Thomas Hilberg

Time-dependent mobilization of circulating progenitor cells during strenuous exercise in healthy individuals

Exercise stimulates the release of hematopoietic and endothelial progenitor cells (EPC) from the bone marrow. However, no data are available concerning the time frame of EPC release during strenuous exercise. The aim of the present study was to investigate the time-dependent release of progenitor cells during strenuous exercise. Eighteen healthy young men cycled for 4 h continuously at 70% of their individual anaerobic threshold. Peripheral blood was drawn at 16 predefined time points during and after finishing cycling. A significant rise in heart rate and leukocytes was obvious, whereas lactate levels and hematocrit did not change. The amount of circulating progenitor cells, EPCs, mature endothelial cells (mECs), and microparticles, quantified by flow cytometry, showed a significant time-dependent increase at 210/240 min. In addition a very early rise in VEGF and later increase in IL-6, both measured by ELISA, were evident. All observed changes were normalized 24 h after finishing the test. In conclusion, strenuous activity in healthy individuals leads to a time-dependent increase in mECs, PCs, and EPCs that may be related to VEGF and IL-6.

Physical training increases isometric muscular strength and proprioceptive performance in haemophilic subjects.

Summary. Sufficient muscular strength and proprioception lessen the risk of joint damage, however, both are impaired in haemophilic subjects. The aim of the study was to investigate proprioceptive performance and isometric muscular strength before and after a specialized training in haemophilic subjects (H) compared with two groups of control subjects (C). Nine subjects with severe haemophilia A, and eight ‘active’ C (AC) without haemophilia took part in a physical training programme over a 6‐month period. Eleven ‘passive’ C (PC) were requested to avoid any additional training during this period. Proprioceptive performance and isometric strength were determined before and after the training programme. The maximal isometric muscular strength in the legs, bilaterally measured by knee extensor (and leg press) was increased (P < 0.05) by 34% (29%) after training in the H and by 20% (28%) in the AC groups while remaining unchanged in the PC group. The performance in one‐leg‐stand tests after training was increased (P < 0.05) in the H and AC groups. An improvement of angle reproduction of 20° and 40° (P < 0.05) in the H compared with the PC groups was seen in the tests. Quantitative sensory testing by the tuning fork showed an increase (P < 0.05) in performance of both H and AC groups. The results of the present study confirm that specific sports therapy focused on proprioceptive function and accompanied by gentle strength training with low resistance and 20–25 repetitions is able to increase proprioceptive performance and muscular strength with a minimal stress to the joints. It is strongly recommended that specialized sports therapy be included as an integral component of the complete treatment regimen of haemophilic subjects.

Circulating microRNA-126 increases after different forms of endurance exercise in healthy adults

Background MicroRNAs (miRNAs) are small non-coding molecules regulating gene expression. Recently circulating miRNAs could be detected in the plasma, serving as novel biomarkers. Different forms of exercise mobilize progenitor cells from the bone marrow, helping in tissue repair. Data of different forms of exercise on endothelial cell damage are lacking. The aim of the study was to evaluate the impact of different exercise modalities on the plasma concentration of miRNA-126, as a marker for endothelial damage. Methods The plasma concentration of miRNA-126 and miRNA-133 (marker for muscle damage) was assessed by qRT-PCR analysis in plasma samples from healthy individuals performing one of the following exercise tests: (1) maximal symptom-limited exercise test, (2) bicycling for 4 h, (3) running a marathon, and (4) resistance exercise. Results A maximal symptom-limited exercise test resulted in a significant increase of circulating miRNA-126 at maximum power (2.1-fold versus begin), whereas the concentration of miRNA-133 remained unchanged. In line, four hours of cycling increased plasma concentration of miRNA-126 with a maximum 30 minutes after begin (4.6-fold versus begin) without an impact on miRNA-133 concentration. Finishing a marathon race resulted in an increase of miRNA-126 and miRNA-133. In contrast, eccentric resistance training led to an isolated increase of miRNA-133 level (2.1-fold versus begin) with unchanged miRNA-126. Conclusion Different endurance exercise protocols lead to damage of the endothelial cell layer as evident by an increase in miRNA-126. On the other hand, resistance exercise has no impact on the endothelial cells, but leads to a destruction of muscular cells.

