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https://www.readbyqxmd.com/read/28325284/type-i-interferons-impede-short-hairpin-rna-mediated-rnai-via-inhibition-of-dicer-mediated-processing-to-small-interfering-rna
#1
Mitsuhiro Machitani, Fuminori Sakurai, Keisaku Wakabayashi, Kosuke Takayama, Masashi Tachibana, Hiroyuki Mizuguchi
RNAi by short hairpin RNA (shRNA) is a powerful tool not only for studying gene functions in various organisms, including mammals, but also for the treatment of severe disorders. However, shRNA-expressing vectors can induce type I interferon (IFN) expression by activation of innate immune responses, leading to off-target effects and unexpected side effects. Several strategies have been developed to prevent type I IFN induction. On the other hand, it has remained unclear whether type I IFNs have effects on shRNA-mediated RNAi...
March 17, 2017: Molecular Therapy. Nucleic Acids
https://www.readbyqxmd.com/read/28323876/pim1-minicircle-as-a-therapeutic-treatment-for-myocardial-infarction
#2
Nan Liu, Bingyan J Wang, Kathleen M Broughton, Roberto Alvarez, Sailay Siddiqi, Rebeca Loaiza, Nicky Nguyen, Pearl Quijada, Natalie Gude, Mark A Sussman
PIM1, a pro-survival gene encoding a serine/ threonine kinase, influences cell proliferation and survival. Modification of cardiac progenitor cells (CPCs) or cardiomyocytes with PIM1 using a lentivirus-based delivery method showed long-term improved cardiac function after myocardial infarction (MI). However, lentivirus based delivery methods have stringent FDA regulation with respect to clinical trials. To provide an alternative and low risk PIM1 delivery method, this study examined the use of a non-viral modified plasmid-minicircle (MC) as a vehicle to deliver PIM1 into mouse CPCs (mCPCs) in vitro and the myocardium in vivo...
2017: PloS One
https://www.readbyqxmd.com/read/28323177/biodegradable-cationic-poly-carbonates-effect-of-varying-side-chain-hydrophobicity-on-key-aspects-of-gene-transfection
#3
Zhan Yuin Ong, Chuan Yang, Wei Cheng, Zhi Xiang Voo, Willy Chin, James L Hedrick, Yi Yan Yang
The degree of hydrophobicity in cationic polymers plays an important but often underappreciated role in the safety and efficacy of gene delivery processes. In order to further elucidate structure-activity relationships of biodegradable cationic poly(carbonate) gene carriers, we synthesized a series of narrowly dispersed homo-polymers via metal-free organocatalytic living ring-opening polymerization (ROP) of cyclic carbonate monomers bearing either alkyl (propyl, hexyl or nonyl) or 4-methyl benzyl halide side chains...
March 17, 2017: Acta Biomaterialia
https://www.readbyqxmd.com/read/28323095/megalin-targeted-enhanced-transfection-efficiency-in-cultured-human-hk-2-renal-tubular-proximal-cells-using-aminoglycoside-carboxyalkyl-polyethylenimine-containing-nanoplexes
#4
Fatemeh Oroojalian, Ali Hossein Rezayan, Wayne Thomas Shier, Khalil Abnous, Mohammad Ramezani
Non-viral vectors are of interest as therapeutic gene delivery agents in gene therapy, because they are simple to prepare, easy to modify and have definable safety profiles compared to viral vectors. The potential of gene therapy in the treatment of renal diseases is limited by a lack of effective kidney-targeted gene delivery systems. Aminoglycoside antibiotics gentamicin and neomycin were connected by amide linkages to carboxyl groups on carboxyalkylated-PEI25 (25kDa PEI) or carboxyalkylated-PEI10 (10kDa PEI)...
