities after alkaline phosphatase treatment. Evidence of intrafibrillar apatite formation could be seen in incompletely resin-infiltrated, STMP-phosphorylated collagen matrices only when PAA was present in the SBF. SIGNIFICANCE: These results reinforce the importance of PAA for sequestration of amorphous calcium phosphate nanoprecursors in the biomimetic remineralization scheme. They also highlight the role of STMP as a templating analog of dentin matrix phosphoproteins for inducing intrafibrillar remineralization of apatite nanocrystals within the collagen matrix of incompletely resin-infiltrated dentin.

Hybrid Root SEAL (MetaSEAL) creates hybrid layers in radicular dentin only when EDTA is used as the final rinse.

PURPOSE: To test if the hybrid layer formation by Hybrid Root SEAL (Sun Medical Co.), a 4-META-containing auto-adhesive self-etching root canal sealer, is affected by the sequence of irrigants employed for removing canal wall smear layers during root canal treatment. METHODS: Single-rooted teeth were shaped and irrigated with EDTA as initial rinse/NaOCl as active final rinse (Group 1), or NaOCl as initial rinse/EDTA as active final rinse (Group 2). All canals were obturated with Hybrid Root SEAL using a single-cone technique. Root slices derived from the coronal, middle and apical thirds of the roots were processed for transmission electron microscopy after removing the gutta-percha, leaving the sealer intact. Additional filled canals from the two groups were evaluated for fluid leakage. RESULTS: Hybrid layer was absent in Group 1 and was present only when a collagen matrix was produced by EDTA demineralization (Group 2). Significantly more leakage (4.03 +/- 1.94 microL min(-1) vs. 1.50 +/- 0.42 microL min(-1); P < 0.05) was observed in the absence of dentin hybridization.

Microporous, demineralized collagen matrices in intact radicular dentin created by commonly used calcium-depleting endodontic irrigants.

The use of calcium-depleting solutions in restorative dentistry results in demineralization of intact coronal dentin beneath smear layers. Likewise, application of calcium-depleting irrigants as final rinses might create demineralized collagen matrices in intact radicular dentin after removal of root canal wall smear layers created during canal shaping. These unsupported collagen matrices collapse on desiccation and might be difficult to detect in scanning electron microscopy dehydration techniques traditionally used in endodontic research. Demineralized collagen matrices were identified, by using a hexamethyldisilazane dehydration protocol, on the surface of smear layer-depleted radicular dentin when NaOCl was used as initial rinse, followed by the use of 17% ethylenediaminetetraacetic acid or BioPure MTAD as the final rinse. Compaction of conventional root filling materials over a demineralized collagen matrix in instrumented dentin is analogous to performing the same procedure against the predentin collagen network in the uninstrumented parts of root canals. The potential consequences of this process are, however, unknown.

Composite-to-composite microtensile bond strength in the repair of a microfilled hybrid resin: effect of surface treatment and oxygen inhibition.

PURPOSE: To compare the 24-h microtensile bond strength of a microfilled hybrid composite to the same material after mechanical and/or chemical treatment and assess the effect of oxygen inhibition on the composite-composite bond. MATERIALS AND METHODS: Forty composite cylinders of Gradia Direct Anterior (GC) were prepared and stored 24 h prior to the following surface treatments: 50-microm aluminum oxide air abrasion and 37% phosphoric acid etching (group 1); hydrochloric acid and 6.9% hydrofluoric acid etching (group 2); diamond bur roughening and 37% phosphoric acid etching (group 3); diamond bur roughening (group 4). In all groups, Prime & Bond NT (Dentsply De Trey) was applied and light cured in air or under a nitrogen atmosphere, prior to layering a buildup of the repairing resin composite. Microtensile bond strength measurements were performed. Data were statistically analyzed with two-way ANOVA and Tukey's test (alpha = 0.05). RESULTS: The curing atmosphere did not significantly influence the interfacial strength (p < 0.05). Surface treatment significantly affected the composite-composite bond (p > 0.05). Air abrasion, regardless of curing atmosphere, resulted in the strongest bond (p < 0.05). The other treatments were comparable. CONCLUSION: Air abrasion and the application of a bonding agent offer satisfactory bond strengths for composite repair. The oxygen inhibition layer on a light-cured adhesive is not crucial to the success of the 24-h composite-composite bond.

The application of hydrogen peroxide in composite repair.

This study examined the influence of different surface conditioning methods on composite-to-composite microtensile bond strength. Thirty two-year old composite resin discs were randomly divided into three groups according to the different mechanical/chemical surface pretreatment tested: (1) 38% hydrogen peroxide (H(2)O(2))-treatment; (2) 50-microm aluminum oxide sandblasting; (3) no treatment. Depending on the intermediate agent applied, two subgroups were created: (A) three-step adhesive system; (B) prehydrolyzed silane coupling agent + three-step adhesive system. Microtensile bond strength measurements were performed and the data were statistically analyzed with Kruskall-Wallis Analysis of Variance and Dunn's multiple range test for post hoc comparisons (p < 0.05). Failure mode was evaluated with a scanning electron microscope. Changes in composite surface topography after H(2)O(2) treatment were also investigated. Composite repair strength did not benefit from H(2)O(2) treatment and adhesive application. Preliminary sandblasting significantly improved interfacial bond strength regardless of the intermediate agent applied. No changes in surface texture were produced after H(2)O(2) treatment. An atypical fracture pattern was detected at the interfacial level between H(2)O(2)-treated composite surfaces and the overlying adhesive and composite. H(2)O(2) treatment affected the composite-to-composite repair strength: a compromised resin polymerization may occur, resulting in a poor interfacial quality and a weak bond. Sandblasting still remains a reliable technique for composite repair.

Have dentin adhesives become too hydrophilic?

This review discusses current trends in the development of dentin adhesives and the possibility that some classes of currently available adhesives are too hydrophilic. Manufacturers have reformulated dentin adhesives to make them more compatible for bonding to intrinsically moist, acid-etched dentin by adding 2-hydroxyethyl methacrylate and other hydrophilic resin monomers. These 3-step adhesives work well but are more time consuming to use and more sensitive to technique than the newer, simplified adhesives. When primers are mixed with adhesives in 2-step single-bottle adhesives and self-etching primers, the adhesives are more permeable to water and hence absorb more water over time than previous generations of adhesives. The most recent single-step self-etching adhesives are even more hydrophilic and hence more permeable to water derived from the underlying bonded dentin. This permeability can lead to a wide variety of seemingly unrelated problems, including incompatibility of chemically or dual-cured composites with simplified adhesives and expedited degradation of resin-dentin bonds.