The cell cycle inhibitor, p21/waf1 regulates cell proliferation. Hyperoxia (Hx), dexamethasone (Dx) and all-trans-retinoic acid (tRa) are therapeutic agents that affect lung cells, specially, fibroblasts, but the role of p21 in mediating their effects are unknown. We hypothesized that these agents modify cell proliferation and apoptosis of lung fibroblasts through changes in p21 expression. Methods: Freshly isolated fetal rat lung fibroblasts were cultured in serum supplemented medium in the presence or absence of Hx (95% oxygen), Dx (0.1-10μM) or tRa (0.01-1μM). Total protein and mRNA were extracted at various time points. The steady state levels of p21 mRNA were determined by Northern analysis and densitometric quantification. Immunoblots from Western analysis of p21 protein were quantified by densitometry. Cell proliferation was assayed by 3H-thymidine incorporation and apoptosis identified by the TUNEL assay. Results: Compared to normal control (NI) conditions, all three agents tested, decreased 3H-thymidine incorporation from 6 h-120 h in fibroblasts (bombesin was used as a positive control). In parallel, the steady state levels of p21(inhibitor of cell cycle) mRNA were significantly increased in Hx and Dx but were unchanged with tRa. Similarly, p21 protein levels were significantly increased in Hx and Dx. There were no changes in p21 protein with tRa(0.01-1μM). Apoptosis (TUNEL) was seen in < 5% of cells after 7 days in culture with Hx, but ≤ 1% apoptosis was seen in controls and cells treated with Dx and tRa for 7 days. Conclusions: Hx and Dx induced suppression of fibroblast proliferation is associated with increased p21 expression at a pretranslational level but not associated with changes in apoptosis. tRA causes decrease in cell proliferation by a mechanism that does not involve increase in p21 or apoptosis. (Graph Presented).
|Original language||English (US)|
|Journal||Journal of Investigative Medicine|
|State||Published - Feb 1999|
All Science Journal Classification (ASJC) codes
- Biochemistry, Genetics and Molecular Biology(all)