Proprioception and isometric muscular strength in haemophilic subjects.

Haemophilia is characterized by intra‐articular bleeding, often requiring immobilization, which may result in muscle atrophy and impaired proprioception. The aim of the study was to investigate differences in proprioceptive performance and isometric muscular strength of the lower limbs in haemophilic subjects compared with control subjects. Twelve subjects with severe haemophilia (11 haemophilia A; one haemophilia B) vs. 12 control subjects were matched for anthropometric data and tested for differences of proprioception (one‐leg‐stand, posturomed, angle‐reproduction, and tuning fork tests) and isometric strength (leg press, knee extensor). The static proprioceptive performance of the haemophilic group, as measured by the one‐leg‐stand test (on hard or soft ground, with open or closed eyes; P < 0.05) was demonstrably impaired (by 41–363%). In contrast, the dynamic proprioceptive perfomance measured by the posturomed test did not show any difference between the groups. The local proprioceptive performance (angle‐reproduction test) of the knee, (the most commonly affected joint in haemophiliacs) showed a trend to impaired function but was not distinctly different from that of controls. The quantitative sensory function (tuning fork) showed significant (P < 0.05) impairment of 9–10% in the haemophilic subjects. Additionally, the isometric muscular strength of the leg extensor was weaker (32–38%) in the haemophilic group when the limbs were tested individually as well as bilaterally (P < 0.05). In conclusion, the results suggest that global proprioceptive performance is impaired and that the isometric strength of the leg extensors is weaker in the haemophilic subjects. Therefore, specialized training for global proprioception would be helpful in order to compensate for proprioceptive deficits. This exercise regimen should also include safe strength‐training for an optimal stabilization of the joints, but must be adapted to the individual needs and situations of the haemophilic subjects.

Variations in the ratio between von Willebrand factor and its cleaving protease during systemic inflammation and association with severity and prognosis of organ failure.

Von Willebrand factor (VWF) and related parameters as well as the protease activity regulating its biological activity were measured in plasma of healthy controls and patients with different cause and severity of systemic inflammation to examine the efficacy of the measures to detect highly prothrombotic states including thrombotic microangiopathy (TMA), one of the sequelae of sepsis. Plasma levels of VWF increased with increasing severity of systemic inflammation, probably due to activation of the endothelium. In parallel, the proteolytic activity of VWF inactivating protease, ADAMTS13, stepwise declined with the severity of inflammation, emphasizing the role of VWF-triggered platelet aggregation on the endothelium subsequently followed by development of TMA. As a consequence, the ratio of VWF antigen level and ADAMTS13 activity was significantly higher in patients with inflammation and sepsis, suggesting that this ratio might be more useful for the diagnosis of highly prothrombotic states including TMA than VWF multimer analysis alone. These findings suggest that ADAMTS13, VWF and related parameters, even in a combined approach, might be useful for the diagnosis and the therapeutic monitoring of patients with sepsis associated thrombotic microangiopathy.

