225 research outputs found
Acute and Chronic Effects of a High-Intensity Interval Training Shock Microcycle on Cell-Free DNA: A Randomized Controlled Trial
No full textAbstract Background This study aimed to evaluate acute and chronic exercise-induced changes in cell-free DNA (cfDNA) concentrations during a 7-day high-intensity interval training (HIIT) shock microcycle in trained endurance athletes. Thirty-five participants were randomly assigned to one of three groups: a HIIT-only group (HSM), a HIIT plus low-intensity training group (HSM + LIT), and a control group maintaining regular training. The intervention included 10 HIIT sessions (5 × 4 min at 90–95% maximum heart rate) over 7 days, with HSM + LIT completing an additional 30 min of low-intensity training after each session. Physiological exercise testing (PET) was conducted at baseline, 3-, 7-, and 14-days post-intervention. On days 2 and 7 during the intervention, HIIT sessions were supervised in both morning and afternoon, and venous blood samples were collected at rest, immediately post-exercise, and 30 min post-exercise to measure cfDNA for 90 and 222 bp fragments. Correlations between cfDNA and physiological exercise variables such as peak power output (PPO), running velocity at lactate threshold (LT), and VO₂max were analyzed. Results cfDNA90 (10.4-fold, p < 0.001) and cfDNA222 (12.4-fold, p < 0.001) increased significantly after PET. In addition, cfDNA90 (17.1-fold, p < 0.001) and cfDNA222 (20.2-fold, p < 0.001) increased after HIIT, both remaining significantly elevated 30 min post-HIIT (both p < 0.001). cfDNA90 concentrations were higher in afternoon (22.4-fold) compared to morning HIIT sessions (17.2-fold, p < 0.001). A significant interaction effect was found between group and measurement point for cfDNA90 (p < 0.001) and cfDNA222 (p < 0.001), with higher concentrations in HSM + LIT compared to HSM 30 min post-HIIT. cfDNA90 showed moderate correlations with PPO (r = 0.48, p < 0.001), LT (r = 0.36, p < 0.001) and VO₂max (r = 0.30, p = 0.01). cfDNA222 correlated moderately with VO₂max (r = 0.34, p = 0.001) and slightly with PPO (r = 0.21, p = 0.05). No chronic changes in cfDNA were observed throughout the study period. Conclusions cfDNA is a reliable marker for detecting acute exercise-induced stress. However, the potential of cfDNA for detecting chronic adaptations in short-term, high-intensity interval training settings, such as a HIIT shock cycle, appears limited thus far. Trial registration clinicaltrials.gov, NCT05067426. Registered 05 October 2021—Retrospectively registered, https://clinicaltrials.gov/ct2/show/NCT05067426 . Graphical AbstractKey Points Acute increases in cfDNA concentrations were observed after physiological exercise testing and high-intensity interval training (HIIT). No chronic cfDNA changes were observed during and after a HIIT shock microcycle. Acute increases in cfDNA concentrations correlated with physiological exercise variables e.g. peak power output, running velocity at lactate threshold and VO2max during acute exercise. Higher increases in cfDNA were found after HIIT compared to physiological exercise testing. Higher increases in cfDNA were found after HIIT in the afternoon compared to morning. Higher increases in cfDNA were observed in males compared to females
Transgene detection by digital droplet PCR
No full textSomatic gene therapy is a promising tool for the treatment of severe diseases. Because of its abuse potential for
performance enhancement in sports, the World Anti-Doping Agency (WADA) included the term ‘gene doping’ in the official
list of banned substances and methods in 2004. Several nested PCR or qPCR-based strategies have been proposed that aim
at detecting long-term presence of transgene in blood, but these strategies are hampered by technical limitations. We
developed a digital droplet PCR (ddPCR) protocol for Insulin-Like Growth Factor 1 (IGF1) detection and demonstrated its
applicability monitoring 6 mice injected into skeletal muscle with AAV9-IGF1 elements and 2 controls over a 33-day period.
