Synthesis and Characterization of Glycol Chitosan DNA Nanoparticles for Retinal Gene Delivery

Given the number of monogenic ocular diseases and the number of non‐monogenic degenerative ocular diseases for which gene therapy is considered as a treatment, the development of effective therapeutic delivery strategies for DNA is a critical research goal. In this work, nonviral nanoparticles (NPs) composed of glycol chitosan (GCS) and plasmid DNA (pDNA) were generated, characterized, and evaluated. These particles are stable, do not aggregate in saline, are resistant to DNases, and have a hydrodynamic diameter of approximately 250 nm. Furthermore, the plasmid in these NPs was shown to maintain its proper conformation and can be released and expressed inside the cell. To determine whether these NPs would be suitable for intraocular use, pDNA carrying the ubiquitously expressed CBA‐eGFP expression cassette was compacted and subretinally injected into adult wild‐type albino mice. At day 14 post‐injection (PI), substantial green fluorescent protein (GFP) expression was observed exclusively in the retinal pigment epithelium (RPE) in eyes treated with GCS NPs but not in those treated with uncompacted pDNA or vehicle (saline). No signs of gross retinal toxicity were observed, and at 30 days PI, there was no difference in electroretinogram function between GCS NP‐, pDNA‐, or vehicle‐treated eyes. These results suggest that with further development, GCS NPs could be a useful addition to the available repertoire of genetic therapies for the treatment of RPE‐associated diseases.     

聚乳酸纳米颗粒的制备与表征

采用改良沉淀法制备了聚乳酸(PLA)纳米颗粒,采用响应面分析法建立数学模型对工艺条件进行优化. 结果表明,影响PLA颗粒制备的主要因素有PLA质量、油相中乙醇含量和油相体积,以平均粒径最小为目标,得到最优工艺条件为：聚乳酸质量76.82 mg,油相中乙醇含量40%(j),油相体积19.49 mL. 按最优工艺条件所制PLA颗粒的粒径为109.1 nm,多分散系数PDI值为0.084,与预测值偏差较小. PLA纳米颗粒为球形实心颗粒,分散性良好且粒径均一. PLA纳米颗粒能被巨噬细胞摄取,有望作为免疫佐剂.     

羟基磷灰石空心纳米球的制备及表征

采用WOW复乳法制备羟基磷灰石,内水相为(NH4)2HPO4水溶液,中间油相为易挥发的环己烷,外水相为Ca(NO3)2.4H2O水溶液。分析了反应中传质机理为Ca2+扩散到内水相,在碱性下与内水相的HPO42-反应生成羟基磷灰石。通过XRD、FT-IR、SEM、TEM分析了产物的成分和形貌,证实产物为300nm左右的羟基磷灰石空心球。讨论了反应温度对纳米球形貌的影响,当温度为10℃时发生界面反应,得到空心的纳米球。     

Carbon Nanotube-Mediated Plasmid DNA Delivery in Rice Leaves and Seeds

CRISPR-Cas gene editing technologies offer the potential to modify crops precisely; however, in vitro plant transformation and regeneration techniques present a bottleneck due to the lengthy and genotype-specific tissue culture process. Ideally, in planta transformation can bypass tissue culture and directly lead to transformed plants, but efficient in planta delivery and transformation remains a challenge. This study investigates transformation methods that have the potential to directly alter germline cells, eliminating the challenge of in vitro plant regeneration. Recent studies have demonstrated that carbon nanotubes (CNTs) loaded with plasmid DNA can diffuse through plant cell walls, facilitating transient expression of foreign genetic elements in plant tissues. To test if this approach is a viable technique for in planta transformation, CNT-mediated plasmid DNA delivery into rice tissues was performed using leaf and excised-embryo infiltration with reporter genes. Quantitative and qualitative data indicate that CNTs facilitate plasmid DNA delivery in rice leaf and embryo tissues, resulting in transient GFP, YFP, and GUS expression. Experiments were also initiated with CRISPR-Cas vectors targeting the phytoene desaturase (PDS) gene for CNT delivery into mature embryos to create heritable genetic edits. Overall, the results suggest that CNT-based delivery of plasmid DNA appears promising for in planta transformation, and further optimization can enable high-throughput gene editing to accelerate functional genomics and crop improvement activities.     

