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Objective The study of the circadian clock and its mechanisms is easily facilitated through clock resetting in cell culture. Among the various established synchronizers of the circadian clock in cell culture (temperature, serum shock, glucocorticoids), the artificial glucocorticoid Dexamethasone (DEX) is the most widely used. DEX treatment as a protocol to reset the circadian clock in culture gives simple readout with minimal laboratory requirements. Even though there are many studies regarding clock resetting in culture using DEX, reference points or expression patterns of core clock genes and their protein products are scarce and sometimes contradict other works with similar methodology. We synchronise a cell line of human origin with DEX to be used for studies on circadian rhythms. Results We treat HEK 293T cells with DEX and describe the patterns of mRNA and proteins of core clock regulators, while making a clear point on how CLOCK is less than an ideal molecule to help monitor rhythms in this cell line.

ObjectiveExposure to engineered nanoparticles (NPs) has been associated with lung and pleura pathology such as inflammation, fibrosis and cancer. Silver (Ag) NPs are used in a large number of consumer products and medical devices increasing therefore the possibility of user exposure. The aim of this study was to investigate the possible effects of engineered AgNPs on the function and expression of ENaC on parietal sheep pleura.Materials and methodsIn the present study the short circuit current (ISC) and the transmesothelial resistance (RTM) of sheep parietal pleura was monitored in Ussing chambers after the preincubation of the tissue with spherical AgNPs of 20nm and 60nm in size for 30 min. The effect of the AgNPs on the function of the epithelial sodium channel (ENaC) was investigated by the addition of the inhibitor amiloride (10‐5M) apically. Moreover, the expression of αENaC subunit was analyzed in these samples by Western Blotting in order to assess effects on the protein level.ResultsAgNPs (20 nm; 2μg/ml) increased the amiloride sensitive ISC indicating that ENaC function is enhanced by this treatment. The RTM of the sheep parietal pleura was not altered, suggestive of no effects on the tight junctions and the tissue integrity. Finally, the levels of αENaC expression were similar in all groups. AgNPs of 60nm had no significant effects in any of the parameters.ConclusionsAgNPs of 20 nm affect sodium permeability of the sheep parietal pleura by increasing ENaC activity. This effect is not due to increased αENaC expression. These data suggest that AgNPs induce changes in the function of pleural ENaC.