Bacteria-mediated delivery of nanoparticles and cargo into cells

Nanoparticles and bacteria can be used, independently, to deliver genes and proteins into mammalian cells for monitoring or altering gene expression and protein production. Here, we show the simultaneous use of nanoparticles and bacteria to deliver DNA-based model drug molecules in vivo and in vitro. In our approach, cargo (in this case, a fluorescent or a bioluminescent gene) is loaded onto the nanoparticles, which are carried on the bacteria surface. When incubated with cells, the cargo-carrying bacteria ('microbots') were internalized by the cells, and the genes released from the nanoparticles were expressed in the cells. Mice injected with microbots also successfully expressed the genes as seen by the luminescence in different organs. This new approach may be used to deliver different types of cargo into live animals and a variety of cells in culture without the need for complicated genetic manipulations.     

FITC delivery into plant cells using magnetic single-walled carbon nanotubes

In this paper, fluorescein isothiocyanate (FITC) was covalently bonded with magnetic single-walled carbon nanotubes (mSWCNTs) that were purified using our previous method. To demonstrate our design, mSWCNT-FITC was delivered into plant cells (canola and carrot cells) driven by external magnetic forces. From FACS results, the FITC delivery efficiency was about 100% for both two canola and carrot protoplasts, which were further confirmed by the confocal and sectional TEM images. Some mSWCNTs were found trapped both inside the endosomes of canola protoplast and outside endosome near the nuclear membrane of carrot protoplast according to the sectional TEM images. All results showed that mSWCNT is a good delivery carrier for biomolecules.     

Mesoporous silica nanoparticles for efficient rivastigmine hydrogen tartrate delivery into SY5Y cells

Rivastigmine hydrogen tartrate (RT) is a molecule with both hydrophilic and hydrophobic properties used for the treatment of the Alzheimer’s disease. In this work, the larger pore size of mesoporous silica nanoparticles (P1-MSN) was synthesized and then, P1-MSN were functionalized by succinic anhydride (S-P1-MSN) and 3-aminopropyltriethoxysilane (APTES) (AP-CO-P1-MSN) using the grafting and co-condensation methods, respectively. A new method was used for the functionalization of P1-MSN by succinic anhydride at room temperature. Nanoparticles were characterized by special instrumental analysis and loaded by RT. Maximum entrapment efficiency and RT loading percentage into P1-MSN, AP-CO-P1-MSN and S-P1-MSN were respectively obtained as 21.26 and 25.5%, 41.5 and 49.8%, and 11.9 and 14.28% for 24?h. In the simulated gastric and body fluids, the release rate of RT-loaded AP-CO-P1-MSN (AP-CO-P1-MSN-RT) was lower than that of other RT-loaded nanoparticles. In oral pathway, the sustained release of RT was observed in AP-CO-P1-MSN-RT. Moreover, no cytotoxicity effect was observed for P1-MSN, but the cells treated by AP-CO-P1-MSN showed a reduction in SY5Y cell viability due to easy entrance of these nanoparticles and their accumulation in different parts of the cell as observed by TEM.     

Rapid magnetic cell delivery for large tubular bioengineered constructs

Delivery of cells into tubular tissue constructs with large diameters poses significant spatial and temporal challenges. This study describes preliminary findings for a novel process for rapid and uniform seeding of cells onto the luminal surface of large tubular constructs. Fibroblasts, tagged with superparamagnetic iron oxide nanoparticles (SPION), were directed onto the luminal surface of tubular constructs by a magnetic field generated by a k4-type Halbach cylinder device. The spatial distribution of attached cells, as measured by the mean number of cells, was compared with a conventional, dynamic, rotational cell-delivery technique. Cell loading onto the constructs was measured by microscopy and magnetic resonance imaging. The different seeding techniques employed had a significant effect on the spatial distribution of the cells (p < 0.0001). The number of attached cells at defined positions within the same construct was significantly different for the dynamic rotation technique (p < 0.05). In contrast, no significant differences in the number of cells attached to the luminal surface were found between the defined positions on the construct loaded with the Halbach cylinder. The technique described overcomes limitations associated with existing cell-delivery techniques and is amenable to a variety of tubular organs where rapid loading and uniform distribution of cells for therapeutic applications are required.     

