Toxicity Effect of Silver Nanoparticles on Photosynthetic Pigment Content, Growth, ROS Production and Ultrastructural Changes of Microalgae Chlorella vulgaris.

Silver nanoparticles (Ag NPs) exhibit antibacterial activity and are extensively used in numerous applications. The aim of this study was to examine the toxic effect of Ag NPs on the marine microalga, Chlorella vulgaris. The microalgae, at the exponential growth phase, were treated with different concentrations of Ag NPs (50 and 100 nm) for 96 h. X-Ray diffraction (XRD) results indicated that the used NPs are single and pure Ag phase with a mean crystallite size of 21 and 32 nm. Ag NPs were found to have a negative effect on viable cell concentration, a variable effect on chlorophyll a concentration, and increased ROS formation. Transmission electron microscopy (TEM) analysis revealed that Ag NPs were present inside the microalgae cells and formed large aggregates in the culture medium. Ag+ ions, in the form of AgNO3, were also assessed at higher concentrations and found to cause inhibitory effects.

Enhanced antitumor activity of carbendazim on HeLa cervical cancer cells by aptamer mediated controlled release.

Carbendazim, is a broad-spectrum fungicide and also a promising experimental antitumor drug as reproduction and developmental toxicant, which is currently under phase II preclinical trials. In this study, an approach based on controlled and targeted release with aptamers and mesoporous silica nanoparticles was investigated to improve the antitumor activity of carbendazim. To this end, we synthesized aptamer conjugated silica nanoparticles for testing cytotoxicity properties in vitro with human cervical adenocarcinoma (HeLa) cultured cells. Nucleolin (AS1411) binding aptamers were used to entrap carbendazim molecules inside nanopores of MCM-41 type silica nanoparticles to obtain a stimuli-dependent release system. The effect of carbendazim loaded aptamer silica complex was tested and compared to free carbendazim treatment on HeLa cells, demonstrating 3.3 fold increase of toxicity on targeted cells with our delivery system. In addition, cytotoxicity of the complex was determined to be mostly due to increased apoptosis and to a less extend necrosis related pathways.

Effects of carbendazim and astaxanthin co-treatment on the proliferation of MCF-7 breast cancer cells.

There has been a controversy in the oncology field about the use of antioxidants along with chemotherapeutics in cancer treatment. This study aimed to investigate the effects of a potent antioxidant (astaxanthin) co-treatment with a promising anti-cancer drug (carbendazim), which is in phase I clinical trials, on MCF-7 breast cancer cell proliferation. MCF-7 cells were treated with carbendazim, astaxanthin, or their combinations and incubated for 24 h. After the incubation, each treatment group was evaluated for proliferation, cell cycle progression, and production of reactive oxygen species (ROS) using WST-1, flow cytometry, and CM-H2DCFDA, respectively. All tested carbendazim and astaxanthin combinations increased the anti-proliferative effect of Carb treatment alone and increased the G2/M phase cell cycle arrest compared to the DMSO-treated control. Astaxanthin, at all concentrations tested, reduced the elevated intracellular ROS levels induced by the carbendazim treatment. Our data suggest that astaxanthin and carbendazim co-treatment enhances the anti-proliferative effect of carbendazim as a single agent, while alleviating the carbendazim treatment-associated ROS production in MCF-7 cells. These findings may contribute to the current debate on the use of antioxidants along with anti-cancer drugs in cancer chemotherapy.

Surface-Enhanced Raman Scattering for Label-Free Living Single Cell Analysis.

Single cell analysis is an active research area with the hope that cellular process can be deciphered from a single living cell other than a cell population. Surface enhanced Raman scattering (SERS) has been increasingly investigated for single cell analysis with its ability to provide information about real-time dynamics of molecular processes taking place in living cells, especially upon external stimulation, in a contactless, noninvasive, and nondestructive way. In this perspective, the fundamental concepts of single cell-SERS analysis including origin of spectral bands and experimental parameters for spectral reproducibility are summarized along with the recent developments.

Investigating the Origins of Toxic Response in TiO2 Nanoparticle-Treated Cells.

Titanium dioxide nanoparticles (TiO2 NPs) are widely used in sunscreens, cosmetics and body implants, and this raises toxicity concerns. Although a large number of reports claim that they are safe to use, others claim that they induce reactive oxygen species formation and can be carcinogenic. In this study, the origins of toxic response to TiO2 NPs were investigated by using Surface-enhanced Raman spectroscopy (SERS) which provides multidimensional information on the cellular dynamics at single cell level without any requirement for cell fixation. Three cell lines of vein (HUVEC), lung carcinoma (A549) and skin (L929) origin were tested for their toxic response upon exposure to 20, 40, 80 and 160 µg/mL anatase-TiO2 NPs for 24 h. It was demonstrated that the level of toxic response is both cell line and dose-dependent. L929 fibroblasts were the most resistant cell line to oxidative stress whereas in HUVEC and A549, cell lines collagen and lipid deformation were observed, respectively.

