Characterization of the Uptake and Intracellular Trafficking of G4 Polyamidoamine Dendrimers

2011 ◽  
Vol 64 (3) ◽  
pp. 302
Author(s):  
Chin-Ling Pai ◽  
Ming-Jium Shieh ◽  
Pei-Jen Lou ◽  
Fei-Hong Huang ◽  
Tzu-Wen Wang ◽  
...  
Keyword(s):  
Intracellular Trafficking ◽  
Fluorescein Isothiocyanate ◽  
Pamam Dendrimers ◽  
Endocytic Pathway ◽  
Lysosomal Ph ◽  
Cellular Environment ◽  
Polyamidoamine Dendrimers ◽  
Energy Dependent ◽  
Endocytic Pathways

Polyamidoamine (PAMAM) dendrimers are highly branched spherical polymers that have emerged as potent synthetic drug and gene carriers; however, much remains to be learned about the mechanism of dendrimer-mediated cellular uptake. In this study, the endocytic pathway and intracellular trafficking of generation 4 (G4) PAMAM dendrimers were evaluated via fluorescein isothiocyanate (FITC) conjugation. We found that the G4-FITC dendrimers were internalized by energy-dependent and non-specific endocytic pathways. Interesting, G4-FITC dendrimers can not only buffer the endosomal/lysosomal pH but also co-localize with lysosomal markers over a period of 3 to 12 h, after which the signal decreased in the lysosomes and began to co-localize with the mitochondrial marker. This study contributes to the understanding of the molecular behaviour of G4 PAMAM dendrimers in a cellular environment and will facilitate the development of more effective PAMAM-mediated drug and gene delivery systems.

Characterization of Endocytic Pathways by Quantitative Fluorescence Microscopy

1998 ◽  
Vol 4 (S2) ◽  
pp. 1024-1025
Author(s):  
Frederick R. Maxfield ◽  
Richik N. Ghosh ◽  
William G. Mallet ◽  
Thwe Thwe Soe ◽  
Philip L. Leopold ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Cell Surface ◽  
Light And Electron Microscopy ◽  
Endocytic Pathway ◽  
Early Endosomes ◽  
Pass Through ◽  
Golgi Network ◽  
Quantitative Fluorescence ◽  
Endocytic Pathways

We have used light and electron microscopy to analyze endocytic trafficking pathways. In one set of studies, we have used fluorescently labeled antibodies to trace an endocytic pathway from the cell surface to the trans- Golgi network (TGN). Cells were transfected with a construct consisting of the transmembrane and cytoplasmic domains of TGN38 and the extracellular domain of Tac. TGN38 is predominantly in the TGN, but a small fraction is found on the cell surface. We used FITC-labeled anti-Tac monoclonal IgG to analyze the pathway from the surface to the TGN. We compared the distribution of internalized Tac-TGN38 to internalized transferrin. We found that most Tac-TGN38 enters the same early endosomes as transferrin. Furthermore, most Tac-TGN38 returns to the cell surface from the endocytic recycling compartment (ERC) at the same rate as transferrin. However, on each pass through the cell approximately 18% of Tac-TGN is retained, and this Tac-TGN38 is delivered to the TGN.

FITC-dextran as a probe for endosome function and localization in kidney

1990 ◽  
Vol 258 (2) ◽  
pp. C309-C317 ◽  
Author(s):  
W. I. Lencer ◽  
P. Weyer ◽  
A. S. Verkman ◽  
D. A. Ausiello ◽  
D. Brown
Keyword(s):  
Epithelial Cells ◽  
Water Channel ◽  
Fluorescein Isothiocyanate ◽  
Endocytic Pathway ◽  
Water Permeability Coefficient ◽  
Osmotic Water ◽  
Endocytic Vesicles ◽  
Excised Tissue ◽  
Specific Membrane ◽  
Endocytic Pathways

Fluorescein isothiocyanate (FITC)-labeled endosomes were localized in kidney epithelial cells after tissue fixation and sectioning, and specific membrane transport properties of isolated endocytic vesicles were measured using the same probe. Rats were infused intravenously with 10 kDa FITC-dextran, and kidneys were fixed with paraformaldehyde lysine periodate. FITC-labeled vesicles were visualized in semithin (1 micron) frozen sections of excised tissue by epifluorescent microscopy and by electron microscopy after a photoconversion reaction. Most FITC-labeled endosomes were apically located in epithelial cells lining the urinary tubules. By immunocytochemistry the anti-lysosomal glycoprotein LGP 120 was absent from most of the FITC-labeled vesicles, although some colocalization was noted. The limiting membrane of FITC-labeled endosomes contained a vacuolar proton pump (pHmin = 6.23 +/- 0.033) and a water channel (osmotic water permeability coefficient, Pf = 0.052 +/- 0.005 cm/s) and was highly permeable to ethylene glycol and urea but relatively impermeable to glucose. Methods allowing the attribution of specific membrane functions to vesicles that can be visualized in the apical endocytic pathway of epithelial cells should be of general use for the study of endocytic pathways in a variety of systems.

