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Cyclic GMP-AMP (cGAMP) synthase (cGAS) is a predominant DNA sensor inducing the activation of the innate immune responses that produce proinflammatory cytokines and type I interferons, which has been well-investigated in mammals. However, chicken cGAS (chcGAS), which participates in avian innate immunity, has not been well-investigated. Here, we cloned the complete open reading frame sequence of chcGAS. Multiple sequence alignment and phylogenetic analysis revealed that chcGAS was homologous to mammalian cGAS. The chcGAS mRNA was highly expressed in the bone marrow and ileum. The subcellular localization of chcGAS was mainly in the cytoplasm, and partial co-localization was observed in the endoplasmic reticulum. Through overexpression and RNA interference, we demonstrated that chcGAS responded to exogenous dsDNA, HS-DNA, and poly(dA:dT), and to self dsDNA from the DNA damage response, thereby triggering the activation of STING/TBK1/IRF7-mediated innate immunity in both chicken embryonic fibroblasts and chicken liver cancer cells. Furthermore, downregulation of chcGAS enhanced the infection of fowl adenovirus serotype 4 in LMH cells. Our results demonstrated that chcGAS was an important cytosolic DNA sensor activating innate immune responses and may shed light on a strategy for preventing infectious diseases in the poultry industry.

Electrochemical tip-enhanced Raman spectroscopy (EC-TERS) appears as a promising in situ nanospectroscopic tool for characterization and understanding of the electrochemical interfacial processes at the nanometer scale and molecular level. However, the wide application of EC-TERS is hampered by its low sensitivity as a result of the optical path distortion due to the refractive index mismatch of the multilayer media (air, glass, and electrolyte). Here, we propose a new side-illumination EC-TERS setup by coupling a water immersion objective with a high numerical aperture to a scanning tunneling microscope scanning head customized with a large open space and a compact spectroelectrochemical cell. It not only effectively eliminates the optical distortion but also increases the sensitivity remarkably, which allows sensitive monitoring of the electrochemical redox processes of anthraquinone molecules. More importantly, EC-TERS is able to independently control the tip position and laser illumination position. By utilizing this feature, we reveal that the irreversible reduction reaction of anthraquinone observed in EC-TERS is induced by the synergistic effect of the negative potential and laser illumination rather than the localized surface plasmon. The highly improved sensitivity and the flexibility to control the tip and laser illumination position on the nanometer scale endows EC-TERS as an important tool for the fundamental understanding of the photo- or plasmon electrochemistry and the interfacial structure-activity relationship of important electrochemical systems.

Purpose: NeosedumosideIII (Neo) is a megastigmanes and belongs to monocyclic sesquiterpenoids compound with antioxidant, anti-inflammatory and other pharmacological activities. In order to explore the anti-cancer effect and possible mechanism of Neo, the study examined the anti-proliferation and apoptosis effect of Neo against human hepatocellular carcinoma HepG2 cells and SMMC-772 cells and related mechanism in vitro. Methods ：The anti-proliferation effect of Neo was detected on HepG2 cells and SMMC-772 cells by MTT assay and IC50 with increasing dose and time. Cell cycle and apoptosis were detected by flow cytometer. The changes of Bcl-2, Bax, Caspase-3, Caspase-8 and Caspase-9 proteins were detected by western blotting.Results ：The results indicated that Neo could inhibited proliferation of HepG2 cells and SMMC-772 cells in vitro and promoted apoptosis, it significantly induced apoptosis of HepG2 cells and SMMC-772 cells arrested cell cycle at G0/G1 phase in a dose-dependent manner, reduce the expression of Bcl-2 protein, and increase the expression of Bax and Caspase-3, Caspase-8 and Caspase-9 proteins. Conclusion：Neo could inhibit proliferation and induce apoptosis of HepG2 cells and SMMC-7721 cells in vivo which suggested that it might be served as a promising candidate for the treatment of liver cancer.