Transcription in response to physical stress— clues to the molecular mechanisms of exercise-induced asthma

To clarify stress‐induced immunological reactions and molecular events during exercise and the potential relevance to exercise‐induced bronchoconstriction, transcriptional responses to standardized physical stress were determined. Six healthy, young volunteers underwent an endurance exercise of 90% of their individual anaerobic threshold for 90 min. Time‐dependent alterations in the expression pattern of leukocytes from healthy, trained subjects were analyzed by DNA microarrays before and 2 h and 6 h after exercise. Starting out from a large collection of cDNA library clones comprising more than 70,000 human expressed sequence tags, we selected, designed, and immobilized oligonucleotide probes (60–70mers) for transcripts of 5000 stress‐and inflammation‐relevant genes. Exercise‐induced stress provoked changes in the expression of 433 gene activities 2 h and/or 6 h after exercise, which could be grouped into six clusters. The most prominent feature was an enhanced transcription of two genes, coding for 5‐lipoxygenase (ALOX5) and ALOX5‐activating protein. Moreover, enhanced levels of leukotriene B4 (LTB4) and LTC4 (P<0.05) were detected in plasma after exercise. Our data demonstrate that exercise alters the activities of a distinct number of genes. In particular, they possibly provide novel insights into the molecular mechanisms of exercise‐induced bronchoconstriction and suggest that enhanced transcription of ALOX5 and its activating protein together with a present predisposition of the subject critically contribute to exercise‐induced asthma.

Blood coagulation and fibrinolysis after extreme short-term exercise

INTRODUCTION Maximal exercise may be a trigger for cardiovascular events. The aim of the study was to investigate changes in blood coagulation and fibrinolysis following maximal short-term exercises with different durations up to 90 s. METHODS A total of 15 healthy nonsmokers underwent three isokinetic maximal tests on an SRM cycle ergometry system with durations of 15, 45, and 90 s. Blood samples were taken after a 30-min rest, immediately before and after exercise, 15 min, and 1 h after completion of exercise. For the investigation of blood coagulation, prothrombin fragment 1+2 (F1+2), thrombin-antithrombin III complex (TAT), intrinsic and extrinsic total (TTPin+ex), and endogenous thrombin potential (ETPin+ex) were measured. For testing fibrinolysis, determinations of plasmin-alpha(2)-antiplasmin complex (PAP), tissue-type plasminogen activator (tPA)-antigen, plasminogen activator inhibitor (PAI)-1-antigen and D-dimer were used. RESULTS Immediately after the exercise tests, only F1+2 (15- and 90-s test) and TTPin (45 and 90 s) showed a moderate increase (p<0.05), while TAT and ETP was unchanged. In contrast, a clear increase in PAP and tPA-antigen already after 15 s maximal exercise in relation to the exercise duration time could be investigated. These effects were not totally reversed to baseline 15 min after exercise; D-dimer and PAI-1-antigen still remained unchanged after these types of exercise. CONCLUSIONS Maximal short-term exercise does not lead to a relevant activation of blood coagulation in healthy young subjects, it is only slightly altered within the normal range. In contrast, fibrinolysis is clearly activated, and the increase is directly dependent on exercise duration. Additionally, it could be shown for the first time that fibrinolysis is already activated after 15 s maximal exercise duration.

Joint pain in people with hemophilia depends on joint status.

Summary People with hemophilia show a clinical severity‐related decrease in mechanical pain thresholds in affected joints, but cope rather well with their chronic pain. ABSTRACT Recurrent joint bleedings in people with hemophilia (PWH) often progress into the full clinical picture of hemophilic arthropathy, accompanied by chronic pain. Although chronic pain is commonly present in PWH, investigations assessing pain thresholds have not been performed yet. Thus, the aim of this study was to obtain objective and subjective measures of joint pain in PWH and to relate these to the severity of joint pathology. Thirty‐six patients (aged 43 ± 11 years) with hemophilia A and B (31 severe A, 1 B; 3 moderate A, 1 B) and 40 healthy control subjects (aged 42 ± 14 years) participated in this study. Mechanical pain thresholds were obtained as objective parameters using an algometer, while subjective pain intensity and quality were assessed using numeric analogue scales. Quality of life was estimated using the Short‐Form Health Survey (SF‐36) questionnaire. Overall, we found reduced mechanical pain thresholds as obtained from the knee (PWH – left 38.1 [28.7/57.7], right 29.5 [20.9/49.3]; control – left 67.4 [56.8/112.6], right 60.9 [42.6/97.2]), and elbow (PWH – left 23.4 [15.3/33.4], right 23.5 [20.1/35.1]; control – left 56.7 [32.6/86.6], right 53.0 [30.7/87.7] in N; median [25th/75th percentile]) joints in PWH. Interestingly, this increased pain sensitivity was related to the severity of clinical joint pathology. In addition, PWH reported their pain in a more descriptive and not affective manner and scored similar to controls in the mental domain of the SF‐36, thereby indicating good coping strategies despite the chronic nature of their complaints. In conclusion, pain sensitivity at the site of the affected joints is increased and closely related to joint pathology in people with hemophilia.

