The immunogenic reaction and bone defect repair function of ε-poly-L-lysine (EPL)-coated nanoscale PCL/HA scaffold in rabbit calvarial bone defect.

Tissue engineering is a promising strategy for bone tissue defect reconstruction. Immunogenic reaction, which was induced by scaffolds degradation or contaminating microorganism, influence cellular activity, compromise the efficiency of tissue engineering, or eventually lead to the failure of regeneration. Inhibiting excessive immune response through modulating scaffold is critical important to promote tissue regeneration. Our previous study showed that ε-poly-L-lysine (EPL)-coated nanoscale polycaprolactone/hydroxyapatite (EPL/PCL/HA) composite scaffold has enhanced antibacterial and osteogenic properties in vitro. However, the bone defect repair function and immunogenic reaction of EPL/PCL/HA scaffolds in vivo remains unclear. In the present study, three nanoscale scaffolds (EPL/PCL/HA, PCL and PCL/HA) were transplanted into rabbit paraspinal muscle pouches, and T helper type 1 (Th1), T helper type 2 (Th2), T helper type 17 (Th17), and macrophage infiltration were analyzed after 1 week and 2 weeks to detect their immunogenic reaction. Then, the different scaffolds were transplanted into rabbit calvarial bone defect to compare the bone defect repair capacities. The results showed that EPL/PCL/HA composite scaffolds decreased pro-inflammatory Th1, Th17, and type I macrophage infiltration from 1 to 2 weeks, and increased anti-inflammatory Th2 infiltration into the regenerated area at 2 weeks in vivo, when compared to PCL and PCL/HA. In addition, EPL/PCL/HA showed an enhanced bone repair capacity compared to PCL and PCL/HA when transplanted into rabbit calvarial bone defects at both 4 and 8 weeks. Hence, our results suggest that EPL could regulate the immunogenic reaction and promote bone defect repair function of PCL/HA, which is a promising agent for tissue engineering scaffold modulation.

Investigating the Correlation between Miscibility and Physical Stability of Amorphous Solid Dispersions Using Fluorescence-Based Techniques.

The purpose of this study was to investigate the feasibility of using a fluorescence-based technique to evaluate drug-polymer miscibility and to probe the correlation between miscibility and physical stability of amorphous solid dispersions (ASDs). Indomethacin-hydroxypropyl methylcellulose (IDM-HPMC), indomethacin-hydroxypropyl methylcellulose acetate succinate, and indomethacin-polyvinylpyrrolidone (IDM-PVP) were used as model systems. The miscibility of the IDM-polymer systems was evaluated by fluorescence spectroscopy, fluorescence imaging, differential scanning calorimetry (DSC), and infrared (IR) spectroscopy. The physical stability of IDM-polymer ASDs stored at 40 °C was evaluated using fluorescence imaging and X-ray diffraction (XRD). The experimentally determined miscibility limit of IDM with the polymers was 50-60%, 20-30%, and 70-80% drug loading for HPMC, HPMCAS, and PVP, respectively. The X-ray results showed that for IDM-HPMC ASDs, samples with a drug loading of less than 50% were maintained in amorphous form during the study period, while samples with drug loadings higher than 50% crystallized within 15 days. For IDM-HPMCAS ASDs, samples with drug loading less than 30% remained amorphous, while samples with drug loadings higher than 30% crystallized within 10 days. IDM-PVP ASDs were found to be resistant to crystallization for all compositions. Thus, a good correlation was observed between phase separation and reduced physical stability, suggesting that miscibility is indeed an important ASDs characteristic. In addition, fluorescence-based techniques show promise in the evaluation of drug-polymer miscibility.

Theoretical prediction of a phase diagram for solid dispersions.

