Silver nanoparticles (AgNPs) have been proven to have potent antibacterial properties, offering an attractive alternative to antibiotics in the treatment of several infections such as otitis media. Concerns have been raised though regarding their toxicity. There are few data regarding the toxic effects of AgNPs in cochlear cells. The aim of our study was to evaluate the effects of AgNPs of four sizes as a function of their size on HEI-OC1 cochlear cells and on HaCaT keratinocytes. The cells were treated with different concentrations of AgNPs. We evaluated silver uptake by atomic absorption spectroscopy and transmission electron microscopy (TEM), cytotoxicity with the alamarBlue test, ROS production with 2′,7′-dichloro-dihydro-fluorescein diacetate, and genotoxicity with the comet assay. Silver intracellular concentration increased proportionally with the incubation time and the size of the NPs. Silver uptake was higher in HEI-OC1 cells compared to HaCaT. While after 4 h exposure, only the 50 nm NPs were observed in both cell lines and only the 5 nm NPs were observed in the HaCaT cells, after 24 h, nanoparticles of all sizes could be visualized in both cell lines. The cells showed signs of distress: vacuolizations, autophagosomes, signs of apoptosis, or cellular debris. AgNPs of all sizes reduced viability proportionally with the concentration, HEI-OC1 cells being more affected. The toxicity of AgNPs decreased with the nanoparticle size, and ROS production was dose and size dependent, mainly in the cochlear cells. Genotoxicity assessed by comet assay revealed a higher level of DNA lesions in HEI-OC1 cells after treatment with small-sized AgNPs. The perspective of using AgNPs in the treatment of otitis media, although very attractive, must be regarded with caution: cochlear cells proved to be more sensitive to the toxic effect of AgNPs compared to other cell lines. Potential treatments must be tailored specifically, choosing NPs with minimum toxicity towards auditory cells.

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BackgroundCancer cells frequently adopt cellular and molecular alterations and acquire resistance to cytostatic drugs. Chemotherapy with oxaliplatin is among the leading treatments for colorectal cancer with a response rate of 50%, inducing intrastrand cross-links on the DNA. Despite of this drug’s efficiency, resistance develops in nearly all metastatic patients. Chemoresistance being of crucial importance for the drug’s clinical efficiency this study aimed to contribute to the identification and description of some cellular and molecular alterations induced by prolonged oxaliplatin therapy. Resistance to oxaliplatin was induced in Colo320 (Colo320R) and HT-29 (HT-29R) colorectal adenocarcinoma cell lines by exposing the cells to increasing concentrations of the drug. Alterations in morphology, cytotoxicity, DNA cross-links formation and gene expression profiles were assessed in the parental and resistant variants with microscopy, MTT, alkaline comet and pangenomic microarray assays, respectively.ResultsMorphology analysis revealed epithelial-to-mesenchymal transition in the resistant vs parental cells suggesting alterations of the cells’ adhesion complexes, through which they acquire increased invasiveness and adherence. Cytotoxicity measurements demonstrated resistance to oxaliplatin in both cell lines; Colo320 being more sensitive than HT-29 to this drug (P < 0.001). The treatment with oxaliplatin caused major DNA cross-links in both parental cell lines; in Colo320R small amounts of DNA cross-links were still detectable, while in HT-29R not. We identified 441 differentially expressed genes in Colo320R and 613 in HT-29R as compared to their parental counterparts (at least 1.5 -fold up- or down- regulation, p < 0.05). More disrupted functions and pathways were detected in HT-29R cell line than in Colo320R, involving genes responsible for apoptosis inhibition, cellular proliferation and epithelial-to-mesenchymal transition. Several upstream regulators were detected as activated in HT-29R cell line, but not in Colo320R.ConclusionsOur findings revealed a more resistant phenotype in HT-29R as compared to Colo320R and different cellular and molecular chemoresistance patterns induced by prolonged treatment with oxaliplatin in cell lines with identical origins (colorectal adenocarcinomas).

Gold nanoparticles (GNPs) were obtained by green synthesis with an extract fromCornus masfruits (GNPs-CM), characterized by several methods, and their biologic effects were evaluated on two cell lines: HaCaT, normal keratinocytes, and A431, epidermoid carcinoma. GNPs were spherical with sizes between 2 and 24 nm. Their optical spectra had a dominant plasmonic band centered at 525 nm; zeta potential distribution was narrow, centered at −19.7 mV, and the mean hydrodynamic diameter was 58 nm. GNPs were visualized in both cell types entering the cells by endocytosis. The amount of gold uptaken by the cells was dose and time dependent. The intracellular concentration of Au ions was higher in HaCaT compared to A431 cells. The toxicity of GNPs-CM was dose dependent being significant only when the highest concentrations were employed. A431 cells were less affected compared to HaCaT cells, but the difference was not statistically significant. ROS production was not significant, except in HaCaT cells at the highest concentration. The comet assay revealed no significant supplementary DNA lesions, while the secretion of inflammatory cytokines was modulated by the presence of GNPs only when the cells were additionally irradiated with UVB. These results recommend GNPs-CM for further testing and possible dermatological applications.