基因治疗用质粒DNA的制备工艺

目的探索基因治疗用质粒DNA的制备工艺。方法以CaCl2沉淀法去除大分子RNA,Q-Sepharose和SOURCE两步离子交换层析法去除染色体DNA、小分子RNA、蛋白质等杂质,并对各步骤样品进行检测。结果质粒pIRVP3HNIL18终产品的产量为1.638mg质粒DNA/g细菌,纯度为1.839,相对回收率为12.55%,蛋白质含量为0.0028mg/ml,未检测到RNA和染色体DNA。结论此工艺可有效去除质粒DNA制备过程中产生的杂质,可应用于药剂水平质粒DNA的制备。     

Efficient Delivery of Plasmid DNA Using Cholesterol-Based Cationic Lipids Containing Polyamines and Ether Linkages

Cationic liposomes are broadly used as non-viral vectors to deliver genetic materials that can be used to treat various diseases including cancer. To circumvent problems associated with cationic liposome-mediated delivery systems such as low transfection efficiency and serum-induced inhibition, cholesterol-based cationic lipids have been synthesized that resist the effects of serum. The introduction of an ether-type linkage and extension of the aminopropyl head group on the cholesterol backbone increased the transfection efficiency and DNA binding affinity compared to a carbamoyl-type linkage and a mono aminopropyl head group, respectively. Under optimal conditions, each liposome formulation showed higher transfection efficiency in AGS and Huh-7 cells than commercially available cationic liposomes, particularly in the presence of serum. The following molecular structures were found to have a positive effect on transfection properties: (i) extended aminopropyl head groups for a strong binding affinity to plasmid DNA; (ii) an ether linkage that favors electrostatic binding to plasmid DNA; and (iii) a cholesterol backbone for serum resistance.     

碳纳米管负载铂-二氧化钌纳米颗粒催化剂的制备及表征

以多壁碳纳米管(MWNTs)为载体,采用化学还原法制备了多壁碳纳米管负载的铂-二氧化钌纳米颗粒催化剂(Pt-RuO2/MWNTs),并利用透射电镜(TEM)、电化学法等技术对该催化剂形貌和电化学性质进行了表征。结果表明,大小约为4～5nm的金属纳米颗粒均匀地分散在碳纳米管上,同时考察了该催化剂在铁氰化钾溶液中的循环伏安行为以及对该催化剂进行了电化学阻抗分析。     

明胶纳米颗粒的制备及表征

以B型明胶为原料,戊二醛为交联剂,通过改进的两步去溶剂法制备了较小粒径（约100nm）的明胶纳米颗粒(Gelatin nanoparticles, GNPs);研究了分散体系浓度、去溶剂化试剂、交联剂用量的影响。结果表明：降低明胶在二次分散体系中的浓度,制得的GNPs的尺寸更小且分布均匀。残留的去溶剂化试剂丙酮必须去除,因为它会使GNPs分散性变差。在明胶质量浓度为8 g/L的分散液中,仅用质量分数为4.5%的戊二醛(与明胶质量比)即可制得稳定的GNPs,其粒径为50~150 nm,PDI （polydispersity index）约为0.14,Zeta电位绝对值< 30 mV。AFM和 TEM测试结果表明GNPs呈光滑球形,为边缘部分比核心更为疏松的非均质结构。     

色氨酸-银纳米微粒的制备与表征

在微乳液中制备出色氨酸-银纳米微粒,并用X射线衍射(XRD),原子力显微镜(AFM),红外光谱(IR)对其结构进行表征。XRD结果表明,色氨酸-银晶粒的平均粒径为27.8nm;从AFM可以看出,色氨酸-银纳米微粒分散性较好;红外可见光谱表明,色氨酸中-COOH,-NH2以及吲哚基上的氮对银(Ⅰ)有直接的配位作用。     

Preparation and Characterization of Soap-free Cationic Fluorinated Poly-styrene-acrylate Latex with Core-shell Structure

Soap-free cationic fluorinated Poly-styrene-acrylate latex particles with core-shell structure were synthesized by seeded semi-continues emulsion polymerization in the presence of water soluble cationic monomer Methacryloxyethyltrimethyl ammonium chloride (MADAC) and using ethanol as co-solvent. The effects of MADAC dosage and ethanol content on the stability of polymerization process and the properties of the latex particles were studied. The chemical component of the polymer was analyzed by Fourier transform infrared (FTIR). The surface element composition of the prepared copolymer film was analyzed by X-ray photoelectron spectroscopy (XPS), the micro-structure of the prepared latex particles were observed by transmission electron microscopy (TEM). The result shows that cationic soap-free fluorinated poly-styrene-acrylate emulsion with a core-shell structure can be prepared when monomer DAMAC is 6.0 wt% and ethanol is 7.5 wt%.     