Self assembly of DNA nanoparticles with polycations for the delivery of genetic materials into cells

Increasing attention has been paid to technology used for the delivery of genetic materials into cells for gene therapy and the generation of genetically engineered cells. So far, viral vectors have been mainly used because of their inherently high transfection efficiency of gene. However, there are some problems to be resolved for the clinical applications, such as the pathogenicity and immunogenicity of viral vectors themselves. Therefore, many research trials with non-viral vectors have been performed to enhance their efficiency to a level comparable to the viral vector. Two directions of these trials exist: Material improvement of non-viral vectors and their combination with various external physical stimuli. In this study gelatin was selected as a non-viral carrier for DNA. To give a positive charge to gelatin, different extents introduction of ethylenediamine (Ed), spermidine (Sd), and spermine (Sm) were reacted with gelatin in the presence of a water-soluble carbodiimide. When positively charged gelatin derivatives (Ed, Sd, and Sm) were mixed with negatively charged DNA, a self assembly of DNA nanoparticle (complex) was formed within few minutes through electrostatic interaction. Irrespective of the type of gelatin derivatives, the apparent molecular size of DNA was reduced by increasing the gelatin/DNA mixing ratio to attain a saturated value of about 150 nm. The condensed gelatin/DNA complexes showed the zeta potential of 10-15 mV. The amount of DNA internalized into the cells was significantly increased by the complexation with every gelatin derivative. The cells incubated with the gelatin/DNA complexes exhibited significantly stronger luciferase activities than naked plasmid DNA. This study clearly demonstrates and self-assembled DNA complexes has potential as a gene delivery vechile and are stable to transfer genetic materials to cells.     

Determining the size and shape dependence of gold nanoparticle uptake into mammalian cells

We investigated the intracellular uptake of different sized and shaped colloidal gold nanoparticles. We showed that kinetics and saturation concentrations are highly dependent upon the physical dimensions of the nanoparticles (e.g., uptake half-life of 14, 50, and 74 nm nanoparticles is 2.10, 1.90, and 2.24 h, respectively). The findings from this study will have implications in the chemical design of nanostructures for biomedical applications (e.g., tuning intracellular delivery rates and amounts by nanoscale dimensions and engineering complex, multifunctional nanostructures for imaging and therapeutics).     

Folic acid-functionalized niosomal nanoparticles for selective dual-drug delivery into breast cancer cells: An in-vitro investigation

In this study, a folic acid-functionalized niosome was formulated and loaded with letrozole and curcumin as a promising drug carrier system for chemotherapy of the breast cancer cells. The formulation process was optimized by varying the type of Span 80 and total lipid to drug ratio, where Span 80 and lipid to drug molar ratio of 10 resulted in the niosomes with maximum encapsulation of both drugs but minimum size. The developed niosomal formulation showed a great storage stability up to one month with the small changes in drug encapsulation efficiency and size during the storage. In addition, they showed a pH-dependent release behaviour with slow drug release at physiological pH (7.4) while considerable drug release in acidic conditions (pH = 3), making it a promising candidate for breast cancer treatment. The cytotoxicity study shows the niosomal formulation has high biocompatibility with HEK-293 healthy cells, while having remarkable inhibitory effects on MCF-7 and MDA-MB-231 breast cancer cells due to the presence of folic acid in formulation, and in turn, selective internalization of the as-developed nanocarrier through folate receptor-mediated endocytosis. The double drug-loaded niosomes affect the gene expression by studied breast cancer cell lines; down-regulates the expression of Bcl2, cyclin D, and cyclin E genes while they up-regulate the expression of p53, Bax, caspase-3, and caspase-9 genes. The flow cytometry results showed that letrozole/curcumin-loaded niosomes enhanced the apoptosis rate in both MCF-7 and MDA-MB-231 cells compared to the mixture of letrozole and curcumin, which was due to the synergic effect between the two drugs as well as higher cell uptake by niosomal formulation. The findings of our study show the importance of developing highly biocompatible niosomal formulations in the future of nanomedicine that enables the co-delivery of two hydrophobic drugs into the cancer cells improves the efficiency of chemotherapy due to the synergic effect between the drugs.     