Active oxygen species (AOS) play key roles in many important catalytic reactions relevant to clean energy and environment. However, it remains challenging to characterize the active sites for producing AOS and to image the surface properties of AOS, especially on multicomponent metallic catalyst surfaces. Herein, we utilize tip-enhanced Raman spectroscopy (TERS) to probe the local generation and diffusion of OH radicals on a Pd/Au(111) bimetallic catalyst surface. The reactive OH radicals can be catalytically generated from hydrogen peroxide (H2O2) at the metal surface, which then oxidizes the surface adsorbed thiolate, a reactant that is used as the TERS probe. By TERS imaging of the spatial distribution of unreacted thiolate molecules, we demonstrate that the Pd surface is active for generation of OH radicals and the Pd step edge shows much higher activity than the Pd terrace, whereas the Au surface is inactive. Furthermore, we find that the locally generated OH radicals at the Pd step edge could diffuse to both the Au and the Pd surface sites to induce oxidative reactions, with a diffusion length estimated to be about 5.4 nm. Our TERS imaging with few-nanometer spatial resolution not only unravels the active sites but also characterizes in real space the diffusion behavior of OH radicals. The results are highly valuable to understand AOS-triggered catalytic reactions. The strategy of using reactants with large Raman cross sections as TERS probes may broaden the application of TERS for studying catalysis with reactive small molecules.

Three new mononuclear lanthanide complexes [Ln(2-Br-5-MOBA)3(terpy)(H2O)] (Ln = Gd (1), Tb (2), Er (3); 2-Br-5-MOBA = 2-bromine-5-methoxybenzoate; terpy = 2,2:6′,2″-terpyridine) have been successfully synthesized and characterized by single-crystal X-ray diffraction. The complexes 1–3 were isostructural and eight-coordinated, which is different from the binuclear structure of the previously reported complexes. The molecular structure of complexes 1–3 is very interesting: two adjacent mononuclear units are connected through hydrogen bonding giving rise to a pseudo binuclear structure. These pseudo binuclear units are further linked via O–H⋯Br hydrogen bonds and slightly offset π–π stacking interactions to from 1D and 2D supramolecular structures. The thermal decomposition mechanism of complexes 1–3 was studied by TG analysis and further authenticated by TG/DSC-FTIR techniques. The solid-state luminescent property of complex 2 was investigated at room temperature. The result indicates that Tb (III) complex appear to be promising candidate for the application as green luminescent material.

Six new lanthanide complexes [Ln(2-Br-5-MOBA)3(2,2′-DIPY)]2 (Ln = Nd(1), Eu(2), Gd(3), Tb(4), Ho(5), Er(6); 2-Br-5-MOBA = 2-bromine-5-methoxybenzoate; 2,2′-DIPY = 2,2′-bipyridine) have been successfully synthesized and characterized. The complexes 1–5 are isostructural and nine-coordinated by the single-crystal X-ray diffraction analyses, while the complex 6 is eight-coordinated. The difference of crystal structure may be the result of the lanthanide contraction effect. The binuclear units were further assembled into 1D, 2D, 3D supramolecular structures by the π-π stacking and C H⋯O hydrogen bonding interactions. The thermal decomposition mechanism of complexes 1–6 was studied by TG analysis and further authenticated by TG/DSC-FTIR techniques. The solid-state luminescence properties of complexes 2 and 4 were investigated at room temperature. The results indicate that complexes 2 and 4 show characteristic emission of Eu3+ ion and Tb3+ ion, respectively. What's more, the title complexes have good antibacterial activities against Candida albicans.

Two novel lanthanide complexes with the general formula [Ln(2-Br-5-MOBA) 3 (2,2′-DIPY)] 2 (Ln = Pr(1), Dy(2); 2-Br-5-MOBA = 2-bromine-5-methoxybenzoate; 2,2′-DIPY = 2,2′-bipyridine) have been successfully synthesized and characterized. The complexes 1 and 2 are isostructural and nine-coordinated with distorted monocapped square antiprismatic coordination geometry by the single-crystal X-ray diffraction analyses. Two complexes are all assembled into the 1D, 2D, 3D supramolecular structures by C H⋯O hydrogen bonding and π–π stacking interactions. The thermal decomposition mechanism for title complexes were obtained by TG/DSC-FTIR technology. The molar heat capacities of the complexes 1 and 2 are measured by a DSC instrument over the temperature range from 263.15 K to 365.15 K. On the basis of the fitted polynomial and thermodynamic equations, the smoothed values of heat capacities and thermodynamic functions ( H T - H 298.15K ), and ( S T - S 298.15K ) of the two complexes are calculated. In addition, the fluorescence spectra indicate that the complex 2 exhibits the characteristic emission of Dy(III) ion ( 4 F 9/2 → 6 H 13/2–15/2 ).

