Wenwen Deng

Encapsulation of plasmid DNA in calcium phosphate nanoparticles: stem cell uptake and gene transfer efficiency.

Background The purpose of this study was to develop calcium phosphate nanocomposite particles encapsulating plasmid DNA (CP-pDNA) nanoparticles as a nonviral vector for gene delivery. Methods CP-pDNA nanoparticles employing plasmid transforming growth factor beta 1 (TGF-β1) were prepared and characterized. The transfection efficiency and cell viability of the CP-pDNA nanoparticles were evaluated in mesenchymal stem cells, which were identified by immunofluorescence staining. Cytotoxicity of plasmid TGF-β1 and calcium phosphate to mesenchymal stem cells were evaluated by MTT assay. Results The integrity of TGF-β1 encapsulated in the CP-pDNA nanoparticles was maintained. The well dispersed CP-pDNA nanoparticles exhibited an ultralow particle size (20–50 nm) and significantly lower cytotoxicity than Lipofectamine™ 2000. Immunofluorescence staining revealed that the cultured cells in this study were probably mesenchymal stem cells. The cellular uptake and transfection efficiency of the CP-pDNA nanoparticles into the mesenchymal stem cells were higher than that of needle-like calcium phosphate nanoparticles and a standard calcium phosphate transfection kit. Furthermore, live cell imaging and confocal laser microscopy vividly showed the transportation process of the CP-pDNA nanoparticles in mesenchymal stem cells. The results of a cytotoxicity assay found that both plasmid TGF-β1 and calcium phosphate were not toxic to mesenchymal stem cells. Conclusion CP-pDNA nanoparticles can be developed into an effective alternative as a nonviral gene delivery system that is highly efficient and has low cytotoxicity.

Seventy-two-hour release formulation of the poorly soluble drug silybin based on porous silica nanoparticles: In vitro release kinetics and in vitro/in vivo correlations in beagle dogs

The objective of this study was to prepare a 72 h-release formulation of silybin (72 h-SLB) using a combination of solid dispersion, gel matrix and porous silica nanoparticles (PSNs) and to investigate the in vitro/in vivo correlations (IVIVCs). The results of scanning electron microscopy and N(2) adsorption demonstrated that empty PSNs possessed a spherical shape, a highly porous structure, a large specific surface area (385.89 ± 1.12 m(2)/g) and a small pore size (2.74 nm on average). The in vitro dissolution profiles of both 72 h-SLB and silybin-loaded PSNs in different concentrations (0.01, 0.06 and 0.08M) of Na(2)CO(3) solutions revealed that 0.06 M Na(2)CO(3) solution was the optimal medium in which silybin could be released from 72 h-SLB with first-order release kinetics and from PSNs with Higuchi kinetics. Furthermore, the IVIVCs of 72 h-SLB and silybin-loaded PSNs in beagle dogs were also established. Using 0.06 M Na(2)CO(3) solution as the in vitro dissolution medium, a good linear relationship could be achieved for both 72 h-SLB and silybin-loaded PSNs. The findings support the fact that the 72 h-SLB (consisting of solid dispersion, regular gel matrix and PSNs) together with Na(2)CO(3) solution as an in vitro dissolution medium can be developed into a promising formulation for poorly soluble drugs, which enjoys a good IVIVC.

Angelica sinensis polysaccharide nanoparticles as novel non-viral carriers for gene delivery to mesenchymal stem cells

UNLABELLED This study centers on the use of a nanoparticle based on the polysaccharide from Angelica sinensis (ASP) as an efficient and safe non-viral gene vector. After modification with branched low molecular weight polyethylenimine (1200 Da), the cationized ASP (cASP) was combined with the plasmid encoding transforming growth factor-beta 1 (TGF-β1) to form a spherical nano-scaled particle (i.e., cASP-pTGF-β1 nanoparticle). This nanoparticle was applied to transfect rat bone marrow mesenchymal stem cells and human umbilical cord mesenchymal stem cells. As a result, nanoparticles (cASP/pDNA weight ratio 10:1) had the greatest transfection efficiency in both cells, which was significantly higher than those of Lipofectamine2000 and PEI (25 kDa). This was in agreement with the findings of the semi-quantitative RT-PCR and live cell imaging. These nanoparticles were also less toxic than Lipofectamine2000 and PEI (25 kDa). Therefore, cASP could be a potential candidate for a novel non-viral gene vector. FROM THE CLINICAL EDITOR These authors demonstrate the use of a nanoparticle-based efficient and safe non-viral gene vector delivery system via a spherical nanoparticle based on a polysaccharide from Angelica sinensis, with parameters superior to Lipofectamine2000.

