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TAT and HA2 facilitate cellular uptake of gold nanoparticles but do not lead to cytosolic localisation.

Cesbron Y, Shaheen U, Free P, Lévy R - PLoS ONE (2015)

Bottom Line: However, conflicting results have been reported on the extent of the cytosolic delivery achieved.To evaluate their potential, we used gold nanoparticles as model cargos and systematically assessed how the functionalisation of their surface by either or both of the viral peptides TAT and HA2 influenced their intracellular delivery.While their uptake increased when the TAT and/or HA2 viral peptides were present on their surface, we did not observe a significant cytosolic delivery of the gold nanoparticles.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Liverpool, Liverpool, United Kingdom; Institute of Integrative Biology, University of Liverpool, Liverpool, United Kingdom; CNRS, UMR 6290, Institute of Genetics and Development of Rennes, Rennes, France; Université de Rennes 1, Université Européenne de Bretagne, Structure fédérative de recherche Biosit, Faculté de Médecine, Rennes, France.

ABSTRACT
The methods currently available to deliver functional labels and drugs to the cell cytosol are inefficient and this constitutes a major obstacle to cell biology (delivery of sensors and imaging probes) and therapy (drug access to the cell internal machinery). As cell membranes are impermeable to most molecular cargos, viral peptides have been used to bolster their internalisation through endocytosis and help their release to the cytosol by bursting the endosomal vesicles. However, conflicting results have been reported on the extent of the cytosolic delivery achieved. To evaluate their potential, we used gold nanoparticles as model cargos and systematically assessed how the functionalisation of their surface by either or both of the viral peptides TAT and HA2 influenced their intracellular delivery. We evaluated the number of gold nanoparticles present in cells after internalisation using photothermal microscopy and their subcellular localisation by electron microscopy. While their uptake increased when the TAT and/or HA2 viral peptides were present on their surface, we did not observe a significant cytosolic delivery of the gold nanoparticles.

No MeSH data available.


Related in: MedlinePlus

Influence of the proportion of CCALNN-dHA2 peptides in the monolayer of gold nanoparticles.HeLa cells were incubated for 4h with 10nm gold nanoparticles (6nM) coated with a monolayer composed of two different proportions of the peptides CCALNN-dHA2 and CALNN, washed thoroughly with PBS, fixed and imaged by electron microscopy. (A-B) TEM images of the cells incubated with nanoparticle with the following peptide monolayer composition: (A) 50% CCALNN-dHA2 and 50% CALNN, (B) 100% CCALNN-dHA2. Arrowheads point toward gold nanoparticles either interacting with vesicular membranes (white) or displaying a cytosolic localisation (black). (C) Density of nanoparticles in endosomes estimated from images shown in (A) (n = 10, middle) and (B) (n = 10, right); quantification for 10% CCALNN-dHA2 (left) is the same as in Fig. 2E (left). The sets of images analysed are available on figshare (10.6084/m9.figshare.875545, 10.6084/m9.figshare.875477).
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pone.0121683.g003: Influence of the proportion of CCALNN-dHA2 peptides in the monolayer of gold nanoparticles.HeLa cells were incubated for 4h with 10nm gold nanoparticles (6nM) coated with a monolayer composed of two different proportions of the peptides CCALNN-dHA2 and CALNN, washed thoroughly with PBS, fixed and imaged by electron microscopy. (A-B) TEM images of the cells incubated with nanoparticle with the following peptide monolayer composition: (A) 50% CCALNN-dHA2 and 50% CALNN, (B) 100% CCALNN-dHA2. Arrowheads point toward gold nanoparticles either interacting with vesicular membranes (white) or displaying a cytosolic localisation (black). (C) Density of nanoparticles in endosomes estimated from images shown in (A) (n = 10, middle) and (B) (n = 10, right); quantification for 10% CCALNN-dHA2 (left) is the same as in Fig. 2E (left). The sets of images analysed are available on figshare (10.6084/m9.figshare.875545, 10.6084/m9.figshare.875477).

Mentions: In order to estimate the effect of CCALNN-dHA2 density in the SAM on the uptake of gold nanoparticles, the mean number of gold nanoparticles per unit area of endosome was calculated. For each nanoparticle-containing endosome present on a given image, a region of interest (ROI) was drawn around the endosome and the number of gold nanoparticles within the ROI was counted. This number was then divided by the surface area of the ROI to obtain the density of gold nanoparticle in that particular endosome. Measurements were repeated for each nanoparticle-containing endosome and for each image analysed. For 10% CCALNN-dHA2-capped gold nanoparticles, 29 and 30 images were analysed, respectively for nanoparticles with and without CCALNN-PEG (Fig. 2E). 10 images each were analysed for 50% and 100% CCALNN-dHA2-capped gold nanoparticles (Fig. 3C).


