Starburst^(TM) Polyamidoamine Dendrimers介导的绿色荧光蛋白基因在小鼠体内的表达分布及对恶性疟DNA疫苗ES312免疫原性的影响

目的研究StarburstTMPolyamidoamine(PAMAM)dendrimers介导绿色荧光蛋白基因在小鼠体内的表达分布及对恶性疟DNA疫苗ES312免疫原性的影响。方法经股四头肌给小鼠注射StarburstTMPAMAMdendrimers与绿色荧光蛋白基因或恶性疟DNA疫苗ES312的复合物。通过流式细胞术、Westernblot和间接免疫荧光方法检测绿色荧光蛋白的表达分布和表达量。用ELISA方法检测恶性疟DNA疫苗ES312的免疫原性。结果在注射StarburstTMPAMAMdendrimers/DNA复合物后2h～7d内,StarburstTMPAMAMdendrimers携带的绿色荧光蛋白基因的表达在小鼠的心、肝、脾、肺、肾、脑、肌肉中均能被检测到,在肾总蛋白中所占百分比最高,且各器官管腔内皮细胞中的绿色荧光蛋白表达量较高。小鼠的脑和肾中,在StarburstTMPAMAMdendrimers的存在下,绿色荧光蛋白基因的表达量明显高于裸DNA的表达量。经StarburstTMPAMAMdendrimers携带的恶性疟DNA疫苗ES312诱导的抗体反应水平明显高于单纯裸DNA疫苗所诱导的抗体水平。结论StarburstTMPAMAMdendrimers作为一种低抗原性、非病毒的新型DNA载体,能够明显提高裸DNA在体内的表达水平,诱导产生很强的体液免疫反应。但是,StarburstTMPAMAMdendrimers/DNA复合物在各组织器官中的广泛分布,尤其是透过血脑屏障的能力,已超出疫苗载体的适用范围。作为DNA疫苗载体,要对现有StarburstTMPAMAMdendrimers分子的结构做进一步的修饰。

Electrochemistry of polyamidoamine dendrimers ester gel electrolytes

This paper described the first example of polyamidoamine dendrimers ester(PAMAM) used as a gel electrolyte with a short-chain polyethylene glycol (MPEG-400) as aplasticizer. The polymer films are solid and sticky. Background cyclic voltammetry (CV) shows apotential window between +0.7 and -0.7 V vs. Ag/AgCl. The voltammetry of ferrocene and7,7,8,8-tetracyanoquinodimethane (TCNQ) indicates that diffusion coefficients are in the range of10^(-8) -10^(-9) cm^2/s. Ionic conductivities are approximately 10^(-6) S/cm. Similar films usingdimethyl sulfoxide (DMSO) as a plasticizer instead of MPEG-400 have demonstrated ionicconductivities of 10^(-4) S/cm and reversible voltammetry. However, UV spectropho-tometry shows that70% of the DMSO is lost under vacuum, indicating the difficulty in quantifying the DMSO contentwhen exposed to vacuum.

A new strategy of enhancing absorptivity and safety for evodiamine: polyamidoamine derivatives modified by polyethylene glycol chain

