Nanoparticles for detection and diagnosis

Nanoparticle based platforms for identification of chemical and biological agents offer substantial benefits to biomedical and environmental science. These platforms benefit from the availability of a wide variety of core materials as well as the unique physical and chemical properties of these nanoscale materials. This review surveys some of the emerging approaches in the field of nanoparticle based detection systems, highlighting the nanoparticle based screening methods for metal ions, proteins, nucleic acids, and biologically relevant small molecules.     

Melatonin-loaded lecithin/chitosan nanoparticles: Physicochemical characterisation and permeability through Caco-2 cell monolayers

In this study, the potential of lecithin/chitosan nanoparticles (NPs) as a mucoadhesive colloidal nanosystem for transmucosal delivery of melatonin was investigated. The size, zeta potential and melatonin loading of the lecithin/chitosan NPs were investigated as a function of lecithin type (Lipoid S45, S75 and S100) and chitosan content in the preparation. The NPs were characterised by mean diameter and zeta potential ranging between 121.6 and 347.5 nm, and 7.5 and 32.7 mV, respectively, and increasing with lecithin-negative charge and chitosan content in the preparation. Melatonin loadings were up to 7.1%. All NPs were characterised by prolonged release profiles with an initial burst (approximately 25%), followed by a slow release phase. Approximately 60–70% of melatonin was released in 4 h. The permeability of melatonin was investigated using Caco-2 cells as an in vitro model of the epithelial barrier. Melatonin permeability from an NP suspension prepared with Lipoid S45 lecithin and a lecithin-to-chitosan weight ratio (L/C) of 20:1 (sample C2) was significantly improved compared to the permeability of melatonin from the solution (P < 0.001) and from all other NPs investigated (P < 0.05). The results obtained by the cell viability studies (MTT and LDH leakage assays) showed that C2 NP suspension did not induce plasma membrane damage or decrease cell viability and could be safely applied to Caco-2 cells in the concentration range tested (<400 μg/ml).     

Dexamethasone acetate encapsulation into Trojan particles

We have combined the therapeutic potential of nanoparticles systems with the ease of manipulation of microparticles by developing a hybrid vector named Trojan particles. We aim to use this new delivery vehicle for intravitreal administration of dexamethasone. Initialy, dexamethasone acetate (DXA) encapsulation into biodegradable poly(d,l-lactide-co-glycolide) (PLGA) nanoparticles was optimized. Then, Trojan particles were formulated by spray drying 1,2-Dipalmitoyl-sn-Glycero-3-Phosphocholine (DPPC), hyaluronic acid (HA) and different concentrations of nanoparticle suspensions. The effect of nanoparticles concentration on Trojan particle physical characteristics was investigated as well as the effect of the spray drying process on nanoparticles size. Finally, DXA in vitro release from nanoparticles and Trojan particles was evaluated under sink condition. SEM and confocal microscopy show that most of Trojan particles are spherical, hollow and possess an irregular surface due to the presence of nanoparticles. Neither Trojan particle tap density nor size distribution are significantly modified as a function of nanoparticles concentration. The mean nanoparticles size increase significantly after spray drying. Finally, the in vitro release of DXA shows that the excipient matrix provides protection to encapsulated nanoparticles by slowing drug release.     

Acceleration of wound healing by a porous collagen/silk fibroin scaffold carrying zinc oxide nanoparticles北大核心

BACKGROUND: Anti-infective ability determine the success or failure of skin grafting. It is one of the commonly used methods to enhance the anti-infective ability of implants by compounding antibacterial materials with scaffolds. OBJECTIVE:To investigate the effect of porous collagen/silk fibroin scaffolds carrying zinc oxide nanoparticles against infection and inflammation, and to evaluate its effect on wound healing. METHODS: Thirty-two Sprague-Dawley rats with a full-thickness wound on the back skin were randomly divided into two groups. In experimental groiup, porous collagen/silk fibroin scaffolds containing zinc oxide nanoparticles were implanted, while only collagen/silk fibroin scaffolds were implanted in control group. Wound healing was compared between the two groups by measuring residual wound area at 1, 2, 4, 8 weeks post implantation. Hematoxylin-eosin and interleukin 6 immumohistochemical staining were performed at 1, 2, 4 weeks post implantation to observe wound morphology and inflammatory reactions. Meanwhile, expression of interleukin 6 and interleukin 1β was detected by real-time PCR. RESULTS AND CONCLUSION: (1) At 2, 4, 8 weeks post implantation, significantly increased healing rate was observed in the experiment group compared with the control group (P<0.05). (2) Findings from the hematoxylin-eosin staining showed that obvious inflammatory cell infiltration was observed in the control group, but less inflammation with vigorous growth of granulation tissues on the wound surface occurred in the experimental group at 1 week after implantation. Then, the wound repair was basically completed in the experimental group presenting with complete and compact epidermal tissue structure, while scar formation with no skin cover was found in the control group at 4 weeks after implantation. (3) Findings from the interleukin 6 immumohistochemical staining showed that there was interleukin 6 positive expression in both two groups to different extents; at 4 weeks after implantation, the expression of interleukin 6 was remarkably reduced in the control group, but it was still a strong positive expression, while week positive expression of interleukin 6 was observed in the experimental group. (4) Compared with the control group, the mRNA expression of interleukin 6 and interleukin 1β was both lower in the experimental group at 1, 2, 4 weeks after implantation, but there was a significant difference between the two groups at 1 and 2 weeks after implantation (P<0.05). Overall, the porous collagen/silk fibroin scaffold carrying zinc oxide nanoparticles can effectively reduce inflammations following skin injury, and accelerate skin wound healing. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

