Delivering DNA into Plant Cell by Gene Carriers of ZnS Nanoparticles

The development of nanotechnology provides a new method for genetic engineering.However,the nanoparticles as gene carriers have been mainly used in the mammalian cells so far.We observed that ZnS nanoparticles modified with positively charged poly-L-lysine（PLL） successfully delivered GUS-encoding plasmid DNA into tobacco cells by means of ultrasound-assisted method.Polymerase chain reaction（PCR） detection,Southern blot analysis and GUS histochemical staining were carried out for the regenerated plants.The stable genetic modified plants mediated by ZnS nanoparticles can be obtained.This article demonstrates the great potential of nanoparticles as gene carrier in plant transformation and proves a novel approach for plant genetic decoration.

Recent progress in cationic polymeric gene carriers for cancer therapy

In recent years,various carriers for gene delivery nave been developed for biomedical applications.Among all kinds of gene carriers,cationic polymeric carriers for delivery therapeutic gene as non-viral carriers have received growing interests due to their improved high transfection efficiency with the relative safety.In particular,the advancement of novel polymeric gene carriers has gained much progress in the development of effective anticancer therapy.Herein,this review focused on the development of cationic polymeric carriers for cancer therapy,including polyethylenimine(PEI),polyamidoamine(PAMAM) dendrimers,polylysine(PLL),chitosan and modified cationic polymers.And recent progresses in the development of novel polymeric carriers for gene delivery,such as targeted gene carriers,responsive gene carriers and multifunctional gene carriers,were summarized.Finally,the future perspectives in the development of novel polymeric carriers for delivery gene were presented.

Exploring Quaternized Hydroxyethylcellulose as Potential Gene Carriers

Cationic polysaccharides have been receiving more attentions and used as nonviral gene delivery vectors. In this paper, quaternized hydroxyethylcellulose （QHEC） derivatives were studied as gene carriers for their efficient DNA binding abilities. All QHECs could form stable QHEC/DNA complexes and resist the degradation of DNase I. And the dynamic light scatter （DLS） results showed that all QHEC/DNA complexes could form compact particles. These QHEC/DNA complexes exhibited effective transfeetion abilities in comparison to the naked DNA. The cytotox- icities of QHEC and QHEC/DNA complexes were also evaluated in four cell lines which were relatively low com- pared with 25 kDa bPEI. All results indicated that these quaternized hydroxyethylcelluloses could be used as poten- tial gene delivery vectors.

Preparation of Superparamagnetic Dextran-coated Iron Oxide Nanoparticles used as a Novel Gene Carrier into Human Bladder Cancer Cells'

Objective: Application of magnetic nanoparticles as gene carrier in gene therapy has developed quickly. This study was designed to investigate the preparation of superparamagnetic dextran-coated iron oxide nanoparticles (SDION) and the feasibility of SDION used as a novel gene carrier for plasmid DNA in vitro. Methods: SDION were prepared by chemical coprecipitation and separated by gel filtration on Sephacryl S-300HR, characterized by TEM, laser scattering system and Vibrating Sample Magnetometer Signal Processor. The green fluorescent protein (pGFP-C2) plasmid DNA was used as target gene. SDION-pGFP-C2 conjugate compounds were produced by means of oxidoreduction reaction. The connection ratio of SDION and pGFP-C2 DNA was analyzed and evaluated by agarose electrophoresis and the concentration of pGFP-C2 in supernatant was measured. Using liposome as control, the transfection efficiency of SDION and liposome was respectively evaluated under fluorescence microscope in vitro. Results: The diameter of SDION ranges from 3 nm to 8 nm, the effective diameter was 59.2 nm and the saturation magnetization was 0.23 emu/g. After SDION were reasonably oxidized, SDION could connect with pGFP-C2 to a high degree. The transfection efficiency of SDION as gene carrier was higher than that of liposome. Conclusion: The successes in connecting SDION with pGFP-C2 plasmid by means of oxidoreduction reaction and in transferring pGFP-C2 gene into human bladder cancer BIU-87 cells in vitro provided the experimental evidence for the feasibility of SDION used as a novel gene carrier.

