Cell-free DNA in plasma as an essential immune system regulator.

The cell-free DNA (cfDNA) is always present in plasma, and it is biomarker of growing interest in prenatal diagnostics as well as in oncology and transplantology for therapy efficiency monitoring. But does this cfDNA have a physiological role? Here we show that cfDNA presence and clearance in plasma of healthy individuals plays an indispensable role in immune system regulation. We exposed THP1 cells to healthy individuals' plasma with (NP) and without (TP) cfDNA. In cells treated with NP, we found elevated expression of genes whose products maintain immune system homeostasis. Exposure of cells to TP triggered an innate immune response (IIR), documented particularly by elevated expression of pro-inflammatory interleukin 8. The results of mass spectrometry showed a higher abundance of proteins associated with IIR activation due to the regulation of complement cascade in cells cultivated with TP. These expression profiles provide evidence that the presence of cfDNA and its clearance in plasma of healthy individuals regulate fundamental mechanisms of the inflammation process and tissue homeostasis. The detailed understanding how neutrophil extracellular traps and their naturally occurring degradation products affect the performance of immune system is of crucial interest for future medical applications.

Increasing Veno-Arterial Extracorporeal Membrane Oxygenation Flow Reduces Electrical Impedance of the Lung Regions in Porcine Acute Heart Failure.

Veno-arterial extracorporeal membrane oxygenation (VA ECMO) is a technique used in patients with severe heart failure. The aim of this study was to evaluate its effects on left ventricular afterload and fluid accumulation in lungs with electrical impedance tomography (EIT). In eight swine, incremental increases of extracorporeal blood flow (EBF) were applied before and after the induction of ischemic heart failure. Hemodynamic parameters were continuously recorded and computational analysis of EIT was used to determine lung fluid accumulation. With an increase in EBF from 1 to 4 l/min in acute heart failure the associated increase of arterial pressure (raised by 44%) was accompanied with significant decrease of electrical impedance of lung regions. Increasing EBF in healthy circulation did not cause lung impedance changes. Our findings indicate that in severe heart failure EIT may reflect fluid accumulation in lungs due to increasing EBF.

Regional tissue oximetry reflects changes in arterial flow in porcine chronic heart failure treated with venoarterial extracorporeal membrane oxygenation.

Venoarterial extracorporeal membrane oxygenation (VA ECMO) is widely used in treatment of decompensated heart failure. Our aim was to investigate its effects on regional perfusion and tissue oxygenation with respect to extracorporeal blood flow (EBF). In five swine, decompensated low-output chronic heart failure was induced by long-term rapid ventricular pacing. Subsequently, VA ECMO was introduced and left ventricular (LV) volume, aortic blood pressure, regional arterial flow and tissue oxygenation were continuously recorded at different levels of EBF. With increasing EBF from minimal to 5 l/min, mean arterial pressure increased from 47+/-22 to 84+/-12 mm Hg (P<0.001) and arterial blood flow increased in carotid artery from 211+/-72 to 479+/-58 ml/min (P<0.01) and in subclavian artery from 103+/-49 to 296+/-54 ml/min (P<0.001). Corresponding brain and brachial tissue oxygenation increased promptly from 57+/-6 to 74+/-3 % and from 37+/-6 to 77+/-6 %, respectively (both P<0.01). Presented results confirm that VA ECMO is a capable form of heart support. Regional arterial flow and tissue oxygenation suggest that partial circulatory support may be sufficient to supply brain and peripheral tissue by oxygen.

Novel porcine model of acute severe cardiogenic shock developed by upper-body hypoxia.

Despite the urgent need for experimental research in the field of acute heart failure and, particularly cardiogenic shock, currently there are only limited options in large animal models enabling research using devices applied to human subjects. The majority of available models are either associated with an unacceptably high rate of acute mortality or are incapable of developing sufficient severity of acute heart failure. The objective of our research was to develop a novel large animal model of acute severe cardiogenic shock. Advanced left ventricular dysfunction was induced by global myocardial hypoxia by perfusing the upper body (including coronary arteries) with deoxygenated venous blood. The model was tested in 12 pigs: cardiogenic shock with signs of tissue hypoxia developed in all animals with no acute mortality. Cardiac output decreased from a mean (+/- SD) of 6.61+/-1.14 l/min to 2.75+/-0.63 l/min, stroke volume from 79.7+/-9.8 ml to 25.3+/-7.8 ml and left ventricular ejection fraction from 61.2+/-4.3 % to 17.7+/-4.8 % (P

Head-up tilt rapidly compromises hemodynamics in healthy anesthetized swine.

