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High content analysis at single cell level identifies different cellular responses dependent on nanomaterial concentrations.

Manshian BB, Munck S, Agostinis P, Himmelreich U, Soenen SJ - Sci Rep (2015)

Bottom Line: A mechanistic understanding of nanomaterial (NM) interaction with biological environments is pivotal for the safe transition from basic science to applied nanomedicine.Upon binning the single cell data into different categories related to NM concentration, this study demonstrates, for the first time, that quantum dots activate both cytoprotective and cytotoxic mechanisms, resulting in a zero net result on the overall cell population, yet with significant effects in cells with higher cellular NM levels.Our results suggest that future NM cytotoxicity studies should correlate NM toxicity with cellular NM numbers on the single cell level, as conflicting mechanisms in particular cell subpopulations are commonly overlooked using classical toxicological methods.

View Article: PubMed Central - PubMed

Affiliation: MoSAIC/Biomedical MRI Unit, Faculty of Medicine, KU Leuven, Herestraat 49, B3000 Leuven, Belgium.

ABSTRACT
A mechanistic understanding of nanomaterial (NM) interaction with biological environments is pivotal for the safe transition from basic science to applied nanomedicine. NM exposure results in varying levels of internalized NM in different neighboring cells, due to variances in cell size, cell cycle phase and NM agglomeration. Using high-content analysis, we investigated the cytotoxic effects of fluorescent quantum dots on cultured cells, where all effects were correlated with the concentration of NMs at the single cell level. Upon binning the single cell data into different categories related to NM concentration, this study demonstrates, for the first time, that quantum dots activate both cytoprotective and cytotoxic mechanisms, resulting in a zero net result on the overall cell population, yet with significant effects in cells with higher cellular NM levels. Our results suggest that future NM cytotoxicity studies should correlate NM toxicity with cellular NM numbers on the single cell level, as conflicting mechanisms in particular cell subpopulations are commonly overlooked using classical toxicological methods.

No MeSH data available.


Related in: MedlinePlus

Cellular distribution of cell-associated fluorescent quantum dots.(a,b) Representative images of (a) MSCs exposed to 10 nM QDots for 24 h or (b) MEFs exposed to 12.5 nMQDots for 24 h. (c,d) Histograms presenting the number of (c) MSC or (d) MEF cells per category when the population is divided into 10 categories spanning the entire range of cellular QDot levels. (e,f) Histograms presenting the percentage of (e) MSC or (f) MEF cells per category for the total population analyzed.
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f2: Cellular distribution of cell-associated fluorescent quantum dots.(a,b) Representative images of (a) MSCs exposed to 10 nM QDots for 24 h or (b) MEFs exposed to 12.5 nMQDots for 24 h. (c,d) Histograms presenting the number of (c) MSC or (d) MEF cells per category when the population is divided into 10 categories spanning the entire range of cellular QDot levels. (e,f) Histograms presenting the percentage of (e) MSC or (f) MEF cells per category for the total population analyzed.

Mentions: When considering average results over the entire visualized population, our data reveal that the effects the QDots induce in MSCs at 10 nM are similar to those observed in MEFs at 12.5 nM. At these concentrations, no significant effects could be observed, apart from the induction of oxidative stress, which does not appear to induce any cell death at these levels due to natural cellular defense mechanisms32. Exposure of both cell types to the QDots (MSC at 10 nM; MEF at 12.5 nM) however resulted in a broad distribution in cellular QDot levels (Fig. 2), as is typical for in vitro cultured cells exposed to NMs31. NM levels between the two cell types were highly similar which explains the high similarity in their toxicity profiles. In order to evaluate the impact of cellular NM distribution on its resulting toxicity, the fluorescent properties of the QDots were exploited to determine relative cellular NM levels, and consequently every cellular parameter was linked to a subpopulation of cells with a certain NM level. NM distribution in cells followed a near-Gaussian profile and cells were subdivided into 10 different categories, category 1 being the lowest NM level and category 10 the highest (Fig. 2c–f). The data reveal that the majority of cells are in the medium categories (c4–c7; 68%). The lower categories (c1–c3) represent merely 12% of the entire population while the higher categories (c8–c10) represent 20% of the population. Interestingly, for MSCs exposed to 10 nM QDots and MEFs exposed to 12.5 nM QDots, the distribution of cellular QDot levels is quite similar, suggesting that the high similarity in cellular effects under these conditions (Fig. 1) is due to the similar levels of intracellular QDots. In general, MEF cells appear to be slightly more resistant to QDot-elicited cytotoxic effects as a secondary mechanism caused by the intrinsic lower internalization efficiency of these cells.