PURPOSE: To predict the temperature-composition phase diagram of solid dispersions (SDs) through theoretical approaches using cinnarizine-Soluplus(®) SD as a model system and evaluate the predicted results. METHODS: A complete phase diagram of cinnarizine-Soluplus(®) SD, including the solubility curve, miscibility curve and glass transition temperature curve, was constructed on the basis of the solid-liquid equilibrium (SLE) equation, Florry-Huggins (F-H) theory and Fox equation. Cinnarizine-Soluplus(®) SDs with different drug loadings were prepared by hot melt extrusion. The extrudates and corresponding physical mixtures were analyzed to check the predicted results. RESULTS: The experimental data revealed a solubility of 7.9 wt% at 110°C and a miscibility level of 65 wt% at room temperature, which were both consistent with predicted values. CONCLUSIONS: The predicted phase diagram agrees well with the experimental results for the non-polar components which mainly interact through dispersion forces, thus facilitating the formulation design of SDs.

Comparison of periodontal ligament anesthesia and submucosal infiltration anesthesia in healthy volunteers.

OBJECTIVE: To compare periodontal ligament anesthesia using a computer-controlled local anesthetic delivery system (C-CLADS) and submucosal infiltration anesthesia using a manually operated syringe in terms of the injection pain, anesthetic effect, anesthetic dose, and complications in healthy volunteers. METHODS: Fifty healthy volunteers, aged 18 to 56 years, were recruited from September 2012 to May 2013 in the Department of Stomatology of Peking Union Medical College Hospital. A randomized self-controlled trial was conducted by applying a periodontal ligament anesthesia on one side and conventional manual submucosal infiltration anesthesia to the other (control) side. The differences in the onset time of anesthesia, drug dosage, anesthetic effect, and the degree of injection pain were compared. The complications associated with the two anesthesia methods were also recorded. RESULTS: When using C-CLADS to perform a periodontal ligament anesthesia, the drug dosage and the severity of injection pain were significantly less than those of conventional manual infiltration anesthesia [drug dosage: (0.34±0.09)ml vs.(0.55±0.13)ml, P<0.01; VRS: 0.42±0.73 vs. 1.38±0.92, P<0.01; VAS: 1.34±1.21 vs. 3.10±1.70, P<0.01]. The anesthesia success rate was approximately 90.0%, showing no significant difference relative to conventional submucosal infiltration anesthesia. Approximately 24% of the volunteers experienced postoperative pain after periodontal ligament anesthesia. CONCLUSION: Compared with conventional submucosal infiltration anesthesia using manual syringes, periodontal ligament anesthesia performed using C-CLADS can reduce the injection pain and drug dosage while achieving a satisfactory anesthetic effect; however, a considerable proportion of cases may experience postoperative pain.

Rational design of antibiotic-free antimicrobial contact lenses: Trade-offs between antimicrobial performance and biocompatibility.

Microbial keratitis associated with contact lenses (CLs) wear remains a significant clinical concern. Antibiotic therapy is the current standard of care. However, the emergence of multidrug-resistant pathogens necessitates the investigation of alternative strategies. Antibiotic-free antimicrobial contact lenses (AFAMCLs) represent a promising approach in this regard. The effectiveness of CLs constructed with a variety of antibiotic-free antimicrobial strategies against microorganisms has been demonstrated. However, the impact of these antimicrobial strategies on CLs biocompatibility remains unclear. In the design and development of AFAMCLs, striking a balance between robust antimicrobial performance and optimal biocompatibility, including safety and wearing comfort, is a key issue. This review provides a comprehensive overview of recent advancements in AFAMCLs technology. The focus is on the antimicrobial efficacy and safety of various strategies employed in AFAMCLs construction. Furthermore, this review investigates the potential impact of these strategies on CLs parameters related to wearer comfort. This review aims to contribute to the continuous improvement of AFAMCLs and provide a reference for the trade-off between resistance to microorganisms and wearing comfort. In addition, it is hoped that this review can also provide a reference for the antimicrobial design of other medical devices.

Increased absorption of mangiferin in the gastrointestinal tract and its mechanism of action by absorption enhancers in rats.

