We tested the hypotheses that: (1) neutrophil accumulation after contraction-induced muscle injury is dependent on the β 2 integrin CD18, (2) neutrophils contribute to muscle injury and oxidative damage after contraction-induced muscle injury, and (3) neutrophils aid the resolution of contraction-induced muscle injury. These hypotheses were tested by exposing extensor digitorum longus (EDL) muscles of mice deficient in CD18 (CD18 −/− ; Itgb2 tm1Bay) and of wild type mice (C57BL/6) to in situ lengthening contractions and by quantifying markers of muscle inflammation, injury, oxidative damage and regeneration/repair. Neutrophil concentrations were significantly elevated in wild type mice at 6 h and 3 days post-lengthening contractions; however, neutrophils remained at control levels at these time points in CD18−/− mice. These data indicate that CD18 is required for neutrophil accumulation after contraction-induced muscle injury. Histological and functional (isometric force deficit) signs of muscle injury and total carbonyl content, a marker of oxidative damage, were significantly higher in wild type relative to CD18−/− mice 3 days after lengthening contractions. These data show that neutrophils exacerbate contraction-induced muscle injury. After statistically controlling for differences in the force deficit at 3 days, wild type mice also demonstrated a higher force deficit at 7 days, a lower percentage of myofibres expressing embryonic myosin heavy chain at 3 and 7 days, and a smaller cross sectional area of central nucleated myofibres at 14 days relative to CD18−/− mice. These observations suggest that neutrophils impair the restoration of muscle structure and function after injury. In conclusion, neutrophil accumulation after contraction-induced muscle injury is dependent on CD18. Furthermore, neutrophils appear to contribute to muscle injury and impair some of the events associated with the resolution of contraction-induced muscle injury.

Pizza, Francis X., Timothy J. Koh, Stephen J. McGregor, and Susan V. Brooks. Muscle inflammatory cells after passive stretches, isometric contractions, and lengthening contractions.

Eight male cross-country runners and five male swimmers were tested four times during their collegiate seasons. Each trial corresponded to a different training load. The runners' trials were conducted before the start of organized practice (RT1), after 3 wk of increased training (RT2), 3 wk prior to the conference championship (pre-taper, RT3), and 4 d after the conference championship (post-taper, RT4). The swimmers' trials were conducted after the first 9 wk of training (ST1), after completing 2 wk of hard training (ST2), after an additional 6 wk of training (pre-taper, ST3) and during a week following the conference championship (post-taper, ST4). Venous blood samples, heart rate (HR) and blood pressure (BP) were obtained after 15 min supine rest (0700 h). Serum was analyzed for cortisol (C), total testosterone (TT), free testosterone (FT), and creatine kinase (CK). Blood samples (lactate), HR and RPE were obtained during a fixed velocity run (75% preseason VO2max) and blood samples and RPE following a 365.8 m swim (90% preseason VO2max). The runners then completed a "performance run" to exhaustion (110% preseason VO2max) and the swimmers completed maximal 22.9 and 365.8 m swims. Serum CK, C, TT, FT, and the TT:C and FT:C ratios were not significantly different among trials for the runners. Serum TT and FT were significantly (P < 0.05) lower for the swimmers at ST2 (TT 16.7 +/- 2.5; FT 85.3 +/- 8.5) compared to ST1 (TT 30.3 +/- 2.8; FT 130.2 +/- 20.9) whereas, C, TT:C or FT:C were not significantly altered.(ABSTRACT TRUNCATED AT 250 WORDS)

Modified muscle use can result in muscle atrophy and impairment. We tested whether inflammatory cell concentrations correlate temporally with muscle impairment during modified loading periods. Rat hindlimbs were unloaded for 10 days followed by reloading. The density of neutrophils and ED1+ macrophages was significantly increased by 16.5- and 9.8-fold, respectively, after 1 day of reloading. ED2+ macrophage concentration was not significantly increased until 3 days of reloading. Maximal isometric tetanic tension (P(o); N/cm2) decreased during hindlimb suspension (HS), which was followed by a second drop in P(o) after 2 h of reloading. This latter loss in muscle force was uncoupled with the significant elevation in muscle inflammatory cell concentrations. Experiments where HS soleus muscles were incubated with caffeine revealed that at least 40% of the P(o) decrement at 2 h could be associated with a loss of efficiency of the excitation-contraction (E-C) coupling process. These data suggest that an important mechanism for the early loss in force is the inability to activate the contractile machinery likely caused by a failure in the E-C coupling process during the reloading period.