Ten novel lanthanide complexes {[Ln(L) 3 phen][Ln(L) 3 phen (C 2 H 5 OH)(H 2 O)]} 2 (Ln = La ( 1 ), Nd ( 2 )) and [Ln(L) 3 phen] 2 (Ln = Pr ( 3 ), Sm ( 4 ), Eu ( 5 ), Gd ( 6 ), Tb ( 7 ), Dy ( 8 ), Ho ( 9 ), Er ( 10 ); HL: 2,3-dimethoxybenzoic acid; phen: 1,10-phenanthroline) have been synthesized and characterized by elemental analyses, IR spectra and powder X-ray diffraction. Single crystals of complexes 1 – 5 and 8 were obtained and their structures have been determined by single crystal X-ray diffraction. The molecular structures of complexes 1 and 2 are very particular. They both contain two structures with different coordination environments but the same geometric configurations in the molecular structures, which has rarely been reported previously. Complexes 3 , 4 , 5 and 8 have similar structures with coordination numbers of nine, forming distorted monocapped square antiprisms, and their binuclear structures are stitched together by π–π intermolecular interactions to form 1D chains. The thermal decomposition mechanisms of complexes 1 – 10 have been studied by the technology of TG and TG-FTIR. The antimicrobial activities of the ten complexes have been explored by the disc diffusion method. The solid-state luminescence of complexes 5 and 7 was also studied, and the two complexes exhibit strong fluorescence, which shows that phen has a better sensitizing effect.

Three novel ternary lanthanide complexes [Ln(2,5-DClBA)3(terpy)(H2O)] (Ln = Eu (1), Tb (2), Ho (3), 2,5-DClBA = 2,5-dichlorobenzoate, terpy = 2,2:6′,2″-terpyridine) have been successfully synthesized and characterized by single-crystal and powder X-ray diffraction. Complexes 1–3 are isostructural, and each metal center is nine-coordinated with distorted monocapped square antiprismatic coordination geometry. The mononuclear units are assembled into 1D, 2D, 3D supramolecular structures by the π–π stacking and halogen–halogen interactions. Thermal decomposition mechanisms of the title complexes were investigated by thermogravimetric analysis and further authenticated by TG/DSC-FTIR techniques. The solid-state fluorescent properties of complexes 1–2 were studied at room temperature, and characteristic emission was observed for all complexes. Complex 1 exhibits the strong red emission, while the complex 2 is green under UV light. Moreover, the antibacterial activities of the complexes against Candida albicans, Escherichia coli and Staphylococcus aureus were also evaluated.

Three novel Ho(III) complexes, [(Ho(2-Br-5-MOBA)3(terpy)(H2O)] (1), [Ho(2-Br-5-MOBA)3(2,2′-bipy)]2 (2) and [Ho(2-Br-5-MOBA)3(5,5′-DM-2,2′-bipy)]2 (3) (2-Br-5-MOBA = 2-bromine-5-methoxybenzoate, terpy = 2,2:6′,2″-terpyridine, 2,2′-bipy = 2,2′-bipyridine, 5,5′-DM-2,2′-bipy = 5,5′-dimethy-2,2′-bipyridine), have been synthesized and characterized by single-crystal X-ray diffraction. Although 1–3 have the same triclinic crystal system and P 1 ¯ space group, they show three different types of molecule structures. For 1, it is mononuclear and fake binuclear units constituted by the mononuclear molecules are stitched together via π–π stacking and C–H⋯O hydrogen bonding interactions to form the 1D, 2D supramolecular structure. Complexes 2 and 3 are binuclear structures with the different coordination numbers, which are assembled into 1D, 2D, 3D supramolecular structures by different intermolecular interactions. This is because of the different auxiliary ligands. The thermal decomposition mechanism of complexes 1–3 were studied by TG analysis and further authenticated by TG/DSC-FTIR techniques, indicating that the difference of the auxiliary ligand affects the thermal stabilities of complexes. The average molar heat capacity values of the complexes gradually increased with an increase of temperature, indicating the complexes have good thermal stability within this range of temperature. In addition, the IR, PXRD, and the antibacterial activities of complexes 1–3 were measured.