March 16, 2017: International Journal of Pharmaceutics
https://www.readbyqxmd.com/read/28322160/an-efficient-transient-expression-system-for-enhancing-generation-of-monoclonal-antibodies-in-293-suspension-cells
#5
Guoquan Zhang, Jie Liu, Wanting Fan, Qianyi Chen, Bin Shi
BACKGROUND: Recombinant monoclonal antibodies (mAbs) are useful in research, diagnosis, and therapy. The increased demands of recombinant mAbs require efficient production systems. A variety of expression vectors have been developed for stable or transient production of mAb in mammalian cells. However, though a few commercial expression systems of mAbs can be listened, the high expense often impedes academia researches. METHODS: In this study, we described the development of a transient mammalian system based on a bicistronic vector, which contains internal ribosome entry site (IRES) and enhancer elements to express IgG1 light chain (LC) and heavy chain (HC) in one transcript...
March 20, 2017: Current Pharmaceutical Biotechnology
https://www.readbyqxmd.com/read/28321779/transfection-of-bovine-fetal-fibroblast-with-polyethylenimine-pei-nanoparticles-effect-of-particle-size-and-presence-of-fetal-bovine-serum-on-transgene-delivery-and-cytotoxicity
#6
D O Forcato, A E Fili, F E Alustiza, J M Lázaro Martínez, S Bongiovanni Abel, M F Olmos Nicotra, A P Alessio, N Rodríguez, C Barbero, P Bosch
The development of efficient transfection protocols for livestock cells is crucial for implementation of cell-based transgenic methods to produce genetically modified animals. We synthetized fully deacylated linear 22, 87 and 217 kDa polyethylenimine (PEI) nanoparticles and compared their transfection efficiency and cytotoxicity to commercial branched 25 kDa PEI and linear 58 kDa poly(allylamine) hydrochloride. We studied the effect of PEI size and presence of serum on transfection efficiency on primary cultures of bovine fetal fibroblasts and established cells lines (HEK 293 and Hep G2)...
March 20, 2017: Cytotechnology
https://www.readbyqxmd.com/read/28320116/in-vitro-non-viral-murine-pro-neurotrophin-3-gene-transfer-into-rat-bone-marrow-stromal-cells
#7
Shahram Darabi, Taki Tiraihi, AliReza Delshad, Majid Sadeghizadeh, Wisam Khalil, Taher Taheri
Neurotrophin 3 (NT-3) is an important factor for promoting prenatal neural development, as well as regeneration, axogenesis and plasticity in postnatal life. Therapy with NT-3 was reported to improve the condition of patients suffering from degenerative diseases and traumatic injuries, however, the disadvantage of NT-3 protein delivery is its short half-life, thus our alternative approach is the use of NT-3 gene therapy. In this study, the bone marrow stromal cells (BMSCs) were isolated from adult rats, cultured for 4 passages and transfected with either pEGFP-N1 or a constructed vector containing murine proNT-3 (pSecTag2/HygroB-murine proNT-3) using Lipofectamine 2000 followed by Hygromycin B (200mg/kg)...
April 15, 2017: Journal of the Neurological Sciences
https://www.readbyqxmd.com/read/28298603/african-swine-fever-virus-np868r-capping-enzyme-promotes-reovirus-rescue-during-reverse-genetics-by-promoting-reovirus-protein-expression-virion-assembly-and-rna-incorporation-into-infectious-virions
#8
Heather E Eaton, Takeshi Kobayashi, Terence S Dermody, Randal N Johnston, Philippe H Jais, Maya Shmulevitz
Reoviruses, like many eukaryotic viruses, contain an inverted 7-methylguanosine (m7G) cap linked to the 5' nucleotide of mRNA. Traditional functions of capping are to promote mRNA stability, protein translation, and concealment from cellular proteins that recognize foreign RNA. To address the role of mRNA capping during reovirus replication, we assessed the benefits of adding the African swine fever virus NP868R capping enzyme. C3P3, a fusion protein containing T7 RNA polymerase and NP868R, was found to increase protein expression 5 to 10-fold, as compared to T7 RNA polymerase alone, while enhancing reovirus rescue from the current reverse genetics system by 100-fold...
