Structural chromosome abnormalities, increased DNA strand breaks and DNA strand break repair deficiency in dermal fibroblasts from old female human donors

Abstract

Dermal fibroblasts provide a paradigmatic model of cellular adaptation to long-term exogenous stress and ageing processes driven thereby. Here we addressed whether fibroblast ageing analysed ex vivo entails genome instability. Dermal fibroblasts from human female donors aged 20–67 years were studied in primary culture at low population doubling. Under these conditions, the incidence of replicative senescence and rates of age-correlated telomere shortening were insignificant. Genome-wide gene expression analysis revealed age-related impairment of mitosis, telomere and chromosome maintenance and induction of genes associated with DNA repair and non-homologous end-joining, most notably XRCC4 and ligase 4. We observed an age-correlated drop in proliferative capacity and age-correlated increases in heterochromatin marks, structural chromosome abnormalities (deletions, translocations and chromatid breaks), DNA strand breaks and histone H2AX-phosphorylation. In a third of the cells from old and middle-aged donors repair of X-ray induced DNA strand breaks was impaired despite up-regulation of DNA repair genes. The distinct phenotype of genome instability, increased heterochromatinisation and (in 30% of the cases futile) up-regulation of DNA repair genes was stably maintained over several cell passages indicating that it represents a feature of geroconversion that is distinct from cellular senescence, as it does not encompass a block of proliferation.

Impaired DNA double-strand break repair contributes to the age-associated rise of genomic instability in humans

“…27,28 Although the previous work greatly advanced our understanding of age-associated changes of DNA DSB repair, due to a lack of proper tools for the analysis of NHEJ and HR efficiency and fidelity separately, and the hardship of acquiring a sufficient number of human samples, whether NHEJ efficiency and fidelity, and HR efficiency change with age in humans and the consequences of any such change, and its underlying molecular mechanism are not well understood. Here, we established 50 eyelids fibroblast cell lines derived from donors who are evenly distributed by age.…”

Ageing as an Important Risk Factor for Cancer

“…This can be also correlated to well-documented clinical differences in wound healing. Similarly, the number of DNA strand breaks and time required for their repair is significantly influenced by ageing (22,23). Double-strand DNA breaks can even participate to the loss of heterozygosity in the elderly and can be important for cancerogenesis (24,25).…”

Emerging metrology for high-throughput nanomaterial genotoxicology

“…As a consequence, mammalian cell-based alternatives to typical animal-based clastogenicity assays are needed for early screening of mammalian genotoxicity (65). The automated FADU assay is a cell-based in vitro system, which is well suited for the measurement of DNA strand breaks in different types of human cell lines (125,126), as well as in primary human cells such as PBMCs, lymphocytes (126–129) and/or fibroblast (130) obtained from different subjects. The automated FADU offers an alternative method for reducing animal use during genotoxicity risk assessments.…”