Background:Traditional Chinese medicine involves complex ingredients and mixtures of ingredients that often exhibit low bioavailability,and excipients are often lacking to increase the absorption-enhancing effects.This study modified the generation 4 polyamidoamine dendrimer with polyethylene glycol of different molecular weights(5000,2000,1000)to form a series of polyamidoamine-co-polyethylene glycol(PAMAM-co-PEG)as a novel class of oral absorption enhancers.Evodiamine,the major alkaloid found in the traditional Chinese medicine Wu Zhu Yu(Fructus Evodiae),was used as a model drug to verify the absorption-enhancing effects and the safety of this alkaloid.Methods:This study utilized the solubility determination method documented in the Pharmacopoeia of the People’s Republic of China(2015 edition)and the D0 values recommended in the US FDA guidelines to comprehensively evaluate the solubility of evodiamine.The permeability of evodiamine was assessed using the apparent permeability coefficient in experiments based on in vitro cell models.Multiple aspects of the biological safety of PAMAM-co-PEG were explored using the MTT assay,LDH assay,and total protein release of the rat intestinal tract.Moreover,the absorption-enhancing effects of PAMAM-co-PEG at different molecular weights on evodiamine were verified via the use of in vitro cell models and in vivo intestinal loop circulation experiments with rats.Results:Evodiamine exhibited low solubility and permeability and was classified into class IV compounds using the biopharmaceutical classification system.PAMAM-co-PEG 2000 demonstrated improvement in the biosafety and absorption-enhancement effect of evodiamine at a specific concentration.This study showed that 0.05%(w/v)of PAMAM-co-PEG 2000 increased the cumulative penetration of evodiamine via cell transport by 1.32 times,and 0.10%(w/v)of PAMAM-co-PEG 2000 increased the area under curve value of evodiamine by 1.31 times.Conclusion:Evodiamine possesses low solubility and permeability and leads to poor oral bioavailability and a certain degree of cytotoxicity.PAMAM-co-PEG 2000 was found to be a potentially safe and efficient oral absorption enhancer.The results of this study might create a foundation for the development of novel excipients suitable for the complex active ingredients of traditional Chinese medicine.

Hyperbranched polyamidoamine protective layer with phosphate and carboxyl groups for dendrite-free Zn metal anodes

Zinc metal is regarded as one of the most promising anodes for Zn-based batteries in next-generation energy storage systems.However,the dendrite growth and interfacial corrosion lead to poor reversibility and cycle life of Zn anodes.Herein,we synthesize a 2-phosphate-1,2,4-butane tricarboxylic acid modified hyperbranched polyamidoamine containing rich terminal groups of phosphate and carboxyl(HPC)as modified layer for the Zn anodes.Importantly,the in situ acid-etching promotes the exposure of(002)Zn plane and the generated salt-polymer complexes could be adhered to the Zn anodes tightly.This greatly favors the uniform deposition of Zn and inhibits interfacial corrosion.Consequently,stable HPC@Zn anode plating/stripping for over 1200 h at a high areal capacity of 4 mAh/cm^(2)and a current density of 4 m A/cm^(2)is obtained.This study provides a new avenue of hyperbranched polymer in interfacial design for highly reversible and stable Zn metal anodes.

Anti-angiogenesis effect of generation 4 polyamidoamine/vascular endothelial growth factor antisense oligodeoxynucleotide on breast cancer in vitro

Objective:To study the effects of the generation 4 polyamidoamine/vascular endothelial growth factor antisense oligodeoxynucleotide(G4PAMAM/VEGFASODN) compound on the expressions of vascular endothelial growth factor(VEGF) and its mRNA of breast cancer cells and on the inhibition of vascular endothelial cells. Methods:We examined the morphology of G4PAMAM/VEGFASODN compound and its pH stability,in vitro transfection efficiency and toxicity,and the expressions of VEGF and its mRNA. Methyl thiazolyl tetrazolium assay was used to detect the inhibitory function of the compound on vascular endothelial cells. Results:The compound was about 10 nm in diameter and was homogeneously netlike. From pH 5 to 10,it showed quite a buffered ability. The 48-h transfection rate in the charge ratio of 1:40 was 98.76%,significantly higher than that of the liposome group(P<0.05). None of the transfection products showed obvious toxicity on the cells. The expressions of both VEGF protein and its mRNA after G4PAMAM/VEGFASODN transfection decreased markedly. Conclusion:With a low toxicity,high safety,and high transfection rate,G4PAMAM/VEGFASODN could be a promising gene vector. Specifically,it inhibits VEGF gene expression efficiently,laying a basis for further in vivo animal studies.