Receptor-mediated gene delivery into antigen presenting cells using mannosylated chitosan/DNA nanoparticles

Dendritic cells (DCs) are professional antigen presenting cells that induce, sustain, and regulate immune responses. Gene modification of DCs is of particular interest for immunotherapy of diseases where the immunes system has failed or is abnormally regulated, such as in cancer or autoimmune disease. Gene transfer using non-viral vectors is a promising approach for the safe delivery of therapeutic DNA. Among various non-viral vectors, chitosan is considered to be a good candidate for gene delivery system, however, lack of cell specificity and low transfection of chitosan need to be overcome prior to clinical use. In this study, mannosylated chitosan (MC) was prepared to induce the receptor-mediated endocytosis and targeting into antigen presenting cells (APCs), especially DCs having mannose receptors. MC showed great ability to form complexes with DNA and showed suitable physicochemical properties for gene delivery system. It had low cytotoxicity and exhibited much enhanced gene transfer efficiency on the macrophage cell line than chitosan itself. Also, MC/DNA complex was more efficient for transferring IL-12 gene into DCs rather than water-soluble chitosan (WSC)/DNA one, which resulted in better induction of INF-gamma from DCs. Therefore, MC is a promising gene delivery system for repeated administration to maintain sustained gene expression, thereby opening the possibility for immunotherapy.     

Polyethyleneimine-functionalized large pore ordered silica materials for poorly water-soluble drug delivery

Four ordered mesoporous silica supports with different pore structure characteristics were investigated for their drug loading and release abilities with regard to their structural variabilities as well as implications of surface modification. The (model) drug molecule in question was the poorly water-soluble glucocorticoid Prednisolone, composed of a steroid skeleton with functional groups in the form of carbonyls and hydroxyls. Under non-aqueous conditions, such as those applied for drug loading, these functional groups are expected to interact with the surface silanols of the silica supports, but this interaction could possibly also be enhanced by introducing amino groups to the silica surfaces. Thus, all four supports were further functionalized by surface hyperbranching of polyethyleneimine), PEI, which was successfully incorporated to all supports in high amounts (>30 wt%). However, the accessibility of the pore system after organic modification was dependent on the pore sizes and structures, highlighting the importance of using large-pore mesophases with adequate structures when aiming for applications involving (bulky) guest molecules. Additionally, after incorporation of large amounts of guest molecules (40 wt%), full water accessibility was retained in that the loaded cargo could be rapidly released from the carrier matrixes, which is a crucial requirement when formulating poorly soluble substances. Results displayed that the release of Prednisolone from the silica supports occurred faster than the dissolution of the pure drug. All silica materials released more than 85 % of the adsorbed drug in 5 h, independently of the support material. Thus, the confinement of Prednisolone inside the mesopores seems to be the main reason for the faster kinetic release rate. These constraints imply that Prednisolone becomes more mobile inside the pores, and therefore more soluble in release medium. These results confirm the potential of silica supports as drug delivery carriers for drugs with limited water solubility such as steroids.     

Photothermal delivery of microscopic objects via convection flows induced by laser beam from fiber tip

We report a photothermal delivery of microscopic objects based on convection flows at the surface of water. The convection flows were induced by photothermal effect through a laser beam of 1.55 μm wavelength from a fiber tip. A 206 μm diameter oil drop was delivered forward and backward by changing the laser beam at a power of 28.5-40 mW. In addition, the delivery has been further demonstrated with a cluster of carbon and red blood cells at the laser powers of 14 and 20 mW, respectively.     

