Figure 4. HP-MR detects changes in ROS driven reducing equivalent levels.

A) IR increases metabolically driven ROS, which are scavenged by thiol moieties. B) Thiol moieties are regenerated through utilization of secondary reducing equivalents (NADH/NADPH). C) Conversion of endogenous pyruvate into lactate (ubiquitous in tumor cells) requires reducing equivalents. The rate of conversion therefore indirectly reflects global levels of reducing equivalents. HP-MR piggybacks on this endogenous reaction by allowing measurement of conversion of ¹³C pyruvate (PYR) into ¹³C (LAC). The rate of conversion is therefore directly related to reducing equivalent levels and indirectly related to intra-cellular ROS levels. Inset illustrates the dependence of the PYRLAC reaction on available NADH concentration (experiment performed in vitro using purified enzyme, NADH and ¹³C HP-PYR).