The aims were to explore the effect of head-up tilt (HUT) to 30 and 60 degrees on hemodynamics and tissue oxygenation in anesthetized healthy swine. The data serve as a reference for a study of resuscitation efficacy at HUT such as during transport. Nine healthy swine (49+/-4 kg) were anesthetized and multiple sensors including myocardial pressure-volume loops catheter, carotid flow probe, blood pressure catheters, near infrared spectroscopy (NIRS) tissue oximetry and mixed venous oximetry (SVO2) catheter were introduced and parameters continuously recorded. Experimental protocol consisted of baseline in supine position (15 min), 30 degrees HUT (15 min), recovery at supine position (15 min) and 60 degrees HUT (5 min). Vacuum mattress was used for body fixation during tilts. We found that 30 and 60 degrees inclination led to significant immediate reduction in hemodynamic and oximetry parameters. Mean arterial pressure (mm Hg) decreased from 98 at baseline to 53 and 39, respectively. Carotid blood flow dropped to 47 % and 22 % of baseline values, end diastolic volume to 49 % and 53 % and stroke volume to 47 % and 45 % of baseline. SVO2 and tissue oximetry decreased by 17 and 21 percentage points. The values are means. In conclusions, within minutes, both 30 and 60 degrees head-up tilting is poorly tolerated in anesthetized swine. Significant differences among individual animals exist.

Direct coupling of a free-flow isotachophoresis (FFITP) device with electrospray ionization mass spectrometry (ESI-MS).

We present the online coupling of a free-flow isotachophoresis (FFITP) device to an electrospray ionization mass spectrometer (ESI-MS) for continuous analysis without extensive sample preparation. Free-flow-electrophoresis techniques are used for continuous electrophoretic separations using an electric field applied perpendicular to the buffer and sample flow, with FFITP using a discontinuous electrolyte system to concurrently focus a target analyte and remove interferences. The online coupling of FFITP to ESI-MS decouples the separation and detection timeframe because the electrophoretic separation takes place perpendicular to the flow direction, which can be beneficial for monitoring (bio)chemical changes and/or extensive MS(n) studies. We demonstrated the coupling of FFITP with ESI-MS for simultaneous concentration of target analytes and sample clean-up. Furthermore, we show hydrodynamic control of the fluidic fraction injected into the MS, allowing for fluidically controlled scanning of the ITP window. Future applications of this approach are expected in monitoring biochemical changes and proteomics.

Are there any differences between stress hormone levels in non-stress conditions and in potentional stress overload (heart catheterisation) in sows?

In order to study a possible effect of mini-invasive heart intervention on a response of hypothalamo-pituitary-adrenal stress axis, we analyzed four stress markers (cortisol, cortisone, DHEA and DHEAS) in 25 sows using minimally invasive heart catheterisation as the stress factor. The marker levels were assessed in four periods of the experiment, (1) the baseline level on the day before intervention, (2) after the introduction of anesthesia, (3) after conducting tissue stimulation or ablation, and (4) after the end of the catheterisation. For statistical analyses we used the non-parametric Friedman test for four dependent samples (including all four stages of the operation) or three dependent samples (influence of operation only, baseline level was excluded). Statistically significant differences in both Friedman tests were found for cortisol and for cortisone. Significant differences for DHEA as well as for DHEAS were found for all tested stages but not for the effect of operation itself. We have concluded that cortisol levels are blunted by the influence of anesthesia after its administration, and therefore decrease back to the baseline at the end of the operation. The other markers (cortisone, DHEA and DHEAS) acted as balanced systems against the injurious stress effect.

From chip-in-a-lab to lab-on-a-chip: towards a single handheld electronic system for multiple application-specific lab-on-a-chip (ASLOC).

We present a portable, battery-operated and application-specific lab-on-a-chip (ASLOC) system that can be easily configured for a wide range of lab-on-a-chip applications. It is based on multiplexed electrical current detection that serves as the sensing system. We demonstrate different configurations to perform most detection schemes currently in use in LOC systems, including some of the most advanced such as nanowire-based biosensing, surface plasmon resonance sensing, electrochemical detection and real-time PCR. The complete system is controlled by a single chip and the collected information is stored in situ, with the option of transferring the data to an external display by using a USB interface. In addition to providing a framework for truly portable real-life developments of LOC systems, we envisage that this system will have a significant impact on education, especially since it can easily demonstrate the benefits of integrated microanalytical systems.

Pig model of pulmonary embolism: where is the hemodynamic break point?