Syntheses of polyamidoamine dendrimers starting from a hexadimensional core and application in gene transfer

Polyamidoamine dendrimers using inositol as a core were designed, synthesized and characterized. The electrostatics interaction of dendrimer with DNA causes the condensation of DNA to form DNA/dendrimer complexes. The sizes of complexes prepared from dendrimer G6 and pRE Luc at N/P ratio 10: 1 range from 80 to 300 nm. Dendrimers G3—G6 are able to efficiently deliver pRE Luc and pSV β-gal gene into HeLa and HEK 293 cells. The transfection efficiencies of G5 and G6 are much higher than that of poly L-lysine (PLL) and comparable to that of the commercial branched polyethylenimine (PEI, Mw 25000), whereas the cytoctoxicities of G5 and G6 are lower than that of PLL.

Multiarm Star Poly(ε-caprolactone) with Hyperbranched Polyamidoamine as Core Capable of Selective Accommodating Cationic or Anionic Guests

Hyperbranched polyamidoamines （HPAs） were directly employed as macroinitiators to initiate the Sn（Oct）2 catalyzed ring-opening polymerization of ε-caprolactone （CL）, resulting in multiarm star copolymers with poly（6- caprolactone） （PCL） as shells and HPA as core （HPA-b-PCL）. From 1H-NMR characterization it was deduced that both the primary amines and the secondary amide groups of HPAs could initiate the CL polymerization, and the initiation efficiency increased when more CL monomers were fed. The average arm-numbers of the obtained stars were in the range of 115-353. Differential scanning calorimetry measurements demonstrated that the melting and crystallization temperatures, fusion and crystallization enthalpy and the degree of crystallinity of the star polymers increased as the PCL arm length increased. HPA- b-PCL stars could be used as nanocarriers to efficiently accommodate anionic dyes at acidic condition, while load cationic dyes at basic condition. Compared with the dye-loading behavior of multiarrn star PCL with the neutral hyperbranched polyglycerol as core, it was deduced that HPA-b-PCL nanocarriers accommodated anionic dyes using the HPA core, while loaded cationic dyes using both the HPA core and the PCL shell. Dynamic light scattering analyses also supported such deduction. Furthermore, HPA-b-PCL nanocarriers could selectively load the anionic Eosin Y or the cationic methylene blue from their mixture at pH = 6 or 9, respectively, realizing their separation.

Low generational polyamidoamine dendrimers to enhance the solubility of folic acid:A‘‘dendritic effect’’investigation

Low generational（G0–G2,G for generation） polyamidoamine（PAMAM） dendrimers were investigated as enhancers to improve the aqueous solubility of folic acid at pH 11 and pH 5.In these two cases,the solubility of folic acid increases with both the dendrimer concentration and generation.However,the solubilization mechanism is different.The electrostatic interaction between the primary amines of dendrimers and the ionized carboxylic groups of folic acid dominates the dissolution process at pH 11 while the increase of the solubility of folic acid at pH 5 is attributed to the hydrophobic encapsulation inside the dendrimer molecules.In addition,for comparison ethylenediamine was used as a small molecule control to examine the ‘‘dendritic effect＇＇ in the dendrimer-related solubilization process.Interestingly,PAMAM dendrimers exhibit,at pH 5,a significant superiority over ethylenediamine in enhancing solubility,whereas this ‘‘dendritic effect＇＇ cannot be observed under the basic condition.

Ab initio Study of Complexation Process between Poly（amido-amine） and Nano-Silicon Dioxide

To understand better the molecular-level details of ≡Si＋ （SC） or ≡SiO- （SOA） ion group to -NH2 teminated poly（amido-amine） dendrimers in the gas phase, density functional theory is used to optimize the minimum energy and transition state structures with UB3LYP/6- 311G（d） and HF/6-31G levels. The tertiary amine nitrogen and the amide oxygen are found to be the most favorable binding sites. The activation energies of the different active sites and the reaction steps of SC and/or SOA ion group and the amide sites are also analyzed. The stable compounds are formed via the electrostatic interaction and the coordination effect. The orientation of the amide O and the rotation of the branches minimizes the energy of the whole system.