The therapeutic efficiency of mangiferin is restricted by its low intestinal permeability. In order to improve the oral absorption of mangiferin, potential of enhancers, including TPGS, sodium deoxycholate and Carbopol 974P, were investigated in a series of in vivo experiments. After administration of mangiferin at a dose of 30 mg/kg combining with sodium deoxycholate, the bioavailability of mangiferin increased four-fold, and this may be due to sodium deoxycholate weakening the compactness between lecithin molecules and increased the paracellular permeability. When Carbopol 974P (100 mg/kg) was combined with mangiferin, the oral bioavailability of it increased seven-fold compared with the control group, and this may be related to the mucoadhesive properties of Carbopol 974P and paracellular drug permeation. However, following coadministration of TPGS (50 mg/kg), the oral absorption of mangiferin increased slightly compared with that of the control group (p > 0.05). The increased oral absorption by the three coexcipients was in the order of Carbopol 974P > sodium deoxycholate > TPGS. The absolute bioavailability of mangiferin in the three different doses following oral administration were evaluated based on the AUC(0-t) of the intravenous dose and there was no increase from low doses to high doses (25 mg/kg to 100 mg/kg). The above results show that the low absorption of mangiferin was due to presence of a narrow absorption window, which may also exist in these compounds, which have structures similar to mangiferin including three fused aromatic rings with polyphenolic hydroxyl groups. Bioadhesion polymers are effective enhancers of the absorption of mangiferin in the gastrointestinal tract.

Supersaturation and phase behavior during dissolution of amorphous solid dispersions.

Amorphous solid dispersion (ASD) is a promising strategy to enhance solubility and bioavailability of poorly water-soluble drugs. Due to higher free energy of ASD, supersaturated drug solution could be generated during dissolution. When amorphous solubility of a drug is exceeded, drug-rich nanodroplets could form and act as a reservoir to maintain the maximum free drug concentration in solution, facilitating the absorption of the drug in vivo. Dissolution behavior of ASD has received increasing interests. This review will focus on the recent advances in ASD dissolution, including the generation and maintenance of supersaturated drug solution in absence or presence of liquid-liquid phase separation. Mechanism of drug release from ASD including polymer-controlled dissolution and drug-controlled dissolution will be introduced. Formation of amorphous drug-rich nanodroplets during dissolution and the underlying mechanism will be discussed. Phase separation morphology of hydrated ASD plays a critical role in dissolution behavior of ASD, which will be highlighted. Supersaturated drug solution shows poor physical stability and tends to crystallize. The effect of polymer and surfactant on supersaturated drug solution will be demonstrated and some unexpected results will be shown. Physicochemical properties of drug and polymer could impact ASD dissolution and some of them even show opposite effect on dissolution and physical stability of ASD in solid state, respectively. This review will contribute to a better understanding of ASD dissolution and facilitate a rational design of ASD formulation.

Effect of the third component on the aging and crystallization of cinnarizine-soluplus® binary solid dispersion.

Multiple carriers may be used to prepare solid dispersions (SDs) for different purposes. The aim of this work is to investigate the effect of the third component on the physical stability and physical aging behavior of cinnarizine-soluplus SDs. HPMC, PVP, sorbitol and citric acid were used as the third component to prepare cinnarizine ternary SDs using hot melt extrusion method. The resultant samples were stored at 25 °C or under stress conditions. Differential scanning calorimetry, powder X-ray diffraction and dissolution tests were performed to investigate the changes of samples during the storage. Infrared spectroscopy was used to evaluate the interactions between drug and carriers. Results showed that the addition of HPMC or PVP enhanced the physical stability of ternary SDs stored at 25 °C rather than those stored under stress conditions. Sorbitol did not show any improvements in physical stability of samples stored at 25 °C or under stress conditions. Surprisingly, the physical stability of samples stored at 25 °C or under stress conditions was enhanced significantly by citric acid due to the ionic and hydrogen bonding interactions. The miscibility between drug and carriers as well as between different carriers should be considered when using multiple carriers. The third component can act as a "linker" by interacting with drug and polymer to enhance the physical stability of SDs effectively.

Synergistic breast tumor cell killing achieved by intracellular co-delivery of doxorubicin and disulfiram via core-shell-corona nanoparticles.

