Concentration-Dependent Pro- and Antitumor Activities of Quercetin in Human Melanoma Spheroids: Comparative Analysis of 2D and 3D Cell Culture Models.

Quercetin, a dietary flavonoid found in fruits and vegetables, has been described as a substance with many anti-cancer properties in a variety of preclinical investigations. In the present study, we demonstrate that 2D and 3D melanoma models exhibit not only different sensitivities to quercetin, but also opposite, cancer-promoting effects when metastatic melanoma spheroids are treated with quercetin. Higher concentrations of quercetin reduce melanoma growth in three tested cell lines, whereas low concentrations induce the opposite effect in metastatic melanoma spheroids but not in the non-metastatic cell line. High (>12.5 µM) or low (<6.3 µM) quercetin concentrations decrease or enhance cell viability, spheroid size, and cell proliferation, respectively. Additionally, melanoma cells cultivated in 2D already show significant caspase 3 activity at very low concentrations (>0.4 µM), whereas in 3D spheroids apoptotic cells, caspase 3 activity can only be detected in concentrations ≥12.5 µM. Further, we show that the tumor promoting or repressing effect in the 3D metastatic melanoma spheroids are likely to be elicited by a precisely controlled regulation of Nrf2/ARE-mediated cytoprotective genes, as well as ERK and NF-κB phosphorylation. According to the results obtained here, further studies are needed to better characterize the mechanisms of action underlying the pro- and anti-carcinogenic effects of quercetin on human melanomas.

A Screening Approach for Identifying Gliadin Neutralizing Antibodies on Epithelial Intestinal Caco-2 Cells.

Celiac disease (CD) is a chronic inflammatory condition caused by the ingestion of gliadin-containing food in genetically susceptible individuals. Undigested peptides of gliadin exert various effects, including increased intestinal permeability and inflammation in the small intestine. Although many therapeutic approaches are in development, a gluten-free diet is the only effective treatment for CD. Affecting at least 1% of the population in industrialized countries, it is important to generate therapeutic options against CD. Here, we describe the establishment of a high-throughput screening (HTS) platform based on AlphaLISA and electrical cell-substrate impedance sensing (ECIS) technology for the identification of anti-inflammatory and barrier-protective compounds in human enterocytes after pepsin-trypsin-digested gliadin (PT-gliadin) treatment. Our results show that the combination of these HTS technologies enables fast, reliable, simple, and label-free screening of IgY antibodies against PT-gliadin. Using this platform, we have identified a new chicken anti-PT-gliadin IgY antibody as a potential anti-CD agent.

Folate deficiency and over-supplementation causes impaired folate metabolism: Regulation and adaptation mechanisms in Caenorhabditis elegans.

SCOPE: Impaired folate metabolism increases the risk of birth defects, neurodegenerative and cardiovascular disease, osteoporosis and cancer. We used Caenorhabditis elegans to investigate impaired folate metabolism by RNA interference of key enzymes in the methionine synthase (MS) and thymidylate synthase (TS) cycle and by folate deficiency and over-supplementation feeding studies. METHODS AND RESULTS: Folate status is influenced by genetic variations (polymorphisms), folate deficiency and supplementation. Single RNAi of dihydrofolate reductase (DHFR), methylenetetrahydrofolate reductase (MTHFR) and MS revealed that gene regulation is largely affected in both folate cycles. Adaptation requires a close transcriptional connection between TS and MS cycle. Coupled DHFR and MS expression is required to balance both cycles, but seems to reduce the overall rate of folate conversion. Feeding studies showed that folate over-supplementation to functioning metabolism inactivates MS and MTHFR expression and enhances TS activity, which favors DNA synthesis over methylation reactions. Folate deficiency disrupted homeostasis by favoring TS cycle and led to malformation in C. elegans offspring. Embryos show aneuploidy and are nonviable lacking DNA repair during meiotic stage of diakinesis. CONCLUSION: Single gene silencing alters gene expression in both cycles and disrupts folate homeostasis. Folate over-supplementation and deficiency favors TS over MS cycle and causes prophase DNA damage.