Silver Nanoparticles Modified by Carbosilane Dendrons and PEG as Delivery Vectors of Small Interfering RNA

The fact that cancer is one of the leading causes of death requires researchers to create new systems of effective treatment for malignant tumors. One promising area is genetic therapy that uses small interfering RNA (siRNA). These molecules are capable of blocking mutant proteins in cells, but require specific systems that will deliver RNA to target cells and successfully release them into the cytoplasm. Dendronized and PEGylated silver nanoparticles as potential vectors for proapoptotic siRNA (siMCL-1) were used here. Using the methods of one-dimensional gel electrophoresis, the zeta potential, dynamic light scattering, and circular dichroism, stable siRNA and AgNP complexes were obtained. Data gathered using multicolor flow cytometry showed that AgNPs are able to deliver (up to 90%) siRNAs efficiently to some types of tumor cells, depending on the degree of PEGylation. Analysis of cell death showed that complexes of some AgNP variations with siMCL-1 lead to ~70% cell death in the populations that uptake these complexes due to apoptosis.     

含氟核壳纳米粒子的制备及其表征

通过半连续的加料工艺制备了聚甲基丙烯酸三氟乙酯（PTFEMA）为壳,以聚丙烯酸甲酯（PMA）为核的纳米核壳乳液。并通过SEM、TEM、DSC、DLS等对合成的乳液进行表征,结果表明：所合成的粒子具有核壳结构,粒子形貌较为规整,且成球效果很好。     

Preparation, Characterization and Catalytic Activity of Palladium Nanoparticles Encapsulated in SBA-15

A simple adsorption method was studied for the preparation of SBA-15-encapsulated palladium nanoparticles. This method employed the SBA-15 with organic template removed by ethanol extraction for the adsorption of cationic Pd precursors in alkaline solution followed by calcination and H₂ reduction. The pH of the solution was found to be a critical factor in determining the palladium content and the ordered mesoporous structure. Our characterizations revealed that the Pd nanoparticles prepared by this method were located inside the mesoporous channels and were quite uniform in size (mostly 3–4 nm). The SBA-15-encapsulated uniform Pd nanoparticles were efficient catalysts for solvent-free aerobic oxidation of alcohols.     

Preparation and Characterization of Gelatin Nanofibers Containing Silver Nanoparticles

Ag nanoparticles (NPs) were synthesized in formic acid aqueous solutions through chemical reduction. Formic acid was used for a reducing agent of Ag precursor and solvent of gelatin. Silver acetate, silver tetrafluoroborate, silver nitrate, and silver phosphate were used as Ag precursors. Ag⁺ ions were reduced into Ag NPs by formic acid. The formation of Ag NPs was characterized by a UV-Vis spectrophotometer. Ag NPs were quickly generated within a few minutes in silver nitrate (AgNO₃)/formic acid solution. As the water content of formic acid aqueous solution increased, more Ag NPs were generated, at a higher rate and with greater size. When gelatin was added to the AgNO₃/formic acid solution, the Ag NPs were stabilized, resulting in smaller particles. Moreover, gelatin limits further aggregation of Ag NPs, which were effectively dispersed in solution. The amount of Ag NPs formed increased with increasing concentration of AgNO₃ and aging time. Gelatin nanofibers containing Ag NPs were fabricated by electrospinning. The average diameters of gelatin nanofibers were 166.52 ± 32.72 nm, but these decreased with the addition of AgNO₃. The average diameters of the Ag NPs in gelatin nanofibers ranged between 13 and 25 nm, which was confirmed by transmission electron microscopy (TEM).     

二聚阳离子表面活性剂改性蒙脱土的制备和表征

合成了二聚阳离子表面活性剂GeminiC12,用核磁共振氢谱(1HNMR)表征了它的结构,并用它作为有机插层剂应用于蒙脱土的改性处理。红外光谱(FTIR)、热重分析(TGA)表明,GeminiC12已插层到蒙脱土片层间。X射线粉末衍射(XRD)表明,插层后蒙脱土层间距从1 19nm增加到3 8nm,是十六烷基三甲基溴化铵(CTAB)处理效果的两倍。沉降实验表明,改性后蒙脱土在苯乙烯和石蜡中形成凝胶体系,表现出很好的相容性和分散性,这种改性效果优于目前常用的CTAB处理效果,更有利于聚合物或其单体进入蒙脱土层间形成纳米复合材料。     