A duplex ddPCR protocol for simultaneous detection of Insulin-Like Growth Factor 1 (IGF1) and Erythropoietin (EPO)
transgenic elements was created. A new DNA extraction procedure with target-orientated usage of restriction enzymes
including on-column DNA-digestion was established. In vivo data revealed that IGF1 transgenic elements could be reliably
detected for a 33-day period in DNA extracted from whole blood. In vitro data indicated feasibility of IGF1 and EPO detection
by duplex ddPCR with high reliability and sensitivity. On-column DNA-digestion allowed for significantly improved target
detection in downstream PCR-based approaches. As ddPCR provides absolute quantification, it ensures excellent day-to-day
reproducibility. Therefore, we expect this technique to be used in diagnosing and monitoring of viral and bacterial infection,
in detecting mutated DNA sequences as well as profiling for the presence of foreign genetic material in elite athletes in the
future
Exercise-induced increases in cell free DNA in human plasma originate predominantly from cells of the haematopoietic lineage
No full textThe role of cell free DNA (cfDNA) has been intensively discussed under various pathological conditions and after acute bouts of exercise. To date, there is still no conclusive evidence concerning the cellular origin of cfDNA and the entire mechanism leading to elevated cfDNA concentrations in human plasma and serum. Here, we investigated the cellular origin of cfDNA in sex-mismatched haematopoietic stem cell transplantation (HSCT) and liver transplantation (LT) patients by determining the relative proportion of Y-chromosomal to total nuclear cfDNA. Total nuclear cfDNA and Y-chromosomal cfDNA concentrations were determined in blood plasma before and after an incremental exercise test via quantitative real-time PCR (qPCR). Female HSCT patients showed high proportions of Y-chromosomal cfDNA. Both total nuclear and Y-chromosomal cfDNA increased significantly and in a highly correlated fashion due to exercise. In male HSCT patients with female donors less than 10% of the cfDNA was of Y-chromosomal origin at any point in time and even though the total amount of cfDNA increased during exercise, no increases in Y-chromosomal DNA could be detected. The percentage of Y-chromosomal cfDNA in female LT patients with male donors was very low and levels remained unchanged during exercise. This indicates that cells not derived from the bone marrow, in this case transplanted liver cells, represented only a minor fraction of cfDNA in blood plasma and were not released during acute physical exercise. Even though many physiological conditions may be altered in transplant patients versus healthy people, our results strongly suggest that cells from the haematopoietic lineage are the main source of cfDNA released during acute bouts of exercise
Erythropoietin receptor is expressed in meningiomas and is downregulated in tumor recurrences
No full textErythropoietin receptor is expressed in meningiomas and is downregulated in tumor recurrences
No full textDie Rolle zellfreier DNA in der Vorhersage des klinischen Outcomes bei Herzinsuffizienz – Ergebnisse der MyoVasc Studie
No full textHeart failure (HF) represents a major cause of mortality with a prevalence of 1-2% in the adult population in developed countries. As populations age, the role of HF is expected to grow. This comes with a steep increase in healthcare costs, placing a substantial burden on society. Identifying HF risk patients earlier, possibly even before symptoms manifest, becomes crucial to initiate interventions promptly, such as lifestyle adjustments or medication. In preventive medicine, the identification of suitable biomarkers plays a key role, as they allow an objective and early disease detection.
Cell-free DNA (cfDNA) is a widely used diagnostic biomarker in clinical fields like oncology or transplantation medicine. In clinical cardiology, however, cfDNA analytics does not yet play a major role. Only a handful of rather small-scale studies have so far investigated the potential of cfDNA diagnostics in HF patients, indicating that cfDNA could be an independent risk factor for cardiovascular disease and overall mortality. The aim of the present study was therefore to evaluate the potential of cfDNA in HF diagnostics in a large cohort of subjects and to compare its predictive power to the currently most often used biomarker, NT-proBNP.
To achieve this, a reliable, reproduceable, and quick high throughput cfDNA quantification method needed to be implemented. The existing manual, time- and labour-consuming qPCR assay was automated by testing and establishing an INTEGRA pipetting robot and tuning its workflow to the special needs of high-viscosity plasma samples. The assay was adjusted to reliably produce the same test results as with the already published qPCR assay established by Neuberger et al. (183). This way a consistent measuring of the study samples was ensured. cfDNA levels were then quantified in 3109 EDTA plasma samples from the prospective MyoVasc study (NCT04064450). Two qPCR assays of different amplicon lengths (cfDNA90 bp/ cfDNA222 bp), both targeting a repetitive LINE1 element, were used for cfDNA quantification and to calculate the cfDNA integrity index, which indicates the fragmentation level of the cfDNA. Competing risk models were applied to investigate the associations of cfDNA with worsening of HF, and Cox proportional hazard regression analyses were used to assess the endpoints of cardiac death and all-cause death. C-statistics were calculated and
compared for each model. The participants were classified as 0 (healthy) or HF stages A (at risk for HF) to D (advanced HF) according to the current Universal Definition of Heart Failure. Analyses were adjusted for age, sex, cardiovascular risk factors (CVRFs) and medication (models 1-3) and additionally for NT-proBNP (model 4). Outcome data were presented as cumulative incidence plots for cfDNA90bp and 222bp levels and for the integrity index.
