Higher DNA repair activity is related with longer replicative life span in mammalian embryonic fibroblast cells

Abstract

Since the detailed comparison of DNA repair activities among mammalian embryonic fibroblast cells with different replicative life spans has not been investigated, we tested DNA repair activities in embryonic fibroblast cells derived from mammals including human, dog, rat, and mouse. The cell viability after treatment of four DNA damage agents appeared to be decreased in the order of human embryonic fibroblasts (HEFs) > dog embryonic fibroblasts (DEFs) > rat embryonic fibroblasts (REFs) > mouse embryonic fibroblasts (MEFs) although statistical significance was lacking. The amounts of strand breaks and AP (apurinic/apyrimidinic) sites also appear to be decreased in the order of HEFs > DEFs > REFs ≥ MEFs after treatment of DNA damage agents. The DNA repair activities and rates including base excision repair (BER), nucleotide excision repair (NER) and double-strand break repair (DSBR) including non-homologous end-joining (NHEJ) decreased again in the order of HEFs > DEFs > REFs ≥ MEFs. BER and NHEJ activities in 3% O(2) also decreased in the order of HEFs > DEFs > REFs > MEFs. This order in DNA repair activity appears to be coincident with that of replicative life span of fibroblasts and that of life span of mammals. These results indicate that higher DNA repair activity is related with longer replicative life span in embryonic fibroblast cells.

Telomere length and telomerase activity are related with immortalization frequency but not with replicative senescence in mammalian embryonic fibroblasts, except human embryonic fibroblasts

“…Senescence-associated β-galactosidase staining (pH 6.0) showed that only senescent cells of these mammalian fibroblasts were stained with blue color at the perinuclear area (data not shown) as reported previously (Dimri et al 1995), suggesting that primary normal mammalian fibroblasts undergo replicative senescence. The growth of human fibroblasts slowed down around 64 PD and ceased at 72 PD ( Figure 1A; Park et al 2011). The life span of the DEFs, REFs, and MEFs was approximately 20-25, 10-15, and 8-10 PDs, respectively (Figure 1B-1D; Park et al 2011).…”

“…The growth of human fibroblasts slowed down around 64 PD and ceased at 72 PD ( Figure 1A; Park et al 2011). The life span of the DEFs, REFs, and MEFs was approximately 20-25, 10-15, and 8-10 PDs, respectively (Figure 1B-1D; Park et al 2011). These results showed that the maximum life spans of the species were proportional with the numbers of PD of fibroblasts.…”

“…Primary dog embryonic fibroblasts (DEFs), primary rat embryonic fibroblasts (REFs), and primary mouse embryonic fibroblasts (MEFs) were prepared from Beagle dog embryos, Sprague-Dawley rat embryos, and BALB/c mouse embryos (embryonic day 13.5), respectively (Park et al 2011). Cells were grown in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS; Biowhittaker, USA) and antibiotics under air atmosphere containing 5% CO 2 at 37°C.…”

“…Replicative senescence of normal somatic cells can be bypassed by overexpression of viral oncogenes, such as SV40 large T antigen (Ide et al 1983;Park et al 2011), E6, E7 (Shay et al 1993), E1A, and E1B (Serrano et al 1997), through the inactivation of the p53 or Rb tumor suppressor pathways. To examine the links between replicative senescence and immortalization process of primary mammalian fibroblasts, primary mammalian fibroblasts were stably transfected with a plasmid containing SV40 large T antigen.…”

“…However, we still do not know how mouse, rat, dog, and human fibroblasts upregulate p21/p16 and replicative senescence at their distinct PD numbers. Since HEFs reportedly have higher DNA repair activity and less susceptibility to DNA damage agent when compared to MEFs, REFs, and DEFs (Park et al 2011), PD numbers of mammalian fibroblasts at replicative senescence may be determined by the extent of accumulated DNA damage or threshold to DNA damage of mammalian fibroblasts. Further detailed study on DNA damage, DNA repair activity, threshold to DNA damage, and other replicative senescence-related processes of mammalian fibroblasts will provide a better understanding of this phenomenon.…”