Time course study of ERK1/2 activity and cell viabilityin lipopolysaccharide challenged PC12

Introduction: Extracellular signal-regulated kinase 1 and 2 (ERK1/2) are important members of mitogen-activated protein kinases (MAPK), which are actively involved in the shaping of cellular responses to different stimuli; however, these responses are somehow contradicting. It is believed that time is a crucial factor in the determination of these effects. Therefore, the present work was designed to obtain a more vivid view about the effect of time on ERK activity and its relation to cell viability. Methods: In the first step, we challenged cultured PC12 cells with different doses of lipopolysaccharides (LPS) for different time intervals (3, 6, 24 and 48h) and the cell viability was checked by MTT test. Thereafter, we cultured the cells in 6-well plates and treated them with the effective dose (10μg/ml) for the abovementioned intervals and the level of ERK phosphorylation, as the active form, was assessed in the Western blotting analysis. Results: The results showed that treating cells with 10μg/ml LPS reduces cell viability after 48h. While being ineffective in shorter periods of time, ERK activity has a fluctuating trend, so that it reaches the highest level at 6h, thereafter it declines to the lowest level at 24h and partially increases again at 48h. Conclusion: These results imply that time is a determinant factor in the activity of ERK and single-point assessments may result in misinterpretation.