A lipidic delivery system of a triple vaccine adjuvant enhances mucosal immunity following nasal administration in mice

We previously developed an highly efficacious combination adjuvant comprised of innate defense regulator (IDR)-1002 peptide, poly(I:C) and polyphosphazene (TriAdj). Here we aimed to design and test the in vivo efficacy of a mucoadhesive nasal formulation of this adjuvant. To determine the physical properties of the formulation, the effect of addition of each individual component was characterised by gel electrophoresis and fluorescence quenching using rhodamine-poly(I:C). Cationic liposomes comprised of didodecyl dimethylammonium bromide (DDAB), dioleoyl phosphatidylethanolamine (DOPE) (50:50 or 75:25?mol:mol) and DDAB, L-α-phosphatidylcholine (egg PC) and DOPE (40:50:10?mol:mol:mol) were prepared by the thin-film extrusion method. The liposomes and TriAdj were combined by simple mixing. The formed complex (L-TriAdj) was characterized by dynamic light scattering, zeta potential, and mucin interactions. We found that IDR-1002 peptide, polyphosphazene and poly(I:C) self-assembled in solution forming an anionic complex. Exposure of RAW267.4 mouse macrophage cells to TriAdj alone vs. L-TriAdj indicated that DDAB/DOPE (50:50) and DDAB/EPC/cholesterol (40:50:10) complexation reduced TriAdj toxicity. Next, TriAdj-containing cationic liposomes were prepared at several molar ratios to determine optimal size, stability and desired positive charge. Transmission electron microscopy showed rearrangement of lipid structures on binding of liposomes to TriAdj and to mucin. Stable particles (<200?nm over 24?h) showed mucin binding of DDAB/DOPE?+?TriAdj was greater than DDAB/EPC/DOPE?+?TriAdj. To verify in vivo efficacy, mice were administered the DDAB/DOPE?+?TriAdj complex intranasally with ovalbumin as the antigen, and the immunogenic response was measured by ELISA (serum IgG1, IgG2a, IgA) and ELISpot assays (splenocyte IL-5, IFN-γ). Mice administered adjuvant showed a significantly greater immune response with L-TriAdj than TriAdj alone, with a dose-response proportionate to the triple adjuvant content, and an overall balanced Th1/Th2 immune response representing both systemic and mucosal immunity.     

聚酰胺-胺树状聚合物靶向修饰及放射性核素标记研究进展

纳米分子在体内具有药物载体的特性,与生物靶向分子结合,可实现药物的靶向传递;与放射性核素的结合,可实现在体内靶向分布与靶向聚集的可视化,从而达到对特定病变的显像或治疗作用。本文对近年来新型树状纳米分子聚乙酰胺-胺（PAMAM）的靶向分子修饰及核素标记的研究进展进行综述。     

Effect of vitamin C and iron chelation on diesel exhaust particle and carbon black induced oxidative damage and cell adhesion molecule expression in human endothelial cells

Exposure to particulate matter is associated with oxidative stress and risk of cardiovascular diseases. We investigated if vitamin C and desferrioxamine (iron chelator) altered the levels of oxidative stress and expression of cell adhesion molecules upon exposure to diesel exhaust particles (DEP) and carbon black in cultured human umbilical vein endothelial cells (HUVECs). We found that the particles were only slightly cytotoxic in the high concentration ranges. Particle-induced intracellular reactive oxygen species (ROS) production was attenuated by vitamin C administration or iron chelation and particularly when combined (p < 0.001). Only desferrioxamine protected the DNA from oxidative damage in terms of strand breaks and formamidopyrimidine DNA glycosylase sensitive sites induced by carbon black (p < 0.01). Carbon black and small sized DEP generated from an Euro4 engine increased the surface expression of VCAM-1 and ICAM-1, whereas DEP from an engine representing an old combustion type engine (SRM2975) with larger particles did not affect the expression of cell adhesion molecules. These effects were also attenuated by desferrioxamine but not vitamin C. The study shows that exposure to carbon black and DEP in HUVECs can generate both oxidative stress and expression of cell surface adhesion molecules and that these effects can in part be attenuated by vitamin C and desferrioxamine.     