Gold Nanoparticles in Single-Cell Analysis for Surface Enhanced Raman Scattering.

The need for new therapeutic approaches in the treatment of challenging diseases such as cancer, which often consists of a highly heterogeneous and complex population of cells, brought up the idea of analyzing single cells. The development of novel techniques to analyze single cells has been intensively studied to fully understand specific alternations inducing abnormalities in cellular function. One of the techniques used for single cell analysis is surface-enhanced Raman spectroscopy (SERS) in which a noble metal nanoparticle is used to enhance Raman scattering. Due to its low toxicity and biocompatibility, gold nanoparticles (AuNPs) are commonly preferred as SERS substrates in single cell analysis. The intracellular uptake, localization and toxicity issues of AuNPs are the critical points for interpretation of data since the obtained SERS signals originate from molecules in close vicinity to AuNPs that are taken up by the cells. In this review, the AuNP-living cell interactions, cellular uptake and toxicity of AuNPs in relation to their physicochemical properties, and surface-enhanced Raman scattering from single cells are discussed.

Surface-Enhanced Raman Scattering to Evaluate Nanomaterial Cytotoxicity on Living Cells.

The increasing number of reports about false positive or negative results from conventional cytotoxicity assays of nanomaterials (NMs) suggests that more reliable NM toxicity assessment methods should be developed. Here, we report a novel approach for nanotoxicity evaluation based on surface-enhanced Raman spectroscopy (SERS). Three model NMs were tested on two model cell lines and the results were validated by WST-1 cytotoxicity assay and annexin V-FITC/propidium iodide (PI) staining as apoptosis-necrosis assay. The localization of nanoparticles (NPs) in the cells and the cellular conditions upon NP incubation were visualized by transmission electron microscopy (TEM) and enhanced dark-field (EDF) microscopy. SERS revealed a broader view on the consequences of cell-NM interactions compared to the conventional cytotoxicity assays where only one aspect of toxicity can be measured by one assay type. The results suggest that SERS can significantly contribute to the cytotoxicity evaluation bypassing NM or assay component-related complications with less effort.

The novel HSP90 inhibitor AT13387 potentiates radiation effects in squamous cell carcinoma and adenocarcinoma cells.

Overexpression of heat shock protein 90 (HSP90) is associated with increased tumor cell survival and radioresistance. In this study we explored the efficacy of the novel HSP90 inhibitor AT13387 and examined its radiosensitizing effects in combination with gamma-radiation in 2D and 3D structures as well as mice-xenografts. AT13387 induced effective cytotoxic activity and radiosensitized cancer cells in monolayer and tumor spheroid models, where low drug doses triggered significant synergistic effects on cell survival together with radiation. Furthermore, AT13387 treatment resulted in G2/M-phase arrest and significantly reduced the migration capacity. The expression of selected client proteins involved in DNA repair, cell-signaling and cell growth was downregulated in vitro, though the expression of most investigated proteins recurred after 8-24 h. These results were confirmed in vivo where AT13387 treated tumors displayed effective downregulation of HSP90 and its oncogenic client proteins.In conclusion, our results demonstrate that AT13387 is a potent new cancer drug and effective radiosensitizer in vitro with an excellent in vivo efficacy. AT13387 treatment has the potential to improve external beam therapy and radionuclide therapy outcomes and restore treatment efficacy in cancers that are resistant to initial therapeutic regimes.

Selection and in vitro characterization of human CD44v6-binding antibody fragments.

The cluster of differentiation (CD) 44v6 antigen has been suggested to be involved in tumor formation, invasion, and metastasis formation, and has been observed in a majority of primary and metastatic squamous cell carcinomas of the head and neck. Probes specifically binding to this region may be utilized as tools for the challenging tasks of early detection and targeted treatments of small residual disease. In this project, an epitope-guided phage display selection of human fragment antigen-binding (Fab) fragments with affinity to the v6 sequence was performed. A selected set of Fab fragments was shown to specifically recognize increasingly complex forms of the target sequence, both in the form of a short synthetic or recombinant peptide and in the context of a purified extracellular domain of CD44. The binding was independent of known v6-sequence variation and posttranslational modifications that are common in the CD44 protein family. Furthermore, real-time interaction measurements on antibody fragments labeled with ¹²5I showed specific and high-affinity binding to the antigen present on cultured head and neck squamous cell carcinoma cells. There was no cross-reactivity toward cells that lack the target protein. As hypothesized, characterization of the interaction between Fab fragments and the targets using the mathematical tool Interaction Map revealed more heterogeneous interactions on cells than with pure proteins analyzed by surface plasmon resonance. One main candidate Fab fragment with optimal affinity for all forms of the target sequence was identified. The flexible recombinant source of the Fab fragments might aid the development of tailored molecules adapted for therapeutic or diagnostic applications in the future.