Cell activity after cell growth inhibition by the IRF-1 system

摘要

A genetic approach based on the activation of interferon-regulated-factor-1 (IRF-1) has been applied to regulate BIlK cell growth. The presence of 100 nM estradiol in the culture medium in agitated flasks leads to the activation of the constitutively expressed IRF-1/estrogen receptor fusion protein (IRF-I-hER) and inhibits cell growth. Two days after estradiol addition cell concentration was still maintained but a significant decrease in cell viability was observed. The IRF-I activation clearly interferes with the cell energetic metabolism, since there is an extra metabolic activity, with higher glucose, glutamine and oxygen consumption rates, while the yield lactate/glucose is maintained Although there is a higher proteolytic activity, the protein content per cell increases significantly, suggesting an overall increase in the protein synthesis. A significant increase in the lactate dehydrogenase enzyme activity and a higher reduction of the MTr were also observed, indicating the increase in cell activity after cell growth inhibition. ATP, ADP and AMP contents were evaluated, and the cells were able to maintain the energy charge (EC) higher than 0.8. The significant increase in the total content on protein per cell can be the reason for the higher energetic needs. Thus, in order to maintain the EC the cells need to increase the ATP production, i.e., increase the catabolic metabolism. Nevertheless, 2 to 3 days after estradiol addition this equilibrium is no longer stable, leading to an increase in the ADP and AMP contents, and at that time the cell viability decreases and the other alterations previously referred start to be significant. The IRF-I activation leads to a significant increase of the cell activity until the cell is no longer able to maintain its metabolic equilibrium and starts to die.