Mitochondrial metabolism in CLL cells. (A) The mitochondrial electrochemical membrane potential (∆ΨM) was semiquantified using the potentiometric dye JC-1 for HD B-cells (n = 10) and CLL cells (n = 10) by FACS. (B) ATP levels were measured in lysates from purified normal B-cells (n = 5) and CLL cells (n = 5) by ELISA. (C) Baseline OCR and ECAR were measured by an XFe96 flux analyzer in purified normal B-cells (n = 4) and CLL cells (n = 4). OCR is an indicator of mitochondrial respiration, and ECAR is predominantly the result of anaerobic glycolysis. (D) Mean ECAR levels on administration of glucose in purified normal B-cells (n = 4) and CLL cells (n = 4) as measured by an XFe96 flux analyzer. (E) The relative gene expression of key molecules of glycolysis (glut1; glut2; hexokinase-2, hk2; lactate dehydrogenase a, ldha; pyruvate dehydrogenase kinase 1, pdk1) is shown comparatively for purified HD B-cells (n = 10) and CLL cells (n = 10) as quantified by qPCR. (F) Respiration (OCR) is measured under basal conditions and in response to the indicated mitochondrial inhibitors in purified normal B-cells (n = 4) and CLL cells (n = 4). The resulting effects on OCR are shown as a percentage of the baseline measurement (set as 100%) for each treatment. Changes after oligomycin (I.) and FCCP (II.) application are indicative for respiration linked to ATP synthesis and the maximal respiratory capacity, respectively. (G.) ECAR is measured under basal conditions and in response to the indicated mitochondrial inhibitors in purified normal B-cells (n = 4) and CLL cells (n = 4). The resulting (compensatory) effects on ECAR are shown as a percentage of the baseline measurement (set as 100%). Bars indicate the standard error mean. Abbreviations: P, P-value; *P < .05; **P < .005; ***P < .001.