Nanoparticles conjugated with bacteria targeting tumors for precision imaging and therapy.

The hypoxic region microenvironment reduces the susceptibility of the cancer cells to radiotherapy and anticancer drugs of the solid tumors. However, the reduced oxygen surroundings provide an appreciable habitat for anaerobic bacteria to colonize and proliferate. Herein, we present a biocompatible bacteriabased system that can deliver poly(lactic-co-glycolic acid)(PLGA) nanoparticles(PLGA NPs) specifically targeting into solid tumor to achieve precision imaging and treatment. In our strategy, anaerobic bacterium Bifidobacterium longum (B. longum) that colonizes selectively in hypoxic regions of animal body was successfully used as a vehicle to conjugate with PLGA NPs and transported into solid tumors. To improve the efficacy and specificity of tumor therapy, low-boiling point perfluorohexane (PFH) liquid was wrapped in the core of PLGA NPs (PFH/PLGA NPs), which could increase the deposition of energy by affecting the acoustic environment of the tumor and destroy cells after liquid-gas phase transition during High Intensity Focused Ultrasound (HIFU) irradiation. This strategy shows an effective diagnosis and treatment integration for giving stronger imaging, longer retention period and more effective tumor therapy.

Encapsulation of Nucleic Acids into Giant Unilamellar Vesicles by Freeze-Thaw: a Way Protocells May Form.

Protocells are believed to consist of a lipid membrane and encapsulated nucleic acid. As the lipid membrane is impermeable to macromolecules like nucleic acids, the processes by which nucleic acids become encapsulated inside lipid membrane compartments are still unknown. In this paper, a freeze-thaw method was modified and applied to giant unilamellar vesicles (GUVs) and deoxyribonucleic acid (DNA) in mixed solution resulting in the efficient encapsulation of 6.4 kb plasmid DNA and similar length linear DNA into GUVs. The mechanism of encapsulation was followed by observing the effect of freeze-thaw temperatures on GUV morphological change, DNA encapsulation and ice crystal formation, and analyzing their correlation. Following ice crystal formation, the shape of spherical GUVs was altered and membrane integrity was damaged and this was found to be a necessary condition for encapsulation. Heating alone had no effects on DNA encapsulation, but was helpful for restoring the spherical shape and membrane integrity of GUVs damaged during freezing. These results suggested that freeze-thaw could promote the encapsulation of DNA into GUVs by a mechanism: the vesicle membrane was breached by ice crystal formation during freezing, DNA entered into damaged GUVs through these membrane gaps and was encapsulated after the membrane was resealed during the thawing process. The process described herein therefore describes a simple way for the encapsulation of nucleic acids and potentially other macromolecules into lipid vesicles, a process by which early protocells might have formed.

[Preparation and Characterization of Manganese and Fluorine Co-Modified Hydroxyapatite Composite Coating].

Titanium and titanium alloys have been widely used as orthopedic, dental implants and cardiovascular stents owing to their superior physical properties. However, titanium surface is inherently bio-inert, thus could not form efficient osseointegration with surrounding bone tissue. Therefore, to improve the surface property of titanium implant is significantly important in clinical application. Manganese and fluorine co-doped hydroxyapatite (FMnHAP) coatings were prepared on titanium substrate by electrochemical deposition technique. The as-prepared coatings were examined by scanning electron microscope (SEM), energy-dispersive X-ray spectroscopy (EDS), Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD) tests. The results indicated that the FMnHAP coatings take the morphology of nanoscale-villous-like, the composite coating becomes more compact. The FTIR test indicated that the symmetry of bending vibration modes of hydroxyl changed, simulated body fluid immersion test proved that the FMnHAP coatings had induce carbonate-apatite formation, indicating that the composite coating possess excellent biocompatibility. In the electrochemical corrosion testing, the FMnHAP coatings showed stronger corrosion resistance than pure Ti.

Giant vesicles form in physiological saline and encapsulate pDNA by the modified electroformation method.

Giant vesicles (GVs) are used to study the structures and functions of cells and cell membranes. Electroformation is the most commonly used method for GV preparation. However, the electroformation of GVs is hindered in highly concentrated ionic solutions, limiting their application as cell models for research under physiological conditions. In this study, giant multilayer vesicles were successfully generated in physiological saline using a modified electroformation device by adding an insulating layer between the two electrode plates. The influence of the electric frequency and strength on the electroformation of GVs in physiological saline was explored, and a possible mechanism for this improvement was assessed. It has been shown that an insulating layer between the two electrodes can improve the electroformation of GVs in physiological saline by increasing the electrical impedance, which is weakened by the saline solution, thereby restoring the reduced effective electric field strength. Furthermore, macromolecular plasmid DNA (pDNA) was successfully encapsulated in the electroformed GVs of the modified device. This modified electroformation method may be useful for generating eukaryotic cell models under physiological conditions.