Preparation, optical properties and cell staining of water soluble amine-terminated PAMAM G2.0-Au nanocomposites

The solution chemical and optical characteristics of formation of amine-terminated polyamidoamine dendrimer G2.0（NH2-PAMAM G2.0）-Au nanocomposites in the aqueous solution of NH2-PAMAM G2.0 at various mole ratios of Au（Ⅲ） to NH2-PAMAM G2.0 were studied by both UV-visible spectrometry and fluorospectrometry. The NH2-PAMAM G2.0-Au nanocomposites, with a type of structure in which one Au nanoparticle is surrounded by several NH2-PAMAM G2.0 dendrimers, emit strong bluish violet fluorescence, and are uniform, water soluble and biocompatible as well as very stable in frozen conditions. The size of gold nanoparticles in the nanocomposites is about 2.5 nm and decreases with the increase of NH2-PAMAM G2.0 concentration. The NH2-PAMAM G2.0 plays an important role in acting as host or micro-reactor for Au（Ⅲ） before Au（Ⅲ） reduction and acting as dispersant and stabilizer for gold nanoparticles after Au（Ⅲ） reduction. Preliminary experiments of cells staining to human embryonic lung fibroblast cell lines show that the NH2-PAMAM G2.0-Au nanocomposites can be used as optical imaging markers for bioanalyses and medical diagnoses.

Poly(styrene-co-divinylbenzene)-PAMAM-IDA chelating resin: Synthesis, characterization and application for Ni(II) removal in aqueous

PS-PAMAM-IDA chelating resins were prepared by low-generations of polyamidoamine(PAMAM) and then chloroacetic acid functionalizing commercially available ammoniated polystyrene matrix, to preconcentrate Ni2+ from synthetic aqueous samples. Different generations of PAMAM were used to obtain different chelating resins, PS-IDA, PS-1.0G PAMAM-IDA and PS-2.0G PAMAM-IDA. The synthesized resins were characterized by FTIR and elemental analysis. The effect of solution pH, kinetic studies, resin loading capacity, matrix effects etc., on metal ion adsorption to adsorbent phase, were studied by batch method. The PS-1.0G PAMAM-IDA resin was the most excellent adsorbents, with a maximum adsorption capacity of(24.09±1.79) mg/g for Ni2+ ion at pH=7. The interpretation of the equilibrium data was given by Langmuir isotherms model, and the correlation coefficient values for PS-IDA, PS-1.0G PAMAM-IDA and PS-2.0G PAMAM-IDA resins were 0.992, 0.994 and 0.987, respectively.

Effect of VEGF-targeted antisense gene therapy on retinoblastoma cell line SO-RB50 in vitro and in vivo

AIM: To evaluate the possibility of generation 4 polyamidoamine (G4PAMAM) dendrimers acting as the delivery system of vascular endothelial growth factor (VEGF) antisense oligodeoxynucleotides (VEGFASODN), and to investigate the anti-tumor effect of G4PAMAM/VEGFASODN complex on the cultured cells and the mouse tumor xenograft model. METHODS: The transfection efficiency was assessed by Row cytometry (FCM). Thiazolyl tetrazolium (MU) assay was performed to determine the relative growth rate (RGR) of the cells after transfection. Then a mouse tumor xenograft model of human retinoblastoma was established. Different interventions were given to the mice by intratumoral injection and the tumor growth was monitored. The expression of VEGF mRNA was detected by reverse transcription PCR (RT-PCR), the expression of VEGF protein was determined by western blot analysis, and the microvessel density (MVD) was measured by immunohistochemistry (IHC) staining. RESULTS: G4PAMAM/VEGFASODN exhibited a high transfection rate in vitro, and the transfection rates of different doses of G4PAMAM/VEGFASODN groups increased with higher doses. This effect was accompanied by a dose-depended reduction in cell viability. The tumor growth in the tumor-bearing athymic mice was significantly inhibited in the G4PAMAM/VEGFASODN group. The expressions of VEGF mRNA and protein were obviously inhibited in the G4PAMAM/VEGFASODN group (p<0.05), and the MVD of the G4PAMAM/VEGFASODN group was lower than that of the other groups(p<0.05). CONCLUSION: VEGFASODN can be delivered into the cultured and transplanted retinoblastoma cells efficiently by G4PAMAM, suppress the expressions of VEGF mRNA and protein, and reduce the MVD of tumor tissues. The G4PAMAM/VEGFASODN complex has antitumor properties vitro and in vivo.