Maximal and submaximal endurance performance in adults with severe haemophilia

Summary.  Maximal exercise testing, including the determination of maximal performance and maximal oxygen uptake (VO2max), is considered the gold standard for assessing maximal endurance performance. The effectiveness of such testing is often reduced in haemophilic adults owing to musculoskeletal impairments or pain rather than because of cardiac exertion. The measurement of submaximal performance parameters overcomes many limitations of maximal exercise testing but a testing standard is still lacking. The aim of this study was to investigate maximal and particularly submaximal endurance performance of adult patients with severe haemophilia A and B. Eleven patients and 11 matched healthy controls were tested by spiroergometry with a specific treadmill test and the power was calculated in Watts. The haemophilic group achieved lower absolute (210 ± 63 W) and weight‐related (2.94 ± 0.98 W kg−1) maximal endurance performance compared with the control group (287 ± 50 W resp. 3.82 ± 0.53 W kg−1; P ≤ 0.05). The patients also showed a lower submaximal endurance performance at the individual anaerobic threshold (IAT = 147 ± 56 W) and fixed lactate values (2 mmol = 98 ± 60 W; 4 mmol = 158 ± 56 W) compared with the healthy controls (IAT = 210 ± 41 W; 2 mmol = 153 ± 30 W; 4 mmol = 223 ± 39 W; all P ≤ 0.05). The heart rate and lactate value at the IAT were not different. The disease‐related musculoskeletal changes in haemophilic adults lead to a reduced maximal and submaximal endurance performance, which can be easily measured by the described test procedure.

Platelet activity, sensitivity to agonist, and platelet-leukocyte conjugate formation after long-term exercise

For rehabilitation training it is recommended that the intensity of exercise should be distinctly below the individual anaerobic threshold (IAT). We investigated platelet activity, reactivity and platelet-leukocyte conjugate formation following a stardardized treadmill (TR) ergometer test at 90% IAT for 60-120 min. Seventeen healthy male non-smokers underwent TR. Blood samples were taken after a 30-min rest, immediately after exercise, and 2 h after exercise completion. Platelets were detected flow cytometrically by CD41 in whole blood, activated platelets by CD62P. In addition, stimulation of platelets in vitro with 7.5 w M TRAP-6 was performed. For testing platelet-leukocyte conjugates, antibodies against CD45 and CD41 were used. After TR the percent of non-stimulated CD62P-positive platelets (%PC) remained unchanged (1.65 - 0.56 to 1.73 - 0.79%PC) (mean - SD). In contrast, an increase ( P <0.05) from 31.9 - 13.5 to 37.4 - 15.0 %PC in CD62P, TRAP-6 stimulated and enhanced (P<0.01) platelet-leukocyte conjugates (11.7 - 3.7 to 16.1 - 6.9, CD41-%PC) after TR were observed. Both changes were independent of thrombin generation measured by F1+2 and TAT, and reversible after 2 h. Long-term exercise (90% IAT) on a treadmill ergometer only leads to a moderate increase of platelet reactivity and platelet-leukocyte conjugates. The determination of platelet-leukocyte conjugates may offer the possibility to detect an early activation stage of platelets in vivo .