March 15, 2017: Journal of Virology
https://www.readbyqxmd.com/read/28297751/preparation-of-vsv-g-viral-envelope-glycoprotein-from-pichia-pastoris-enhances-transfection-of-dna-into-animal-cells
#9
Xin Liu, Ying Dong, Jingquan Wang, Long Li, Zhenmin Zhong, Yun-Pan Li, Shao-Jun Chen, Yu-Cai Fu, Wen-Can Xu, Chi-Ju Wei
Vesicular stomatitis virus G glycoprotein (VSV-G) has been widely used for pseudotyping retroviral, lentiviral and artificial viral vectors. The objective of this study was to establish a potential approach for large scale production of VSV-G. To this end, VSV-G was cloned with a N-terminal His-tag into Pichia pastoris expression vector pPIC3.5K. Three clones (Mut(s)) containing the VSV-G expression cassette were identified by PCR. All clones proliferated normally in expansion medium, while the proliferation was reduced significantly under induction conditions...
March 15, 2017: Journal of Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28293718/transfection-of-primary-brain-capillary-endothelial-cells-for-protein-synthesis-and-secretion-of-recombinant-erythropoietin-a-strategy-to-enable-protein-delivery-to-the-brain
#10
Annette Burkhart, Thomas Lars Andresen, Achim Aigner, Louiza Bohn Thomsen, Torben Moos
Treatment of chronic disorders affecting the central nervous system (CNS) is complicated by the inability of drugs to cross the blood-brain barrier (BBB). Non-viral gene therapy applied to brain capillary endothelial cells (BCECs) denotes a novel approach to overcome the restraints in this passage, as turning BCECs into recombinant protein factories by transfection could result in protein secretion further into the brain. The present study aims to investigate the possibility of transfecting primary rat brain endothelial cells (RBECs) for recombinant protein synthesis and secretion of the neuroprotective protein erythropoietin (EPO)...
March 14, 2017: Cellular and Molecular Life Sciences: CMLS
https://www.readbyqxmd.com/read/28292248/surface-engineered-dendrimers-in-sirna-delivery-and-gene-silencing
#11
Vishakha Tambe, Shreya Thakkar, Nidhi Raval, Dilip Sharma, Kiran Kalia, Rakesh Kumar Tekade
BACKGROUND: Therapeutic efficacy of dreadful diseases like cancer, HIV (Human Immunodeficiency Virus) can be enhanced by delivering molecules which regulate function at gene level rather than at receptor level. Silencing RNA is one such approach recently used to silence target gene expressed diseases; and thereby reduce target protein levels. Many of the non-viral vectors are proved to act as carriers for silencing RNA. Dendrimers being one of them have less size, low poly dispersibility index, water solubility, multivalence, and easy surface modification...
March 14, 2017: Current Pharmaceutical Design
https://www.readbyqxmd.com/read/28282145/targeted-ultrasound-triggered-phase-transition-nanodroplets-for-her2-overexpressing-breast-cancer-diagnosis-and-gene-transfection
#12
Di Gao, Jinbiao Gao, Ming Xu, Zhong Cao, Luyao Zhou, Yingqin Li, Xiaoyan Xie, Qing Jiang, Wei Wang, Jie Liu
For successful gene therapy, it is imperative to accumulate therapeutic gene in tumor tissues followed by efficiently delivering gene into targeted cells. Ultrasound irradiation, as a noninvasive and cost-effective external stimulus, has been proved to be one of the most potential external-stimulating gene delivery strategies recently in further improving gene transfection. In this study, we developed tumor-targeting ultrasound-triggered phase-transition nanodroplets AHNP-PFP-TNDs comprising a perfluorinated poly(amino acid) C11F17-PAsp (DET) as a core for simultaneously loading perfluoropentane (PFP) and nucleic acids, and a polyanionic polymer PGA-g-PEG-AHNP as the shell for not only modifying the surface of nanodroplets but also introducing an anti-Her2/neu peptide (AHNP) aiming to targeted treatment of Her2-overexpressing breast cancer...