Combination therapy with different functional chemotherapeutic agents based on nano-drug delivery systems is an effective strategy for the treatment of breast cancer. However, co-delivery of drug molecules with different physicochemical properties still remains a challenge. In this study, an amphiphilic poly (ε-caprolactone)-b-poly (l-glutamic acid)-g-methoxy poly (ethylene glycol) (PCL-b-PGlu-g-mPEG) copolymer was designed and synthesized to develop a nanocarrier for the co-delivery of hydrophilic doxorubicin (DOX) and hydrophobic disulfiram (DSF). The amphiphilic copolymer self-assembled into core-shell-corona structured nanoparticles with the hydrophobic PCL core for DSF loading (hydrophobic interaction) and anionic poly (glutamic acid) shell for DOX loading (electrostatic interaction). DSF and DOX co-loaded nanoparticles (Co-NPs) resulted in high drug loading and precisely controlled DSF/DOX ratio via formulation optimization. Compared with free drug solutions, DSF and DOX delivered by the Co-NPs were found to have improved intracellular accumulation. Results of cytotoxicity assays showed that DSF/DOX delivered at the weight ratio of 0.5 and 1 could achieve a synergistic cytotoxic effect on breast cancer cell lines (MCF-7 and MDA-MB-231). In vivo imaging confirmed that the core-shell-corona nanoparticles could efficiently accumulate in tumors. In vivo anti-tumor effect results indicated that Co-NPs showed an improved drug synergistic effect on antitumor activity compared with the free drug combination. Therefore, it can be concluded that core-shell-corona nanoparticles prepared by PCL-b-PGlu-g-mPEG could be a promising co-delivery system for drug combination therapy in the treatment of breast cancer.

Strategies for improving the payload of small molecular drugs in polymeric micelles.

In the past few years, substantial efforts have been made in the design and preparation of polymeric micelles as novel drug delivery vehicles. Typically, polymeric micelles possess a spherical core-shell structure, with a hydrophobic core and a hydrophilic shell. Consequently, poorly water-soluble drugs can be effectively solubilized within the hydrophobic core, which can significantly boost their drug loading in aqueous media. This leads to new opportunities for some bioactive compounds that have previously been abandoned due to their low aqueous solubility. Even so, the payload of small molecular drugs is still not often satisfactory due to low drug loading and premature release, which makes it difficult to meet the requirements of in vivo studies. This problem has been a major focus in recent years. Following an analysis of the published literature in this field, several strategies towards achieving polymeric micelles with high drug loading and stability are presented in this review, in order to ensure adequate drug levels reach target sites.

Extruded Soluplus/SIM as an oral delivery system: characterization, interactions, in vitro and in vivo evaluations.

The aim of this study was to obtain a stable, amorphous solid dispersion (SD) with Soluplus, prepared by hot-melt extrusion (HME) as an effective and stable oral delivery system to improve the physical stability and bioavailability of the poorly water-soluble simvastatin (SIM), a drug with relatively low Tg. The drug was proved to be miscible with Soluplus by calculation and measurements. The solubility, dissolution, thermal characteristics, interactions and physical stability of the SIM/Soluplus SDs were investigated. The crystal state of simvastatin in the SD was found to change from crystalline to amorphous form during the HME process and also hydrogen bonds were observed between SIM and the extruded Soluplus. The phase solubility showed the solubilization effect of Soluplus was strong and spontaneous. The equilibrium solubility illustrated that Soluplus/SIM SDs gained much higher solubility than its corresponding physical mixtures (PMs). Both of the dissolution profiles and in-vivo performance showed that the SIM/Soluplus SD obtained a marked enhancement, compared with the PM. There was a little change in the SIM/Soluplus SD during a 3-month storage period (40 °C, 75%), indicating the good physicochemical stability. The extruded Soluplus system prepared by HME is a good alternative for the water-insoluble SIM to improve the stability and bioavailability.

A Copper-Mediated Disulfiram-Loaded pH-Triggered PEG-Shedding TAT Peptide-Modified Lipid Nanocapsules for Use in Tumor Therapy.