Cellular uptake mechanism and knockdown activity of siRNA-loaded biodegradable DEAPA-PVA-g-PLGA nanoparticles

Efficient downregulation of gene expression depends on the uptake, intracellular distribution and efficient release of siRNA from their carrier. Therefore, the cellular uptake behavior and mechanism and intracellular localization of siRNA-loaded biodegradable nanoparticles were investigated.A biodegradable polymer, composed of poly(vinyl alcohol) (PVA) modified with diamine moieties and grafted with PLGA, abbreviated as DEAPA-PVA-g-PLGA, was used for the preparation of siRNA-loaded nanoparticles by solvent displacement. Particle sizes and morphology were determined by dynamic light scattering (DLS) and scanning electron microscopy (SEM). The dependence of particle uptake into H1299-EGFP cells (lung cancer cells expressing green fluorescent protein) on both incubation time and temperature was studied by flow cytometry. Inhibition experiments focusing on clathrin- or caveolae-mediated uptake or uptake by macropinocytosis were performed. The intracellular localization was investigated by confocal laser scanning microscopy. The GFP knockdown efficiency was determined in vitro to establish the potential of the nanoparticles for the downregulation of gene expression.Nanoparticles with diameters of 120-180 nm were successfully generated. In contrast to the uptake of standard PEI-polyplexes, which increased continuously over a period of 4 h, nanoparticle uptake was complete within 2 h. A decrease in particle uptake at 4 °C (in comparison with 37 °C) suggests an active uptake process. Inhibition experiments revealed the predominance of clathrin-mediated uptake for siRNA-loaded nanoparticles. The siRNA-loaded nanoparticles could be clearly located within cells, mainly in intracellular vesicles. Particle uptake could be increased by the addition of lung surfactant to the formulation. Bioactivity in terms of successful GFP knockdown in vitro was demonstrated and could be further optimized by the use of surfactant-modified particles.In conclusion, a high and rapid cellular uptake was shown for siRNA-loaded nanoparticles. Cell internalization is based on an energy-dependent and predominantly clathrin-mediated process. Particle localization in endosomes and lysosomes was demonstrated. Evidence for the efficient delivery of bioactive siRNA and specific GFP knockdown provides a solid basis for the application of DEAPA-PVA-g-PLGA-based particles for gene silencing in vivo.     

因子设计-效应面法优化紫杉醇聚乳酸-羟基乙酸纳米粒处方和制备工艺

目的:优化紫杉醇聚乳酸-羟基乙酸(PLGA)纳米粒处方和制备工艺.方法:以PLGA为载体,采用溶剂扩散法制备紫杉醇PLGA纳米粒,用32满因子设计实验,考察因素PLGA在有机相中的浓度和理论载药量对纳米粒的粒径、载药量和包封率的影响,实验数据分别采用线性方程和二次多项式拟合,根据最佳数学模型绘制效应面并选出最优处方.结果:2个影响因素和3个评价指标之间存在定量关系,最优处方为:紫杉醇的理论载药量为9.09%、有机相中PLGA浓度为2%,制备得到的纳米粒粒径为281 nm,实际载药量为7.73%,包封率为57.43%.结论:采用因子设计-效应面法完成了紫杉醇纳米给药系统的多目标同步优化.     

Distinct immunomodulatory effects of a panel of nanomaterials in human dermal fibroblasts

There are many efforts in understanding the effects of nanoparticles on cell viability and metabolism, however, not much is known regarding the distinct molecular mechanisms of inflammation and cellular stress using low dosing concentrations. To address this gap in the literature, we utilized a novel experimental design that specifically probes the effects of a panel of commonly studied engineered nanomaterials along immunomodulatory pathways, including NF-κB. The panel of particles selected for this study included quantum dot nanocrystals, titanium dioxide, hydroxylated fullerenes, and silver nanoparticles. Cell viability, antioxidant activity, select messenger RNA, and protein modulation were studied in primary human dermal fibroblasts (HDF) and NF-κB knockdown HDF cells. Inflammatory and non-inflammatory immune responses were measured using protein and real-time PCR array analysis from HDF cells exposed to sub-lethal concentrations of nanoparticles. Differences in cellular response to nanoparticles in protein and antioxidant experiments were evident in NF-κB knockdown cells. The methods used in the study, along with the resultant data sets, serve as a potential model for studying the complex pathway-specific biochemical responses in cell and tissue systems associated with nanoparticle exposures.