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BMC Proceedings 2013, Volume 7 Suppl 6<br />
http://www.biomedcentral.com/bmcproc/supplements/7/S6<br />
Page 24 of 151<br />
Figure 1(abstract P9) One input mask of the seed train starting conditions as an example for the tool’s user interface and courses of Space-<br />
Time-Yield (STY) and viability over time during growth for flask scale 2.<br />
specialized microfold (M) cells, able to increase the absorption of micro- and<br />
nanoparticles [2,3].<br />
In the current study, different aspects of the toxicity of Ag-NPs on the cell of<br />
intestinal epithelium were studied, i.e. cytotoxicity, inflammatory response<br />
and barrier integrity of the epithelial monolayer.<br />
Materials and methods: The cytotoxic effect of AgNPs < 20 nm (10-90<br />
μg/ml, Mercator GmbH, DE) was assessed by MTT assay on Caco-2 cells<br />
(clone 1, from Dr. M. Rescigno, University of Milano-Bicocca, IT). The<br />
co-culture model was received by co-culturing Caco-2 cells with RajiB cells<br />
(ATCC, Manassas, VA) in Transwell permeable supports (Corning Inc., NY)<br />
[1,2]. The inflammatory mediators chemokine IL-8 and nitric oxide (NO)<br />
levels were analysed in both apical (AP) and basolateral (BL) compartments<br />
by ELISA (BD Biosciences Pharmingen, San Diego, CA) and by Nitrate/Nitrite<br />
Colorimetric Assay Kit (Cayman Chemical Company, Ann Arbor, MI),<br />
respectively, according to the manufacturer’s instructions.<br />
The expression levels of the IL-8 and iNOS (inducible Nitric Oxide Synthase)<br />
genes were evaluated by quantitative real-time PCR (qRT-PCR), where the<br />
primers used were: for IL-8 CTGGCCGTGGCTCTCTTG (sense) and GGGT<br />
GGAAAGGTTTGGAGTATG (antisense) and for iNOS - TGTGCCACCTC<br />
CAGTCCAGT (sense) CTTATGGTGAAGTGTGTCTTGGAA (antisense). Levels of<br />
individual transcripts were normalized to those of glyceraldehyde-3-<br />
phosphate dehydrogenase (GAPDH). Relative quantification (RQ) values -<br />
fold change of the target gene expression compared to the untreated<br />
sample, were calculated by 2 -ΔΔCt method [4].<br />
The barrier integrity of the cell monolayers of mono- and co-cultures under<br />
the influence of AgNPs was evaluated on 21 days fully differentiated