TAT and HA2 facilitate cellular uptake of gold nanoparticles but do not lead to cytosolic localisation.

Cesbron Y, Shaheen U, Free P, Lévy R - PLoS ONE (2015)

Influence of the proportion of CCALNN-dHA2 peptides in the monolayer of gold nanoparticles.HeLa cells were incubated for 4h with 10nm gold nanoparticles (6nM) coated with a monolayer composed of two different proportions of the peptides CCALNN-dHA2 and CALNN, washed thoroughly with PBS, fixed and imaged by electron microscopy. (A-B) TEM images of the cells incubated with nanoparticle with the following peptide monolayer composition: (A) 50% CCALNN-dHA2 and 50% CALNN, (B) 100% CCALNN-dHA2. Arrowheads point toward gold nanoparticles either interacting with vesicular membranes (white) or displaying a cytosolic localisation (black). (C) Density of nanoparticles in endosomes estimated from images shown in (A) (n = 10, middle) and (B) (n = 10, right); quantification for 10% CCALNN-dHA2 (left) is the same as in Fig. 2E (left). The sets of images analysed are available on figshare (10.6084/m9.figshare.875545, 10.6084/m9.figshare.875477).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4383524&req=5

pone.0121683.g003: Influence of the proportion of CCALNN-dHA2 peptides in the monolayer of gold nanoparticles.HeLa cells were incubated for 4h with 10nm gold nanoparticles (6nM) coated with a monolayer composed of two different proportions of the peptides CCALNN-dHA2 and CALNN, washed thoroughly with PBS, fixed and imaged by electron microscopy. (A-B) TEM images of the cells incubated with nanoparticle with the following peptide monolayer composition: (A) 50% CCALNN-dHA2 and 50% CALNN, (B) 100% CCALNN-dHA2. Arrowheads point toward gold nanoparticles either interacting with vesicular membranes (white) or displaying a cytosolic localisation (black). (C) Density of nanoparticles in endosomes estimated from images shown in (A) (n = 10, middle) and (B) (n = 10, right); quantification for 10% CCALNN-dHA2 (left) is the same as in Fig. 2E (left). The sets of images analysed are available on figshare (10.6084/m9.figshare.875545, 10.6084/m9.figshare.875477).
Mentions: In order to estimate the effect of CCALNN-dHA2 density in the SAM on the uptake of gold nanoparticles, the mean number of gold nanoparticles per unit area of endosome was calculated. For each nanoparticle-containing endosome present on a given image, a region of interest (ROI) was drawn around the endosome and the number of gold nanoparticles within the ROI was counted. This number was then divided by the surface area of the ROI to obtain the density of gold nanoparticle in that particular endosome. Measurements were repeated for each nanoparticle-containing endosome and for each image analysed. For 10% CCALNN-dHA2-capped gold nanoparticles, 29 and 30 images were analysed, respectively for nanoparticles with and without CCALNN-PEG (Fig. 2E). 10 images each were analysed for 50% and 100% CCALNN-dHA2-capped gold nanoparticles (Fig. 3C).

Bottom Line: However, conflicting results have been reported on the extent of the cytosolic delivery achieved.To evaluate their potential, we used gold nanoparticles as model cargos and systematically assessed how the functionalisation of their surface by either or both of the viral peptides TAT and HA2 influenced their intracellular delivery.While their uptake increased when the TAT and/or HA2 viral peptides were present on their surface, we did not observe a significant cytosolic delivery of the gold nanoparticles.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Liverpool, Liverpool, United Kingdom; Institute of Integrative Biology, University of Liverpool, Liverpool, United Kingdom; CNRS, UMR 6290, Institute of Genetics and Development of Rennes, Rennes, France; Université de Rennes 1, Université Européenne de Bretagne, Structure fédérative de recherche Biosit, Faculté de Médecine, Rennes, France.

ABSTRACT
The methods currently available to deliver functional labels and drugs to the cell cytosol are inefficient and this constitutes a major obstacle to cell biology (delivery of sensors and imaging probes) and therapy (drug access to the cell internal machinery). As cell membranes are impermeable to most molecular cargos, viral peptides have been used to bolster their internalisation through endocytosis and help their release to the cytosol by bursting the endosomal vesicles. However, conflicting results have been reported on the extent of the cytosolic delivery achieved. To evaluate their potential, we used gold nanoparticles as model cargos and systematically assessed how the functionalisation of their surface by either or both of the viral peptides TAT and HA2 influenced their intracellular delivery. We evaluated the number of gold nanoparticles present in cells after internalisation using photothermal microscopy and their subcellular localisation by electron microscopy. While their uptake increased when the TAT and/or HA2 viral peptides were present on their surface, we did not observe a significant cytosolic delivery of the gold nanoparticles.

No MeSH data available.


Related in: MedlinePlus