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Silver and Gold Nanoparticles Alter Cathepsin Activity In vitro

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ABSTRACT

Nanomaterials are being incorporated into many biological applications for use as therapeutics, sensors, or labels. Silver nanomaterials are being utilized for biological implants and wound dressings as an antiviral material, whereas gold nanomaterials are being used as biological labels or sensors due to their surface properties and biocompatibility. Cytotoxicity data of these materials are becoming more prevalent; however, little research has been performed to understand how the introduction of these materials into cells affects cellular processes. Here, we demonstrate the impact that silver and gold nanoparticles have on cathepsin activity in vitro. Cathepsins are important cellular proteases that are imperative for proper immune system function. We have selected to examine gold and silver nanoparticles due to the increased use of these materials in biological applications. This manuscript depicts how both of these types of nanomaterials affect cathepsin activity, which could impact the host's immune system and its ability to respond to pathogens. Cathepsin B activity decreases in a dose-dependent manner with all nanoparticles tested. Alternatively, the impact of nanoparticles on cathepsin L activity depends greatly on the type and size of the material.

No MeSH data available.


Cathepsin B confocal imaging in Ag-NP-treated Vero cells. A fluorescent substrate cleaved by active cathepsin B was detected using confocal microscopy in Vero cells treated with Ag-NPs or left untreated. a Negative control (Vero cells alone), b Positive control (Vero cells + CV-(FR)2), c 10-nm uncoated Ag-NP 10 μg/ml, d 10-nm uncoated Ag-NP 50 μg/ml, e 10-nm PS-Ag-NP 10 μg/ml, f 10-nm PS-Ag-NP 50 μg/ml, g 25-nm uncoated Ag-NP 10 μg/ml, h 25-nm uncoated Ag-NP 50 μg/ml, i 25-nm PS-Ag-NP 10 μg/ml, j 25-nm PS-Ag-NP 50 μg/ml. Red fluorescent intensity was normalized to Vero cells exposed to the substrate (b). The table below represents a quantitative assessment of the confocal images as determined using a fluorescent plate reader. The values indicate the mean percent of control +/- SEM (n = 6).
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Figure 2: Cathepsin B confocal imaging in Ag-NP-treated Vero cells. A fluorescent substrate cleaved by active cathepsin B was detected using confocal microscopy in Vero cells treated with Ag-NPs or left untreated. a Negative control (Vero cells alone), b Positive control (Vero cells + CV-(FR)2), c 10-nm uncoated Ag-NP 10 μg/ml, d 10-nm uncoated Ag-NP 50 μg/ml, e 10-nm PS-Ag-NP 10 μg/ml, f 10-nm PS-Ag-NP 50 μg/ml, g 25-nm uncoated Ag-NP 10 μg/ml, h 25-nm uncoated Ag-NP 50 μg/ml, i 25-nm PS-Ag-NP 10 μg/ml, j 25-nm PS-Ag-NP 50 μg/ml. Red fluorescent intensity was normalized to Vero cells exposed to the substrate (b). The table below represents a quantitative assessment of the confocal images as determined using a fluorescent plate reader. The values indicate the mean percent of control +/- SEM (n = 6).

Mentions: A significant decrease in red fluorescent intensity, indicating a decrease in cathepsin B activity, was observed in the 50 μg/ml doses of 10 nm both uncoated and PS-coated and 25-nm uncoated Ag-NPs (Figure 2d, f, h) over the untreated control (Figure 2b). There was little visual difference in red fluorescence intensity between the 10 μg/ml treated groups (Figure 2c, e, g, i) and the 25-nm PS-Ag at 50 μg/ml (Figure 2j) from the untreated control (Figure 2b), although the 10-nm PS-Ag and 25-nm uncoated Ag-NPs did have a significant decline in fluorescence intensity (Figure 2 table). The decrease in cathepsin B activity in Vero cells treated with Ag-NPs was confirmed via fluorescent quantification in a fluorescent plate reader and a dose-dependent decrease in cathepsin B activity is observed in all treatment groups except for the 25-nm PS-Ag, which interestingly had no effect on cathepsin B activity (Figure 2 table).


