Here we show that replicative senescence in normal human diploid IMR90 fibroblasts is accompanied by altered expression of a set of microRNAs (miRNAs) (senescence-associated miRNAs), with 14 and 10 miRNAs being either up or downregulated (42-fold), respectively, in senescent with respect to young cells. The expression of most of these miRNAs was also deregulated upon senescence induced by DNA damage (etoposide) or mild oxidative stress (diethylmaleate). Four downregulated miRNAs were part of miRNA family-17, recently associated to human cell and tissue aging. Moreover, eight upregulated and six downregulated miRNAs mapped in specific chromosomal clusters, suggesting common transcriptional regulation. Upon adoptive overexpression, seven upregulated miRNAs induced the formation of senescence-associated heterochromatin foci and senescence-associated b-galactosidase staining (Po0.05), which was accompanied, in the case of five of them, by reduced cell proliferation. Finally, miR-210, miR-376a*, miR-486-5p, miR-494, and miR-542-5p induced double-strand DNA breaks and reactive oxygen species accumulation in transfected cells. In conclusion, we have identified a set of human miRNAs induced during replicative and chemically induced senescence that are able to foster the senescent phenotype by prompting DNA damage. Replicative or cellular senescence, a state of irreversible arrest of cell division, was first described in cultures of human fibroblasts. 1 Since then, replicative senescence has been described in various mammalian cells. 2 The mechanisms underlying senescence include telomere shortening, upregulation of the CDKN1A (p21WAF1) and CDKN2A (p16INK4a and p14ARF) loci, and accumulation of DNA damage. 3 Telomeres become progressively shorter at every round of cell division and this leads to critically short telomere length sensed as double-strand DNA breaks. 4 DNA damage and DNA-damage response (DDR) could be common events to cellular senescence programs initiated by telomere dysfunction and aberrant oncogene activation. 5 Senescent cells are marked by lack of DNA replication; expression of senescence-associated b-galactosidase (SA-b-gal); accumulation of discrete nuclear foci that are termed senescence-associated heterochromatin foci (SAHFs); and senescence-associated DNA-damage foci (SDFs). SAHFs are detected by preferential binding of DNA dyes, such as 4 0 ,6-diamidino-2-phenylindole (DAPI), and the presence of certain heterochromatin-associated histone modifications (trimethyl-Lys9 Histone H3). SDFs are nuclear foci containing proteins that are associated to DNA damage (Ser139-phosphorylated histone H2AX -g-H2AX-and p53-binding protein-1-53BP1). 6 Senescent cells show striking changes in gene expression, including upregulation of cell-cycle inhibitors (p21WAF1 and p16INK4a) and secreted proteins involved in microenvironment remodeling (IL-6), 7 and downregulation of genes that facilitate cell-cycle progression (c-FOS, cyclin-A, cyclin-B, PCNA) 8 or that are involved in cell-cycle execution (FOXM1, UBE2C, TYMS). ...