Two unprecedented Ln-based metal–organic frameworks (MOFs) based on 5-nitroisophthalic acid (H2L) and 2,4,5-tri(4-pyridyl)-imidazole (Htpim), namely {[H3tpim][Ln2(L)4(H2O)]}n [Ln = La (I), Nd (II)], were prepared by solvothermal reactions and characterized by IR, elemental analysis and X-ray diffraction analysis. X-ray single crystal diffraction indicates that complexes I and II display similar 2D 44 frameworks with similar tetranuclear [Ln4(COO)16(H2O)2] secondary building units due to the effect of Ln contraction and the same coordination modes of L ligands as well as the template effect of Htpim. Moreover, thermal stabilities and luminescent properties of I and II were also investigated.

Three-dimensional (3D) α -Fe 2 O 3 nanoparticle anchored graphene aerogel (Fe 2 O 3 @GA) composites were assembled by a hydrothermal method using Fe(OH) 3 colloids and graphene oxides as starting materials. It was found that the Fe 2 O 3 nanoparticles were uniformly embedded into the 3D networks of graphene aerogels and the resulting composites contained meso- and macro-scale pores. Remarkably, the composites possessed much higher surface area (S BET = 212.5 m 2 g −1 ) and larger pore volume (V p = 0.2073 cm 3 g −1 ) than those of pure Fe 2 O 3 (S BET = 19.8 m 2 g −1 , V p = 0.1770 cm 3 g −1 ). The Fe 2 O 3 @GA composites used as electrode materials for lithium ion batteries were demonstrated to exhibit high reversible capacity at large current densities and excellent cycling stabilities.

Aluminum-substituted MCM-41 (Al-MCM-41) were hydrothermally synthesised and used as supports to fabricate the mesostructured H3PW12O40 (HPW) solid acid catalysts via an impregnation method. The influences of various HPW loadings on the structures of the catalysts were verified by X-ray diffraction (XRD), nitrogen physisorption, scanning electron microscopy (SEM), transmission electron microscopy (TEM), ultraviolet–visible (UV–Vis) spectroscopy and Fourier-transform infrared spectroscopy (FT-IR) measurements and their acidities were evaluated by infrared of pyridine adsorption (Py-IR) and temperature-programmed desorption of ammonia (NH3-TPD) measurements. The optimisation of the reaction conditions for levulinic acid esterification with n-hexanol such as the reaction temperature and time, the reactant molar ratio, and the catalyst dosages was performed to maximise the conversion of levulinic acid. The effects of various HPW loadings on the conversion of levulinic acid were investigated under the o...

Two novel mononuclear lanthanide complexes, [Ln(2,4-DClBA)3(terpy)(H2O)]·H2O (Ln = Dy(1), Er(2)); 2,4-DClBA: 2,4-dichlorobenzoate; terpy: 2,2′:6′,2″-terpyridine; have been synthesized and characterized by single-crystal and powder X-ray diffraction. The results reveal that complexes 1–2 are isomorphous, and each Ln3+ ion is nine coordinated adopting a distorted monocapped square antiprismatic molecular geometry. Mononuclear complexes 1–2 are stitched together via Cl–π and hydrogen bonding interactions to form the 1D, 2D, 3D supramolecular structures, which the 3D is rarely observed. Thermal decomposition mechanism is determined by TG/DSC-FTIR, and the 3D stacked plots for the FTIR spectra of the evolved gases were recorded. Heat capacities of complexes 1–2 were measured, and the derived thermodynamic functions (H T − H 298.15), (S T − S 298.15) and (G T − G 298.15) of the complexes relative to the standard reference temperature 298.15 K were obtained. The activation energy E values of the first decomposition stage for title complexes were calculated by integral isoconversional nonlinear (NL-INT) and Starink methods. Complex 1 shows the characteristic emission of Dy3+ with strong yellow emission.