Design of Dendrimer Modified Carbon Nanotubes for Gene Delivery

To investigate the efficiency of polyamidoamine dendrimer grafted carbon nanotube （dendrimer-CNT） mediated entrance of anti-survivin oligonucleotide into MCF-7 cells, and its effects on the growth of MCF-7 cells. Methods： Antisense survivin oligonucleotide was anchored onto polyamidoamine dendrimer grafted carbon nanotubes to form dendrimer-CNT-asODN complex and the complex was characterized by Zeta potential, AFM, TEM, and 1% agarose gel electrophoresis analysis. Dendrimer-CNT-asODN complexes were added into the medium and incubated with MCF-7 cells. MTT method was used to detect the effects of asODN and dendrimer-CNT-asODN on the growth of MCF-7 cells. TEM was used to observe the distribution of dendrimer-CNT-asODN complex within MCF-7 cells. Results： Successful synthesis of dendrimer-CNT-asODN complexes was proved by TEM, AFM and agarose gel electrophoresis. TEM showed that the complexes were located in the cytoplasm, endosome, and lysosome within MCF-7 cells. When dendrimer-CNT-asODN （1.0 μmol/L） and asODN （1.0 μmol/L） were used for 120 h incubation, the inhibitory rates of MCF-7 cells were （28.22±3.5）% for dendrimer-CNT-asODN complex group, （9.23±0.56）% for only asODN group, and （3.44±0.25）% for dendrimer-CNT group. Dendrimer-CNT-asODN complex at 3.0 μmol/L inhibited MCF-7 cells by （30.30±10.62）%, and the inhibitory effects were in a time- and concentration-dependent manner. Conclusion： Dendrimer-CNT nanoparticles may serve as a gene delivery vector with high efficiency, which can bring foreign gene into cancer cells, inhibiting cancer cell proliferation and markedly enhancing the cancer therapy effects.

TGA/Chemometrics addressing innovative preparation strategies for functionalized carbon nanotubes

In this work,functionalized carbon nanotubes(CNTs)using two polyamine polymers,polyethyleneimine(PEI)and polyamidoamine dendrimer(PAMAM),were investigated by thermal analysis in order to address preparation strategies to obtain low cytotoxic compounds with the ability to conjugate micro-RNAs and,at the same time,to transfect efficiently endothelial cells.Thermogravimetric analysis(TGA)was coupled to chemometrics as a novel analytical strategy to characterize functionalized CNTs from different preparation conditions.In particular,two starting materials were considered:very small CNTs and carboxylated CNTs(CNT-COOH)in order to examine the affinity with polymers.Chemometrics permitted to compare results from TGA and to investigate the effect of a number of factors affecting the synthesis of coated nanotubes including a different amount of involved polymer and the time required for the suspension for a satisfactory and reproducible preparation procedure.The results demonstrated the effectiveness of TGA as a tool able to address synthesis of coated CNTs to be employed as efficient drug delivery vectors in biomedical applications.

The Modification of Poly amidoamine (PAMAM-G0.5) by Cytosine

The half generation Poly(amidoamine) (PAMAM) dendrimers was synthesized and characterized and modification by heterocyclic DNA base (Cytosine) . The conjugated Cytosine? modified detected by FT-IR, 1H NMR, 13C NMR and Mass spectroscopy analysis.

