Cell specific cytotoxicity and uptake of graphene nanoribbons

Abstract

The synthesis of oxidized graphene nanoribbons (O-GNR) via longitudinal unzipping of carbon nanotubes opens avenues for their further development for a variety of biomedical applications. Evaluation of the cyto- and bio-compatibility is necessary to develop any new material for in vivo biomedical applications. In this study, we report the cytotoxicity screening of O-GNRs water-solubilized with PEG-DSPE (1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)]), using six different assays, in four representative cell lines; Henrietta Lacks cells (HeLa) derived from cervical cancer tissue, National Institute of Health 3T3 mouse fibroblast cells (NIH-3T3), Sloan Kettering breast cancer cells (SKBR3) and Michigan cancer foundation-7 breast cancer cells (MCF7). These cell lines significantly differed in their response to O-GNR-PEG-DSPE formulations; assessed and evaluated using various endpoints (lactate dehydrogenase (LDH) release, cellular metabolism, lysosomal integrity and cell proliferation) for cytotoxicity. In general, all the cells showed a dose-dependent (10–400 μg/ml) and time-dependent (12–48 h) decrease in cell viability. However, the degree of cytotoxicity was significantly lower in MCF7 or SKBR3 cells compared to HeLa cells. These cells were 100% viable upto 48 hours, when incubated at 10μg/ml O-GNR-PEG-DSPE concentration, and showed decrease in cell viability above this concentration with ~78% of cells viable at the highest concentration (400 μg/ml). In contrast, significant cell death (5–25% cell death depending on the time point, and the assay) was observed for HeLa cells even at a low concentration of 10μg/ml. The decrease in cell viability was steep with increase in concentration with the CD50 values ≥ 100μg/ml depending on the assay, and time point. Transmission electron microscopy of the various cells treated with the O-GNR solutions show higher uptake of the O-GNR-PEG-DSPEs into HeLa cells compared to other cell types. Additional analysis indicates that this increased uptake is the dominant cause of the significantly higher toxicity exhibited by HeLa cells. The results suggest that water-solubilized O-GNR-PEG-DSPEs have a heterogeneous cell-specific cytotoxicity, and have significantly different cytotoxicity profile compared to graphene nanoparticles prepared by the modified Hummer’s method (graphene nanoparticles prepared by oxidation of graphite, and its mechanical exfoliation) or its variations.

Silver Nanoparticles Induced RNA Polymerase-Silver Binding and RNA Transcription Inhibition in Erythroid Progenitor Cells

“…Alamar Blue assay is an ideal approach to quantitatively measure proliferation and metabolic status of cells, especially for cells cultured in suspension (such as MEL cells used in the current study). 27,28 Similar to the results of cell number counting, the Alamar Blue assay suggested that the concentration of half maximal inhibition on cell viability was greater than 10 μg/mL ( Figure 5B, P < 0.01). In contrast to nAg, MEL cells are fairly vulnerable to Ag ions (in AgNO 3 ) at even very low concentrations (0.08 μg/mL), with significant reduction in cell viability in comparison to the control ( Figure 5C, P < 0.05), and cell viability was dramatically impaired by Ag ions at 1.2 μg/mL ( Figure 5C, P < 0.001).…”

Graphene nanoribbons elicit cell specific uptake and delivery via activation of epidermal growth factor receptor enhanced by human papillomavirus E5 protein

“…In vitro drug delivery studies were conducted using 50 µl of O-GNR-PEG-DSPE solution at a potential therapeutic dosage of 50 µg/ml (determined from previous cytotoxicity studies for O-GNR-PEG-DSPE[10]) loaded with 40% Dox by weight (20 µg/ml); treatment duration was 24 hours and the solution was studied in 11 cell lines (Table 1, cell line selection criteria for these studies were based on previous results). Since HeLa is a cervical cancer cell line with an integrated HPV genome, we chose cell lines comprised of cervical cancers cells with or without an HPV genome, and non-cervical cancer cells with low, normal or high EGFR expression[18–28].…”

“…We used a lactate dehydrogenase (LDH) assay to assess cytotoxicity due to cellular delivery of Dox. LDH, a cytoplasmic marker for membrane integrity, provides an indirect means of assessing cytotoxicity[10]. Leaky membranes of damaged or dead cells show increased LDH release into surrounding media compared to normal intact cells.…”

“…O-GNRs (Figure 1A and B) water-solubilized with the amphiphilic polymer PEG-DSPE to form a supramolecular complex (O-GNR-PEG-DSPE) were incubated at various concentrations (0–400µg/ml) and time points (24–72 hours) in four different cell lines (HeLa, MCF7, SKBR 3 and NIH3T3). O-GNR-PEG-DSPEs were more cytotoxic to HeLa cells compared to other cell lines[10]. These studies provided preliminary indication that enhanced uptake of O-GNR-PEG-DSPEs into HeLa cells was an important reason for the observed differences in cytotoxicity.…”

“…Recently, we evaluated the cytotoxicity of oxidized graphene nanoribbon (O-GNR)-based formulations[10]. O-GNRs were synthesized in macroscopic amounts using an oxidative method pioneered by Kosynkin, Tour, and co-workers that longitudinally “unzips” carbon nanotubes[11].…”