In vitro release and in vitro–in vivo correlation for silybin meglumine incorporated into hollow-type mesoporous silica nanoparticles

Background The purpose of this study was to develop a sustained drug-release model for water-soluble drugs using silica nanoparticles. Methods Hollow-type mesoporous silica nanoparticles (HMSNs) were prepared using Na2CO3 solution as the dissolution medium for the first time. The water-soluble compound, silybin meglumine, was used as the model drug. The Wagner–Nelson method was used to calculate the in vivo absorption fraction. Results The results of transmission electron microscopy and nitrogen adsorption revealed that the empty HMSNs had uniformly distributed particles of size 50–100 nm, a spherical appearance, a large specific surface area (385.89 ± 1.12 m2/g), and ultralow mean pore size (2.74 nm). The highly porous structure allowed a large drug-loading rate (58.91% ± 0.39%). In 0.08 M Na2CO3 solution, silybin meglumine-loaded HMSNs could achieve highly efficacious and long-term sustained release for 72 hours in vitro. The results of in vitro–in vivo correlation revealed that HMSNs in 0.08 M Na2CO3 solution had a correlation coefficient R2 value of 0.9931, while those of artificial gastric juice and artificial intestinal juice were only 0.9287 and 0.7689, respectively. Conclusion The findings of in vitro–in vivo correlation indicate that HMSNs together with Na2CO3 solution could achieve an excellent linear relationship between in vitro dissolution and in vivo absorption for 72 hours, leading to a promising model for sustained release of water-soluble drugs.

Efficient Gene Delivery to Mesenchymal Stem Cells by an Ethylenediamine‐Modified Polysaccharide from Mulberry Leaves

This study investigates the use of a natural polysaccharide isolated from mulberry leaves as a nonviral gene vector. Ethylenediamine is chemically grafted to the backbone of a polysaccharide from mulberry leaves (MPS) to acquire nucleic acid binding affinity. A particle-size observation indicates that the cationic mulberry leaf polysaccharide (CMPS) can efficiently combine with plasmid transforming growth factor β1 (TGF-β1) to form nanoscaled particles. In addition, the electrophoresis assay indicates a retarded plasmid migration when the CMPS/pTGF-β1 weight ratio is increased to 30:1. The in vitro cell transfection experiment is performed based on bone marrow mesenchymal stem cells (MSCs) derived from rat femurs and tibias, and the findings reveal that the complex with a CMPS/pTGF-β1 weight ratio of 50:1 exhibits the highest cell transfection effect, which is significantly higher than that of branched poly(ethyleneimine) (PEI) (25 kDa; p = 0.001, Students t-test) and slightly higher than Lipofectamine 2000. Moreover, the cytotoxicity assay also demonstrates that all of these tested complexes and the plasmid TGF-β1 are nontoxic to mesenchymal stem cells (MSCs). The results of the living cell imaging confirm that more of the CMPS/plasmid TGF-β1 nanoparticles can be taken up and at a faster rate by the MSCs than by the positive control Lipofectamine 2000; these data are consistent with the transfection efficiency data. Together, these results suggest that the CMPS/pTGF-β1 nanoparticle can potentially be developed into a promising alternative for the transfer of therapeutic genes into cells.

Oral bioavailability of silymarin formulated as a novel 3-day delivery system based on porous silica nanoparticles.

The purpose of this study was to develop porous silica nanoparticles (PSNs) as a carrier to improve oral bioavailability of poorly water-soluble drugs, using silymarin as a model. PSNs were synthesized by reverse microemulsion and ultrasonic corrosion methods. A 3-day release formulation consisting of a silymarin solid dispersion, a hydrophilic gel matrix and silymarin-loaded PSNs was prepared. In vitro release studies indicated that both the silymarin-loaded PSNs and the 3-day release formulation showed a typical sustained-release pattern over a long period, about 72 h. The in vivo studies revealed that the 3-day release formulation gave a significantly higher plasma concentration and larger area under the concentration-time curves than commercial tablets when orally administered to beagle dogs. This implies that the prepared 3-day release formulation significantly enhanced the oral bioavailability of silymarin, suggesting that PSNs can be used as promising drug carriers for oral sustained release systems. Thus providing a technically feasible approach for improving the oral bioavailability and long-term efficacy of poorly soluble drugs.

Oral delivery of capsaicin using MPEG-PCL nanoparticles

Aim:To prepare a biodegradable polymeric carrier for oral delivery of a water-insoluble drug capsaicin (CAP) and evaluate its quality.Methods:CAP-loaded methoxy poly (ethylene glycol)-poly(ε-caprolactone) nanoparticles (CAP/NPs) were prepared using a modified emulsification solvent diffusion technique. The quality of CAP/NPs were evaluated using transmission electron microscopy, powder X-ray diffraction, differential scanning calorimetry and Fourier transform infrared techniques. A dialysis method was used to analyze the in vitro release profile of CAP from the CAP/NPs. Adult male rats were orally administered CAP/NPs (35 mg/kg), and the plasma concentrations of CAP were measured with a validated HPLC method. The morphology of rat gastric mucosa was studied with HE staining.Results:CAP/NPs had an average diameter of 82.54±0.51 nm, high drug-loading capacity of 14.0%±0.13% and high stability. CAP/NPs showed a biphasic release profile in vitro: the burst release was less than 25% of the loaded drug within 12 h followed by a more sustained release for 60 h. The pharmacokinetics study showed that the mean maximum plasma concentration was observed 4 h after oral administered of CAP/NPs, and approximately 90 ng/mL of CAP was detected in serum after 36 h. The area under the curve for the CAP/NPs group was approximately 6-fold higher than that for raw CAP suspension. Histological studies showed that CAP/NPs markedly reduced CAP-caused gastric mucosa irritation.Conclusion:CAP/NPs significantly enhance the bioavailability of CAP and markedly reduce gastric mucosa irritation in rats.