The cohort included 3109 study participants with an age between 34 to 85 years and 35.7% females. cfDNA concentration was lowest in stage 0/A subjects (n=534) with 10.99 (8.70/13.93) ng/ml (median (Q1/Q3)). Stage B (pre-HF) (n=923) or stage C/D subjects (n=1652) showed elevated cfDNA90bp concentrations with 13.37 (10.35/18.11) or 17.11 (12.56/22.80) ng/ml, respectively. Cox proportional hazard regression analyses indicated that the concentration of cfDNA90bp is a relevant prognostic marker for all-cause death, adjusted for age, sex, CVRFs and medication (HR = 1.312 [1.205-1.430], p < 0.0001). After additional adjustment for NT-proBNP, the effect estimates were lower, but still statistically significant (HR = 1.173 [1.073-1.282], p = 0.00046). Regarding the endpoints worsening of HF and cardiac death, the effect estimates were no longer significant after adjustment for NT-proBNP. A C-
index comparison showed the same tendency, with a significant added value of testing cfDNA additionally to NT-proBNP only when looking at all-cause death (C = 0.807 vs. C = 0.805; p = 0.050). However, cumulative incidence plots for dichotomised values of NT-proBNP and cfDNA showed the highest incidence rates for all three outcomes in patients with elevations in both biomarkers, significantly higher than in patients with elevations of NT-pro BNP alone.
The present results indicate that cfDNA is a risk factor, which independently of NT-proBNP contributes to the prediction of overall mortality (all-cause death) in the study cohort. cfDNA also appears to possess additional information value to NT-proBNP for predicting worsening of HF and cardiac death.Herzinsuffizienz (HI) ist mit einer Prävalenz von 1 bis 2 % in der erwachsenen Bevölkerung eine der Haupttodesursachen in den Industrieländern. Mit zunehmender Alterung der Bevölkerung wird auch die Bedeutung der HI in den kommenden Jahren zunehmen. Dies geht mit einem starken Anstieg der Gesundheitskosten einher und stellt eine erhebliche Belastung für die Gesellschaft dar. Die Identifizierung von HI-Risikopatienten zu einem früheren Zeitpunkt, möglicherweise sogar noch vor Auftreten von Symptomen, ist von entscheidender Bedeutung, um rechtzeitig Maßnahmen wie Lebensstiländerungen oder eine medikamentöse Therapie einleiten zu können. In der Präventivmedizin spielt die Identifizierung geeigneter Biomarker eine Schlüsselrolle, da sie eine objektive und frühzeitige Krankheitserkennung ermöglichen.
Zellfreie DNA (cfDNA) ist ein weit verbreiteter diagnostischer Biomarker in klinischen Bereichen wie der Onkologie oder der Transplantationsmedizin. In der klinischen Kardiologie spielt die cfDNA-Analytik jedoch bislang noch keine große Rolle. Nur eine Handvoll eher kleinerer Studien hat bisher das Potenzial der cfDNA-Diagnostik bei HI-Patienten untersucht. Diese Arbeiten deuten jedoch darauf hin, dass cfDNA ein unabhängiger Risikofaktor für Herz-Kreislauf-Erkrankungen und die Gesamtmortalität sein könnte. Ziel der vorliegenden Arbeit war es daher, das Potenzial von cfDNA in der HI-Diagnostik in einer größeren Studienkohorte zu untersuchen und ihre Vorhersagekraft mit dem derzeit am häufigsten verwendeten Biomarker, NT-proBNP, zu vergleichen.
Um dies zu erreichen, musste eine zuverlässige, reproduzierbare und schnelle Hochdurchsatzmethode zur Quantifizierung von cfDNA implementiert werden. Der bestehende manuelle, zeit- und arbeitsaufwändige qPCR-Assay wurde durch die Erprobung und Etablierung eines INTEGRA-Pipettierroboters automatisiert und dessen Arbeitsablauf auf die speziellen Bedürfnisse hochviskoser Plasmaproben hin abgestimmt. Der Assay wurde so angepasst, dass er zuverlässig die gleichen Testergebnisse liefert wie der bereits publizierte qPCR-Assay von Neuberger et al. (182). Auf diese Weise wurde eine konsistente Messung der Studienproben sichergestellt. Die cfDNA-Konzentration wurde in 3109 EDTA-Plasmaproben der prospektiven MyoVasc-Studie (NCT04064450) analysiert. Zur Quantifizierung der cfDNA und zur Berechnung des cfDNA-Integritätsindex wurden zwei qPCR-Assays mit unterschiedlichen Amplikonlängen (cfDNA90 bp/ cfDNA222 bp) verwendet, die beide auf ein repetitives LINE1-Element abzielen. Um die Assoziationen von cfDNA mit der Verschlechterung von HI zu untersuchen, wurden “Competing risk models“ angewandt.