High content analysis at single cell level identifies different cellular responses dependent on nanomaterial concentrations.

Manshian BB, Munck S, Agostinis P, Himmelreich U, Soenen SJ - Sci Rep (2015)

Cellular distribution of cell-associated fluorescent quantum dots.(a,b) Representative images of (a) MSCs exposed to 10 nM QDots for 24 h or (b) MEFs exposed to 12.5 nMQDots for 24 h. (c,d) Histograms presenting the number of (c) MSC or (d) MEF cells per category when the population is divided into 10 categories spanning the entire range of cellular QDot levels. (e,f) Histograms presenting the percentage of (e) MSC or (f) MEF cells per category for the total population analyzed.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4561960&req=5

f2: Cellular distribution of cell-associated fluorescent quantum dots.(a,b) Representative images of (a) MSCs exposed to 10 nM QDots for 24 h or (b) MEFs exposed to 12.5 nMQDots for 24 h. (c,d) Histograms presenting the number of (c) MSC or (d) MEF cells per category when the population is divided into 10 categories spanning the entire range of cellular QDot levels. (e,f) Histograms presenting the percentage of (e) MSC or (f) MEF cells per category for the total population analyzed.
Mentions: When considering average results over the entire visualized population, our data reveal that the effects the QDots induce in MSCs at 10 nM are similar to those observed in MEFs at 12.5 nM. At these concentrations, no significant effects could be observed, apart from the induction of oxidative stress, which does not appear to induce any cell death at these levels due to natural cellular defense mechanisms32. Exposure of both cell types to the QDots (MSC at 10 nM; MEF at 12.5 nM) however resulted in a broad distribution in cellular QDot levels (Fig. 2), as is typical for in vitro cultured cells exposed to NMs31. NM levels between the two cell types were highly similar which explains the high similarity in their toxicity profiles. In order to evaluate the impact of cellular NM distribution on its resulting toxicity, the fluorescent properties of the QDots were exploited to determine relative cellular NM levels, and consequently every cellular parameter was linked to a subpopulation of cells with a certain NM level. NM distribution in cells followed a near-Gaussian profile and cells were subdivided into 10 different categories, category 1 being the lowest NM level and category 10 the highest (Fig. 2c–f). The data reveal that the majority of cells are in the medium categories (c4–c7; 68%). The lower categories (c1–c3) represent merely 12% of the entire population while the higher categories (c8–c10) represent 20% of the population. Interestingly, for MSCs exposed to 10 nM QDots and MEFs exposed to 12.5 nM QDots, the distribution of cellular QDot levels is quite similar, suggesting that the high similarity in cellular effects under these conditions (Fig. 1) is due to the similar levels of intracellular QDots. In general, MEF cells appear to be slightly more resistant to QDot-elicited cytotoxic effects as a secondary mechanism caused by the intrinsic lower internalization efficiency of these cells.

Bottom Line: A mechanistic understanding of nanomaterial (NM) interaction with biological environments is pivotal for the safe transition from basic science to applied nanomedicine.Upon binning the single cell data into different categories related to NM concentration, this study demonstrates, for the first time, that quantum dots activate both cytoprotective and cytotoxic mechanisms, resulting in a zero net result on the overall cell population, yet with significant effects in cells with higher cellular NM levels.Our results suggest that future NM cytotoxicity studies should correlate NM toxicity with cellular NM numbers on the single cell level, as conflicting mechanisms in particular cell subpopulations are commonly overlooked using classical toxicological methods.

View Article: PubMed Central - PubMed

Affiliation: MoSAIC/Biomedical MRI Unit, Faculty of Medicine, KU Leuven, Herestraat 49, B3000 Leuven, Belgium.

ABSTRACT
A mechanistic understanding of nanomaterial (NM) interaction with biological environments is pivotal for the safe transition from basic science to applied nanomedicine. NM exposure results in varying levels of internalized NM in different neighboring cells, due to variances in cell size, cell cycle phase and NM agglomeration. Using high-content analysis, we investigated the cytotoxic effects of fluorescent quantum dots on cultured cells, where all effects were correlated with the concentration of NMs at the single cell level. Upon binning the single cell data into different categories related to NM concentration, this study demonstrates, for the first time, that quantum dots activate both cytoprotective and cytotoxic mechanisms, resulting in a zero net result on the overall cell population, yet with significant effects in cells with higher cellular NM levels. Our results suggest that future NM cytotoxicity studies should correlate NM toxicity with cellular NM numbers on the single cell level, as conflicting mechanisms in particular cell subpopulations are commonly overlooked using classical toxicological methods.

No MeSH data available.


Related in: MedlinePlus