Bunc, M;
Hanning, SM;
Prieto Garcia, J;
Kristl, J;
Orlu, M;
(2019)
Characterisation of an in vitro aged cell culture model.
Farmacia
, 67
(2)
pp. 240-247.
10.31925/farmacia.2019.2.7.
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Abstract
We here aim to provide a detailed protocol to obtain an aged cell culture model, which is conveniently characterised in terms of morphology, metabolism, and key biochemical markers of senescence. Commercial Human Dermal Fibroblasts (HDF) (67-year-old female Asian donor) were cultured over a span of 35 passagesto determine at what passages they express the phenotypes found in vivo in the geriatric population. The growth rate and density (counting, Trypan Blue, SRB) at confluence declined 4- and 3-fold, respectively. During 15 passages in vitro ageing the fibroblast metabolic capacity (XTT) whas heavily reduced (a by 75 %), while conversely the content of cellular proteins (SRB) has slightly increased (2 fold). Senescent cells (SA-β-gal positive) were continuously emerging from 0 to 20%. Freezing/thawing at this stage restores restored metabolic performance to original values which declines declined at a similar rate as before. Passages 5 – 10 of our selected aged HDFs were found to mimic in vivo tissue of older adults to the greatest extent.
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