In aquatic ecosystems, mercuric compounds are considered to be highly toxic to a great variety of aquatic biota ranging from bacteria to fishes. In the present study, the effects of different concentrations (0, 2.0, 6.0, 10.0, 14.0 and 18.0 μg·L − 1 ) of Hg 2 + on the life table demography of Brachionus calyciflorus were investigated at three densities (1.0 × 10 6 , 2.0 × 10 6 and 4.0 × 10 6 cells·mL − 1 ) of Scenedesmus obliquus . The results showed that compared to the controls, Hg 2 + at 14.0 and 18.0 μg·L − 1 significantly decreased both the average lifespan and the life expectancy at hatching, Hg 2 + at 2.0–18.0 μg·L − 1 shortened the generation time, and Hg 2 + at 18.0 μg·L − 1 decreased the net reproductive rate of B. calyciflorus cultured at 1.0 × 10 6 cells·mL − 1 of S. obliquus . Increase in algal food level reduced the negative effects of high concentrations of Hg 2 + on survival, reproduction and population growth of the rotifers. Hg 2 + at 6.0 μg·L − 1 , and 6.0 and 18.0 μg·L − 1 increased the intrinsic rate of population increase of the rotifers cultured at 1.0 × 10 6 and 2.0 × 10 6 cells·mL − 1 of S. obliquus , respectively. But Hg 2 + at 18.0 μg·L − 1 decreased the intrinsic rate of population increase of the rotifers cultured at 4.0 × 10 6 cells·mL − 1 of S. obliquus . Significantly nominal concentration–response relationships existed between Hg 2 + concentration and each of all the five life history variables of B. calyciflorus cultured at 1.0 × 10 6 cells·mL − 1 of S. obliquus , the generation time of the rotifers cultured at 2.0 × 10 6 cells·mL − 1 of S. obliquus , as well as the intrinsic rate of population increase of the rotifers cultured at 4.0 × 10 6 cells·mL − 1 of S. obliquus . Among all the life history variables, generation time was the most sensitive to mercury pollution. Since algal levels chosen in the present study are much higher than those available in natural waters, except in eutrophic water bodies, the effects of mercury on survival, reproduction and population growth of the rotifers in natural waters may be more severe than those extrapolated from the present study.

MCM-41 was synthesized with a non-hydrothermal procedure and then used to assemble a series of supported H3PW12O40 (HPW) mesoporous catalysts. The optimization of the reaction conditions for LA esterification with ethanol, such as the reaction temperature, reaction time, the reactant molar ratio and the catalyst dosages was performed to maximize the conversion of LA over 25 wt% HPW/MCM-41 catalysts. The catalytic performance of the resultant HPW/MCM-41 catalysts was investigated. The results show that these catalysts retain the hexagonal phase of MCM-41 and the Keggin characteristic of HPW. Although their textural parameters decrease with increasing HPW loading, these catalysts still exhibit much higher specific surface area and pore volume (SBET ≥ 446 m2 g−1, VP ≥ 0.345 cm3 g−1) than that of HPW (SBET = 5 m2 g−1, VP = 0.010 cm3 g−1). LA conversion increases with increasing HPW loading and the value may reach up to 83.7 % over 45 wt% HPW/MCM-41 under the present conditions. The resultant ethyl levulinate is promising in reducing the consumption of petroleum-derived fossil fuels.

Four complexes based on 2,4,5-tri(4-pyridyl)-imidazole (Htpim), such as [Co(Htpim)2(SO4)(H2O)]n (1), [Co(tpim)2(H2O)2]n (2), [Cu(Htpim)2(SO4)]n (3), and [Cu2(Htpim)(tpim)(NO3)3(DMAC)]n (4), have been synthesized and characterized. Different structures, including two similar 1D linear chains in 1 and 2, a 2D 44 layer in 3, and a 2D double layer {4.62}2{42.64.82.102}{62.8} network in 4, were attributed to different kinds and quantities of inorganic acids under the reaction condition. Hereinto, inorganic acid was not only used to adjust the pH value of the reaction system, but also to act as a coordination component in 1, 3, and 4. Moreover, thermal stabilities and luminescent properties of 1–4 were also investigated.

