Figure 3: RAW 264.7 and Maf-DKO cells are highly glycolytic. Glucose consumption (A) and lactate production (B) in RAW 264.7 and Maf-DKO cells in the presence and absence of the OXPHOS inhibitor oligomycin. (C) Expression levels of four metabolic enzymes that were noticeably different between the two cell lines. Protein expression (relative to tubulin) was calculated from the band intensities of three different blots and normalized to the expression in RAW 264.7 (n=3 different assays). D, Zymogram analysis of LDH-isoenzyme expression in RAW 264.7 and Maf- DKO cells. To verify the gel’s capacity to separate LDH1-LDH5 isoforms of mouse and human origin, a lysate of the human glioma cell line U87 (which expresses all five isoforms) was run in parallel as a control (according to Eyben et al. [58]). Note the presence of low levels of the LDH3 and LDH4 tetramers, containing two or one H-subunits from the LDH-B gene in the Maf-DKO lysate. E, Mitochondrial respiration measured as the basal oxygen consumption (Basal), the leak respiration (Leak), maximal oxygen consumption (Max), and residual oxygen consumption (Res). Data in (C) represent band intensities of HXK I, HXK II, LDH-A, and PKM2 of one blot, relative to tubulin. Data in A and B represent means ± SEM of three independent experiments performed in triplicate, and in E means ± SEM of four experiments performed with RAW 264.7 and Maf-DKO cells in parallel.