Early recognition of collapsing hemodynamics in pulmonary embolism is necessary to avoid cardiac arrest using aggressive medical therapy or mechanical cardiac support. The aim of the study was to identify the maximal acute hemodynamic compensatory steady state. Overall, 40 dynamic obstructions of pulmonary artery were performed and hemodynamic data were collected. Occlusion of only left or right pulmonary artery did not lead to the hemodynamic collapse. When gradually obstructing the bifurcation, the right ventricle end-diastolic area expanded proportionally to pulmonary artery mean pressure from 11.6 (10.1, 14.1) to 17.8 (16.1, 18.8) cm(2) (p<0.0001) and pulmonary artery mean pressure increased from 22 (20, 24) to 44 (41, 47) mmHg (p<0.0001) at the point of maximal hemodynamic compensatory steady state. Similarly, mean arterial pressure decreased from 96 (87, 101) to 60 (53, 78) mmHg (p<0.0001), central venous pressure increased from 4 (4, 5) to 7 (6, 8) mmHg (p<0.0001), heart rate increased from 92 (88, 97) to 147 (122, 165) /min (p<0.0001), continuous cardiac output dropped from 5.2 (4.7, 5.8) to 4.3 (3.7, 5.0) l/min (p=0.0023), modified shock index increased from 0.99 (0.81, 1.10) to 2.31 (1.99, 2.72), p<0.0001. In conclusion, instead of continuous cardiac output all of the analyzed parameters can sensitively determine the individual maximal compensatory response to obstructive shock. We assume their monitoring can be used to predict the critical phase of the hemodynamic status in routine practice.

TUGENDHAT: a pilot randomized study on effects of biventricular pacing in patients with bradycardia pacing indication and normal systolic function on heart failure, atrial fibrillation and quality of life (results of 12 month follow-up).

BACKGROUND: Since the late 1990s, a growing number of clinical studies have indicated that long-term permanent right ventricular (RV) apical pacing will induce severe complications such as development of heart failure, increased burden of atrial fibrillation leading to decreased quality of life. AIM OF THE STUDY: To investigate whether cardiac resynchronization therapy (CRT) using biventricular (BiV) pacing can prevent the development of left ventricular (LV) dysfunction, LV remodelling, worsening of the clinical status and quality of life in chronically RV paced patients with normal LV ejection fraction (EF). METHODS AND RESULTS: A total of 127 patients with Class I indication for permanent cardiac pacing and without established indication for CRT were subjected to 6 months of RV and BiV pacing in a patient-blinded, randomized crossover trial. Treatment effects of BiV pacing were evaluated for LV function, LV remodelling and clinical status. As compared with RV pacing, BiV pacing did not significantly prevent the decrease of LV function [LVEF 61.0 % (36.0; 68.0) vs 60.5 % (38.5; 67.5) in RV pacing], did not change the functional class according to the New York Heart Association [52 % in Class II vs 53.9 % in Class II in RV pacing, and 3.9 % in Class III vs 6.9 % in Class III in RV pacing], and did not present any changes in quality of life [32.5 (18.0; 80.0) vs 32.0 (21.0; 47.0) indexes in RV pacing]. CONCLUSION: BiV pacing, compared to RV pacing, did not change LV function and quality of life in patients with the absence of LV dysfunction or remodelling, standard bradycardia pacing indications in a pilot phase (12- month follow-up) of the TUGENDHAT trial. The final report will be published after 60-month follow-up termination (Tab. 5, Fig. 3, Ref. 30).

Protein-carbohydrate complex reveals circulating metastatic cells in a microfluidic assay.

Advances in carbohydrate sequencing technologies reveal the tremendous complexity of the glycome and the role that glycomics might have to bring insight into the biological functions. Carbohydrate-protein interactions, in particular, are known to be crucial to most mammalian physiological processes as mediators of cell adhesion and metastasis, signal transducers, and organizers of protein interactions. An assay is developed here to mimic the multivalency of biological complexes that selectively and sensitively detect carbohydrate-protein interactions. The binding of ß-galactosides and galectin-3--a protein that is correlated to the progress of tumor and metastasis--is examined. The efficiency of the assay is related to the expression of the receptor while anchoring to the interaction's strength. Comparative binding experiments reveal molecular binding preferences. This study establishes that the assay is robust to isolate metastatic cells from colon affected patients and paves the way to personalized medicine.

Hemodynamic and metabolic parameters during prolonged cardiac arrest and reperfusion by extracorporeal circulation.