Synthesis and Characterization of Ferrocenyl Carboxilic Surface-Functionalized Resorcinaren-PAMAM Dendrimers

2015 ◽  
Vol 19 (19) ◽  
pp. 1954-1960 ◽  
Author(s):  
Sandra Cortez-Maya ◽  
Elena Klimova ◽  
R. I. Puente Lee ◽  
Andres Borja-Miranda ◽  
Marcos Martinez-Garcia
Keyword(s):  
Pamam Dendrimers ◽  
Synthesis And Characterization

scholarly journals Characterization of Two Protein Disulfide Isomerases from the Endocytic Pathway of Bloodstream Forms of Trypanosoma brucei

2005 ◽  
Vol 280 (11) ◽  
pp. 10410-10418 ◽  
Author(s):  
Joyce Rubotham ◽  
Katherine Woods ◽  
Jose A. Garcia-Salcedo ◽  
Etienne Pays ◽  
Derek P. Nolan
Keyword(s):  
Trypanosoma Brucei ◽  
Endocytic Pathway ◽  
Protein Disulfide Isomerases ◽  
Disulfide Isomerases ◽  
Protein Disulfide

In-Situ Characterization of Growth and Intermixing at a Heteroepitaxial Interface: Fe on Au(001)

1993 ◽  
Vol 318 ◽  
Author(s):  
Q. Jiang ◽  
A. Chan ◽  
Y.-L. He ◽  
G.-C. Wang
Keyword(s):  
Elevated Temperatures ◽  
Chemical Elements ◽  
Background Intensity ◽  
In Situ Characterization ◽  
Initial Stage ◽  
Monolayer Growth ◽  
Low Energy Electron ◽  
Energy Dependent

ABSTRACTThe growth and chemical intermixing of submonolayer and a few monolayer thick Fe films on a Au(001) surface was studied by High Resolution Low Energy Electron Diffraction (HRLEED) technique. Through the analysis of the energy dependent angular profiles as a function of time, we obtained the distribution of islands and distribution of spacings during submonolayer growth. The interference of electron waves from different chemical elements in terraces at different heights in the surface contributes to the background intensity and broadening in the angular profiles of diffraction beams. A subsurface Fe capped by Au islands as a result of atomic place exchange was observed at the initial stage of monolayer growth. From the energy dependent angular profiles as a function of temperature, we determine the quantitative change of inhomogeneity length (∼20 Å) at the interface of ultrathin films at elevated temperatures due to intermixing.

scholarly journals The Spike Protein VP4 Defines the Endocytic Pathway Used by Rotavirus To Enter MA104 Cells

2012 ◽  
Vol 87 (3) ◽  
pp. 1658-1663 ◽  
Author(s):  
Marco A. Díaz-Salinas ◽  
Pedro Romero ◽  
Rafaela Espinosa ◽  
Yasutaka Hoshino ◽  
Susana López ◽  
...  
Keyword(s):  
Surface Protein ◽  
Endocytic Pathway ◽  
Spike Protein ◽  
Single Amino Acid ◽  
Cellular Receptors ◽  
Hypertonic Medium ◽  
Bovine Rotavirus ◽  
Single Amino Acid Change ◽  
Interfering Rna

ABSTRACTRotaviruses are internalized into MA104 cells by endocytosis, with different endocytic pathways used depending on the virus strain. The bovine rotavirus UK strain enters cells through a clathrin-mediated endocytic process, while the simian rhesus rotavirus (RRV) strain uses a poorly defined endocytic pathway that is clathrin and caveolin independent. The viral surface protein VP7 and the spike protein VP4 interact with cellular receptors during cell binding and penetration. To determine the viral protein that defines the mechanism of internalization, we used a panel of UK × RRV reassortant viruses having different combinations of the viral structural proteins. Characterization of the infectivities of these reassortants in MA104 cells either transfected with a small interfering RNA (siRNA) against the heavy chain of clathrin or incubated with hypertonic medium that destabilizes the clathrin coat clearly showed that VP4 determines the pathway of virus entry. Of interest, the characterization of Nar3, a sialic acid-independent variant of RRV, showed that a single amino acid change in VP4 shifts the route of entry from being clathrin dependent to clathrin independent. Furthermore, characterizations of several additional rotavirus strains that differ in their use of cellular receptors showed that all entered cells by clathrin-mediated endocytosis, suggesting that diverse VP4-cell surface interactions can lead to rotavirus cell entry through this endocytic pathway.

scholarly journals Characterization of ATP7A missense mutants suggests a correlation between intracellular trafficking and severity of Menkes disease

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Tina Skjørringe ◽  
Per Amstrup Pedersen ◽  
Sidsel Salling Thorborg ◽  
Poul Nissen ◽  
Pontus Gourdon ◽  
...  
Keyword(s):  
Intracellular Trafficking ◽  
Menkes Disease

scholarly journals A systematic characterization of intrinsically formed microglia-like cells during retinal organoid differentiation.

2022 ◽  
Author(s):  
Katarina Bartalska ◽  
Verena Hübschmann ◽  
Medina Korkut-Demirbaş ◽  
Ryan John Abat Cubero ◽  
Alessandro Venturino ◽  
...  
Keyword(s):  
Protein Composition ◽  
Human Microglia ◽  
Development Organization ◽  
Cellular Environment ◽  
Preferential Localization ◽  
Circuit Development ◽  
And Migration ◽  
Induced Pluripotent ◽  
Insight Into

Brain organoids differentiated from human induced pluripotent stem cells provide a unique opportunity to investigate the development, organization and connectivity of neurons in a complex cellular environment. However, organoids usually lack microglia, brain-resident immune cells which are both present in the early human embryonic brain and participate in neuronal circuit development. Here, we find that microglia innately develop in unguided retinal organoid differentiation between week 3 and 4 in 2.5D culture and appear later in floating, non-pigmented, 3D-cystic compartments. We enriched for cystic structures using a low-dosed BMP4 application and performed mass spectrometry, thus defining the protein composition of microglia-containing compartments. We found that cystic compartments expressed both mesenchymal and epithelial markers with microglia enriched in the mesenchymal region. Interestingly, microglia-like cells started to express the border-associated macrophage marker CD163. The preferential localization of human microglia to a mesenchymal compartment provides insight into the behavior and migration of microglia. The model will ultimately allow detailed study of these enigmatic cells and how they enter and distribute within the human brain.

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