Fig. 5.

Conversion of pyruvate to acetate in cell-culture medium. (A) Carbon atom transition map depicting labeling patterns of metabolites derived from [1, 6-¹³C2]-d-glucose. (B) Levels of ¹³C-labeled acetate in the culture medium from cells labeled with [3-¹³C]-pyruvate. At 48 hpi, mock- and HCMV-infected HFs were labeled with medium containing 1 mM [3-¹³C]-pyruvate but no glucose. Cultured medium was collected at 0, 10, 60, and 120 min to measure [2-¹³C]-acetate by NMR. Data are shown as the mean ± SEM of triplicates. (C) Acetate levels in fresh serum-free DMEM without glucose in the presence or absence of 1 mM pyruvate. ¹²C-acetate levels were measured by NMR. Data are shown as the mean ± SEM of triplicates. (D) [2-¹³C]-acetate is produced from [3-¹³C]-pyruvate added into fresh serum-free medium without glucose. [3-¹³C]-Pyruvate was freshly added to water or serum-free DMEM without glucose, and the levels of [2-¹³C]-acetate in water or serum-free medium were measured directly by NMR without cell culture. [2-¹³C]-acetate is produced from [3-¹³C]-pyruvate only in medium but not in water. Data are shown as the mean ± SEM of triplicates. (E) Levels of [2-¹³C]-acetate produced from mock- or HCMV-infected cells labeled with [1, 6-¹³C2]-d-glucose. At 24 hpi, mock- and HCMV-infected cells were replaced with serum-free DMEM containing 5.6 mM [1, 6-¹³C2]-d-glucose; medium was collected at 48 hpi and analyzed for [2-¹³C]-acetate levels. Data are shown as the mean ± SD of triplicates. (F) Levels of [3-¹³C]-pyruvate in the same media collected in E. Data are shown as the mean ± SD of triplicates.