Cox-Proportional-Hazard-Regressionsanalysen wurden verwendet, um die Endpunkte Herztod und Gesamtmortalität zu bewerten. Zusätzlich wurden C-Statistiken für jedes Modell berechnet und verglichen. Die Teilnehmer wurden als 0 (gesund) oder als HI-Stadien A (Risiko für HI) bis D (fortgeschrittene HI) gemäß der aktuellen universellen Definition der Herzinsuffizienz eingestuft. Die Analysen wurden für Alter, Geschlecht, kardiovaskuläre Risikofaktoren (CVRF) und Medikamente (Modelle 1-3) sowie zusätzlich für NT-proBNP (Modell 4) adjustiert. Die Ergebnisse wurden als kumulative Inzidenzdiagramme für die cfDNA-Assays 90bp und 222bp sowie für den Integritätsindex, der den Fragmentierungsgrad der cfDNA beschreibt, dargestellt.
Die Kohorte umfasste 3109 Studienteilnehmer mit einem Alter zwischen 34 und 85 Jahren bei einem Frauen-Anteil von 35,7 %. Personen im Stadium 0/A (n=534) zeigten mit 10,99 (8,70/13,93) ng/ml (Median (Q1/Q3)) die niedrigsten cfDNA90bp-Konzentrationen. Probanden im Stadium B (prä-HI) (n=923) oder im Stadium C/D (n=1652) wiesen erhöhte cfDNA90bp-Konzentrationen von 13,37 (10,35/18,11) bzw. 17,11 (12,56/22,80) ng/ml auf. Cox-Proportional-Hazard-Regressionsanalysen zeigten, dass die cfDNA90bp-Konzentration ein relevanter prognostischer Marker für die Gesamtmortalität ist, sofern eine Adjustierung für Alter, Geschlecht, CVRF und Medikation vorgenommen wurde (HR = 1,312 [1,205-1,430], p < 0,0001). Nach zusätzlicher Adjustierung für den etablierten Marker NT-proBNP waren die Effektschätzungen geringer, aber immer noch statistisch signifikant (HR = 1,173 [1,073-1,282], p = 0,00046). Für die Endpunkte „Verschlechterung der HF“ und „Herztod“ waren die
Effektschätzer nach Adjustierung für NT-proBNP hingegen nicht mehr statistisch signifikant. Ein C-Index-Vergleich zeigte ebenfalls einen statistisch signifikanten Zusatznutzen der cfDNA zusammen mit der Bestimmung von NT-proBNP bei Betrachtung der Gesamtmortalität (C = 0,807 vs. C = 0,805; p = 0,050). Diagramme für kumulative Inzidenzen von NT-proBNP- und cfDNA-Werten (dichotomisiert) zeigten die höchsten Inzidenzraten hingegen sogar für alle drei Endpunkte bei Patienten mit gleichzeitiger Erhöhung beider Biomarker. Die Inzidenzen waren signifikant höher als bei Patienten mit einer reinen Erhöhung von NT-pro BNP.
Die Ergebnisse deuten darauf hin, dass cfDNA ein Risikofaktor ist, der unabhängig von NT-proBNP zur Vorhersage der Gesamtmortalität in der Studienkohorte beiträgt. cfDNA besitzt offenbar auch einen zusätzlichen Nutzen zu NT-proBNP für die Vorhersage einer Verschlechterung der HI und des Herztodes.X, 100 Seiten : Illustrationen, Diagramm
Multimodal Web-Based Telerehabilitation for Patients With Post–COVID-19 Condition: Protocol for a Randomized Controlled Trial
No full textBackgroundPatients with post–COVID-19 condition (PCC) experience persistent, long-term health consequences following SARS-CoV-2 infection, including fatigue, hyperventilation, cognitive impairment, and limitations in daily activities. There is emerging evidence suggesting that exercise and respiratory therapy–based telerehabilitation is safe and could potentially improve physical capacity while reducing health care costs.