The primary purpose of the study was to examine circulating neutrophils and monocytes and their plasma membrane expression of CD64, CD11b, and CD18 after two bouts (B1 and B2) of eccentric exercise. Subjects (n = 10) performed 25 forced-lengthened contractions of the forearm flexors on two occasions separated by 3 wk. Blood samples were obtained before exercise and at 1.5, 6, 12, 24, 48, 72, and 96 h of recovery. CD64, CD11b, and CD18 expression was determined via direct immunofluorescence and used as an indicator of neutrophil and monocyte activation. Creatine kinase activity (B1 = 1,390, B2 = 108 U/l), myoglobin (B1 = 163, B2 = 41, ng/dl), and muscle soreness and tenderness were higher (P < 0.01) after B1 compared with B2. Neutrophils at 6, 12, and 96 h were higher (P < 0.05) for B1 vs. B2. CD11b expression on neutrophils was 2.7-fold higher at 72 h for B1 vs. B2. CD64 expression on neutrophils at 72 and 96 h was 1.4- and 1.9-fold higher, respectively, for B1 vs. B2. At 72 and 96 h, CD18 and CD64 expression on monocytes was 1.3-fold higher for B1 vs. B2. The observed changes were not significantly correlated with changes in creatine kinase activity or myoglobin. In conclusion, the adaptation to eccentric arm exercise was associated with a reduction in circulating neutrophils and a lower state of neutrophil and monocyte activation.

We tested the contribution of beta(2)-integrins, which are important for normal function of neutrophils and macrophages, to skeletal muscle hypertrophy after mechanical loading. Using the synergist ablation model of hypertrophy and mice deficient in the common beta-subunit of beta(2)-integrins (CD18(-/-)), we found that overloaded muscles of wild-type mice had greater myofiber size, dry muscle mass, and total protein content compared with CD18(-/-) mice. The hypertrophy in wild-type mice was preceded by elevations in neutrophils, macrophages, satellite cell/myoblast proliferation (5'-bromo-2'-deoxyuridine- and desmin-positive cells), markers of muscle differentiation (MyoD1 and myogenin gene expression and formation and size of regenerating myofibers), signaling for protein synthesis [phosphorylation of Akt and 70-kDa ribosomal protein S6 kinase (p70S6k)], and reduced signaling for protein degradation (decreased gene expression of muscle atrophy F box/atrogin-1). The deficiency in beta(2)-integrins, however, altered the accumulation profile of neutrophils and macrophages, disrupted the temporal profile of satellite cell/myoblast proliferation, reduced the markers of muscle differentiation, and impaired the p70S6k signaling, all of which could serve as mechanisms for the impaired hypertrophy in overloaded CD18(-/-) mice. In conclusion, our findings indicate that beta(2)-integrins contribute to the hypertrophic response to muscle overload by temporally regulating satellite cells/myoblast proliferation and by enhancing muscle differentiation and p70S6k signaling.

To determine the effect of carbohydrate (CHO) status on immune responses after long-duration exercise, on two occasions, 10 men completed a glycogen-depleting bout of cycle ergometry followed by 48 h of either a high-CHO diet (HiCHO; 8.0 g CHO/kg) or a low-CHO diet (LoCHO; 0.5 g CHO/kg). After the 48 h, subjects completed a 60-min ride at 75% maximal O2 uptake (EX). Blood samples were taken predepletion, pre-EX, post-EX, and 2 and 24 h post-EX and were assayed for leukocyte number and function, glucose, glutamine, and cortisol. The glucose responses were significantly higher in the HiCHO (4.62 +/- 0.26 mM) vs. the LoCHO (3.19 +/- 0.15 mM) condition post-EX, and glutamine was significantly higher in the HiCHO (0.472 +/- 0.036 mM) vs. the LoCHO (0.410 +/- 0.025 mM) condition throughout. Cortisol levels were significantly greater in the LoCHO (587 +/- 50 nM) vs. the HiCHO (515 +/- 62 nM) condition throughout the trial. Lymphocyte proliferation (phytohemagglutinin) was significantly depressed after exercise. However, there was no difference between conditions, and the depression was not correlated with elevations in cortisol. Circulating numbers of leukocytes, neutrophils, lymphocytes, and lymphocyte subsets were significantly greater in the LoCHO vs. the HiCHO condition at the post-EX and 2 h post-EX time points. These data indicate that the exercise and diet manipulation altered the number of circulating leukocytes but did not affect the decrease in lymphocyte proliferation that occurred after exercise.

The accumulation of neutrophils and macrophages, as well as the activation of satellite cells, are early events following skeletal muscle injury. We examined the temporal relationship between changes in neutrophils, macrophages, and MyoD protein, a marker of satellite cell activation, after injurious exercise. Male rats ( n=47) performed an intermittent downhill (-16% grade) running (17 m/min) protocol and the solei were obtained at 0, 2, 6, 24, 48, or 72 h post-exercise. Neutrophils, macrophages (ED1 and ED2), and MyoD+ cells were determined in muscle cross sections using immunohistochemistry. Downhill running increased ( P