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Alkyl-capped silicon nanocrystals lack cytotoxicity and have enhanced intracellular accumulation in malignant cells via cholesterol-dependent endocytosis.

Alsharif NH, Berger CE, Varanasi SS, Chao Y, Horrocks BR, Datta HK - Small (2009)

Bottom Line: The internalization of the nanocrystals by Hela and SW1353 cells is almost completely blocked by the pinocytosis inhibitors filipin, cytochalasin B, and actinomycin D.The internalization process is not associated with any surface change in the nanoparticles, as their luminescence spectrum is unaltered upon transport into the cytosol.The observed dramatic difference in the rate and extent of internalization of the nanocrystals between malignant and non-malignant cells therefore offers potential application in the management of human neoplastic conditions.

View Article: PubMed Central - PubMed

Affiliation: Musculoskeletal Research Group, Institute of Cellular Medicine Newcastle University, Framlington Place Newcastle upon Tyne, UK.

ABSTRACT
Nanocrystals of various inorganic materials are being considered for application in the life sciences as fluorescent labels and for such therapeutic applications as drug delivery or targeted cell destruction. The potential applications of the nanoparticles are critically compromised due to the well-documented toxicity and lack of understanding about the mechanisms involved in the intracellular internalization. Here intracellular internalization and toxicity of alkyl-capped silicon nanocrystals in human neoplastic and normal primary cells is reported. The capped nanocrystals lack cytotoxicity, and there is a marked difference in the rate and extent of intracellular accumulation of the nanoparticles between human cancerous and non-cancerous primary cells, the rate and extent being higher in the malignant cells compared to normal human primary cells. The exposure of the cells to the alkyl-capped nanocrystals demonstrates no evidence of in vitro cytotoxicity when assessed by cell morphology, apoptosis, and cell viability assays. The internalization of the nanocrystals by Hela and SW1353 cells is almost completely blocked by the pinocytosis inhibitors filipin, cytochalasin B, and actinomycin D. The internalization process is not associated with any surface change in the nanoparticles, as their luminescence spectrum is unaltered upon transport into the cytosol. The observed dramatic difference in the rate and extent of internalization of the nanocrystals between malignant and non-malignant cells therefore offers potential application in the management of human neoplastic conditions.

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Screening for alkyl-SiNC cytotoxicity. a) Hela and SW1353 cells were exposed to 2 µL of nanocrystals or vehicle in 1 mL culture medium for 30 min (white columns), and 1 h (dark columns), and observed under a phase-contrast microscope. There was no evidence of acute cytotoxicity, as evident from apoptosis (colorimetric TUNEL) assay performed on SW1353 and Hela cell-lines. b) The possible cytotoxic effect of NCs was excluded by the lack of effect in the cell proliferation and viability assays. SW1353 and Hela cells that had been starved overnight were exposed to medium containing dots or vehicle for variable times (1 h (white column), 4 h (gray column), and 24 h (dark column)), and cell viability assays were carried out using the CellTiter-Blue Cell Viability Assay. c) For the MTT assay, cells were exposed for to NCs for 2 h (white column), 4 h (gray column), and 24 h (dark column). The cells were exposed to either NCs suspended in 2 µL ether and controls were exposed to vehicle (2 µL ether) alone. All experiments ranged between n = 5 to 8 and were repeated on at least three different occasions.
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fig03: Screening for alkyl-SiNC cytotoxicity. a) Hela and SW1353 cells were exposed to 2 µL of nanocrystals or vehicle in 1 mL culture medium for 30 min (white columns), and 1 h (dark columns), and observed under a phase-contrast microscope. There was no evidence of acute cytotoxicity, as evident from apoptosis (colorimetric TUNEL) assay performed on SW1353 and Hela cell-lines. b) The possible cytotoxic effect of NCs was excluded by the lack of effect in the cell proliferation and viability assays. SW1353 and Hela cells that had been starved overnight were exposed to medium containing dots or vehicle for variable times (1 h (white column), 4 h (gray column), and 24 h (dark column)), and cell viability assays were carried out using the CellTiter-Blue Cell Viability Assay. c) For the MTT assay, cells were exposed for to NCs for 2 h (white column), 4 h (gray column), and 24 h (dark column). The cells were exposed to either NCs suspended in 2 µL ether and controls were exposed to vehicle (2 µL ether) alone. All experiments ranged between n = 5 to 8 and were repeated on at least three different occasions.

