Reconstitution of active telomerase in primary human foreskin fibroblasts: effects on proliferative characteristics and response to ionizing radiation

Continuous growth of telomerase-immortalised fibroblasts: How long do cells remain normal?

“…No changes in phenotype were reported, and hTERT immortalised cells appeared to provide an infinite supply of normal human cells (Bodnar et al, 1998;Jiang et al, 1999;Morales et al, 1999;Nakamura et al, 2002). We recently reported our results with hTERT immortalised human foreskin fibroblasts (VH25) in response to ionising radiation (Kampinga et al, 2004) and showed that these cells retained their normal phenotype for the first few 100 population doublings (PDs) while VH25 fibroblasts senesce at PD 40-45. However, we observed one subclone at PD 310 which showed tetraploidy and altered response in radiation sensitivity (Kampinga et al, 2004).…”

“…We recently reported our results with hTERT immortalised human foreskin fibroblasts (VH25) in response to ionising radiation (Kampinga et al, 2004) and showed that these cells retained their normal phenotype for the first few 100 population doublings (PDs) while VH25 fibroblasts senesce at PD 40-45. However, we observed one subclone at PD 310 which showed tetraploidy and altered response in radiation sensitivity (Kampinga et al, 2004). To further investigate the frequency of occurrence of an abnormal phenotype, we continued to grow seven additional independent hTERT immortalised clones during 80-150 weeks, corresponding to 250-550 PDs.…”

“…Although there is no significant difference in growth rate seen between the normal clones and the clones which developed abnormalities in cell cycle control (Group B) or ploidy (Group C) we observed an increase in growth rate before the onset of detectable abnormalities. We previously determined (Kampinga et al, 2004) that the mitotic index for one strain (K20) increased from about 85% in growing VH25 cells to about 95% in K20 cells at PD 300, which may be one explanation for the www.elsevier.com/locate/mechagedev Mechanisms of Ageing and Development 127 (2006) [85][86][87] observed increase in growth rate of the cultures. The increasing growth rate during prolonged culture did not result in loss of contact inhibition in any of the eight clones as determined by the soft agar assay (data not shown).…”

“…Tetraploidy without cell cycle control changes was detected in only one clone (Group C). Ploidy was tested by FACS analysis and survival after 6 Gy of g-rays was assayed using the clonogenic assay (Kampinga et al, 2004). Fig.…”

Actions of human telomerase beyond telomeres

“…However, several other studies found no correlation between telomerase expression and DNA repair capacity [49,50]. In summary, direct molecular evidence explaining the involvement of telomerase in DNA repair activity is still lacking.…”