Di[12]aneN3-Functionalized Green Fluorescent Protein Chromophore for GFP Luminescence Simulation and Efficient Gene Transfection In Vitro and In Vivo.

Although a plethora of gene carriers have been developed for potential gene therapy, imageable stimuli-responsive gene vectors with fast access to the nucleus, high biocompatibility, and transfection efficiency are still scarce. Herein, we report the design and synthesis of four dendrite-shaped cationic liposomes, MPA-HBI-R/DOPE (R: n-butyl, 1; n-octyl, 2; n-dodecyl, 3; palmyl, 4), prepared via esterification of 4-alkoxybenzylideneimidazolinone containing aliphatic chains of different lengths (HBI-R), the green fluorescent protein (GFP) chromophore, with a di[12]aneN3 unit. Liposomes were fabricated via the self-assembly of MPA-HBI-R, assisted with 1,2-dioleoyl-sn-glycerol-3-phosphorylethanolamine (DOPE). These liposomes (MPA-HBI-R/DOPE) exhibited efficient DNA condensation, pH-responsive degradation, excellent cellular biocompatibility (up to 150 µM), and high transfection efficiency. Molecular docking experiments were also used to verify the optimal interaction between MPA-HBI-R and DNA, as well as the fluorescence enhancements. In particular, MPA-HBI-2/DOPE delivered DNA into the nucleus in less than an hour, and its luciferase transfection activity was more than 10 times that by Lipo2000, across multiple cell lines. The GFP chromophore conjugation allowed trackable intracellular delivery and release of DNA in real time via fluorescence imaging. Furthermore, efficient red fluorescent protein (RFP) transfection in zebrafish, with an efficiency of more than 6 times that by Lipo2000, was also achieved. The results not only realized, for the first time, the combination of gene delivery and GFP-simulated light emission, allowing fluorescent tracking and highly efficient gene transfection, but also offered valuable insights into the use of biomimetic chromophore for the development of the next-generation nonviral vectors.

Focused characteristics and effects of light reflected from spherical lipid membrane of giant unilamellar vesicles.

Lipid vesicle is spherical membranous structure with a concave surface on the inside. When a beam of light illuminates a lipid vesicle, the light reflected from the vesicular concave membrane can be focused to have higher intensity and generate enhanced effects. By observing and simulating light reflected from giant unilamellar vesicles (GUVs), the intensity distribution of the light reflected from a spherical concave lipid membrane was investigated. The reflected light had focused characteristics. Its intensity was concentrated 10,000 times and even exceeded the intensity of incident light in a confined region, creating another effective light source in the lipid vesicle. The fluorescence quenching of sulfo-Cy5 encapsulated in spherical GUVs was stronger than that of the outside solution when irradiated by a 632.8 nm laser. When irradiated with ultraviolet light C (UVC), the damage to plasmid DNA encapsulated with spherical GUVs was greater than that of pure plasmid DNA solution and plasmid DNA mixed with lipid membrane fragments. Therefore, in addition to the effects of incident light, the focused light reflected from GUVs could generate incremental effects on encapsulated photoreactive materials if the spherical structure of the lipid membrane was maintained. These results proved that concave lipid membranes of spherical vesicles can focus light and utilize it to generate enhanced effects. The capability of light focusing and its influence on DNA may provide new insights for understanding the function of lipid membranes in cellular life.

Experimental Study of Retention on the Combination of Bifidobacterium with High-Intensity Focused Ultrasound (HIFU) Synergistic Substance in Tumor Tissues.

High intensity focused ultrasound (HIFU) has been recently regarded to be a new type of technique for non-invasive ablation of local tumors and HIFU synergists could significantly improve its therapeutic efficiency. The therapeutic efficiency of HIFU is greatly limited by the low retention of HIFU synergists in the target area and short residence time. This study aimed to explore a method to increase the deposition of HIFU synergists in tumors. Cationic lipid nanoparticle can be used to enhance the HIFU ablation effect, but there is still a problem for it that the deposition amount in the tumor tissue is small and the residence time is short. Bifidobacterium is highly biosafe and can be selectively colonized in the hypoxic zone of tumor tissue. Cationic lipid nanoparticles can be observed in vitro by attachment to bifidobacterium by electrostatic adsorption. And the effect of the proliferation of bifidobacterium in tumor tissues on the retention amount and retention time of cationic lipid nanoparticles in vivo was evaluated. Results showed that the cationic lipid nanoparticles were linked to the surface of Bifidobacterium effectively in vitro, while in vivo, the retention amount and retention time of cationic lipid nanoparticles could be increased by Bifidobacterium in tumor tissues, which provided a new method for improving the therapeutic efficiency of HIFU.