The strawberry vein banding virus (SVBV) open reading frame (ORF) VI encodes a P6 protein known as the RNA silencing suppressor. This protein is known to form inclusion like granules of various sizes and accumulate in both the nuclei and the cytoplasm of SVBV-infected plant cells. In this study, we have determined that the P6 protein is the only trans-activator (TAV) encoded by SVBV, and can efficiently trans-activate the translation of downstream gfp mRNA in a bicistron derived from the SVBV. Furthermore, the P6 protein can trans-activate the expression of different bicistrons expressed by different caulimovirus promoters. The P6 protein encoded by SVBV from an infectious clone can also trans-activate the expression of bicistron. Through protein-protein interaction assays, we determined that the P6 protein could interact with the cell translation initiation factor FveIF3g of Fragaria vesca and co-localize with it in the nuclei of Nicotiana benthamiana cells. This interaction reduced the formation of P6 granules in cells and its trans-activation activity on translation.

The SiO/C composite materials were prepared via a solution route and subsequent thermal treatment. The effects of carbon source on the particle morphology of composite, the graphitization degree of carbon component and the electrochemical performance of the prepared SiO/C composites were investigated by scanning electron microscopy (SEM), Raman spectrometer and electrochemical charge/discharge tests. The results reveal that the SiO/C composite synthesized with the epoxy resin as carbon source exhibits the higher graphitic degree and higher reversible specific capacity (~690 mAhg-1) and excellent cycling stability.

Spatial distribution characteristics of available phosphorus (AP) and total phosphorus (TP) in wetland soils withPhragmites australis,Suaeda salsaandTamarix chinnensiswere investigated and their influencing factors was also indentified using principal correspondence analysis for ordination in the Yellow River Delta of China. Our results showed that TP and AP contents in both sites withPhragmites australisandSuaeda salsa, and the horizontal distributions of AP content in three sites exhibited similar distribution characteristics to that of AP:TP ratios. Additionally, soil properties such as soil moisture, salinity, total nitrogen, total carbon and pH values were the important influencing factors of higher spatial variability of AP and TP.

Spatial variability of soil nitrogen and phosphorus in the Yellow River Delta was investigated using geostatistical method. Our results showed moderate variation in TN and TP and high variations in NH4+-N and AP. The best semi-variogram model for each nutrient was identified. The model parameters suggested that the structure variance dominated the total variance of TN, TP and NH4+-N, while the spatial variability of AP was relatively random. The spatial variation scales of soil nitrogen and phosphorus are similar.

A dendritic cell (DC) networking system has become an attractive approach in cancer immunotherapy. Successful DC gene engineering depends on the development of transgene vectors. A cationic polymer, chitosan-linked polyethylenimine (PEI) (CP), possessing the advantages of both PEI and chitosan, has been applied in nonviral transfection of DCs. Physicochemical evaluation showed that CP/DNA complexes could form cationic nanoparticles. Compared with DCs transfected with commercial reagent, Lipofectamine2000, it showed higher transfection efficiency and lower cytotoxicity when DCs were transfected with CP/DNA complexes. A nuclear trafficking observation of CP/DNA complexes by a confocal laser scanning microscope further revealed that the CP could help DNA enter into the cytoplasm and finally into the nucleus of a DC. Finally, vaccination of DCs transfected with CP/DNA encoding gp100 slightly improved resistance to the B16BL6 melanoma challenge. This is the first report that CP polymer is used as a nonviral vector for DC gene delivery and DC vaccine. Essentially, these results might be helpful to design a promising nonviral vector for DC gene delivery.

PEI and chitosan are considered to be promising non-viral gene delivery vectors. To improve the transfection efficiency of chitosan, we linked chitosan with polyethylenimine (PEI, Mw = 1.8 kDa) by 1,1′-carbonyldiimidazole to form a complex. The composition, particle size, as well as the zeta potential of this chitosan-linked-PEI (CP) complex were measured. And the DNA binding ability, cytotoxicity, and gene transfection efficiency of CP complex were also investigated in cancer cells. In HepG2, A549 and HeLa cells, CP complex exhibited lower cytotoxicity as compared with PEI25KDa (Mw = 25 kDa), a positive control proved to be an efficient gene transfection polymer. Likewise, it showed good transfection efficiency in these cancer cell lines. Specifically, the long-term transfection efficiency of CP was higher than PEI25KDa as demonstrated by the in vitro cancer cell model. The confocal laser scanning microscopy data showed the time for CP to enter the nucleus was 4 h, which was longer than that of PEI25KDa but shorter than that of chitosan. Furthermore, CP complexes were used as a gene carrier to deliver the CCL22 gene into H22 cells. When these gene-altered cells were inoculated in mice, the tumor growth rate was significantly decreased, indicating the CP copolymer was a promising vector for the therapeutic gene delivery.

