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https://www.readbyqxmd.com/read/28453254/gene-transfection-mediated-by-catiomers-requires-free-highly-charged-polymer-chains-to-overcome-intracellular-barriers
#1
Lindomar Jose Calumby Albuquerque, Carlos E de Castro, Karin A Riske, Maria Cristina Carlan da Silva, Paulo I R Muraro, Vanessa Schmidt Giacomelli, Cristiano Giacomelli, Fernando Carlos Carlos Giacomelli
The prospective use of the block copolymers PEO113-b-PDEA50 and POEGMA70-b-P(OEGMA10-co-DEA47-co-DPA47) as non-viral gene vectors was evaluated. The polymers are able to properly condense DNA into nano-sized particles (RH ~ 75 nm) which are marginally cytotoxic and can be uptaken by cells. However, the GFP expression assays evidenced that DNA delivery is essentially negligible meaning that intracellular trafficking hampers efficient gene release. Subsequently we demonstrate that cellular uptake and particularly the quantity of GFP-positive cells are substantially enhanced when the block copolymer polyplexes are produced and further supplemented by BPEI chains (branched polyethyleneimine)...
April 28, 2017: Biomacromolecules
https://www.readbyqxmd.com/read/28449222/tspan33-is-expressed-in-transitional-and-memory-b-cells-but-is-not-responsible-for-high-adam10-expression
#2
César Augusto Pérez-Martínez, José Luis Maravillas-Montero, Iván Meza-Herrera, Felipe Vences-Catalán, Albert Zlotnik, Leopoldo Santos-Argumedo
Tetraspanins are a family of transmembrane proteins that form membrane microdomains. They play important roles in migration, adhesion, and other cellular processes. TspanC8, a subfamily of tetraspanins, were found to associate and promote ADAM10 trafficking and cell surface localization. One of its members, Tspan33, is expressed in activated B cells. Using RT-PCR and flow cytometry, we analyzed the pattern of expression of Tspan33 in B cells from healthy donors. We found Tspan33 expression in early and late stages of B cell development...
April 27, 2017: Scandinavian Journal of Immunology
https://www.readbyqxmd.com/read/28449134/characterizing-exogenous-mrna-delivery-trafficking-cytoplasmic-release-and-rna-protein-correlations-at-the-level-of-single-cells
#3
Jonathan L Kirschman, Sushma Bhosle, Daryll Vanover, Emmeline L Blanchard, Kristin H Loomis, Chiara Zurla, Kathryn Murray, Blaine C Lam, Philip J Santangelo
The use of synthetic messenger ribonucleic acid (mRNA) to express specific proteins is a highly promising therapeutic and vaccine approach that avoids many safety issues associated with viral or DNA-based systems. However, in order to optimize mRNA designs and delivery, technology advancements are required to study fundamental mechanisms of mRNA uptake and localization at the single-cell and tissue level. Here, we present a single RNA sensitive fluorescent labeling method which allows us to label and visualize synthetic mRNA without significantly affecting function...
April 26, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28448737/involvement-of-arhgef10-gef-for-rhoa-in-rab6-rab8-mediating-membrane-traffic
#4
Satoshi Shibata, Yui Teshima, Kenta Niimi, Shinobu Inagaki
Small GTPases play crucial roles in the maintenance of a homeostatic environment and appropriate movements of the cell. In these processes, the direct or indirect interaction between distinct small GTPases could be required for regulating mutual signaling pathways. In our recent study, ARHGEF10, known as a guanine nucleotide exchange factor (GEF) for RhoA, was indicated to interact with Rab6A and Rab8A, which are known to function in the exocytotic pathway, and colocalized with these Rabs at exocytotic vesicles...
April 27, 2017: Small GTPases
https://www.readbyqxmd.com/read/28448055/transient-expression-and-cellular-localization-of-recombinant-proteins-in-cultured-insect-cells
#5
Jeffrey A Fabrick, J Joe Hull
Heterologous protein expression systems are used for the production of recombinant proteins, the interpretation of cellular trafficking/localization, and the determination of the biochemical function of proteins at the sub-organismal level. Although baculovirus expression systems are increasingly used for protein production in numerous biotechnological, pharmaceutical, and industrial applications, nonlytic systems that do not involve viral infection have clear benefits but are often overlooked and underutilized...
