Fig. 2
From: Metabolic changes associated with methionine stress sensitivity in MDA-MB-468 breast cancer cells
Homocysteine media induces a metabolic shift from oxidative phosphorylation to glycolysis. a MB468 and MB468res-R8 cells cultured in Met+ or Met-Hcy+ growth media for 48 h with or without 4 mM KCN were imaged using two-photon microscopy at 740 nm. NADH free/bound fluorescence lifetime imaging microscopy (FLIM) maps of cells are shown using the color spectrum in the FLIM phasor histogram (b). c The phasor histogram represents relative fractions of free NADH (cyan-white) and protein-bound NADH (red-purple). d Cluster analysis using the phasor histogram to separate signals of high free/bound (black square) and low free/bound (pink square) NADH ratios. e Quantitation of NADH ratios from cells in a is shown. Two fields for each sample with a minimum of 15 cells per field were analyzed by cluster analysis and normalized to total pixels measured. The black dotted line indicates the average percentage of lower free/bound NADH ratios of MB468 (80 %) and MB468res-R8 (79 %) cells cultured in in Met+, KCN(−). f MB468 and MB468res-R8 cells cultured in Met+ media for 12 h or Met-Hcy+ media for 2, 4, 8, and 12 h were analyzed by Seahorse Bioscience XF analyzer. MB468 (blue line with squares) and MB468res-R8 (black line with circles) basal oxygen consumption rates (OCR) are shown normalized to the Met+-treated sample. Error bars represent normalized standard deviation, n = 4 replicates; statistical differences between Met+ media and Met-Hcy+ media treatments are indicated by asterisk where p ≤ 5.0 × 10−7