To investigate the time course of leukocyte accumulation in eccentric exercised human muscles and its relation to recovery of muscle function and soreness. Eleven young males performed 300 unilateral, maximal voluntary, eccentric actions with the musculus quadriceps femoris (30 degrees x s(-1)). Before and at regular intervals for 7 d after exercise, force-generating capacity was measured with maximal concentric knee extensions (60 degrees x s(-1)). Accumulation of radiolabeled (autologous) leukocytes was measured with scintigraphy. Biopsies from musculus vastus lateralis were obtained 0.5, 4, 8, 24, 96, and 168 h after exercise from both the exercised leg and the control leg. Muscle cross-sections were stained with antibodies against leukocytes (CD16 and CD68). Muscle soreness was rated on a visual analog scale. Immediately after exercise, the subjects' ability to generate force was reduced by 47 +/- 5%. Muscle function recovered slowly and was not fully restored after 1 wk. Radiolabeled leukocytes accumulated in the muscles during the first hour (3-24 h) after exercise, and leukocytes were at the same time observed histologically, primarily in the endomysium and perimysium. A part of the accumulated radiolabeled leukocytes appeared to be located within local blood vessels. The highest numbers of CD16(+) and CD68(+) cells were found 4 and 7 d after exercise. There was a positive correlation between accumulation of radiolabeled leukocytes and muscle weakness measured 1-3 d after exercise (r = 0.8, P < 0.05) and, surprisingly, a negative correlation between radiolabeled leukocyte accumulation and muscle soreness (r = -0.96, P < 0.01). Exercise-induced muscle damage initiated a rapid local inflammatory response that gradually increased over the next days. Halted recovery of muscle function was associated with local accumulation of leukocytes, whereas muscle soreness could not be explained by the presence of leukocytes.

In this study the stress protein response to unaccustomed maximal eccentric exercise in humans was investigated. Eleven healthy males performed 300 maximal eccentric actions with the quadriceps muscle. Biopsies from vastus lateralis were collected at 30 min and 4, 8, 24, 96, and 168 h after exercise. Cellular regulation and localization of heat shock protein (HSP) 27, αB-crystallin, and HSP70 were analyzed by immunohistochemistry, ELISA technique, and Western blotting. Additionally, mRNA levels of HSP27, αB-crystallin, and HSP70 were quantified by Northern blotting. After exercise (30 min), 81 ± 8% of the myofibers showed strong HSP27 staining ( P < 0.01) that gradually decreased during the following week. αB-Crystallin mimicked the changes observed in HSP27. After exercise (30 min), the ELISA analysis showed a 49 ± 13% reduction of the HSP27 level in the cytosolic fraction ( P < 0.01), whereas Western blotting revealed a 15-fold increase of the HSP27 level in the myofibrillar fraction ( P < 0.01). The cytosolic HSP70 level increased to 203 ± 37% of the control level 24 h after exercise ( P < 0.05). After 4 days, myofibrillar-bound HSP70 had increased ∼10-fold ( P < 0.01) and was accompanied by strong staining on cross sections. mRNA levels of HSP27, αB-crystallin, and HSP70 were all elevated the first day after exercise ( P < 0.01); HSP70 mRNA showed the largest increase (20-fold at 8 h). HSP27 and αB-crystallin seemed to respond immediately to maximal eccentric exercise by binding to cytoskeletal/myofibrillar proteins, probably to function as stabilizers of disrupted myofibrillar structures. Later, mRNA and total HSP protein levels, especially HSP70, increased, indicating that HSPs play a role in skeletal muscle recovery and remodeling/adaptation processes to high-force exercise.

The aims of this study were to investigate the sarcomeric accumulation and expression of heat shock proteins (HSPs) after two bouts of maximal eccentric exercise. Twenty-four subjects performed two bouts of 70 maximal voluntary eccentric actions using the elbow flexors in one arm. The bouts were separated by 3 wk. The changes in concentric (60°/s) and isometric (90°) force-generating capacity were monitored for 9 days after each bout, and biopsies were taken 1 and 48 h and 4 and 7 days after bout 1 and 1 and 48 h after bout 2. The content of HSP27, αB-crystallin, HSP70, and desmin in the cytosolic and cytoskeleton/myofibrillar fractions of homogenized muscle samples was determined by immunoassays, and the cellular and subcellular localization of the HSPs in the myofibrillar structure was analyzed by conventional and confocal immunofluorescence microscopy and quantitative electron microscopy. The force-generating capacity was reduced by ∼50% and did not recover completely during the 3 wk following bout 1. After bout 2, the subjects recovered within 4 days. The HSP levels increased in the cytosolic fraction after bout 1, especially HSP70 (∼300% 2–7 days after exercise). Increased levels of HSP27, αB-crystallin, and HSP70 were found in the cytoskeletal/myofibrillar fraction after both bouts, despite reduced damage after bout 2. At the ultrastructural level, HSP27 and αB-crystallin accumulated in Z-disks, in intermediate desmin-like structures (αB-crystallin), and in areas of myofibrillar disruption. In conclusion, HSP27 and αB-crystallin accumulated in myofibrillar structures, especially in the Z-disks and the intermediate structures (desmin). The function of the small HSPs is possibly to stabilize and protect the myofibrillar structures during and after unaccustomed eccentric exercise. The large amount of HSP27, αB-crystallin, and HSP70 in the cytoskeletal/myofibrillar fraction after a repeated bout of exercise suggests a protective role as part of the repeated-bout effect.