Disulfiram, which exhibits marked tumor inhibition mediated by copper, was encapsulated in lipid nanocapsules modified with TAT peptide (TATp) and pH-triggered sheddable PEG to target cancer cells on the basis of tumor environmental specificity. PEG-shedding lipid nanocapsules (S-LNCs) were fabricated from LNCs by decorating short PEG chains with TATp (HS-PEG(1k)-TATp) to form TATp-LNCs and then covered by pH-sensitive graft copolymers of long PEG chains (PGA-g-PEG(2k)). The DSF-S-LNCs had sizes in the range of 60-90 nm and were stable in the presence of 50% plasma. DSF-S-LNCs exhibited higher intracellular uptake and antitumor activity at pH 6.5 than at pH 7.4. The preincubation of Cu showed that the DSF cytotoxicity was based on the accumulation of Cu in Hep G2 cells. Pharmacokinetic studies showed the markedly improved pharmacokinetic profiles of DSF-S-LNCs (AUC= 3921.391 µg/L·h, t(1/2z) = 1.294 h) compared with free DSF (AUC = 907.724 µg/L·h, t(1/2z) = 0.252 h). The in vivo distribution of S-LNCs was investigated using Cy5.5 as a fluorescent probe. In tumor-bearing mice, the delivery efficiency of S-LNCs was found to be 496.5% higher than that of free Cy5.5 and 74.5% higher than that of LNCs in tumors. In conclusion, DSF-S-LNCs increased both the stability and tumor internalization and further increased the cytotoxicity because of the higher copper content.

Improving cabazitaxel chemical stability in parenteral lipid emulsions using cholesterol.

Intravenous lipid emulsions of cabazitaxel (CLEs) with a high stability were prepared by adding cholesterol (CH) to provide a new and more suitable delivery system for its administration. The factors affecting CLEs, such as the solubility of cabazitaxel in various oils, different kinds of lecithin, pH, different types of oil phases, and different concentrations of lipoid E80®, CH and poloxamer 188 were investigated systematically. The degradation of cabazitaxel in aqueous solution and lipid emulsion both followed pseudo first-order kinetics. A degradation mechanism was suggested by the U-shaped pH-rate profile of cabazitaxel. A formulation containing 0.5% (w/v) CH and another formulation without CH were made to investigate the protective influence of CH on the chemical stability of CLEs. The activation energy of the two formulations was calculated to be 65.74±6.88 and 54.24±1.43 kJ/mol (n=3), respectively. Compared with the untreated CH, the shelf-life of cabazitaxel with added CH was longer, namely 134.0±23.4 days versus 831.4±204.4 days (n=3) at 4 °C. This indicates that the addition of CH significantly improved the lifetime of cabazitaxel in intravenous lipid emulsions. The hydrogen bonding that takes place between cabazitaxel and CH accounts for the protective effect of CH on the chemical stability of CLEs in two ways: preventing cabazitaxel from leaking and hydrolyzing in aqueous solution and hindering hydrolysis in the oil phase. Finally, the hypothesis was confirmed by LC/TOFMS and Fourier-transform infrared-spectroscopy. As a result, CLEs were obtained successfully by the addition of CH and were stable enough to allow further research.

A comparison of the effect of temperature and moisture on the solid dispersions: aging and crystallization.

The purpose of this work was to compare the effect of temperature and relative humidity (RH) on the physical stability and dissolution of solid dispersions. Cinnarizine-Soluplus(®) solid dispersions (SDs) at three different drug loadings (10, 20 and 35 wt%) were prepared by hot melt extrusion and exposed to stress conditions: high temperatures (40 and 60 °C), high relative humidities (75% and 94% RH) and accelerated conditions (40 °C/75% RH) for 30 days, or stored at 25 °C for up to 5 months. Changes in solid state and dissolution of SDs were investigated by differential scanning calorimetry (DSC), powder X-ray diffraction (PXRD) and dissolution testing. For samples under stress conditions, the results showed a reduced dissolution and a recrystallization of the drug with an increased crystallinity in the order of 40 °C/75% RH, >60 °C/0% RH, >25 °C/94% RH, >40 °C/0% RH, >25 °C/75% RH. For samples stored at 25 °C, nonlinear physical aging was observed and the dissolution also decreased although the SDs were still amorphous. The results indicated that temperature and humidity seemed to have comparable effects on the crystallization of cinnarizine-Soluplus(®) SDs. It is not reasonable to regard recrystallization as a sign of reduced dissolution, and glass transition temperature (Tg) may be a good indicator of the changes in dissolution.