The present paper examines one of the multiple uses of the CellScan technique, its utilization in the biological evaluation of novel compounds, in order to improve the methods concerning the prediction of their anti-proliferative effects and application as anti-cancer drugs. The CellScan is a laser scanning static cytometer, enabling repetitive spectroscopic measurements in intact living cells. The detection of cell viability and apoptosis is possible, based on the transformations that occur in the cytoplasm matrix of tumour cells influenced by cytotoxic compounds. The measurement of fluorescence changes, due to this phenomenon, is possible with the CellScan system. The potential of this technology to detect the in vitro effects of the inhibitory molecules on tumour cells was demonstrated, making this method a valuable tool for chemosensitivity tests. We synthesized and fully characterised three novel platinum complexes of tertiary arsine ligands: trans-[PtI2(2-iPrOC6H4AsPh2)2] (), trans-[PtCl2(2-MeOC6H4AsPh2)2] () and cis-[PtCl2(2-HOC6H4AsPh2)2] (). The three compounds are biologically active against tumour cells and their cytotoxicity is comparable with standard drugs. Measurements using the CellScan technology correlate well with the results provided by other bioassay methods.

Despite the notable efficacy of oxaliplatin in the treatment of colorectal cancers, the metastatic tumours ultimately become resistant to the drug. This study investigated whether the oxaliplatin-resistant cells display different behaviour to this drug versus the sensitive cells and if this difference may be further exploited into the clinical treatments improvement. In order to establish a stable cell line resistant to oxaliplatin, a human colorectal cancer cell line (Colo320) was exposed to increasing doses of the drug up to the clinically relevant plasma concentration. Four cell groups with different levels of chemoresistance were subjected to additional doses of oxaliplatin, and their cytotoxicity, apoptosis and DNA damage production were assessed. Cells selected for resistance to oxaliplatin reacted differently to the application of additional doses of the drug, displaying lower toxicity and cellular death and fewer DNA cross-links formation, in accordance with the extent of the oxaliplatin pretreatments. As the cross-links formation by oxaliplatin being the main cause for cytotoxicity of this drug and a correlation between cytotoxicity and clinical outcome being shown repeatedly, we consider that the evaluation of oxaliplatin-induced cytotoxicity, apoptosis and DNA damage could be a valuable tool to assess the tumour cells sensitivity and thus to predict the response to chemotherapy.

The reactive oxygen species (ROS) production due to ultraviolet B (UV-B) exposure is extremely harmful to the skin. It causes lesions of DNA, proteins and lipids and leads to cellular death. In the present study the UV-B-induced ROS and subsequent apoptosis in the human keratinocyte cell line (HaCaT) were counterbalanced by a plant extract with antioxidant capacity. Some molecules modulated by common heather (Calluna vulgaris) (CV) extract through which this may exert its photoprotective effects were also identified. The ROS were evaluated with CM-H2DCFDA assay, while apoptosis and Bax-α/Bcl-xL molecules with ELISA. The extract was standardized according to its polyphenolic content and the most important biologically active compounds, such as hyperozid, quercetin, isoquercetin, kampferol were evidenced by high-performance liquid chromatography. The UV-B induced ROS production occurred at its highest level at 2 h after the exposure of the HaCaT cells, while apoptosis later, at 4 h. The most significant changes in Bax-α and Bcl-XL proteins induced by UV-B, as well as the highest effect of the extract on apoptosis, were both registered at 4 h. The CV extract decreased concentration- and time-dependently the UV-B-induced ROS production and prevented apoptosis. These effects of CV occurred, at least to a certain extent, due to the modulation of Bax-α/Bcl-XL proteins. These findings suggest that skin cells could be protected from some of the UV-B-induced harmful effects by the administration of the CV extract, which may be further exploited as a potential photoprotective agent.

This review focuses on the clinical translation of preclinical studies, especially those that have used stem cells in the treatment of glaucoma, with an emphasis on optic nerve regeneration. The studies referred to in the review aim to treat optic nerve atrophy, while cell therapies targeting other sites in the eye, such as the trabecular meshwork, have not been addressed. Such complex and varied pathophysiological mechanisms that lead to glaucoma may explain the fact that although stem cells have a high capacity of neuronal regeneration, the treatments performed did not have the expected results and the promise offered by animal studies was not achieved. By analyzing the facts associated with failure, important lessons are to be learned: the type of stem cells that are used, the route of administration, the selection of patients eligible for these treatments, additional therapies that support stem cells transplantation and their mode of action, methods of avoiding the host’s immune response. Many of these problems could be solved using exosomes (EV), but also miRNA, which allows more targeted approaches with minimal side effects.