Extracorporeal membranous oxygenation (ECMO) is increasingly used in the management of refractory cardiac arrest. Our aim was to investigate early effects of ECMO after prolonged cardiac arrest. In fully anesthetized swine (48 kg, N=18) ventricular fibrillation (VF) was induced and untreated period (20 min) of cardiac arrest commenced, followed by 60 min extracorporeal reperfusion (ECMO flow 100 ml/kg.min). Hemodynamics, arterial blood gasses, plasma potassium, tissue oximetry (StO(2)) and cardiac (EGM) and cerebral (BIS) electrophysiological parameters were continuously recorded and analyzed. Within 3 minutes of VF hemodynamic and oximetry parameters fall abruptly while metabolic parameters destabilize gradually over 20 minutes peaking at pH 7.04 ± 0.05, pCO(2) 89 ± 14 mmHg, K(+) 8.5 ± 1.6 mmol/l. During reperfusion most parameters restore rapidly: within 3-5 minutes mean arterial pressure reaches >40 mmHg, StO(2)>50 %, paO(2)>100 mmHg, pCO(2)<50 mmHg, K(+)<5 mmol/l. EGMs mean amplitude peaks at 4.5 ± 2.4 min. Cerebral activity (BIS>60) reappeared in 5 animals after 87 ± 21 min. In 12/18 animals return of spontaneous circulation was achieved. In conclusions, ECMO provides rapid restitution of internal milieu even after prolonged arrest. However, despite normalization of global parameters full recovery was not guaranteed since cardiac and cerebral electrical activities were sufficiently restored only in some animals. More sensitive and organ specific indicators need to be identified in order to estimate adequacy of cardiac support devices.

[Deficits in lipid lowering therapy and achieving target lipid concentrations in patients with high risk of a cardiovascular event]. / Rezervy v hypolipidemické terapii a dosazení cílových koncentrací lipidu u pacientu s vysokým rizikem kardiovaskulárních príhod.

Lipid concentrations were analysed in 529 consecutive patients, 190 females (34%) and 339 males (66%) with mean age 66 years, at high cardiovascular risk, undergoing elective coronary angiography between 1st January and 31st May 2010. LDL-cholesterol level < 2,5 mmol/l was identified only in 36% of patients in primary prevention, LDL-cholesterol level < 2,0 mmol/l was identified only in 28% of subjects in secondary prevention of cardiovascular events. The study proves inadequate control of dyslipidaemia in patients at high cardiovascular risk.

Might cardiac catheterization influence diurnal rhythm of the steroid stress hormones secretion?

Glucocorticoids (GCs) are steroid hormones produced by the adrenal cortex in reaction to stress stimuli. GCs production is not stable over a 24-hour period; the plasma concentration peaks in the morning (approximately upon awakening) and then the plasma levels decrease, reaching the nadir in the evening. In our experiments, the levels of cortisol, cortisone, DHEA and DHEAS were tested in young female pigs (n=23) during heart catheterization at two different day times (in the morning and in the afternoon). The non-parametric Mann-Whitney test for statistical analysis was used. We found only minimal statistical differences in studied markers between the morning and afternoon group (p>0.05). The absence of circadian variation in GCs levels could originate either at an early age of our experimental pigs, or in stressful conditions on the experiment day, or most likely the day before (e.g. social isolation, fasting, transport, and catheterization), respectively. We can conclude there is no difference in the stress load between morning and afternoon experiments, and therefore we can assume the stress load is not a limiting factor for the timing when catheterization should be preferably performed.

Critical evaluation of the optimal medical therapy in the cardiac resynchronization therapy candidates--single centre experience.

UNLABELLED: The aim of the present study was to evaluate the optimal medical therapy in the chronic heart failure (CHF) patients referred from the comunity centres and the outpatients cardiology clinics for the cardiac resynchronization therapy with defibrilator (CRTD) to the Department of Cardiology, Na Homolce Hospital with the device implantation between 1st January 2008 and 30st September 2009. METHODS: The optimal medical therapy was analysed retrospectively from the medical records of 179 consecutive CHF patients NYHA class III-IV. Beta-blockers (BB) were used only in 81% subjects referred for CRTD, ACE inhibitors (ACEI) were used only in 68% patients Angiotensin receptor blockers (ARB) were used in 18% subjects. ACEI or ARB were used in 81%, spironolacton was use in 59%. Recommended target DD for BB (carvedilol 25 mg bid) was used only in 13% subjects, recommended target DD for ACEI (enalapril 10 mg bid) was used only in 9.4% patients. RESULTS: In the Department of Cardiology, the optimal medical therapy was changed after CRTD, BB were used in 95% subjects at discharge (p < 0.01) and the number of patients reaching at least of 50% of recommended daily dose (DD) of BB increased (p < 0.05). ACEI were recommended after CRTD in 80% subjects after implantation (p < 0.05), the number of patients reaching at least of 50% of recommended DD for ACEIs increased too (p < 0.05). There was no significant difference in ARB use recomended in the hospital (19% after CRTD - NS). ACEI or ARB were used in 98% patients after the device implanted (p < 0.05) and spironolacton in 77% after CRTD (p < 0.05). CONCLUSIONS: Despite optimal composition of the optimal medical heart failure therapy only small number of CRTD candidates are reaching recommended drug dose. The optimization of the medical therapy in the specialized center lead to significantly higher proportion of CHF using the optimal therapy with the increased dose of BB and ACEI.