Mentions: In vitro cytotoxicity investigations, comprising apoptosis, cell viability, and proliferation assays, were carried out on Hela and SW1353 cells (human chondrosarcoma). In all these studies, solutions of alkyl-SiNCs in ether or vehicle alone (2 µL) were added to a Hela cell monolayer culture in 1 mL of culture medium for a variable period ranging between 1 h and 24 h. When the cells were observed under a phase contrast microscope there was no evidence of acute cytotoxicity, of cell necrosis, or of plasma membrane blebbing. The lack of observed cellular toxicity was further confirmed by the apoptosis assay (colorimetric TdT-mediated dUTP Nick-End Labeling (TUNEL) assay) performed on Hela and SW1353 cell-lines (Figure 3a). There were no significant differences in the rate of apoptosis between cells exposed to alkyl-SiNCs and those exposed to vehicle alone. A possible chronic cytotoxic effect of alkyl-SiNCs was investigated by exposing cells to alkyl-SiNCs for up to 24 h. These assays determined changes in the rate of cell proliferation and cell viability in the presence of constant exposure to alkyl-SiNCs in the culture medium (Figure 3b). It was found to show no effect on the rate of cell proliferation (the number of NCs in the medium was ∼3 × 1012 or 5 pmol). No effect on cell proliferation was seen even after cells had been exposed to alkyl-SiNCs for 24 h (Figure 3c).


Alkyl-capped silicon nanocrystals lack cytotoxicity and have enhanced intracellular accumulation in malignant cells via cholesterol-dependent endocytosis.

Alsharif NH, Berger CE, Varanasi SS, Chao Y, Horrocks BR, Datta HK - Small (2009)