March 21, 2017: Molecular Pharmaceutics
https://www.readbyqxmd.com/read/28281780/fibroblast-growth-factor-1-transfected-adipose-derived-mesenchymal-stem-cells-promote-angiogenic-proliferation
#13
Seyed Javad Hoseini, Hamed Ghazavi, Fatemeh Forouzanfar, Baratali Mashkani, Ahmad Ghorbani, Elahe Mahdipour, Faezeh Ghasemi, Hamid Reza Sadeghnia, Majid Ghayour-Mobarhan
The aim of this study was to investigate, for the first time, the effects of using adipose-derived mesenchymal stem cells (AD-MSCs) transfected with an episomal plasmid encoding fibroblast growth factor 1 (FGF1) (AD-MSCs(FGF1)), in providing the microenvironment required for angiogenic proliferation. The isolated rat AD-MSCs were positive for mesenchymal (CD29 and CD90) and negative for hematopoietic (CD34 and CD45) surface markers. Adipogenic and osteogenic differentiation of the AD-MSCs also occurred in the proper culture media...
March 10, 2017: DNA and Cell Biology
https://www.readbyqxmd.com/read/28279738/modifying-plasmid-loaded-hsa-nanoparticles-with-cell-penetrating-peptides-cellular-uptake-and-enhanced-gene-delivery
#14
J Mesken, A Iltzsche, D Mulac, K Langer
Gene therapy bears great potential for the cure of a multitude of human diseases. Research efforts focussed on the use of viral delivery vectors in the past decades, neglecting non-viral gene therapies of physical or chemical origin due to low transfection efficiency. However, side effects such as activation of oncogenes and inflammatory reactions upon immune cell activation are major obstacles impeding the clinical applicability of viral gene therapy vectors. The aim of this study was the development of a non-viral gene delivery system based on plasmid-loaded human serum albumin nanoparticles, which are biocompatible, biodegradable, and non-toxic in relevant concentrations...
March 6, 2017: International Journal of Pharmaceutics
https://www.readbyqxmd.com/read/28278583/hydrophobically-modified-polyethylenimine-based-ternary-complexes-for-targeting-brain-tumor-stability-in-vitro-and-in-vivo-studies
#15
Brahmanand Dube, Abhijeet Pandey, Ganesh Joshi, Krutika Sawant
Hydrophobic modification of low molecular weight polyethylenimine (PEI 2 kDa) by cholic acid (ChA) was done to obtain PEI2-ChA. The nanoplexes of PEI2-ChA with gWIZ-GFP demonstrated increase transfection efficiency (∼27%) in NT8e cell lines. The cell-cycle analysis of NT8e cells (p53 mutant) treated with transferrin containing nanoplexes showed increased apoptosis of cells. In vitro protein expression revealed expression of exogenous p53 protein. In vivo imaging of mice showed localized signal for GFP protein in brain region...
February 17, 2017: Artificial Cells, Nanomedicine, and Biotechnology
https://www.readbyqxmd.com/read/28274742/a-hyaluronic-acid-based-hydrogel-enabling-cd44-mediated-chondrocyte-binding-and-gapmer-oligonucleotide-release-for-modulation-of-gene-expression-in-osteoarthritis
#16
Yunpeng Cai, Elena López-Ruiz, Jesper Wengel, Laura B Creemers, Kenneth A Howard
Hyaluronic acid (HA) is an attractive biomaterial for osteoarthritis (OA) treatment due to inherent functional and compatibility properties as an endogenous knee joint component. In this work, we describe a HA-based hydrogel with the dual functionality of increased CD44-dependent chondrocyte binding and controlled release of gapmer antisense oligonucleotides for unassisted cellular entry and subsequent gene silencing activity. A Schiff base-mediated gelation method was used to produce a panel of hydrogels varying in the aldehyde-modified HA (900kDa) to chitosan ratios (3:7, 5:5 and 7:3) for identifying designs displaying optimal engagement of OA patient-derived CD44-expressing chondrocytes...
