Figure 5.

MKK6 expression accelerates cell metabolism. U2OS cells expressing a Tet-regulated construct were either mock treated (control) or treated with tetracycline for the indicated times to induce the expression of constitutively active MKK6. (a) GSH content (total and reduced) were analysed in control and MKK6 expressing cells at 24 h and presented as μmol/10³ cells. (b) ROS levels in control and MKK6 expressing cells treated with NAC (5 mM) and GSH (5 mM) were analysed at 24 h after MKK6 induction using the DCFH-DA probe. (c) ROS levels in control and MKK6 expressing cells treated with MitoQ (500 nM) were analysed at 24 h using the DCFH-DA probe. (d) Control and MKK6 expressing cells were treated with MitoQ (500 nM) for 2 days and cell survival was analysed by Annexin V/PI staining. Data were taken from 3 independent experiments. (e) The oxygen consumption rate was analysed 24 h after the induction of MKK6 expression in routine conditions and after addition of MitoQ (500 nM), as indicated. Results are presented as pmoles of O₂ consumed per 10⁵ cells.