PEG-PLGA copolymers: their structure and structure-influenced drug delivery applications.

In the paper, we begin by describing polyethylene glycol-poly lactic acid-co-glycolic acid (PEG-PLGA) which was chosen as a typical model copolymer for the construction of nano-sized drug delivery systems and also the types of PEG-PLGA copolymers that were eluted. Following this we examine the structure-influenced drug delivery applications including nanoparticles, micelles and hydrogels. After that, the preparation methods for nano-sized delivery systems are presented. In addition, the drug loading mode of PEG-PLGA micelles is divided into three aspects. Finally, the drug release profiles of PEG-PLGA micelles, both in terms of their in vitro and in vivo characteristics, are represented. PEG-PLGA copolymers are very suitable for the construction of micelles as carriers for insoluble drugs. This article reviews the structure and the different structure-influenced applications of PEG-PLGA copolymers, concentrating on the application of PEG-PLGA micelles.

Investigation of a nanosuspension stabilized by Soluplus® to improve bioavailability.

The purpose of this work was to explore the feasibility of using Soluplus(®) in preparing a fenofibrate (FBT) nanosuspension adopting wet media milling technology. HPMC and Soluplus(®) were used as stabilizers to prepare FBT/HPMC nanosuspension (F1) and FBT/Soluplus(®) nanosuspension (F2), respectively. The nanosuspensions were subjected to evaluations involving particle size, dissolution, preliminary stability and pharmacokinetic behavior. A marked reduction in particle size was achieved by nanosuspensions (from 17.55 µm to 642 nm (F1) and 344 nm (F2)). The nanosuspensions displayed almost complete dissolution while percentages of 30% and 13% were obtained by physical mixtures and coarse FBT separately. Soluplus(®) could stabilize the nanosuspension more effectively due to a weaker Ostwald ripening effect resulting from a slower diffusion of micelles formed by Soluplus(®) entrapping dissolved FBT than FBT exposed to pure water directly. In the in vivo evaluation, larger AUC0-72h and Cmax, and shorter Tmax were obtained by the nanosuspensions. Significant differences were observed between the physical mixtures. The phenomenon of double peaks was present in this study. The major factor may be the multiple absorption sites of FBT. The current work indicated that Soluplus(®) is well suited for preparation of a nanosuspension with good stability and improved dissolution and bioavailability.

Clarithromycin-loaded liposomes offering high drug loading and less irritation.

The aim of this study was to develop an efficient method of preparing less irritant clarithromycin-loaded liposomes (CLA-Lip) for injection with a high drug loading and to evaluate their physicochemical characteristics before and after lyophilization. CLA-Lip were prepared using the film-dispersion method with sodium cholesterol sulfate (SCS) and n-hexyl acid as the regulators and then lyophilized. The liposomes were characterized in terms of their size, size distribution, zeta potential, morphology, in vitro release, haemolysis, and lyophilization and irritation testing was carried out. The TEM images revealed that the structure of the CLA-Lip were multilamellar and of a regular size of around 100 nm. In addition, the lyophilized CLA-Lip were characterized by DSC and Infrared spectroscopy to confirm the structure. H-bonding and salt-forming reactions were used to ensure that clarithromycin (CLA) was stably encapsulated in the liposomes. This method provided a 30-fold increase in the concentration of clarithromycin relative to that in aqueous solution. Sucrose was found to be the best protective agent and was added in an amount of 12.5% (w/v). According to the mouse scratch test and the rat paw lick test, the pain of CLA-Lip was significantly reduce by approximately 80% compared with the solution of clarithromycin phosphate. In addition, rabbit ear vein experiments produced similar results. These findings suggested that CLA-Lip was a stable delivery system with less irritation, which should be extremely suitable for clinical application.