March 5, 2017: Journal of Controlled Release: Official Journal of the Controlled Release Society
https://www.readbyqxmd.com/read/28271570/single-domain-antibodies-for-the-knockdown-of-cytosolic-and-nuclear-proteins
#17
REVIEW
Thomas Böldicke
Single domain antibodies (sdAbs) from camels or sharks comprise only the variable heavy chain domain. Human sdAbs comprise the variable domain of the heavy chain (VH) or light chain (VL) and can be selected from human antibodies. SdAbs are stable, non aggregating molecules in vitro and in vivo compared to complete antibodies and scFv fragments. They are excellent novel inhibitors of cytosolic/nuclear proteins because they are correctly folded inside the cytosol in contrast to scFv fragments. SdAbs are unique because of their excellent specificity and possibility to target posttranslational modifications such as phosphorylation sites, conformers or interaction regions of proteins that cannot be targeted with genetic knockout techniques and are impossible to knockdown with RNAi...
March 8, 2017: Protein Science: a Publication of the Protein Society
https://www.readbyqxmd.com/read/28256721/a-functional-microrna-screening-method-for-organ-morphogenesis
#18
Ivan T Rebustini
The increasing repertoire of microRNAs expressed during organ development and their role in regulating organ morphogenesis provide a compelling need to develop methods to assess microRNA function using various in vitro and in vivo experimental models. Methods to assess microRNA function during organ morphogenesis include transfection of microRNA inhibitors (antagomirs) and activators (mimics) into mouse embryonic explanted organs using liposomes, which can potentially result in low efficiency of transfection and off-target effects...
March 3, 2017: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/28247930/identification-of-murine-phosphodiesterase-5a-isoforms-and-their-functional-characterization-in-hl-1-cardiac-cell-line
#19
Federica Campolo, Alessandra Zevini, Silvia Cardarelli, Lucia Monaco, Federica Barbagallo, Manuela Pellegrini, Marisa Cornacchione, Antonio Di Grazia, Valeria De Arcangelis, Daniele Gianfrilli, Mauro Giorgi, Andrea Lenzi, Andrea M Isidori, Fabio Naro
Phosphodiesterase 5A (PDE5A) specifically degrades the ubiquitous second messenger cGMP and experimental and clinical data highlight its important role in cardiac diseases. To address PDE5A role in cardiac physiology, three splice variants of the PDE5A were cloned for the first time from mouse cDNA library (mPde5a1, mPde5a2 and mPde5a3). The predicted amino acidic sequences of the three murine isoforms are different in the N-terminal regulatory domain. mPDE5A isoforms were transfected in HEK293T cells and they showed high affinity for cGMP and similar sensitivity to sildenafil inhibition...
March 1, 2017: Journal of Cellular Physiology
https://www.readbyqxmd.com/read/28239294/transfection-studies-with-colloidal-systems-containing-highly-purified-bipolar-tetraether-lipids-from-sulfolobus-acidocaldarius
#20
Konrad H Engelhardt, Shashank Reddy Pinnapireddy, Elias Baghdan, Jarmila Jedelská, Udo Bakowsky
Lipid vectors are commonly used to facilitate the transfer of nucleic acids into mammalian cells. In this study, two fractions of tetraether lipids from the archaea Sulfolobus acidocaldarius were extracted and purified using different methods. The purified lipid fractions polar lipid fraction E (PLFE) and hydrolysed glycerol-dialkyl-nonitol tetraether (hGDNT) differ in their structures, charge, size, and miscibility from conventional lipids. Liposomes were prepared by mixing tetraether lipids with cholesterol (CH) and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) resulting in stable vectors for gene delivery...
2017: Archaea: An International Microbiological Journal
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