Screening for alkyl-SiNC cytotoxicity. a) Hela and SW1353 cells were exposed to 2 µL of nanocrystals or vehicle in 1 mL culture medium for 30 min (white columns), and 1 h (dark columns), and observed under a phase-contrast microscope. There was no evidence of acute cytotoxicity, as evident from apoptosis (colorimetric TUNEL) assay performed on SW1353 and Hela cell-lines. b) The possible cytotoxic effect of NCs was excluded by the lack of effect in the cell proliferation and viability assays. SW1353 and Hela cells that had been starved overnight were exposed to medium containing dots or vehicle for variable times (1 h (white column), 4 h (gray column), and 24 h (dark column)), and cell viability assays were carried out using the CellTiter-Blue Cell Viability Assay. c) For the MTT assay, cells were exposed for to NCs for 2 h (white column), 4 h (gray column), and 24 h (dark column). The cells were exposed to either NCs suspended in 2 µL ether and controls were exposed to vehicle (2 µL ether) alone. All experiments ranged between n = 5 to 8 and were repeated on at least three different occasions.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig03: Screening for alkyl-SiNC cytotoxicity. a) Hela and SW1353 cells were exposed to 2 µL of nanocrystals or vehicle in 1 mL culture medium for 30 min (white columns), and 1 h (dark columns), and observed under a phase-contrast microscope. There was no evidence of acute cytotoxicity, as evident from apoptosis (colorimetric TUNEL) assay performed on SW1353 and Hela cell-lines. b) The possible cytotoxic effect of NCs was excluded by the lack of effect in the cell proliferation and viability assays. SW1353 and Hela cells that had been starved overnight were exposed to medium containing dots or vehicle for variable times (1 h (white column), 4 h (gray column), and 24 h (dark column)), and cell viability assays were carried out using the CellTiter-Blue Cell Viability Assay. c) For the MTT assay, cells were exposed for to NCs for 2 h (white column), 4 h (gray column), and 24 h (dark column). The cells were exposed to either NCs suspended in 2 µL ether and controls were exposed to vehicle (2 µL ether) alone. All experiments ranged between n = 5 to 8 and were repeated on at least three different occasions.
Mentions: In vitro cytotoxicity investigations, comprising apoptosis, cell viability, and proliferation assays, were carried out on Hela and SW1353 cells (human chondrosarcoma). In all these studies, solutions of alkyl-SiNCs in ether or vehicle alone (2 µL) were added to a Hela cell monolayer culture in 1 mL of culture medium for a variable period ranging between 1 h and 24 h. When the cells were observed under a phase contrast microscope there was no evidence of acute cytotoxicity, of cell necrosis, or of plasma membrane blebbing. The lack of observed cellular toxicity was further confirmed by the apoptosis assay (colorimetric TdT-mediated dUTP Nick-End Labeling (TUNEL) assay) performed on Hela and SW1353 cell-lines (Figure 3a). There were no significant differences in the rate of apoptosis between cells exposed to alkyl-SiNCs and those exposed to vehicle alone. A possible chronic cytotoxic effect of alkyl-SiNCs was investigated by exposing cells to alkyl-SiNCs for up to 24 h. These assays determined changes in the rate of cell proliferation and cell viability in the presence of constant exposure to alkyl-SiNCs in the culture medium (Figure 3b). It was found to show no effect on the rate of cell proliferation (the number of NCs in the medium was ∼3 × 1012 or 5 pmol). No effect on cell proliferation was seen even after cells had been exposed to alkyl-SiNCs for 24 h (Figure 3c).

Bottom Line: The internalization of the nanocrystals by Hela and SW1353 cells is almost completely blocked by the pinocytosis inhibitors filipin, cytochalasin B, and actinomycin D.The internalization process is not associated with any surface change in the nanoparticles, as their luminescence spectrum is unaltered upon transport into the cytosol.The observed dramatic difference in the rate and extent of internalization of the nanocrystals between malignant and non-malignant cells therefore offers potential application in the management of human neoplastic conditions.

View Article: PubMed Central - PubMed

Affiliation: Musculoskeletal Research Group, Institute of Cellular Medicine Newcastle University, Framlington Place Newcastle upon Tyne, UK.

ABSTRACT
Nanocrystals of various inorganic materials are being considered for application in the life sciences as fluorescent labels and for such therapeutic applications as drug delivery or targeted cell destruction. The potential applications of the nanoparticles are critically compromised due to the well-documented toxicity and lack of understanding about the mechanisms involved in the intracellular internalization. Here intracellular internalization and toxicity of alkyl-capped silicon nanocrystals in human neoplastic and normal primary cells is reported. The capped nanocrystals lack cytotoxicity, and there is a marked difference in the rate and extent of intracellular accumulation of the nanoparticles between human cancerous and non-cancerous primary cells, the rate and extent being higher in the malignant cells compared to normal human primary cells. The exposure of the cells to the alkyl-capped nanocrystals demonstrates no evidence of in vitro cytotoxicity when assessed by cell morphology, apoptosis, and cell viability assays. The internalization of the nanocrystals by Hela and SW1353 cells is almost completely blocked by the pinocytosis inhibitors filipin, cytochalasin B, and actinomycin D. The internalization process is not associated with any surface change in the nanoparticles, as their luminescence spectrum is unaltered upon transport into the cytosol. The observed dramatic difference in the rate and extent of internalization of the nanocrystals between malignant and non-malignant cells therefore offers potential application in the management of human neoplastic conditions.

Show MeSH
Related in: MedlinePlus