In vitro and in vivo evaluation of capsaicin‐loaded microemulsion for enhanced oral bioavailability

BACKGROUND Capsaicin, as a food additive, has attracted worldwide concern owing to its pungency and multiple pharmacological effects. However, poor water solubility and low bioavailability have limited its application. This study aims to develop a capsaicin-loaded microemulsion to enhance the oral bioavailability of the anti-neuropathic-pain component, capsaicin, which is poorly water soluble. RESULTS In this study, the microemulsion consisting of Cremophor EL, ethanol, medium-chain triglycerides (oil phase) and water (external phase) was prepared and characterized (particle size, morphology, stability and encapsulation efficiency). The gastric mucosa irritation test of formulated capsaicin was performed in rats to evaluate its oral feasibility, followed by the pharmacokinetic study in vivo. Under these conditions, the encapsulated capsaicin revealed a faster capsaicin release in vitro coupled with a greater absorption in vivo when compared to the free capsaicin. The oral bioavailability of the formulated capsaicin-loaded microemulsions was 2.64-fold faster than that of free capsaicin. No significant irritation was observed on the mucosa from the pathological section of capsaicin-loaded microemulsion treated stomach. CONCLUSION These results indicate that the developed microemulsion represents a safe and orally effective carrier for poorly soluble substances. The formulation could be used for clinical trials and expand the application of capsaicin.

Hypolipidemic potential of perillaldehyde-loaded self-nanoemulsifying delivery system in high-fat diet induced hyperlipidemic mice: Formulation, in vitro and in vivo evaluation

This study reports the hypolipidemic effects of perillaldehyde-loaded self-nanoemulsifying delivery system (PAH-SNEDS) developed with D-optimal experimental design based on a three component system: 40% w/w drug-oil phase, X1 (a mixture of perillaldehyde-isopropyl myristate/medium chain triglyceride, 1:1, w/w); 48% surfactant, X2 (Kolliphor EL); and 12% co-surfactant, X3 (PEG 200). The design space was navigated using a linear model to produce spherical and homogenous droplets which were observed under TEM, with mean size, polydispersity index (PDI) and zeta potential of 32.8 ± 0.1 nm, 0.270 ± 0.029 and -10.14 ± 0.66 mV, respectively. PAH-SNEDS demonstrated significant increase in dissolution in vitro compared to the free PAH, and further yielded an oral relative bioavailability of about 206.18% in vivo which suggested a promising formulation design for potential liquid bioactive compounds. Oral administration of PAH-SNEDS (240 mg/kg per body weight) in high-fat induced hyperlipidemia in mice, also significantly decreased serum total cholesterol (TC), triglyceride (TG) and low-density lipoprotein cholesterol (LDL-C) while increasing high-density lipoprotein cholesterol (HDL-C) level. The improved bioavailability and functional application of PAH via SNEDDS suggested a suitable approach to promote hypolipidemic effect of the drug. Perillaldehyde, therefore, promises to be a useful bioactive compound to prevent high-fat diet induced hyperlipidemia.

MicroRNA Replacing Oncogenic Klf4 and c-Myc for Generating iPS Cells via Cationized Pleurotus eryngii Polysaccharide-based Nanotransfection

Induced pluripotent stem cells (iPSCs), resulting from the forced expression of cocktails out of transcription factors, such as Oct4, Sox2, Klf4, and c-Myc (OSKM), has shown tremendous potential in regenerative medicine. Although rapid progress has been made recently in the generation of iPSCs, the safety and efficiency remain key issues for further application. In this work, microRNA 302-367 was employed to substitute the oncogenic Klf4 and c-Myc in the OSKM combination as a safer strategy for successful iPSCs generation. The negatively charged plasmid mixture (encoding Oct4, Sox2, miR302-367) and the positively charged cationized Pleurotus eryngii polysaccharide (CPEPS) self-assembled into nanosized particles, named as CPEPS-OS-miR nanoparticles, which were applied to human umbilical cord mesenchymal stem cells for iPSCs generation after characterization of the physicochemical properties. The CPEPS-OS-miR nanoparticles possessed spherical shape, ultrasmall particle size, and positive surface charge. Importantly, the combination of plasmids Oct4, Sox2, and miR302-367 could not only minimize genetic modification but also show a more than 50 times higher reprogramming efficiency (0.044%) than any other single or possible double combinations of these factors (Oct4, Sox2, miR302-367). Altogether, the current study offers a simple, safe, and effective self-assembly approach for generating clinically applicable iPSCs.