Metabolic changes during cell growth inhibition by the IRF-1 system

A genetic approach based on the activation of interferon-regulated-factor-1 (IRF-1) has been applied to arrest BHK cell growth in spinner flasks through the presence of estradiol in the culture medium, leading to the activation of the constitutively expressed IRF-1/estrogen receptor fusion protein (IRF-1-hER). Two days after estradiol addition cell proliferation was inhibited; cell concentration being kept for the next 6 days, although a decrease in cell viability was observed. IRF-1 activation alters the cell energetic metabolism, as there is extra metabolic activity with higher glucose, glutamine and oxygen consumption rates. Although the proteolytic activity is higher than in the control cells, the cell protein content increases significantly after IRF-1 activation, suggesting an overall increase in protein synthesis. A significant increase in lactate dehydrogenase activity and a higher reduction of MTT (3-(4,5-dimethylthiazol-2-yl)-2–5-diphenyl tetrazolium bromide) were also observed, showing an increase in cell activity during cell growth inhibition. Changes in ATP and in cellular phospholipid content as well in glucose metabolism were detected, in real time by 31P and 13C nuclear magnetic resonance spectroscopy (NMR). The levels of ATP were unaffected but changes in cellular phospholipid content were observed. Although an increase on ADP and AMP content was observed 48 to 72 h after IRF-1 activation, both the ATP content and the energy charge (EC) remained constant. It can be concluded that the IRF-1 activation leads to a significant increase of the cell activity until the cell is no longer able to maintain the same energetic metabolism and starts to lose viability.