Silver and Gold Nanoparticles Alter Cathepsin Activity In vitro
Cathepsin B confocal imaging in Ag-NP-treated Vero cells. A fluorescent substrate cleaved by active cathepsin B was detected using confocal microscopy in Vero cells treated with Ag-NPs or left untreated. a Negative control (Vero cells alone), b Positive control (Vero cells + CV-(FR)2), c 10-nm uncoated Ag-NP 10 μg/ml, d 10-nm uncoated Ag-NP 50 μg/ml, e 10-nm PS-Ag-NP 10 μg/ml, f 10-nm PS-Ag-NP 50 μg/ml, g 25-nm uncoated Ag-NP 10 μg/ml, h 25-nm uncoated Ag-NP 50 μg/ml, i 25-nm PS-Ag-NP 10 μg/ml, j 25-nm PS-Ag-NP 50 μg/ml. Red fluorescent intensity was normalized to Vero cells exposed to the substrate (b). The table below represents a quantitative assessment of the confocal images as determined using a fluorescent plate reader. The values indicate the mean percent of control +/- SEM (n = 6).
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Figure 2: Cathepsin B confocal imaging in Ag-NP-treated Vero cells. A fluorescent substrate cleaved by active cathepsin B was detected using confocal microscopy in Vero cells treated with Ag-NPs or left untreated. a Negative control (Vero cells alone), b Positive control (Vero cells + CV-(FR)2), c 10-nm uncoated Ag-NP 10 μg/ml, d 10-nm uncoated Ag-NP 50 μg/ml, e 10-nm PS-Ag-NP 10 μg/ml, f 10-nm PS-Ag-NP 50 μg/ml, g 25-nm uncoated Ag-NP 10 μg/ml, h 25-nm uncoated Ag-NP 50 μg/ml, i 25-nm PS-Ag-NP 10 μg/ml, j 25-nm PS-Ag-NP 50 μg/ml. Red fluorescent intensity was normalized to Vero cells exposed to the substrate (b). The table below represents a quantitative assessment of the confocal images as determined using a fluorescent plate reader. The values indicate the mean percent of control +/- SEM (n = 6).
Mentions: A significant decrease in red fluorescent intensity, indicating a decrease in cathepsin B activity, was observed in the 50 μg/ml doses of 10 nm both uncoated and PS-coated and 25-nm uncoated Ag-NPs (Figure 2d, f, h) over the untreated control (Figure 2b). There was little visual difference in red fluorescence intensity between the 10 μg/ml treated groups (Figure 2c, e, g, i) and the 25-nm PS-Ag at 50 μg/ml (Figure 2j) from the untreated control (Figure 2b), although the 10-nm PS-Ag and 25-nm uncoated Ag-NPs did have a significant decline in fluorescence intensity (Figure 2 table). The decrease in cathepsin B activity in Vero cells treated with Ag-NPs was confirmed via fluorescent quantification in a fluorescent plate reader and a dose-dependent decrease in cathepsin B activity is observed in all treatment groups except for the 25-nm PS-Ag, which interestingly had no effect on cathepsin B activity (Figure 2 table).

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Nanomaterials are being incorporated into many biological applications for use as therapeutics, sensors, or labels. Silver nanomaterials are being utilized for biological implants and wound dressings as an antiviral material, whereas gold nanomaterials are being used as biological labels or sensors due to their surface properties and biocompatibility. Cytotoxicity data of these materials are becoming more prevalent; however, little research has been performed to understand how the introduction of these materials into cells affects cellular processes. Here, we demonstrate the impact that silver and gold nanoparticles have on cathepsin activity in vitro. Cathepsins are important cellular proteases that are imperative for proper immune system function. We have selected to examine gold and silver nanoparticles due to the increased use of these materials in biological applications. This manuscript depicts how both of these types of nanomaterials affect cathepsin activity, which could impact the host's immune system and its ability to respond to pathogens. Cathepsin B activity decreases in a dose-dependent manner with all nanoparticles tested. Alternatively, the impact of nanoparticles on cathepsin L activity depends greatly on the type and size of the material.

No MeSH data available.