| External factors: | Ionizing radiation |
| Aging type: | Accelerate |
| Aging characteristic: |
| Category: | Other |
| Phenotype: | Aging |
| Experiment: | SA-β-gal activity assay//Western blot//Cell morphological analysis//Cell counting |
| Description: | The cytosol stained positive for senescent cells (blue), and 60 and 90% of cells displayed SA-β-gal activity at 48 and 72 h, respectively.p53 accumulation and phosphorylation at Ser-15 peaked 3 h after IR, and p21 was activated by a p53-dependent pathway. IR treatment resulted in induction of p16, a positive regulator of pRb, and thus caused hypophosphorylation and activation of pRb. Consistent with SA-β-gal activity, characteristics of senescence, such as large and flat morphology, were identified in cells treated with IR in a dose-dependent manner. Various doses of IR ranging from 0 to 10 Gy additionally caused a decrease in number of chondrocytes in a concentration-dependent manner at 48 h after irradiation , and this phenomenon was also dependent upon the time after irradiation with 10 Gy. |
| Target gene: | SIRT1 |
| R-EF-Target gene: | Downregulation |
| Official symbol(s): | SIRT1 |
| Target gene experiment: | Western blot |
| Target gene description: | IR treatment with 10 Gy resulted in a time-dependent reduction in the SIRT1 protein level after irradiation and increased acetylation level of histone H3, a major deacetylation substrate of SIRT1 . |
| Regulatory pathway: | ROS-p38//ERK |
| R-EF-Pathway: | --//-- |
| Official symbol(s): | MAPK14//ERK |
| Pathway experiment: | Flow cytometry//SA-β-gal activity assay//Western blot |
| Pathway description: | Consistent with these findings, at 6 and 12 h after irradiation, ROS levels increased significantly with the radiation dose in primary cultured articular chondrocytes. ROS production was continuously accumulated up to 8 h and then slightly reduced at 12 h after IR treatment. Conversely, phosphorylation of p38 kinase was continuous and lasted up to 12 h after IR treatment.To establish the feedback linkage between elevation of the intracellular ROS level and MAPK activation in IRtreated articular chondrocytes, cells were preincubated with specific MAPK inhibitors.Pretreatment of cells with the p38 kinase-specific inhibitor, SB203580, did not affect IR-induced ROS generation at the early time point (90 min; top) but led to the complete inhibition of ROS production at a later time point (12 h; middle) with concomitant suppression of SA-β-gal activity after IR treatment (48 h; bottom). The results support the formation of a ROS-p38 positive feedback loop to sustain down-stream signaling. In contrast, the ERK inhibitor, PD98059, significantly suppressed IR-induced ROS generation at the early and late time points (top and middle) as well as SA-β-gal activity (bottom), indicating that the ERK pathway acts upstream of ROS generation. |
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