Fig. 3
Selective inhibition of NTPDase3 with PSB 06126 (3 µM) or the monoclonal antibody hN3-B3S (0.5 µg/ml) increases the extracellular ATP accumulation in Pm BM-MSCs cultures, but not in those obtained from younger females, when cells were allowed to grow for 7 days in an osteogenic-inducing medium (first subculture). In panel A, ordinates are the difference between endogenously released ATP during 30 min by cells of the same individual incubated in the presence and the absence of NTPDase3 inhibitors (Drug-Ctr, respectively); zero represents the identity between ATP values obtained in the absence and the presence of the inhibitors (horizontal dashed line). The ATP content of the samples was measured using the luciferin-luciferase bioluminescence assay (75 µl/well; see Materials and Methods for details). Boxes and whiskers represent pooled data from three younger females (21 ± 7 years old; ATP release in Ctr: 90 ± 20 fmol/well) and six Pm women (68 ± 5 years old; ATP release in Ctr: 180 ± 40 fmol/well); two to four replicates were performed per individual. ***P < 0.001 and ****P < 0.0001 (Wilcoxon signed rank test for comparing medians with a hypothetical null variation) represent significant differences. Panel B shows that BM-MSCs from a Pm woman undergoing osteogenic differentiation exhibit high amounts of ATP inside intracellular granules stained with quinacrine. The scale bar is 50 µm