A green light-enhanced cytosolic protein delivery platform based on BODIPY-protein interactions

Development of cytosolic protein delivery platforms brings new possibilities for various incurable diseases.Strategies based on polymer/protein self-assembly have shown their potential in protein delivery.However,versatile photocontrolled platforms based on self-assembly for protein delivery are seldom reported.Herein,we report a boron-dipyrromethene(BODIPY)-modified polyamidoamine(PAMAM)with excellent photo-controllability and efficiency for the cytosolic delivery of various proteins.High serum stability was achieved by coating hyaluronic acid and human serum albumin on the surface of BODIPY-modified PAMAM/protein nanoparticles.The nanoparticles under green light irradiation allowed efficient intracellular delivery of multiple cargo proteins with different charges and molecular weights and promoted endosome escape.The study provides valuable guidance for the development of BODIPY derivative-based protein delivery systems and advances the research in intracellular protein delivery.

Efficient GSH delivery using PAMAM-GSH into MPP-induced PC12 cellular model for Parkinson’s disease

Glutathione(GSH)depletion has been an important contributor to the dysfunction of dopamine neurons.Polyamidoamine-GSH(PAMAM-GSH)was synthesized and the delivery effect of GSH into PC12 cells was tested.MTT assessment for cytotoxicity and reactive oxygen species(ROS)as well as nitrite oxide(NO)and intracelluar superoxide dismutase(SOD)detection for antioxidative ability were performed.Furthermore,the antiapoptotic ability was analysed by assessing caspase-3,JNK1/2 and Erk1/2 expression.Our data indicated that PAMAM-GSH is an effective agent to replenish GSH into PC12 cells.PAMAM-GSH developed its antioxidative and protective ability for 1-methyl-4-phenylpyridinium(MPP)-induced PC12 cells by reducing the intracellular levels of ROS and SOD activity as well as decreasing the release of NO.Meanwhile,PAMAM-GSH could inhibit caspase-3 activation and might show its antiapoptotic ability to MPP-induced PC12 cells through JNK2/Erk1/2 pathway.In summary,these studies suggest that PAMAM-GSH conjugate has an intrinsic ability to penetrate PC12 cells and deliver GSH into these cells which may provide a new strategy for clinical applications in the treatment of Parkinson’s disease.

Polymer nanoparticles with dendrimer-Ag shell and its application in catalysis

Polymer nanoparticles with dendrimer-Ag shell were prepared and their application in catalytic reduction of 4-nitrophenol （4-NP） was investigated. Cross-linked polystyrene （PS） microspheres were prepared through dispersion copolymerization of styrene, acrylic acid and crosslinking monomer 1, 2- divinylbenzene. PS microspheres with average size of 450 nm and narrow size distribution were used as support for the immobilization of dendrimer-Ag shell, The polyamidoamine （PAMAM） dendrimer shell was successively grafted onto the surface ofPS microspheres through repetitive Michael addition reaction of methyl acrylate （MA） and amidation of the obtained esters with large excess of ethylenediamine （EDA）. Silver nanoparticles were formed directly inside the PAMAM shell through reduction with NaBH4. The resulting PS@PAMAM-Ag nanopartides were packed in a stainless steel column and used successfully for catalytic reduction of 4-NP. This technique for packing catalytic polymer particles in a column could imnrove the efficiency of using the metal catalyst and the tedious seuaration in catalytic reaction.

SYNTHESIS AND CHARACTERIZATION OF SURFACE-HYPERBRANCHED MAGNETITE NANOPARTICLE FOR BOVINE SERUM ALBUMIN IMMOBILIZATION

A hyperbranched polyamidoamine polymer was synthesized on the surface of magnetite nanoparticles to enhance bovine serum albumin (BSA) immobilization efficiency. The amount of immobilized bovine serum albumin (BSA) on the surface-hyperbranched magnetite nanoparticle was up to 2.5 times as much as that of magnetite nanoparticle modified with only amino silane.