ObjectiveThis study aims to evaluate the superiority of a multimodal, symptom-titrated telerehabilitation program over standard care in patients with PCC who are severely affected, using the highest oxygen uptake rate (VO2peak [mL/min/kg]) achieved during the cardiopulmonary exercise test (CPET) and minute ventilation/carbon dioxide production slope (VE/VCO2 [full slope]) as primary outcomes. In addition, this study seeks to provide novel insights into the clinical and physiological adaptations associated with PCC, informing future rehabilitation strategies.
MethodsThis prospective, randomized, waitlist-controlled trial was approved by the Rhineland-Palatinate Medical Association ethics committee. All procedures comply with the Declaration of Helsinki. This study comprises 3 examination time points, which include patient-reported outcomes, clinical assessments, and a CPET. It is structured into an 8-week intervention phase followed by an 8-week follow-up phase. Following baseline assessment, patients will be randomly assigned to either the intervention group (IG) or the control group (CG). During the intervention phase, IG participants will receive a web-based, multimodal, symptom-titrated telerehabilitation program consisting of sports medicine consultations, weekly teleconsultations, a structured pacing approach, and exercise and respiratory therapy. In contrast, CG participants will receive treatment as usual, which includes a single sports medicine consultation on healthy habits and a self-directed pacing approach for managing symptoms and daily activities. During the follow-up phase, IG participants will continue training independently without teleconsultations, whereas CG participants will undergo the same telerehabilitation intervention as the IG. A follow-up assessment will be conducted for both groups to evaluate long-term effects. This study adheres to the SPIRIT (Standard Protocol Items: Recommendations for Interventional Trials) guidelines and follows the Consensus on Exercise Reporting Template.
ResultsRecruitment began in August 2023 and was extended until March 2025. As of March 2025, 80 participants have been recruited, and data analysis is ongoing. Final results are expected by December 2025, with a cross-sectional analysis of baseline data anticipated by July 2025.
ConclusionsThis study is the first randomized controlled trial investigating the effectiveness of multimodal and symptom-titrated telerehabilitation in patients with PCC who are severely affected. The integration of various objective diagnostic systems will provide valuable insights into emerging postviral fatigue syndromes, supporting the development of CPET-based diagnostics, personalized rehabilitation strategies, and future research on long-term telerehabilitation effectiveness. The findings will be disseminated through peer-reviewed publications, professional networks, and patient advocacy groups to ensure scientific, clinical, and public impact.
Trial RegistrationGerman Clinical Trials Register (DRKS) DRKS00032394; https://drks.de/search/de/trial/DRKS00032394
International Registered Report Identifier (IRRID)DERR1-10.2196/6504
Platelets, endothelial cells and leukocytes contribute to the exercise-triggered release of extracellular vesicles into the circulation
No full textPhysical activity initiates a wide range of multi-systemic adaptations that promote mental and physical health. Recent work demonstrated that exercise triggers the release of extracellular vesicles (EVs) into the circulation, possibly contributing to exercise-associated adaptive systemic signalling. Circulating EVs comprise a heterogeneous collection of different EV-subclasses released from various cell types. So far, a comprehensive picture of the parental and target cell types, EV-subpopulation diversity and functional properties of EVs released during exercise (ExerVs) is lacking. Here, we performed a detailed EV-phenotyping analysis to explore the cellular origin and potential subtypes of ExerVs.Healthy male athletes were subjected to an incremental cycling test until exhaustion and blood was drawn before, during, and immediately after the test. Analysis of total blood plasma by EV Array suggested endothelial and leukocyte characteristics of ExerVs. We further purified ExerVs from plasma by size exclusion chromatography as well as CD9-, CD63- or CD81-immunobead isolation to examine ExerV-subclass dynamics. EV-marker analysis demonstrated increasing EV-levels during cycling exercise, with highest levels at peak exercise in all EV-subclasses analysed. Phenotyping of ExerVs using a multiplexed flow-cytometry platform revealed a pattern of cell surface markers associated with ExerVs and identified lymphocytes (CD4, CD8), monocytes (CD14), platelets (CD41, CD42, CD62P), endothelial cells (CD105, CD146) and antigen presenting cells (MHC-II) as ExerV-parental cells. We conclude that multiple cell types associated with the circulatory system contribute to a pool of heterogeneous ExerVs, which may be involved in exercise-related signalling mechanisms and tissue crosstalk
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