Is there circadian variation in cortisol levels in young sows in heart catheterization?

Cortisol is the main glucocorticoid (GC) hormone in pigs associated with stress response. It is well known that GCs levels are not stable during the day; their concentration is a circadian variable with the peak in the morning and the nadir in the night (in diurnal animals). Circadian variation is present during postnatal ontogeny. The onset of the circadian fluctuation occurs in pigs at the age of 3 to 20 weeks (according to the literature). The aim of our pilot study was to determine if young sows (used in cardiosurgical experiments) already developed the circadian variation. Twelve-week-old sows were used in the heart catheterization experiment. Cortisol was measured during four different stages of the experiment at two different times of the day (the operation was performed in the morning or afternoon). To determine circadian variation the Mann-Whitney test was used; to determine changes in cortisol levels within the experiment the Friedman test was performed. We didn't find any circadian variation (p>0.05) or statistical significant variation in the Friedman test (p>0.05). We assumed that our pigs are too young to have circadian rhythm present. Our findings are in accordance with many authors.

[Embolic complication of the postcathetrization femoral artery pseudoaneurysm treatment using percutaneous thrombotization. A case review]. / Embolická komplikace lécby poskatetrizacního pseudoaneuryzmatu arteria femoralis perkutánní trombotizací. Kazuistika.

Considering frequencies of the heart cathetrization procedures, including treatment of pseudoaneurysms of the femoral artery using percutaneous thrombotization, a possibility of peripheral arterial emobolization should be considered. The authors describe management of the relatively rare complication in the treatment of postcathetrization pseudoaneurysms. According to the case review, clearing the obliterating arterial embolus of the tissue glue is technically feasible even more than 24 hours folowing the embolization.

[Infectious endocarditis as a complication of cardiac pacemakers requiring emergency removal of the stimulating electrode]. / Infekcní endokarditida jako komplikace trvalé kardiostimulace s nutností chirurgické extrakce stimulacní elektrody.

Case report describes a patient with permanent pacemaker (implanted in childhood) who has developed infectious endocarditis with bacterial vegetation on the stimulation electrode while no febrile condition occurred and laboratory tests showed no inflammation. The course of the disease was affected by the previous inappropriate outpatient treatment of an infection in the suture after the pacemaker exchange. It should be noted that echocardiographic examination was essential for both the diagnostics and the choice of strategy. The therapeutic approach consisted of the combination of long-term antibiotic therapy and the explantation of the infected stimulation system. As it was not possible to accomplish by endovasal methods only, cardiosurgery for extraction of the lead was chosen.

[Cardiac bradyarrhythmia ]. / Srdecní bradyarytmie.

Permanent cardiac pacing was introduced in 1958 and till the end of 70s this method saved lives in particular of the patients with advanced atrioventricular block. The implantation technique has changed from complicated thoracotomy to endovasal approaches. The introduction of physiological AV sequenced atrioventricular pacing marked a significant progress in this field. Acute haemodynamic studies documented positive effect of the atrial contribution. Numerous studies subsequently analyzed the influence of different pacing regimes on total and specific cardiovascular mortality and morbidity. It can be concluded that on the basis of present evidence-based medicine the use of physiological pacing is clearly indicated in the patients with expressed sinus bradycardia and AV block of a higher degree. Atrial pacing remains an ideal solution for the patients with isolated sinus node dysfunction and sufficient atrioventricular conduction capacity. Research is continued in order to clarify how to influence the occurrence of ventricular fibrillation by permanent cardiac pacing including the use of preventive algorithms. This topic has not yet been reliably and unambiguously concluded. Biventricular pacing is currently established and recognized not only for typical indications in cases of bradyarrhythmias but also to solve primary haemodynamic problems in the patients with advanced heart failure and evidence of ventricular dyssynchrony.