April 20, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28448009/using-primary-neurosphere-cultures-to-study-primary-cilia
#6
Issei S Shimada, Hemant Badgandi, Bandarigoda N Somatilaka, Saikat Mukhopadhyay
The primary cilium is fundamentally important for the proliferation of neural stem/progenitor cells and for neuronal differentiation during embryonic, postnatal, and adult life. In addition, most differentiated neurons possess primary cilia that house signaling receptors, such as G-protein-coupled receptors, and signaling molecules, such as adenylyl cyclases. The primary cilium determines the activity of multiple developmental pathways, including the sonic hedgehog pathway during embryonic neuronal development, and also functions in promoting compartmentalized subcellular signaling during adult neuronal function...
April 14, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28447987/live-cell-imaging-and-3d-analysis-of-angiotensin-receptor-type-1a-trafficking-in-transfected-human-embryonic-kidney-cells-using-confocal-microscopy
#7
Parnika Kadam, Ryan McAllister, Jeffrey S Urbach, Kathryn Sandberg, Susette C Mueller
Live-cell imaging is used to simultaneously capture time-lapse images of angiotensin type 1a receptors (AT1aR) and intracellular compartments in transfected human embryonic kidney-293 (HEK) cells following stimulation with angiotensin II (Ang II). HEK cells are transiently transfected with plasmid DNA containing AT1aR tagged with enhanced green fluorescent protein (EGFP). Lysosomes are identified with a red fluorescent dye. Live-cell images are captured on a laser scanning confocal microscope after Ang II stimulation and analyzed by software in three dimensions (3D, voxels) over time...
March 27, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28447703/conserved-effects-and-altered-trafficking-of-cetuximab-antibodies-conjugated-to-gold-nanoparticles-with-precise-control-of-their-number-and-orientation
#8
Lorena García-Fernández, Javier Garcia-Pardo, Olivia Tort, Ian Prior, Mathias Brust, Eudald Casals, Julia Lorenzo, Victor F Puntes
Gold nanoparticles (17 nm) have been functionalized with the antiangiogenic monoclonal antibody drug Cetuximab at a well-defined orientation and coverage density of antibodies. Functionalization has been carried out through site-directed chemistry via the selective oxidation of the carbohydrate moiety of antibodies linked to a thiolated hydrazide. A431 tumor cells have been exposed to these conjugates for in vitro evaluation of their effects. In addition to epithelial growth factor receptor blocking, trafficking and signaling alterations were also observed...
April 27, 2017: Nanoscale
https://www.readbyqxmd.com/read/28446566/induction-of-adaptive-immunity-leads-to-nigrostriatal-disease-progression-in-mptp-mouse-model-of-parkinson-s-disease
#9
Goutam Chandra, Avik Roy, Suresh B Rangasamy, Kalipada Pahan
Although the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model is the most widely used animal model for Parkinson's disease (PD), it is known that nigrostriatal pathologies do not persist in the acute MPTP mouse model. This study highlights the importance of adaptive immunity in driving persistent and progressive disease in acute MPTP-intoxicated mice. Although marked infiltration of T cells into the nigra was found on 1 d of MPTP insult, T cell infiltration decreased afterward, becoming normal on 30 d of insult...
April 26, 2017: Journal of Immunology: Official Journal of the American Association of Immunologists
https://www.readbyqxmd.com/read/28446186/fingolimod-attenuates-experimental-autoimmune-neuritis-and-contributes-to-schwann-cell-mediated-axonal-protection
#10
Björn Ambrosius, Kalliopi Pitarokoili, Lisa Schrewe, Xiomara Pedreiturria, Jeremias Motte, Ralf Gold
BACKGROUND: Fingolimod, a sphingosine-1-phosphate receptor modulator with well-described immunomodulatory properties involving peripheral immune cell trafficking, was the first oral agent approved for the treatment of relapsing remitting multiple sclerosis. Analogous immunomodulatory treatment options for chronic peripheral autoimmune neuropathies are lacking. METHODS: We tested fingolimod in the animal model of experimental autoimmune neuritis in Lewis rat. Six to eight-week-old female rats were immunized with P2 peptide and from this day on treated with fingolimod...
