Fig. 5
From: Myoglobin expression by alternative transcript in different mesenchymal stem cells compartments
Metabolic characterization of PAM- and MAT-MSCs. Seahorse metabolic flux analyzer was used to assess the oxygen consumption rate (OCR) and the glycolysis-linked extracellular acidification rate (ECAR) in adherent PAM- and MAT-MSCs following the manufacturer protocols (see Materials and Methods). A and B show representative experimental outcomes, which indicated that oligomycin (Olig.), the uncoupler (FCCP), rotenone plus antimycin A (R + A) and 2-deoxy-glucose (2-DG) were sequentially added. C and D show the averaged values ± S.E.M. of three experiments each performed in triplicate of three independent MAT- and PAM-MSCs preparations. In (C) OCRs under basal/routine conditions (Basal), after addition of oligomycin (+ Olig.), after addition of FCCP (Uncoupl.) are shown as corrected for the R + A-insensitive OCR; ***, P < 0.001. In (D) glycolysis (i.e., basal ECAR) and glycolytic reserve (i.e., the difference between ECAR before and after addition of 2DG) are shown; ***, P < 0.001. E Bioenergetic map obtained combining basal OCR vs ECAR. F Relative contributions to the OCR of oxidizable substrates. The oxidative capacities of long-chain fatty acid oxidation (FAO), pyruvate oxidation (PyrOX) and glutamine oxidation (GlnOX) were assessed following the Seahorse Xf Mito Fuel Flex Test kit using combination of specific inhibitors of the three oxidative pathways as described in Materials and Methods; the capacities of each substrate were normalized to the sum of the capacities of the three substrates and the normalized capacities measured in PAM-MSCs divided by those in MAT-MSCs; the bars show the mean values ± S.E.M. of two experiments with independent MSCs preparation each performed in triplicate; **, P < 0.05]. G Representative experimental polarographic outputs of the basal O2 changes elicited by MAT- and PAM-MSCs after reducing the dissolved O2 concentration to about 20–25 µM (see Materials and Methods for details). H Averaged values ± S.E.M. of three experiments each performed in duplicate of three independent MAT- and PAM-MSCs preparations whose OCR was assessed as shown in panel (G); **, P < 0.01