Increased dissolution and oral absorption of itraconazole/Soluplus extrudate compared with itraconazole nanosuspension.

The purpose of this article was to compare the in vitro and in vivo profiles of itraconazole (ITZ) extrudates and nanosuspension separately prepared by two different methods. And it was proved truly to form nanocrystalline and amorphous ITZ characterized by differential scanning calorimetry (DSC), X-ray powder diffraction (XRD) analysis, Fourier transform infrared spectrum (FTIR), transmission electron microscope (TEM), and scanning electron microscope (SEM). The release of ITZ/Soluplus solid dispersions with amorphous ITZ was almost complete while only 40% release was obtained with ITZ nanocrystals. The amorphous state need not to cross over the crystal lattice energy upon dissolution while the crystalline need to overcome it. In the in vivo assay, the AUC(0-t) and C(max) of ITZ/Soluplus were 6.9- and 11.6-time higher than those of pure ITZ. The formulation of the extrudate had an AUC(0-t) and C(max) similar to those of ITZ and also OH-ITZ compared with the commercial capsule (Sporanox®). The relative bioavailability values with their 95% confidence limit were calculated to be 98.3% (92.5-104.1%) and 101.3% (97.9-104.1%), respectively. The results of this study showed increased dissolution and bioavailability of the solid dispersion of Soluplus-based carrier loading ITZ prepared by HME compared with the ITZ nanosuspension prepared by wet milling.

Pluronic F127 nanomicelles engineered with nuclear localized functionality for targeted drug delivery.

PKKKRKV (Pro-Lys-Lys-Lys-Arg-Lys-Val, PV7), a seven amino acid peptide, has emerged as one of the primary nuclear localization signals that can be targeted into cell nucleus via the nuclear import machinery. Taking advantage of chemical diversity and biological activities of this short peptide sequence, in this study, Pluronic F127 nanomicelles engineered with nuclear localized functionality were successfully developed for intracellular drug delivery. These nanomicelles with the size ~100 nm were self-assembled from F127 polymer that was flanked with two PV7 sequences at its both terminal ends. Hydrophobic anticancer drug doxorubicin (DOX) with inherent fluorescence was chosen as the model drug, which was found to be efficiently encapsulated into nanomicelles with the encapsulation efficiency at 72.68%. In comparison with the non-functionalized namomicelles, the microscopic observation reveals that PV7 functionalized nanomicelles display a higher cellular uptake, especially into the nucleus of HepG2 cells, due to the nuclear localization signal effects. Both cytotoxicity and apoptosis studies show that the DOX-loaded nanomicelles were more potent than drug nanomicelles without nuclear targeting functionality. It was thus concluded that PV7 functionalized nanomicelles could be a potentially alternative vehicle for nuclear targeting drug delivery.

Biochemical characterization of a cinnamoyl-CoA reductase from wheat.

Cinnamoyl-CoA reductase (CCR) is responsible for the CoA ester-->aldehyde conversion in monolignol biosynthesis, which can divert phenylpropanoid-derived metabolites into the biosynthesis of lignin. To gain a better understanding of lignin biosynthesis in wheat (Triticum aestivum L.), a cDNA encoding CCR was isolated and named Ta-CCR2. DNA hybridization analyses demonstrated that the Ta-CCR2 gene exists in three copies in the wheat genome. RNA blot hybridization indicated that Ta-CCR2 was expressed most abundantly in root and stem tissues that were in the process of lignification. The secondary and three-dimensional structures of Ta-CCR2 were analyzed by molecular modeling. Recombinant Ta-CCR2 protein purified from E. coli converted feruloyl CoA, 5-OH-feruloyl CoA, sinapoyl CoA and caffeoyl CoA with almost similar efficiency, suggesting that it is involved in both G and S lignin synthesis. Ta-CCR2 had a very low V max value for 4-coumaroyl CoA, which may serve as a mechanism to control metabolic flux to H lignin in vivo . Furthermore, the reaction mechanism of Ta-CCR2 was analyzed in relation to its possible three-dimensional structure. The activity of Ta-CCR2 in relation to lignin biosynthesis is discussed.