Co-Formulation of Amphiphilic Cationic and Anionic Cyclodextrins Forming Nanoparticles for siRNA Delivery in the Treatment of Acute Myeloid Leukaemia

Non-viral delivery of therapeutic nucleic acids (NA), including siRNA, has potential in the treatment of diseases with high unmet clinical needs such as acute myeloid leukaemia (AML). While cationic biomaterials are frequently used to complex the nucleic acids into nanoparticles, attenuation of charge density is desirable to decrease in vivo toxicity. Here, an anionic amphiphilic CD was synthesised and the structure was confirmed by Fourier-transform infrared spectroscopy (FT-IR), Nuclear Magnetic Resonance (NMR), and high-resolution mass spectrometry (HRMS). A cationic amphiphilic cyclodextrin (CD) was initially used to complex the siRNA and then co-formulated with the anionic amphiphilic CD. Characterisation of the co-formulated NPs indicated a significant reduction in charge from 34 ± 7 mV to 24 ± 6 mV (p < 0.05) and polydispersity index 0.46 ± 0.1 to 0.16 ± 0.04 (p < 0.05), compared to the cationic CD NPs. Size was similar, 161–164 nm, for both formulations. FACS and confocal microscopy, using AML cells (HL-60), indicated a similar level of cellular uptake (60% after 6 h) followed by endosomal escape. The nano co-formulation significantly reduced the charge while maintaining gene silencing (21%). Results indicate that blending of anionic and cationic amphiphilic CDs can produce bespoke NPs with optimised physicochemical properties and potential for enhanced in vivo performance in cancer treatment.     

溶胶凝胶法制备Ni掺杂AZO纳米粉体及其表征

利用溶胶凝胶法制备Ni掺杂AZO(NAZO)纳米粉体。通过TG-DSC、XRD、TEM、XPS等分析手段,研究掺杂浓度和煅烧温度对纳米粉体结构和电学性能影响规律。结果表明:在600℃下煅烧2 h,NAZO纳米粉体具有六角纤锌矿结构,粉体的粒径约25~35 nm,主要呈现球状和棒状结构。当铝掺杂摩尔百分比为1 mol%,镍掺杂摩尔百分比为1.5 mol%时,纳米粉体的电阻率为4.309 M?·cm。     

Kidney-Protector Lipidic Cilastatin Derivatives as Structure-Directing Agents for the Synthesis of Mesoporous Silica Nanoparticles for Drug Delivery

Mesoporous silica nanomaterials have emerged as promising vehicles in controlled drug delivery systems due to their ability to selectively transport, protect, and release pharmaceuticals in a controlled and sustained manner. One drawback of these drug delivery systems is their preparation procedure that usually requires several steps including the removal of the structure-directing agent (surfactant) and the later loading of the drug into the porous structure. Herein, we describe the preparation of mesoporous silica nanoparticles, as drug delivery systems from structure-directing agents based on the kidney-protector drug cilastatin in a simple, fast, and one-step process. The concept of drug-structure-directing agent (DSDA) allows the use of lipidic derivatives of cilastatin to direct the successful formation of mesoporous silica nanoparticles (MSNs). The inherent pharmacological activity of the surfactant DSDA cilastatin-based template permits that the MSNs can be directly employed as drug delivery nanocarriers, without the need of extra steps. MSNs thus synthesized have shown good sphericity and remarkable textural properties. The size of the nanoparticles can be adjusted by simply selecting the stirring speed, time, and aging temperature during the synthesis procedure. Moreover, the release experiments performed on these materials afforded a slow and sustained drug release over several days, which illustrates the MSNs potential utility as drug delivery system for the cilastatin cargo kidney protector. While most nanotechnology strategies focused on combating the different illnesses this methodology emphasizes on reducing the kidney toxicity associated to cancer chemotherapy.     

Structure and Property of Cationic Hydroxyethyl Cellulose

Through the macromolecule reaction, hydroxyethyl cellulose (HEC) was modified by quaternary ammonium monomer (GTAC) and the cationic HEC(C-HEC) with optimized viscosity was synthesized successfully. The structure of C-HEC was characterized by FTIR and Kjeldahl nitrogen content measurement. The introduction of cationic group onto the molecules of HEC resulted in the increase of the hydrophilicity of HEC. The dependence of the aqueous solution properties of C-HEC on the polymer concentration, GTAC dosage, additional electrolyte, temperature and shear rate were studied comprehensively.