Apoptosis or senescence-like growth arrest: influence of cell-cycle position, p53, p21 and bax in H2O2 response of normal human fibroblasts

Abstract

Early-passage human diploid fibroblasts (HDFs) undergo senescence-like growth arrest in response to sublethal concentrations of H(2)O(2) [Chen and Ames (1994) Proc. Natl. Acad. Sci. USA. 95, 4130-4134]. We determine here whether H(2)O(2) can cause apoptosis in HDFs and the molecular changes that differ between apoptosis and senescence-like growth arrest. When exponentially growing early-passage IMR-90 cells were treated for 2 h with 50-200 microM (or 0.25-1 pmol/cell) H(2)O(2), a fraction of cells detached at 16-32 h after the treatment. The cells remaining attached were growth-arrested and developed features of senescence in 1 week. The detached cells showed caspase-3 activation and typical morphological changes associated with apoptosis. Caspase-3 activation was H(2)O(2) dose-dependent and preceded nuclear condensation or plasma membrane leakage. Apoptotic cells were mainly distributed in the S-phase of the cell cycle, while growth-arrested cells exhibited predominantly G1- and G2/M-phase distributions. H(2)O(2) pretreatment induced G1 arrest and prohibited induction of apoptosis by a subsequent H(2)O(2) challenge. The p53 protein showed an average 6.1-fold elevation in apoptotic cells and a 3.5-fold elevation in growth-arrested cells. Reduction of p53 levels with human papillomavirus E6 protein prohibited the activation of caspase-3 and decreased the proportion of apoptotic cells. Growth-arrested cells had elevated p21, while p21 was absent in apoptotic cells. Bcl-2 was elevated in both growth-arrested and apoptotic cells. Finally, although the overall level of bax did not change in growth-arrested or apoptotic cells, the solubility of bax protein increased in apoptotic cells. Our data suggest that in contrast with growth-arrested cells, apoptotic cells show an S-phase cell cycle distribution, a higher degree of p53 elevation, an absence of p21 protein and increased solubility of bax protein.

Beneficial Effects of Fluvastatin on Liver Microcirculation and Regeneration After Massive Hepatectomy in Rats

“…Liver regeneration also might be affected by ROS via indirect or direct effects [29][30][31][32][33][34][35]. As an indirect effect, it has been reported that stellate cells activated via the ROS system produce excessive transforming growth factor beta (TGF-b), a potent inhibitory cytokine for liver regeneration [29,30].…”

Hallmarks for senescence in carcinogenesis: novel signaling players

“…It was found that Bcl-2 levels are positively regulated by the transcription factor CREB (cAMP responsive element binding protein). A reduction in phospho-CREB (Ser-133) levels was observed in young but not in senescent HDFs in response to apoptotic stimuli [53,55,77,82]. The phosphatase PP2A was responsible for inactivation of CREB in young HDFs.…”

“…These findings were subsequently extended to primary human fibroblasts undergoing replicative-as well as oxidative stress-induced senescence [81,82]. Furthermore, up-regulation of Bcl-2 in senescent cells proved to be a general mechanism of resistance to a variety of apoptotic stimuli [77,81,82]. It was found that Bcl-2 levels are positively regulated by the transcription factor CREB (cAMP responsive element binding protein).…”

“…The resistance of senescent fibroblasts to apoptosis has been linked to elevated expression levels of the pro-survival protein Bcl-2 [80]. These findings were subsequently extended to primary human fibroblasts undergoing replicative-as well as oxidative stress-induced senescence [81,82]. Furthermore, up-regulation of Bcl-2 in senescent cells proved to be a general mechanism of resistance to a variety of apoptotic stimuli [77,81,82].…”

Positive and negative effects of cellular senescence during female reproductive aging and pregnancy

“…For example, low dose of the DNA-damaging agent doxorubicin leads to cellular senescence, while high dose will induce apoptosis in rat cardiomyocytes (Spallarossa et al 2009, Altieri et al 2012. Low dose of H 2 O 2 also induces cellular senescence in human lung and skin fibroblasts, while high dose promotes apoptosis in these cells (Chen & Ames 1994, Chen et al 2000. Some DNAdamaging agents, such as busulfan, preferentially cause human lung fibroblasts to undergo cellular senescence instead of apoptosis (Probin et al 2006).…”