Enhancement of Immunological Activity of CpG ODN by Chitosan Gene Carrier

To investigate the enhancement of immunological activity of CpG ODN by chitosan gene carrier in mice, the effect of lymphocyte proliferation was detected in mice by using MTT, the levels of IgG and cytokines （IL-2 and IL-12） in serum were measured by ELISA and peripheral blood T lymphocyte subsets CD4^＋, CD8^＋ were analyzed by flow cytometry. Our results showed that spleen lymphocytes isolated from the CS-CpG ODN group of mice showed the strongest proliferation （SI =1.551）, and the levels of IgG, IL-2 and IL-12 in serum were higher than those of other groups. Compared with the immunization with CpG ODN, the immunization with CS-CpG ODN gene carrier was more efficient in up-regulating the percentage of CD4^＋T cells and the ratio of CD4^＋/CD8^＋ of mice, It was concluded that CS gene carrier of CpG ODN was much more effective in improving immunity of CpG ODN in mice.

Genomic Analysis of Mitochondrial Carrier Genes in the Bombyx mori

This study is performed to investigate the mitochondrial carrier gene family in silkworm genome. In total, 30 genes are identified and claded into eight well-conserved groups. Gene duplication contributes to the expansion and complexity of this family. Diverse expression patterns suggest their functional differentiation. Analyses of the sitespecific profiles reveal critical amino acid residues for functional divergence. This study highlights the molecular evolution of the mitochondrial carrier gene family in silkworm and may provide a starting point for further experimental verification.

Modulating the Oxidation Degree of Linear Polyethyleneimine for Preparation of Highly Efficient and Low-cytotoxicity Degradable Gene Delivery Carriers

Polyethyleneimine(PEI),as a widely used polymer material in the field of gene delivery,has been extensively studied for modification and shielding to reduce its cytotoxicity.However,research aimed at preparing degradable PEI is scarce.In this work,the hydrogen peroxide(H_(2)O_(2))oxidation method was used to introduce degradable amide groups in the PEI and a series of oxidized PEI22k(oxPEI22k)with different degrees of oxidation were synthesized by regulating the dosage of H_(2)O_(2).The relationship between the oxidation degree of oxPEI22k and the gene transfection efficiency of oxPEI22k was studied in detail,confirming that the oxPEI22k with oxidation degrees of 16.7%and 28.6%achieved improved transfection efficiency compared to unmodified PEI.These oxPEI22k also proved reduced cytotoxicity and improved degradability.Further,this strategy was extended to the synthesis of low-molecular-weight oxPEI1.8k.The oxPEI1.8k with suitable oxidation degree also achieved improved transfection efficiency and reduced cytotoxicity.In brief,this work provided high-efficiency and low-cytotoxicity degradable gene delivery carriers by regulating the oxidation degree of PEI,which was of great significance for promoting clinical applications of PEI.

Neuroimaging in Huntington's disease

Huntington's disease(HD) is a progressive and fatal neurodegenerative disorder caused by an expanded tri-nucleotide CAG sequence in huntingtin gene(HTT) on chromosome 4. HD manifests with chorea, cognitive and psychiatric symptoms. Although advances in genetics allow identification of individuals carrying the HD gene, much is still unknown about the mechanisms underly-ing the development of overt clinical symptoms and the transitional period between premanifestation and mani-festation of the disease. HD has no cure and patients rely only in symptomatic treatment. There is an urgent need to identify biomarkers that are able to monitor disease progression and assess the development and efficacy of novel disease modifying drugs. Over the past years, neuroimaging techniques such as magnetic resonance imaging(MRI) and positron emission tomog-raphy(PET) have provided important advances in our understanding of HD. MRI provides information about structural and functional organization of the brain, while PET can detect molecular changes in the brain. MRI and PET are able to detect changes in the brains of HD gene carriers years ahead of the manifestation of the dis-ease and have also proved to be powerful in assessingdisease progression. However, no single technique hasbeen validated as an optimal biomarker. An integrativemultimodal imaging approach, which combines differ-ent MRI and PET techniques, could be recommendedfor monitoring potential neuroprotective and preventivetherapies in HD. In this article we review the currentneuroimaging literature in HD.

Development of novel gene carrier using modified nano hydroxyapatite derived from equine bone for osteogenic differentiation of dental pulp stem cells