高光热效应碳球的快速制备及肿瘤细胞光热治疗

光热治疗是一种非侵入式的新型肿瘤治疗手段,可弥补传统治疗方式的不足。碳纳米材料作为一种高效的光热剂,在肿瘤光热治疗中表现出巨大的应用潜力。本研究采用超声辅助法使邻苯三酚与甲醛5 min快速聚合,经煅烧处理制备了单分散、粒径均一的碳球。该碳球兼具优良的细胞生物相容性和高光热转换效率。在808 nm近红外光照射下,碳球呈现良好的光热效应和光热稳定性,光热转换效率达到41.4%。细胞实验表明,碳球无明显细胞毒性,对肿瘤细胞具有显著的光热杀伤效果。制备的高光热效应碳球光热剂有望用于肿瘤光热治疗。     

Insight into a model for large strain anisotropic elasto-plasticity

Efficient and accurate simulation of the deformations in anisotropic metallic sheets requires a constitutive model and an accompanying algorithm at large strains which take into account the anisotropy of both the elastic and plastic material behaviors, as well as their evolution with plastic strains. Recently we proposed such a constitutive model based on continuum energy considerations, the Lee decomposition and an anisotropic stored energy function of the logarithmic strains in which the rotation of the orthotropic axes is also considered. We obtained a framework similar to the one used in isotropic elasto-plasticity. In the present work we give some physical insight into the parameters of the model and their effects on the predictions, both in proportional and in non-proportional loading problems. We also present a procedure to obtain the spin parameter of the model from Lankford R-values.     

Iontophoresis for drug delivery into the nail apparatus: exploring hyponychium as the site of delivery

In present studies, a hyponychium pathway (from ventral side of the nail plate) was investigated as a potential route of drug delivery into the nail apparatus using iontophoresis as an active physical method. In vitro transport studies were performed across the human nail plate using sodium fluorescein as a marker substrate for 24?h. After transport studies, the amount of sodium fluorescein extracted from an active diffusion area of the nail plate in case of iontophoresis was found to be ～54-folds more to that of passive. The amount of sodium fluorescein retained in the peripheral area of the nail plate after application of iontophoresis was found to be ～30-folds more relative to passive. Ex vivo transport studies were performed on excised human cadaver toe using terbinafine hydrochloride as a model drug for three days (8?h/day). The amount of terbinafine retained in the nail plate after application of iontophoresis (3.43?±?1.34?µg/mg) was ～20-folds more when compared with passive (0.17?±?0.10?µg/mg). The amount of drug extracted from the nail bed and nail matrix was 1.73?±?0.12?µg/mg and 0.55?±?0.22?µg/mg, respectively. On the other hand, there was no detectable amount of terbinafine found in the nail bed and nail matrix in case of control (passive delivery). These studies show that the iontophoretic drug delivery through hyponychium region to other parts of the nail apparatus could be a potential way of onychomycosis treatment.     

Catalytic motors for transport of colloidal cargo

Autonomous micro- and nanomotors should, in principle, deliver materials in a site-directed fashion, powering the assembly of dynamic, nonequilibrium superstructures. Here we demonstrate that catalytic Pt-Au nanomotors can transport a prototypical cargo: polystyrene microspheres. In addition, motors with Ni segments can overcome both Brownian orientational fluctuations and biased rotation of the rod-sphere doublet to enable persistent steerable uniaxial motion in an external magnetic field. Assuming a cargo-independent motive force, the speeds are inversely proportional to the Stokes resistance, which we compute using a completed double-layer boundary integral equation. In addition, we demonstrate motors transporting cargo via chemotaxis toward a H2O2 fuel source.