Figure 2.

Replicative senescence in HMECs is accompanied by significant alterations in metabolite pool sizes. A, glucose consumption, lactate secretion, and ratio of glucose consumption to lactate secretion are unchanged in senescent HMECs. Metabolite extracts from blank and conditioned media were analyzed by LC-MS. Secretion or uptake is calculated as follows: conditioned medium − blank medium, normalized to integrated cell number. Secreted metabolites have positive values, and consumed metabolites have negative values. B, volcano plot of intracellular pool sizes for all measured metabolites. Data represent average weighted log₂ -fold change (senescent/proliferating) and FDR-corrected Fisher's combined p value from five independent experiments. Depicted metabolites were measured in at least two independent experiments. C, depletion of dNDPs and dNTPs and accumulation of nucleobases and nucleosides in senescent HMECs (same data as in B for selected dNDPs, dNTPs, guanine, and uridine). * and **, FDR-corrected Student's t test p values less than 0.02 and 0.002, respectively. D, extracellular secretion and consumption measured by LC-MS as in A for the nucleobases adenine, guanine, and uracil and the nucleoside uridine. Secreted metabolites have positive values, and consumed metabolites have negative values. *, FDR-corrected Student's t test p < 0.001. E, metabolic pathway map depicting the log₂ -fold change (senescent/proliferating) intracellular pool sizes of metabolites in glycolysis, pentose phosphate pathway, nucleotide synthesis, and TCA cycle using the indicated color scale. Metabolites that were not measured are shown as small circles with gray color. Isomers that were not resolved with LC-MS are shown as diamonds. Error bars, S.D.