April 26, 2017: Journal of Neuroinflammation
https://www.readbyqxmd.com/read/28445739/autophagy-associated-cytotoxicity-and-cellular-uptake-mechanisms-of-bismuth-nanoparticles-in-human-kidney-cells
#11
Yongming Liu, Jing Zhuang, Xihui Zhang, Cong Le, Ning Zhu, Liecheng Yang, Yong Wang, Tao Chen, Yangyun Wang, Leshuai W Zhang
Bismuth compounds have been used for treatment of bacterial infection, and recently bismuth nanoparticles (BiNP) were synthesized for imaging and diagnostic purpose, while safety concern of bismuth cannot be ignored. Here, we prepared ultrasmall BiNP and showed an enhanced tumor imaging, but BiNP revealed a differentiated cytotoxicity in human embryonic kidney 293 cells (HEK293) compared to other cell types. For the first time, we found that BiNP can induce autophagy, shown as the increase of monodansylcadaverine fluorescence staining and the amount of LC3II that can be inhibited by 3-MA...
April 23, 2017: Toxicology Letters
https://www.readbyqxmd.com/read/28445535/microrna-125a-3p-is-regulated-by-myd88-in-legionella-pneumophila-infection-and-targets-ntan1
#12
Elisa Jentho, Malena Bodden, Christine Schulz, Anna-Lena Jung, Kerstin Seidel, Bernd Schmeck, Wilhelm Bertrams
BACKGROUND: Legionella pneumophila (L. pneumophila) is a causative agent of severe pneumonia. It is highly adapted to intracellular replication and manipulates host cell functions like vesicle trafficking and mRNA translation to its own advantage. However, it is still unknown to what extent microRNAs (miRNAs) are involved in the Legionella-host cell interaction. METHODS: WT and MyD88-/- murine bone marrow-derived macrophages (BMM) were infected with L. pneumophila, the transcriptome was analyzed by high throughput qPCR array (microRNAs) and conventional qPCR (mRNAs), and mRNA-miRNA interaction was validated by luciferase assays with 3´-UTR mutations and western blot...
2017: PloS One
https://www.readbyqxmd.com/read/28444985/membrane-traffic-control-by-hypoxia
#13
EDITORIAL
Joachim Fandrey
Cells and tissues have to adapt to adverse changes in their environment to survive and continue to function. The lack of oxygen, hypoxia, is one of the fundamental challenges to which cells need to respond in a coordinate manner. In this issue of Acta Physiologica Wottawa et al.(1) studied hypoxia induced membrane trafficking. This article is protected by copyright. All rights reserved.
April 26, 2017: Acta Physiologica
https://www.readbyqxmd.com/read/28444310/arl3-and-rp2-regulate-the-trafficking-of-ciliary-tip-kinesins
#14
Nele Schwarz, Amelia Lane, Katarina Jovanovich, David A Parfitt, Monica Aguila, Clare L Thompson, Lyndon da Cruz, Peter J Coffey, J Paul Chapple, Alison J Hardcastle, Michael E Cheetham
Ciliary trafficking defects are the underlying cause of many ciliopathies, including Retinitis Pigmentosa (RP). Anterograde intraflagellar transport (IFT) is mediated by kinesin motor proteins; however, the function of the homodimeric Kif17 motor in cilia is poorly understood, whereas Kif7 is known to play an important role in stabilising cilia tips. Here we identified the ciliary tip kinesins Kif7 and Kif17 as novel interaction partners of the small GTPase Arl3 and its regulatory GTPase activating protein (GAP) Retinitis Pigmentosa 2 (RP2)...
April 21, 2017: Human Molecular Genetics
https://www.readbyqxmd.com/read/28442598/ethylene-regulates-differential-growth-via-big-arf-gef-dependent-post-golgi-secretory-trafficking-in-arabidopsis
#15
Kristoffer Jonsson, Yohann Boutté, Rajesh Kumar Singh, Delphine Gendre, Rishikesh Bhalerao
During early seedling development, the shoot apical meristem is protected from damage as the seedling emerges from soil by the formation of apical hook. Hook formation requires differential growth across the epidermis below the meristem in the hypocotyl. The plant hormones ethylene and auxin play key roles during apical hook development by controlling differential growth. We provide genetic and cell biological evidence for the role of ADP-ribosylation factor 1 (ARF1)-GTPase and its effector ARF-guanine-exchange factors (GEF)s of the BFA-inhibited GEF (BIG) family and GNOM in ethylene- and auxin-mediated control of hook development...