In this study, the cytotoxicity and transfection efficiency of using chitosan/DNA complex combined with poly(ethylenimine) (PEI) were investigated. The combination of PEI with the chitosan/DNA complex markedly enhanced the gene expression of HeLa cells to 1000-fold of that induced by chitosan alone. PEI's cytotoxicity was considerably decreased upon combination with the chitosan/DNA complex. Furthermore, the PEI/chitosan/DNA could maintain the gene expression efficiency in the presence of serum.

For designing a complex vector that has the advantages of both polyethylenimine (PEI) and chitosan for gene delivery, a PEI/chitosan/DNA complex was constructed at various N/P ratios (the ratios of moles of the amine groups of cationic polymers to those of the phosphate ones of DNA) and both the cytotoxicity and the transfection efficiency of the vector were evaluated. The results demonstrated that the chitosan/DNA binding degree was depended on the N/P ratio. The mean size of the complex vector was between 100 nm and 150 nm. Compared with PEI/DNA, the complex vector (PEI/chitosan/DNA with chitosan/DNA N/P=4, PEI/DNA N/P=10) appeared to have low cytotoxicity, which maintained the cell survival rate at greater than 80%, and showed higher transfection efficiency of nearly 1000 fold compared with that using chitosan/DNA alone. Furthermore, the expression efficiency of the complex vector carrying enhanced green fluorescent protein was not inhibited in the presence of serum in both HeLa cells and A549 cells. The PEI/chitosan complex may be a promising gene carrier that has high transfection efficiency as well as low cytotoxicity.

Qing-Qing Zhao,1,2 Yu-Lan Hu,1 Yang Zhou,3 Ni Li,1 Min Han,1 Gu-Ping Tang,4 Feng Qiu,2 Yasuhiko Tabata,5 Jian-Qing Gao,11Institute of Pharmaceutics, Zhejiang University, Hangzhou, China; 2Department of Pharmacy, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China; 3Institute of Biochemistry, Iowa State University, Ames, IA, USA; 4Institute of Chemical Biology and Pharmaceutical Chemistry, Zhejiang University, Hangzhou, China; 5Institute for Frontier Medical Sciences, Kyoto University, Kyoto, JapanBackground: The success of gene transfection is largely dependent on the development of a vehicle or vector that can efficiently deliver a gene to cells with minimal toxicity.Methods: A liver cancer-targeted specific peptide (FQHPSF sequence) was successfully synthesized and linked with chitosan-linked polyethylenimine (CP) to form a new targeted gene delivery vector called CPT (CP/peptide). The structure of CPT was confirmed by 1H nuclear magnetic resonance spectroscopy and ultraviolet spectrophotometry. The particle size of CPT/DNA complexes was measured using laser diffraction spectrometry and the cytotoxicity of the copolymer was evaluated by methylthiazol tetrazolium method. The transfection efficiency evaluation of the CP copolymer was performed using luciferase activity assay. Cellular internalization of the CP/DNA complex was observed under confocal laser scanning microscopy. The targeting specificity of the polymer coupled to peptide was measured by competitive inhibition transfection study. The liver targeting specificity of the CPT copolymer in vivo was demonstrated by combining the copolymer with a therapeutic gene, interleukin-12, and assessed by its abilities in suppressing the growth of ascites tumor in mouse model.Results: The results showed that the liver cancer-targeted specific peptide was successfully synthesized and linked with CP to form a new targeted gene delivery vector called CPT. The composition of CPT was confirmed and the vector showed low cytotoxicity and strong targeting specificity to liver tumors in vitro. The in vivo study results showed that interleukin-12 delivered by the new gene vector CPT/DNA significantly enhanced the antitumor effect on ascites tumor-bearing imprinting control region mice as compared with polyethylenimine (25 kDa), CP, and other controls, which further demonstrate the targeting specificity of the new synthesized polymer.Conclusion: The synthesized CPT copolymer was proven to be an effective liver cancer-targeted vector for therapeutic gene delivery, which could be a potential candidate for targeted cancer gene therapy.Keywords: targeting, peptide, polyethylenimine, chitosan, antitumor