Hydroxyapatite(HA)is a representative substance that induces bone regeneration.Our research team extracted nanohydroxyapatite(EH)from natural resources,especially equine bones,and developed it as a molecular biological tool.Polyethylenimine(PEI)was used to coat the EH to develop a gene carrier.To verify that PEI is well coated in the EH,we first observed the morphology and dispersity of PEI-coated EH(pEH)by electron microscopy.The pEH particles were well distributed,while only the EH particles were not distributed and aggregated.Then,the existence of nitrogen elements of PEI on the surface of the pEH was confirmed by EDS,calcium concentration measurement and fourier transform infrared spectroscopy(FT-IR).Additionally,the pEH was confirmed to have a more positive charge than the 25 kD PEI by comparing the zeta potentials.As a result of pGL3 transfection,pEH was better able to transport genes to cells than 25 kD PEI.After verification as a gene carrier for pEH,we induced osteogenic differentiation of DPSCs by loading the BMP-2 gene in pEH(BMP-2/pEH)and delivering it to the cells.As a result,it was confirmed that osteogenic differentiation was promoted by showing that the expression of osteopontin(OPN),osteocalcin(OCN),and runt-related transcription factor 2(RUNX2)was significantly increased in the group treated with BMP-2/pEH.In conclusion,we have not only developed a novel nonviral gene carrier that is better performing and less toxic than 25 kD PEI by modifying natural HA(the agricultural byproduct)but also proved that bone differentiation can be effectively promoted by delivering BMP-2 with pEH to stem cells.

Silica Nanoparticles-mediated Stable Genetic Transformation in Nicotiana tabacum

Nanoparticles as gene carriers become popular in the mammalian cells, whereas the application of them in plant cells is still very limited. Herein lies a report on silica nanoparticles（SiNPs） modified with positively charged poly-L-lysine（PLL） successfully delivering plasmid-encoded β-glucuronidase（GUS） gene into tobacco with the help of gene gun. The stable transgenic tobacco plants mediated by SiNPs can be obtained. Furthermore, we revealed the quantity of gene and types of receptor materials could affect the expression efficiency. In comparison to conven- tional gold particles-mediated transformation, the silica nanoparticles-mediated stable genetic transformation enhances transformation efficiency, potentially overcoming transgenic silencing. Our results demonstrate the great potential of SiNPs as gene carrier in plant genetic transformation and prove a novel approach for plant genetic decoration.

Polylysine-modified Polyethylenimines as siRNA Carriers for Effective Tumor Treatment

Polyethylenimine-poly（L-lysine） （PEI-PLL） copolymer was synthesized via ring-opening polymerization of L-lysine N-carboxyanhydride （Lys（Z）-NCA） initiated by PEI. The complexation of PEI-PLL with siRNA was studied by particle size and zeta potential measurements. The flow cytometric analysis and confocal imaging showed its excellent intracellular trafficking ability. PEI-PLL displayed higher gene silencing efficiency and lower cytotoxicity than commercial PEI-25k in vitro. In the antitumor study, PEI-PLL was further combined with siVEGF and showed obviously tumor inhibition effect for the treatment of CT26 tumor model. Therefore, PEI-PLL is a promising siRNA carrier candidate for further antitumor treatment in vivo.

纳米材料助力合成生物学的生物医学应用

合成生物学是借助基因工具改造活细胞和生物体的学科,其经历了蓬勃发展阶段,在生物医学领域得到广泛应用,包括新型生物传感器和生物材料设计,以及最新临床治疗药物和疫苗的开发.纳米材料具有独特的物理、化学和生物特性,在拓展合成生物学的生物医学应用场景上具有巨大的潜力,可以提高治疗剂的生产效率,并增强改造生物体的功能.在此,我们简要概述了纳米材料应用于合成生物学领域的最新进展,主要涵盖纳米材料作为递送载体、信号转导器和功能增强剂的角色及其制备过程和性能特点.本综述首先强调了纳米材料作为基因载体的最新成果,包括有机、无机和仿生递送系统;其次,回顾了纳米材料介导的刺激响应型基因表达调控的研究;第三,总结了在可编程的杂化生物活性材料方面的开创性工作,包括纳米材料/细菌杂合系统和纳米材料/哺乳动物细胞杂合系统;最后,讨论了该领域当前面临的挑战,并从个人观点展望了未来的发展前景.

Versatile functionalization of amylopectin for effective biomedical applications

Successful gene vectors should be with high transfection efficiency and minimal cytotoxicity. Natural polysaccharides, due to their good biocompatibility and biodegradability, have been widely studied and applied. Amylopectin is one of polysaccharides with dendritic structure and numerous hydroxyl groups that could be used for subsequent modification. In this work, a series of dendritic cationic gene vectors comprising amylopectin backbones and poly(2-(dimethylamino) ethyl methacrylate)(PDMAEMA) side chains with different lengths(termed as AMY-PDs) were readily prepared by atom transfer radical polymerization(ATRP). The gene condensation ability, cytotoxicity and gene transfection of AMY-PDs carriers were investigated. In comparison with "gold-standard" poly(ethyleneimine)(PEI, 25 k Da), the AMY-PDs exhibited higher transfection efficiency with lower cytotoxicity. AMY-PDs could be further modified with Au nanoparticles(termed as AMY-PD@Au). The potential of the AMY-PD@Au vectors to be utilized as a CT contrast agent for imaging of cancer cells was investigated. Such AMY-PD@Au vectors may realize gene therapy with the ability of real-time imaging.

