Growth Inhibition by Bupivacaine Is Associated with Inactivation of Ribosomal Protein S6 Kinase 1

Figure 2

Concentration and time dependence of S6K1 inhibition by bupivacaine. (a) NIH-3T3 cells were allowed to grow for 12 hrs in the absence (control) or presence of indicated concentrations of bupivacaine (BPV) and S6K1 was immunoprecipitated, subjected to kinase assays, and then probed with indicated antibodies. (b) Densitometric analysis of anti-pS6K signals (normalized to total S6K levels) from three independent experiments. Data are relative S6K phosphorylation levels with control set as 100%, presented as mean ± SEM. (c) NIH-3T3 cells were allowed to grow in the absence or presence of bupivacaine (1 mM) for indicated time intervals and processed similarly as above. (d) Densitometric analysis of anti-pS6K signals (normalised to total S6K levels) from three independent experiments. (e) NIH-3T3 cells were incubated with inhibitory concentrations (1 mM) of bupivacaine as described above. Cells were processed for treatment with S6K Phospho-T412 and S6K Phospho-T252 antibodies and imaged using LI-COR infrared imager. (f) Average florescent intensity of each well was calculated in arbitrary units (AU) using LI-COR ODYSSEY software.