Figure 1.
Oxygen consumption, lactate and creatine kinase (CK) levels in the blood.
Overall experimental procedures (A). Fatigue was induced by spontaneous running on an electric treadmill that elevated the oxygen consumption (B). At different time-points after the fatiguing exercise blood samples were collected for measurement of lactate (C), creatine kinase (D) and glucose level (E). In this study WT C57Bl/6 mice were used. These results C, D and E are presented as the mean ± SEM (n = 5). *P<0.05 when compared with the control group.
Figure 2.
Number of leukocytes in the peripheral blood.
Fatigue was induced by spontaneous running on an electric treadmill. At different time-points after the exercise protocol blood samples were collected and numbers of total cell (A) neutrophils (B), monocytes (C) and lymphocytes (D) were evaluated. In this study WT C57Bl/6 mice were used. The results are presented as the mean ± SEM (n = 4–6). *P<0.05 when compared with the control group.
Figure 3.
Recruitment of neutrophil into quadriceps muscle.
Fatigue was induced by spontaneous running on an electric treadmill. At different time-points after the exercise protocol, mice were anesthetized and muscle postcapillary venules selected for counting numbers of rolling (A), adherent (B) and transmigrating (C) leukocytes using epifluorescence and confocal intravital microscopy, respectively. The time course on the x-axis in the 3C figure represents the time of recording. It was recorded for 20 minutes. After intravital microscopy, quadriceps muscle samples were removed for histological analyses. Tissue section from control (D) and 12 hours after the exercise (E–F) mice are shown for illustration. Neutrophil infiltration is shown by arrows and inflammatory foci consisting predominantly of neutrophils are shown by asterisks. Original scale bars 50 µm for all panels. In this study WT C57Bl/6 and LysM-eGFP C57Bl/6 mice were used. The results are presented as the mean ± SEM (n = 5). *P<0.05 when compared with the control group.
Figure 4.
Leukocytes-endothelium interaction in quadriceps muscle after exercise and adhesion molecules expression.
Fatigue was induced by spontaneous running on an electric treadmill. Quadriceps muscle was exposed to visualize the vasculature 12(A) and exercised wild-type mice (B). Confocal intravital microscopy images are shown in control (C) and exercised LysM-eGFP mice (D). Using qPCR technique, the molecules expression was quantified: E-selectin (E), L-selectin (F) and PECAM (G) were evaluated 12 hours after the exercise. APO = apocynin and dotted line represents the control group. The results are presented as the fold increase (n = 4, control group and n = 6, exercised group). *P<0.05 when compared with the control group.
Figure 5.
ROS production from NADPH oxidase on leukocyte recruitment.
Fatigue was induced by spontaneous running on an electric treadmill. GSH levels were measured at different time-points after exercise protocol (A). Mice received an injection of apocynin (10 mg/kg, i.p.) 30 minutes before exercise and numbers of rolling (B), adherent (C) and transmigrating (D) leukocytes was evaluated 6 and 12 hours after the exercise. In another experimental setting, numbers of rolling (E) and adherent (F) leukocytes was evaluated 12 hours after the exercise using mutant mice (gp91phox-/-). The results are presented as the mean ± SEM (n = 5). *P<0.05 when compared with the control and # when compared with exercised mice.
Figure 6.
Superoxide dismutase (SOD) exacerbated adhesion and transmigration of neutrophil.
Fatigue was induced by spontaneous running on an electric treadmill. Superoxide dismutase (3 mg/kg, i.p.) was injected 30 minutes before exercise. Using LysM-eGFP mice, numbers of rolling (A), adherent (B) and transmigrating (C) neutrophil was evaluated 12 hours after the exercise. These results were confirmed by 3D images from exercised (D) and exercised + SOD (E) mice. The results are presented as the mean ± SEM (n = 4, control group and n = 6, exercised and exercised + SOD group). *P<0.05 when compared with the control and # when compared with exercised mice. (red = vessels; green = neutrophils).