April 24, 2017: Plant Cell
https://www.readbyqxmd.com/read/28442548/keratin-8-18-regulation-of-insulin-receptor-signaling-and-trafficking-in-hepatocytes-through-a-concerted-phosphoinositide-dependent-akt-and-rab5-modulation
#16
Alexandra Roux, Anne Loranger, Josée N Lavoie, Normand Marceau
Keratins (Ks) are epithelial cell intermediate filament (IF) proteins that are expressed as pairs in a differentiation-regulated manner. Hepatocyte IFs are made only of K8/K18 pairs, which means that a K8 loss in K8-null mice leads to degradation of K18. Functionally, there is accumulating evidence that IFs contribute to signaling platforms. Here, we investigate the role of K8/K18 IFs in the regulation of insulin receptor (IR) signaling and trafficking in hepatocytes. We find that the IR substrate 1 (IRS1)/PI3K/Akt signaling cascade-downstream of IR-display prolonged activation in K8-null compared with wild-type hepatocytes...
April 25, 2017: FASEB Journal: Official Publication of the Federation of American Societies for Experimental Biology
https://www.readbyqxmd.com/read/28442538/peripheral-tumor-necrosis-factor-alpha-tnf-%C3%AE-modulates-amyloid-pathology-by-regulating-blood-derived-immune-cells-and-glial-response-in-the-brain-of-ad-tnf-transgenic-mice
#17
Evi Paouri, Ourania Tzara, Georgia-Ioanna Kartalou, Sofia Zenelak, Spiros Georgopoulos
Increasing evidence has suggested that systemic inflammation along with local brain inflammation can play a significant role in Alzheimer's disease (AD) pathogenesis. Identifying key molecules that regulate the crosstalk between the immune and the central nervous system can provide potential therapeutic targets. TNF-α is a pro-inflammatory cytokine implicated in the pathogenesis of systemic inflammatory and neurodegenerative diseases such as rheumatoid arthritis (RA) and AD. Recent studies have reported that anti-TNF-α therapy or RA itself can modulate AD pathology, although the underlying mechanism is unclear...
April 25, 2017: Journal of Neuroscience: the Official Journal of the Society for Neuroscience
https://www.readbyqxmd.com/read/28442379/peripheral-membrane-associations-of-matrix-metalloproteinases
#18
REVIEW
Steven R Van Doren, Tara C Marcink, Rama K Koppisetti, Alexander Jurkevich, Yan G Fulcher
Water soluble matrix metalloproteinases (MMPs) have been regarded as diffusing freely in the extracellular matrix. Yet multiple MMPs are also observed at cell surfaces. Their membrane-proximal activities include sheddase activities, collagenolysis, bacterial killing, and intracellular trafficking reaching as far as the nucleus. The catalytic domains of MMP-7 and MMP-12 bind bilayers peripherally, each in two different orientations, by presenting positive charges and a few hydrophobic groups to the surface. Related peripheral membrane associations are predicted for other soluble MMPs...
April 22, 2017: Biochimica et Biophysica Acta
https://www.readbyqxmd.com/read/28441569/the-cytoskeleton-autophagy-connection
#19
REVIEW
David J Kast, Roberto Dominguez
Actin cytoskeleton dynamics play vital roles in most forms of intracellular trafficking by promoting the biogenesis and transport of vesicular cargoes. Mounting evidence indicates that actin dynamics and membrane-cytoskeleton scaffolds also have essential roles in macroautophagy, the process by which cellular waste is isolated inside specialized vesicles called autophagosomes for recycling and degradation. Branched actin polymerization is necessary for the biogenesis of autophagosomes from the endoplasmic reticulum (ER) membrane...
April 24, 2017: Current Biology: CB
https://www.readbyqxmd.com/read/28441135/proteomic-mapping-of-cytosol-facing-outer-mitochondrial-and-er-membranes-in-living-human-cells-by-proximity-biotinylation
#20
Victoria Hung, Stephanie S Lam, Namrata D Udeshi, Tanya Svinkina, Gaelen Guzman, Vamsi K Mootha, Steven A Carr, Alice Y Ting
The cytosol-facing membranes of cellular organelles contain proteins that enable signal transduction, regulation of morphology and trafficking, protein import and export, and other specialized processes. Discovery of these proteins by traditional biochemical fractionation can be plagued with contaminants and loss of key components. Using peroxidase-mediated proximity biotinylation, we captured and identified endogenous proteins on the outer mitochondrial membrane (OMM) and endoplasmic reticulum membrane (ERM) of living human fibroblasts...
April 25, 2017: ELife
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