Fig. 4.

Unraveling the rate-limiting step of the pyruvate-to-lactate conversion monitored by NMR studies in T47D cells. (A) Effects observed upon adding quercetin, a naturally occurring inhibitor of pyruvate's transport into cells, on the kinetics of lactate's build-up. These measurements involved injecting repeated aliquots of 5.9 mM hyperpolarized ¹³C₁-pyruvate into batches of T47D cells, to whose perfusing media the indicated concentrations of inhibitor were coadded 8 min before the NMR acquisitions. Red lines indicate the build-up best fits, leading to the indicated k values. (B) ³¹P NMR spectra showing the constant viability displayed by cells throughout these experiments, despite the marked changes experienced by the pyruvate-to-lactate transformation rates. (C) Washout experiments illustrating the time dependencies of the pyruvate and lactate ¹³C₁ NMR peak intensities, upon suddenly removing the extracellular medium. Shown by the various continuous curves are expectations from a pyruvate-to-lactate conversion whose rate remains unaffected by the washing out (k_wash = k₀, gray Upper and red Lower); a conversion whose rate is halved vis-à-vis its original level (k_wash = k₀/2, green Lower); or a conversion whose rate goes suddenly to zero upon restarting the perfusion (k_wash = 0, blue Lower). The latter's curves best data fit suggests that a sudden disappearance of the pyruvate leads to a full stop of the pyruvate-to-lactate conversion, and to an ensuing purely longitudinal decay of the latter's hyperpolarization.