Multifunctional peptide conjugated amphiphilic cationic copolymer for enhancing ECs targeting, penetrating and nuclear accumulation

Gene therapy has drawn great attention in the treatments of many diseases,especially for cardiovascular diseases.However,the development of gene carriers with low cytotoxicity and multitargeting function is still a challenge.Herein,the multitargeting REDV-G-TATG-NLS peptide was conjugated to amphiphilic cationic copolymer poly(e-caprolactone-co-3(S)-methyl-morpho-line-2,5-dione)-g-polyethyleneimine(PCLMD-g-PEI)via a heterobifunctional orthopyridyl disulfde-poly(ethylene glycol)-N-hydroxysuccinimide(OPSS-PEG-NHS)linker to prepare PCLMD-g-PEI-PEG-REDV-G-TAT-G-NLS copolymers with the aim to develop the gene carriers with low cytotoxicity and high transfection efficiency.The multitargeting micelles were prepared from PCLMD-g-PEI-PEG-REDV-G-TAT-G-NLS copolymers by self-assembly method and used to load pEGFP-ZNF580 plasmids(pDNA)to form gene complexes for enhancing the proliferation and migration of endothelial cells(ECs).The loading pDNA capacity was proved by agarose gel electrophoresis assay.These multitargeting gene com-plexes exhibited low cytotoxicity by 3-(4,-dimethylthia-zol-2-yl)-2,5-diphenyltetrazolium bromide assay.The high internalization efficiency of these gene complexes was confirmed by flow cytometry.The results of in vitro transfection demonstrated that these multitargeting gene complexes possessed relatively high transfection effi-ciency.The rapid migration of ECs transfected by these gene complexes was verified by wound healing assay.Owing to ECs-targeting ability,cell-penetrating ability and nuclear targeting capacity of REDV-G-TAT-G-NLS pep-tide,the multitargeting polycationic gene carrier with low cytotoxicity and high transfection efficiency has great potential in gene therapy.

Identification of a Novel Mutation in Solute Carrier Family 29, Member 3 in a Chinese Patient with H Syndrome

Background：H syndrome （OMIM 612391） is a recently described autosomal recessive genodermatosis characterized by indurated hyperpigmented and hypertrichotic skin,as well as other systemic manifestations.Most of the cases occurred in the Middle East areas or nearby countries such as Spain or India.The syndrome is caused by mutations in solute carrier family 29,member 3 （SLC29A3）,the gene encoding equilibrative nucleoside transporter 3.The aim of this study was to identify pathogenic SLC29A 3 mutations in a Chinese patient clinically diagnosed with H syndrome.Methods：Peripheral blood samples were collected from the patient and his parents.Genomic DNA was isolated by the standard method.All six SLC29A3 exons and their flanking intronic sequences were polymerase chain reaction （PCR）-amplified and the PCR products were subjected to direct sequencing.Results：The patient,an 18-year-old man born to a nonconsanguineous Chinese couple,had more extensive cutaneous lesions,involving both buttocks and knee.In his genomic DNA,we identified a novel homozygous insertion-deletion,c.1269_1270delinsA,in SLC29A3.Both of his parents were carriers of the mutation.Conclusions：We have identified a pathogenic mutation in a Chinese patient with H syndrome.

A review on exosome-based cancer therapy

Cancer is one of the leading causes of mortality and morbidity globally.Many types of cancer treatments have been developed,such as chemotherapy,surgery,radiotherapy,and immunotherapy.However,these therapies can also kill healthy cells and lead to severe side effects.Therefore,scientists are looking for new strategies to eliminate cancerous cells specifically.Exosomes,nanometer-sized lipid bilayer-enclosed vesicles secreted from various cell types,exist in nearly all body fluids,including blood,breast milk,saliva,urine,bile,pancreatic juice,cerebrospinal,and peritoneal fluids.They carry myriad donor cell-derived bioactive molecules such as proteins,lipids,and RNAs(including microRNA and lncRNA)and can deliver them to both nearby and distant recipient cells.Due to these characteristics,exosomes have attracted great interest in cancer treatment(especially serving as a biological carrier for some drugs,microRNA,lncRNA,inhibitors,and antibodies).In this paper,we will review the current knowledge of exosome therapeutic applications in cancer.